U.S. patent application number 14/773683 was filed with the patent office on 2016-02-18 for crystals of laquinimod sodium and improved process for the manufacture thereof.
This patent application is currently assigned to Teva Pharmaceutical Industries Ltd.. The applicant listed for this patent is Anton FEWNKEL, Ulf Tomas FRISTEDT, Vladimir IOFFE, Karl-Erik JANSSON, Avital LAZER. Invention is credited to Anton FRENKEL, Ulf Tomas FRISTEDT, Vladimir IOFFE, Karl-Erik JANSSON, Avital LAXER.
Application Number | 20160046582 14/773683 |
Document ID | / |
Family ID | 50631081 |
Filed Date | 2016-02-18 |
United States Patent
Application |
20160046582 |
Kind Code |
A1 |
FRENKEL; Anton ; et
al. |
February 18, 2016 |
CRYSTALS OF LAQUINIMOD SODIUM AND IMPROVED PROCESS FOR THE
MANUFACTURE THEREOF
Abstract
The subject invention provides a mixture of Crystalline
Laquinimod sodium particles, wherein (i) .gtoreq.90% of the total
amount by volume of the laquinimod sodium particles have a size of
.ltoreq.40 .mu.m or (ii) .gtoreq.50% of the total amount by volume
of the laquinimod sodium particles have a size of .ltoreq.15 .mu.m
and wherein: a) the mixture has a bulk density of 0.2-0.4 g/mL; b)
the mixture has a tapped density of 0.40-0.7 g/mL; c) an amount of
heavy metal in the mixture is no more than 20 ppm relative to the
amount by weight of laquinimod sodium; d) an amount of MCQ in the
mixture is no more than 0.15% relative to the amount of laquinimod
sodium as measured by HPLC; e) an amount of MCQCA in the mixture is
no more than 0.15% relative to the amount of laquinimod sodium as
measured by HPLC; or f) an amount of MCQME in the mixture is no
more than 0.12% relative to the amount of laquinimod sodium as
measured by HPLC. The subject invention also provides a
pharmaceutical composition comprising an amount of laquinimod and
at least one of BH-3-HKAQ, MCQ, MCQCA, MCQME, NEA, and MCQEE. The
subject invention also provides processes for preparing BH-3-HLAQ,
MCQ, MCQCA, MCQEE, and compounds prepared by said processes.
Further provided is a process for testing whether a sample of
laquinimod contains an undesirable impurity. Further provided is a
process for preparing a validated pharmaceutical composition
comprising laquinimod, for preparing a pharmaceutical composition
comprising laquinimod, or for distributing a validated batch of a
pharmaceutical composition comprising laquinimod, for validating a
batch of a pharmaceutical product containing laquinimod and a
pharmaceutically acceptable carrier for distribution, and for
preparing a packaged pharmaceutical composition comprising
laquinimod, each comprising determining the amount of at least one
of BH-3-HLAO, MCQ, MCQCA, MCQME<NEA, and MCQEE in a sample or
batch. The subject invention further provides use of BH-3-HLAQ,
MCQ, MCQCA, MCQME, MCQEE as a reference standard to detect trace
amounts of the impurity in a pharmaceutical composition composing
laquinimod. Finally, the subject invention provides methods of
ddenrrining the concentration of BH-3-HLAQ, MCQ, MCQCA, MCQME,
MCQE<5-HLAQ, SPIRO-LAQ or H-LAQ in a pharmaceutical composition
composing laquinimod.
Inventors: |
FRENKEL; Anton; (Netanya,
IL) ; LAXER; Avital; (Tel Aviv, IL) ; IOFFE;
Vladimir; (Kfar Saba, IL) ; JANSSON; Karl-Erik;
(Dalby, SE) ; FRISTEDT; Ulf Tomas; (Helsingborg,
SE) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
FEWNKEL; Anton
LAZER; Avital
IOFFE; Vladimir
JANSSON; Karl-Erik
FRISTEDT; Ulf Tomas |
Netanya
Tel Aviv
Kfar Saba
Dalby
Helsingborg |
|
IL
IL
IL
SE
SE |
|
|
Assignee: |
Teva Pharmaceutical Industries
Ltd.
Petach Tikva
IL
|
Family ID: |
50631081 |
Appl. No.: |
14/773683 |
Filed: |
March 14, 2014 |
PCT Filed: |
March 14, 2014 |
PCT NO: |
PCT/US2014/029292 |
371 Date: |
September 8, 2015 |
Current U.S.
Class: |
424/489 ;
428/402; 514/312; 546/155; 562/455; 73/61.52; 73/61.55 |
Current CPC
Class: |
A61K 31/47 20130101;
C07D 215/56 20130101; A61P 29/00 20180101; A61P 25/24 20180101;
G01N 33/15 20130101; C07B 2200/13 20130101; G01N 30/06 20130101;
A61P 25/00 20180101; A61P 13/12 20180101; A61P 27/02 20180101; A61P
37/06 20180101; A61P 1/04 20180101; A61P 19/02 20180101 |
International
Class: |
C07D 215/56 20060101
C07D215/56; G01N 30/06 20060101 G01N030/06; G01N 33/15 20060101
G01N033/15 |
Claims
1. A mixture of crystalline laquinimod sodium particles, wherein
(i) 90% or more of the total amount by volume of the laquinimod
sodium particles have a size of 40 microns or less or (ii) 50% or
more of the total amount by volume of the laquinimod sodium
particles have a size of 15 microns or less, and wherein: a. the
mixture has a bulk density of 0.2 g/mL to 0.4 g/mL; b. the mixture
has a tapped density of 0.40 g/mL to 0.7 g/mL; c. an amount of
heavy metal which is present in the mixture is no more than 20 ppm
of heavy metal relative to the amount by weight of laquinimod
sodium; d. an amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) which is
present in the mixture is no more than 0.15% relative to the amount
of laquinimod sodium as measured by HPLC; e. an amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) which is present in the mixture is no more than 0.15%
relative to the amount of laquinimod sodium as measured by HPLC; f.
an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyla-
te (MCQME) which is present in the mixture is no more than 0.12%
relative to the amount of laquinimod sodium as measured by HPLC; g.
an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyla-
te (MCQME) which is present in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC; h.
an amount of N-ethylaniline (NEA) which is present in the mixture
is no more than 0.10% relative to the amount of laquinimod sodium
as measured by HPLC; or i. an amount of
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide (5-HLAQ) which is present in the mixture is no more than
0.10% relative to the amount of laquinimod sodium as measured by
HPLC.
2. The mixture of claim 1, wherein (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
less than 40 microns or (ii) 50% or more of the total amount by
volume of the laquinimod sodium particles have a size of less than
15 microns.
3. The mixture of claim 1 or 2, wherein 10% or more of the total
amount by volume of the laquinimod sodium particles have a size of
5 microns or less and wherein: a. the mixture has a tapped density
of 0.40 g/mL to 0.7 g/mL; or b. an amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) which is
present in the mixture is no more than 0.15% relative to the amount
of laquinimod sodium as measured by HPLC.
4. A mixture of crystalline laquinimod sodium particles, wherein
(i) 90% or more of the total amount by volume of the laquinimod
sodium particles have a size of less than 40 microns, (ii) 50% or
more of the total amount by volume of the laquinimod sodium
particles have a size of less than 15 microns, and (iii) 10% or
more of the total amount by volume of the laquinimod sodium
particles have a size of less than 5 microns and wherein: a. the
mixture has a bulk density of 0.2 g/mL to 0.4 g/mL; b. the mixture
has a tapped density of 0.40 g/mL to 0.7 g/mL; c. an amount of
heavy metal which is present in the mixture is no more than 20 ppm
of heavy metal relative to the amount by weight of laquinimod
sodium; d. an amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) which is
present in the mixture is no more than 0.15% relative to the amount
of laquinimod sodium as measured by HPLC; e. an amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) which is present in the mixture is no more than 0.15%
relative to the amount of laquinimod sodium as measured by HPLC; f.
an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyla-
te (MCQME) which is present in the mixture is no more than 0.12%
relative to the amount of laquinimod sodium as measured by HPLC; g.
an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyla-
te (MCQME) which is present in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC; h.
an amount of N-ethylaniline (NEA) which is present in the mixture
is no more than 0.10% relative to the amount of laquinimod sodium
as measured by HPLC; or i. an amount of
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide (5-HLAQ) which is present in the mixture is no more than
0.10% relative to the amount of laquinimod sodium as measured by
HPLC.
5. The mixture of any one of claims 1-4 prepared in a single batch
comprising 2.5 kg or more of laquinimod sodium.
6. The mixture of any one of claims 1-5, wherein the laquinimod
sodium particles are determined based on an unmilled sample of the
mixture or wherein the size and amount by volume of laquinimod
sodium particles are determined based on a milled sample of the
mixture.
7. The mixture of any one of claims 1-6, having a bulk density of
0.2 g/mL to 0.4 g/mL, and/or having a tapped density of 0.40 g/mL
to 0.7 g/mL.
8. The mixture of any one of claims 1-7, wherein: a. an amount of
aluminum in the mixture is less than 5 ppm or less than 2 ppm
relative to the amount by weight of laquinimod sodium; b. an amount
of calcium in the mixture is less than 60 ppm or less than 25 ppm
relative to the amount by weight of laquinimod sodium; c. an amount
of calcium in the mixture is less than 25 ppm relative to the
amount by weight of laquinimod sodium; d. an aunt of copper in the
mixture is less than 1 ppm or less than 0.6 ppm relative to the
amount by weight of laquinimod sodium; and/or e. an amount of zinc
in the mixture is less than 7 ppm or less than 4 ppm relative to
the amount by weight of laquinimod sodium.
9. The mixture of any one of claims 1-8, wherein an amount of heavy
metal in the mixture is no more than 20 ppm relative to the amount
by weight of laquinimod sodium.
10. The mixture of any one of claims 1-9, wherein a total amount of
polar impurities in the mixture is no more than 1.00% relative to
the amount of laquinimod sodium as measured by HPLC.
11. The mixture of any one of claims 1-10, wherein: a. an amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) in the mixture
is no more than 0.15% relative to the amount of laquinimod sodium
as measured by HPLC; b. an amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the mixture is no more than 0.15% relative to the
amount of laquinimod sodium as measured by HPLC; c. an amount of
methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) in the mixture is no more than 0.12% relative to the amount
of laquinimod sodium as measured by HPLC; d. an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) which is present in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC;
and/or e. an amount of ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxy-
late (MCQEE) in the mixture is no more than 0.10% relative to the
amount of laquinimod sodium as measured by HPLC.
12. The mixture of any one of claims 1-11, wherein a total amount
of non-polar impurities in the mixture is no more than 0.50%
relative to the amount of laquinimod sodium as measured by
HPLC.
13. The mixture of any one of claims 1-12, wherein: a. an amount of
N-ethylaniline (NEA) in the mixture is no more than 0.10% relative
to the amount of laquinimod sodium as measured by HPLC; b. an
amount of
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide (5-HLAQ) in the mixture is no more than 0.10% relative to
the amount of laquinimod sodium as measured by HPLC; and/or c. an
amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxa-
mide (DELAQ) in the mixture is no more than 0.10% relative to the
amount of laquinimod sodium as measured by HPLC.
14. The mixture of any one of claims 1-13, wherein: a. an amount of
laquinimod acid in the mixture is no more than 1.00% relative to
the amount of laquinimod sodium as measured by HPLC; b. an amount
of dimethyl malonate in the mixture is no more than 0.10% relative
to the amount of laquinimod sodium as measured by HPLC; c. an
amount of diethyl malonate in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC; d.
an amount of dimethyl sulfate in the mixture is no more than 1 ppm
relative to the amount by weight of laquinimod sodium; e. an amount
of water in the mixture is no more than 1.5% by weight relative to
the amount of laquinimod sodium as measured by K.F. coulometric
titration; and/or f. an amount of sodium from 5.8% to 6.4% relative
to the amount by weight of laquinimod sodium.
15. The mixture of any one of claims 1-14, wherein: a. an amount of
ethanol in the mixture is no more than 5000 ppm relative to the
amount by weight of laquinimod sodium; b. an amount of n-heptane in
the mixture is no more than 5000 ppm relative to the amount by
weight of laquinimod sodium; c. an amount of n-octane in the
mixture is no more than 2000 ppm relative to the amount by weight
of laquinimod sodium; d. an amount of n-octane in the mixture is no
more than 200 ppm relative to the amount by weight of laquinimod
sodium; e. an amount of n-octane in the mixture is no more than 20
ppm relative to the amount by weight of laquinimod sodium; f. an
amount of methanol in the mixture is no more than 3000 ppm relative
to the amount by weight of laquinimod sodium; g. an amount of
acetone in the mixture is no more than 5000 ppm relative to the
amount by weight of laquinimod sodium; h. an amount of dioxane in
the mixture is no more than 380 ppm relative to the amount by
weight of laquinimod sodium; and/or i. an amount of dimethyl
formamide in the mixture is no more than 880 ppm relative to the
amount by weight of laquinimod sodium.
16. A pharmaceutical composition comprising the mixture of any one
of claims 1-15 and a pharmaceutically acceptable carrier.
17. The pharmaceutical composition of claim 16, wherein; a. a total
amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the pharmaceutical composition is no more than
0.50% relative to the amount of laquinimod sodium as measured by
HPLC; b. a total amount of polar impurities in the pharmaceutical
composition is no more than 2.00% relative to the amount of
laquinimod sodium as measured by HPLC; c. an amount of
N-ethylaniline (NEA) in the pharmaceutical composition is no more
than 0.50% relative to the amount of laquinimod sodium as measured
by HPLC; d. an amount of
5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide (3-HLAQ) in the pharmaceutical composition
is no more than 0.50% relative to the amount of laquinimod sodium
as measured by HPLC; e. a total amount of non-polar impurities in
the pharmaceutical composition is no more than 1.00% relative to
the amount of laquinimod sodium as measured by HPLC; and/or f. an
amount of
N-ethyl-4-hydroxy-1-methyl-5-(methyl(2,3,4,5,6-pentahydroxyhexyl)amino)-2-
-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide (MEG-LAQ) in the
pharmaceutical composition is no more than 1.00% relative to the
amount of laquinimod sodium as measured by HPLC.
18. The pharmaceutical composition of claims 16 or 17, wherein: a.
an amount of water in the pharmaceutical composition is no more
than 1.50% relative to the amount of laquinimod sodium as measured
by K.F. coulometric titration; b. an amount of water in the
pharmaceutical composition is no more than 0.80% relative to the
amount of laquinimod sodium as measured by K.F. coulometric
titration; and/or c. the pharmaceutical composition comprises an
amount of sodium from 5.8% to 6.4% relative to the amount by weight
of laquinimod sodium.
19. The pharmaceutical composition of any one of claims 16-18,
wherein: a. an amount of ethanol in the pharmaceutical composition
is no more than 5000 ppm relative to the amount by weight of
laquinimod sodium; b. an amount of n-heptane in the pharmaceutical
composition is no more than 5000 ppm relative to the amount by
weight of laquinimod sodium; c. an amount of n-octane in the
pharmaceutical composition is no more than 2000 ppm relative to the
amount by weight of laquinimod sodium; d. an amount of n-octane in
the pharmaceutical composition is no more than 200 ppm relative to
the amount by weight of laquinimod sodium; e. an amount of n-octane
in the pharmaceutical composition is no more than 20 ppm relative
to the amount by weight of laquinimod sodium; f. an amount of
methanol in the pharmaceutical composition is no more than 380 ppm
relative to the amount by weight of laquinimod sodium; g. an amount
of acetone in the pharmaceutical composition is no more than 880
ppm relative to the amount by weight of laquinimod sodium; h. an
amount of dioxane in the pharmaceutical composition is no more than
380 ppm relative to the amount by weight of laquinimod sodium;
and/or i. an amount of dimethyl formamide in the pharmaceutical
composition is no more than 880 ppm relative to the amount by
weight of laquinimod sodium.
20. A method of treating a subject afflicted with, or alleviating a
symptom of, a form of multiple sclerosis, lupus nephritis, lupus
arthritis, rheumatoid arthritis, a BDNF-related disorder, Crohn's
disease, a GABA-related disorder, a cannabinoid receptor type 1
(CB1) mediated disorder, or an ocular inflammatory disorder
comprising administering to the subject the mixture of any one of
claims 1-15 or the pharmaceutical composition of any one of claims
16-20 so as to thereby treat the subject or alleviate the symptom
in the subject.
21. The mixture of any one of claims 1-15 or the pharmaceutical
composition of any one of claims 16-19 for use in the treatment of,
or alleviation of symptoms of, a form of multiple sclerosis, lupus
nephritis, lupus arthritis, rheumatoid arthritis, a BDNF-related
disorder, Crohn's disease, a GABA-related disorder, a cannabinoid
receptor type 1 (CB1) mediated disorder, or an ocular inflammatory
disorder.
22. Use of the mixture of any one of claims 1-15 or the
pharmaceutical composition of any one of claims 16-19 for the
manufacture of a medicament for treating, or alleviating a symptom
of, a form of multiple sclerosis, a GABA-related disorder, a
cannabinoid receptor type 1 (CB1) mediated disorder, lupus
nephritis, lupus arthritis, rheumatoid arthritis, a BDNF-related
disorder, Crohn's disease, or an ocular inflammatory disorder.
23. A process of recrystallization of laquinimod sodium comprising:
a. dissolving an amount of laquinimod sodium in water to form an
aqueous solution; b. concentrating the aqueous solution to form a
concentrated solution comprising approximately 1.7-1.8 mL of water
per gram of laquinimod sodium; c. adding acetone to the
concentrated solution of step b); and d. isolating recrystallized
laquinimod sodium.
24. The process of claim 23, wherein: a. the amount of laquinimod
sodium in step a) is 2.5 kg or greater; b. step a) is performed
with 10-12 mL of water per gram of laquinimod sodium, preferably
step a) is performed with approximately 11 mL of water per gram of
laquinimod sodium; and/or c. step a) is performed by heating the
aqueous solution to a temperature of 58-75.degree. C., or wherein
step a) is performed by heating the aqueous solution to a
temperature of 60-73.degree. C.
25. The process of claims 23 or 24, wherein crystallization occurs
after step a) and before step c), preferably wherein
crystallization: a. is induced by rapid stirring during or after
the concentrating step b); b. is induced by addition of a seed
crystal during or after the concentrating step b); or c. occurs
without addition of a seed crystal.
26. The process of any one of claims 23-25, wherein: a. step b) is
performed under conditions appropriate to induce crystallization at
the concentration of 1.7-1.8 mL of water per gram of laquinimod
sodium; and/or b. step b) is performed at 28-45.degree. C. or at
30-40.degree. C.
27. The process of any one of claims 23-26, wherein: a. step c) is
performed with the concentrated solution at 40-55.degree. C. or at
45-50.degree. C.; b. step c) is performed with 6-12 mL of acetone
per gram of laquinimod sodium, preferably step c) is performed with
approximately 10 mL of acetone per gram of laquinimod sodium; c.
step c) is performed over a period of 1-4 hours or over a period of
1.2-2.5 hours; and/or d. step c) is followed by cooling the
solution to a temperature no less than -14.degree. C. and no more
than 6.degree. C. or wherein step c) is followed by cooling the
solution to a temperature no less than -4.degree. C. and no more
than 4.degree. C.
28. The process of any one of claims 23-27, wherein the solution is
cooled over a period of 3-5 hours or over a period of 3.5-4.5
hours.
29. The process of any one of claims 23-28, wherein a. step d)
further comprises washing the recrystallized laquinimod sodium with
1-4 mL of acetone per gram of crude laquinimod sodium used in step
a), preferably step d) further comprises washing the recrystallized
laquinimod sodium with approximately 3 mL of acetone per gram of
crude laquinimod sodium used in step a); b. step d) further
comprises drying the recrystallized laquinimod sodium for no less
than one hour at 30-40.degree. C. under a vacuum of no more than 50
mmHg; and/or c. the isolated recrystallized laquinimod sodium in
step d) is a mixture of crystalline laquinimod sodium particles
having a particle size distribution such that (i) 90% or more of
the total amount by volume of the laquinimod sodium particles have
a size of 40 microns or less, (ii) 50% or more of the total amount
by volume of the laquinimod sodium particles have a size of 15
microns or less, and (iii) 10% or more of the total amount by
volume of the laquinimod sodium particles have a size of less than
5 microns or less.
30. A mixture of crystalline laquinimod sodium particles prepared
by any one of claims 23-29.
31. The mixture of claim 30, wherein (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
40 microns or less, (ii) 50% or more of the total amount by volume
of the laquinimod sodium particles have a size of 15 microns or
less, and (iii) 10% or more of the total amount by volume of the
laquinimod sodium particles have a size of 5 microns or less.
32. The mixture of claim 30 or 31, wherein: a. the mixture has a
bulk density of 0.2 g/mL to 0.4 g/mL; b. the mixture has a tapped
density of 0.40 g/mL to 0.7 g/mL; c. an amount of aluminum in the
mixture is less than 5 ppm relative to the amount by weight of
laquinimod sodium; d. an amount of calcium in the mixture is less
than 60 ppm relative to the amount by weight of laquinimod sodium;
e. an amount of copper in the mixture is less than 1 ppm relative
to the amount by weight of laquinimod sodium; f. an amount of zinc
in the mixture is less than 7 ppm relative to the amount by weight
of laquinimod sodium; g. an amount of heavy metal in the mixture is
no more than 20 ppm relative to the amount by weight of laquinimod
sodium; h. a total amount of polar impurities in the mixture is no
more than 1.00% relative to the amount of laquinimod sodium as
measured by HPLC; i. an amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) in the mixture
is no more than 0.15% relative to the amount of laquinimod sodium
as measured by HPLC; j. an amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the mixture is no more than 0.15% relative to the
amount of laquinimod sodium as measured by HPLC; k. an amount of
methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) in the mixture is no more than 0.12% relative to the amount
of laquinimod sodium as measured by HPLC; l. an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) in the mixture is no more than 0.10% relative to the amount
of laquinimod sodium as measured by HPLC; m. an amount of ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the mixture is no more than 0.10% relative to the amount
of laquinimod sodium as measured by HPLC; n. a total amount of
non-polar impurities in the mixture is no more than 0.50% relative
to the amount of laquinimod sodium as measured by HPLC; o. an
amount of N-ethylaniline (NEA) in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC; p.
an amount of laquinimod acid in the mixture is no more than 1.00%
relative to the amount of laquinimod sodium as measured by HPLC; q.
an amount of dimethyl malonate in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC; r.
an amount of diethyl malonate in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC; s.
an amount of dimethyl sulfate in the mixture is no more than 1 ppm
relative to the amount by weight of laquinimod sodium; t. an amount
of water in the mixture is no more than 1.5% by weight relative to
the amount of laquinimod sodium as measured by K.F. coulometric
titration; u. the mixture comprises an amount of sodium from 5.8%
to 6.4% relative to the amount by weight of laquinimod sodium; v.
an amount of ethanol in the mixture is no more than 5000 ppm
relative to the amount by weight of laquinimod sodium; w. an amount
of n-heptane in the mixture is no more than 5000 ppm relative to
the amount by weight of laquinimod sodium; x. an amount of n-octane
in the mixture is no more than 2000 ppm relative to the amount by
weight of laquinimod sodium; y. an amount of n-octane in the
mixture is no more than 200 ppm relative to the amount by weight of
laquinimod sodium; z. an amount of n-octane in the mixture is no
more than 20 ppm relative to the amount by weight of laquinimod
sodium; aa. an amount of methanol in the mixture is no more than
3000 ppm relative to the amount by weight of laquinimod sodium; bb.
an amount of acetone in the mixture is no more than 5000 ppm
relative to the amount by weight of laquinimod sodium; cc. an
amount of dioxane in the mixture is no more than 380 ppm relative
to the amount by weight of laquinimod sodium; or dd. an amount of
dimethyl formamide in the mixture is no more than 880 ppm relative
to the amount by weight of laquinimod sodium.
33. The mixture of claim 30 or 31, wherein: a. the mixture has a
bulk density of 0.2 g/mL to 0.4 g/mL; b. the mixture has a tapped
density of 0.40 g/mL to 0.7 g/mL; c. an amount of aluminum in the
mixture is less than 5 ppm relative to the amount by weight of
laquinimod sodium; d. an amount of calcium in the mixture is less
than 60 ppm relative to the amount by weight of laquinimod sodium;
e. an amount of copper in the mixture is less than 1 ppm relative
to the amount by weight of laquinimod sodium; f. an amount of zinc
in the mixture is less than 7 ppm relative to the amount by weight
of laquinimod sodium; g. an amount of heavy metal in the mixture is
no more than 20 ppm relative to the amount by weight of laquinimod
sodium; h. a total amount of polar impurities in the mixture is no
more than 1.00% relative to the amount of laquinimod sodium as
measured by HPLC; i. an amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) in the mixture
is no more than 0.15% relative to the amount of laquinimod sodium
as measured by HPLC; j. an amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the mixture is no more than 0.15% relative to the
amount of laquinimod sodium as measured by HPLC; k. an amount of
methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) in the mixture is no more than 0.12% relative to the amount
of laquinimod sodium as measured by HPLC; l. an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) in the mixture is no more than 0.10% relative to the amount
of laquinimod sodium as measured by HPLC; m. an amount of ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the mixture is no more than 0.10% relative to the amount
of laquinimod sodium as measured by HPLC; n. a total amount of
non-polar impurities in the mixture is no more than 0.50% relative
to the amount of laquinimod sodium as measured by HPLC; o. an
amount of N-ethylaniline (NEA) in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC; p.
an amount of laquinimod acid in the mixture is no more than 1.00%
relative to the amount of laquinimod sodium as measured by HPLC; q.
an amount of dimethyl malonate in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC; r.
an amount of diethyl malonate in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC; s.
an amount of dimethyl sulfate in the mixture is no more than 1 ppm
relative to the amount by weight of laquinimod sodium; t. an amount
of water in the mixture is no more than 1.5% by weight relative to
the amount of laquinimod sodium as measured by K.F. coulometric
titration; u. the mixture comprises an amount of sodium from 5.8%
to 6.4% relative to the amount by weight of laquinimod sodium; v.
an amount of ethanol in the mixture is no more than 5000 ppm
relative to the amount by weight of laquinimod sodium; w. an amount
of n-heptane in the mixture is no more than 5000 ppm relative to
the amount by weight of laquinimod sodium; x. an amount of n-octane
in the mixture is no more than 2000 ppm, 200 ppm or 20 ppm relative
to the amount by weight of laquinimod sodium; y. an amount of
methanol in the mixture is no more than 3000 ppm relative to the
amount by weight of laquinimod sodium; z. an amount of acetone in
the mixture is no more than 5000 ppm relative to the amount by
weight of laquinimod sodium; aa. an amount of dioxane in the
mixture is no more than 380 ppm relative to the amount by weight of
laquinimod sodium; and bb. an amount of dimethyl formamide in the
mixture is no more than 880 ppm relative to the amount by weight of
laquinimod sodium.
34. A pharmaceutical composition comprising the mixture of any one
of claims 30-33 and a pharmaceutically acceptable carrier.
35. The pharmaceutical composition of claim 34, wherein: a. a total
amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the pharmaceutical composition is no more than
0.50% relative to the amount of laquinimod sodium as measured by
HPLC; b. a total amount of polar impurities in the pharmaceutical
composition is no more than 2.00% relative to the amount of
laquinimod sodium as measured by HPLC; c. an amount of
N-ethylaniline (NEA) in the pharmaceutical composition is no more
than 0.50% relative to the amount of laquinimod sodium as measured
by HPLC; d. a total amount of non-polar impurities in the
pharmaceutical composition is no more than 1.00% relative to the
amount of laquinimod sodium as measured by HPLC; e. an amount of
water in the pharmaceutical composition is no more than 0.80%
relative to the amount of laquinimod sodium as measured by K.F.
coulometric titration; f. the pharmaceutical composition comprises
an amount of sodium from 5.8% to 6.4% relative to the amount by
weight of laquinimod sodium; g. an amount of ethanol in the
pharmaceutical composition is no more than 5000 ppm relative to the
amount by weight of laquinimod sodium; h. an amount of n-heptane in
the pharmaceutical composition is no more than 5000 ppm relative to
the amount by weight of laquinimod sodium; i. an amount of n-octane
in the pharmaceutical composition is no more than 2000 ppm relative
to the amount by weight of laquinimod sodium; j. an amount of
n-octane in the pharmaceutical composition is no more than 200 ppm
relative to the amount by weight of laquinimod sodium; k. an amount
of n-octane in the pharmaceutical composition is no more than 20
ppm relative to the amount by weight of laquinimod sodium; l. an
amount of methanol in the pharmaceutical composition is no more
than 380 ppm relative to the amount by weight of laquinimod sodium;
m. an amount of acetone in the pharmaceutical composition is no
more than 880 ppm relative to the amount by weight of laquinimod
sodium; n. an amount of dioxane in the pharmaceutical composition
is no more than 380 ppm relative to the amount by weight of
laquinimod sodium; or o. an amount of dimethyl formamide in the
pharmaceutical composition is no more than 880 ppm relative to the
amount by weight of laquinimod sodium.
36. The pharmaceutical composition of claim 34, comprising: a. a
total amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ)
and
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the pharmaceutical composition is no more than
0.50% relative to the amount of laquinimod sodium as measured by
HPLC; b. a total amount of polar impurities in the pharmaceutical
composition is no more than 2.00% relative to the amount of
laquinimod sodium as measured by HPLC; c. an amount of
N-ethylaniline (NEA) in the pharmaceutical composition is no more
than 0.50% relative to the amount of laquinimod sodium as measured
by HPLC; d. a total amount of non-polar impurities in the
pharmaceutical composition is no more than 1.00% relative to the
amount of laquinimod sodium as measured by HPLC; e. an amount of
water in the pharmaceutical composition is no more than 0.80%
relative to the amount of laquinimod sodium as measured by K.F.
coulometric titration; f. the pharmaceutical composition comprises
an amount of sodium from 5.8% to 6.4% relative to the amount by
weight of laquinimod sodium; g. an amount of ethanol in the
pharmaceutical composition is no more than 5000 ppm relative to the
amount by weight of laquinimod sodium; h. an amount of n-heptane in
the pharmaceutical composition is no more than 5000 ppm relative to
the amount by weight of laquinimod sodium; i. an amount of n-octane
in the pharmaceutical composition is no more than 2000 ppm, 200 ppm
or 20 ppm relative to the amount by weight of laquinimod sodium; an
amount of methanol in the pharmaceutical composition is no more
than 380 ppm relative to the amount by weight of laquinimod sodium;
j. an amount of acetone in the pharmaceutical composition is no
more than 880 ppm relative to the amount by weight of laquinimod
sodium; k. an amount of dioxane in the pharmaceutical composition
is no more than 380 ppm relative to the amount by weight of
laquinimod sodium; and l. an amount of dimethyl formamide in the
pharmaceutical composition is no more than 880 ppm relative to the
amount by weight of laquinimod sodium.
37. A mixture of crystalline laquinimod sodium particles, wherein
(i) 90% or more of the total amount by volume of the laquinimod
sodium particles have a size of 40 microns or less or (ii) 50% or
more of the total amount by volume of the laquinimod sodium
particles have a size of 15 microns or less, and wherein a. an
amount of aluminium in the mixture is less than 5 ppm relative to
the amount by weight of laquinimod sodium; b. an amount of calcium
in the mixture is less than 60 ppm relative to the amount by weight
of laquinimod sodium; c. an amount of copper in the mixture is less
than 1 ppm relative to the amount by weight of laquinimod sodium;
or d. an amount of zinc in the mixture is less than 7 ppm relative
to the amount by weight of laquinimod sodium.
38. A pharmaceutical composition comprising the mixture of claim 37
and a pharmaceutically acceptable carrier.
39. An isolated compound having the structure: ##STR00037## or a
salt thereof.
40. A composition comprising a compound having the structure:
##STR00038## or a salt thereof, wherein the composition is free of
laquinimod or a salt thereof.
41. A pharmaceutical composition comprising an amount of laquinimod
and at least one of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE), wherein a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount not more than 10%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount not more than 10%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount not more than 10%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or d)
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 10%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or e) N-Ethylaniline
(NEA) is present in the pharmaceutical composition in an amount not
more than 10%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or f) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
not more than 10%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
42. The pharmaceutical composition of claim 41, wherein a)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount not more than 0.15%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount not more than 0.15%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or c)
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.15%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or d) N-Ethylaniline
(NEA) is present in the pharmaceutical composition in an amount not
more than 0.1%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
not more than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
43. The pharmaceutical composition of claim 42, wherein methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.12%, relative to the concentration of laquinimod,
based on a determination by an HPLC method; and/or wherein methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
44. The pharmaceutical composition of any one of claims 41-43,
wherein a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is
present in the pharmaceutical composition in an amount less than
0.05%, relative to the concentration of laquinimod, based on a
determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or c) Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
less than 0.05%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is
present in the pharmaceutical composition in an less than 0.06%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
less than 0.05%, relative to the concentration of laquinimod, based
on a determination by an HPLC method.
45. The pharmaceutical composition of claim 44, wherein
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.05%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
46. The pharmaceutical composition of claims 44 or 45, wherein a)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount less than 0.02%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.03%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or c)
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
less than 0.02%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is
present in the pharmaceutical composition in an less than 0.02%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
less than 0.02%, relative to the concentration of laquinimod, based
on a determination by an HPLC method.
47. The pharmaceutical composition of claim 46, wherein
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.02%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
48. The pharmaceutical composition of claim 42, wherein a)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount greater than 0.02%, and
not more than 0.15%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.02%, and not more than 0.15%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or c) Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
greater than 0.02%, and not more than 0.15%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical
composition in an greater than 0.02%, and not more than 0.1%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
greater than 0.02%, and not more than 0.1%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method.
49. The pharmaceutical composition of claim 48, wherein a. methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.12%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or wherein methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, and/or b.
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.03%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
50. The pharmaceutical composition of claims 48 or 49, wherein a)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount greater than 0.05%, and
not more than 0.15%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.05%, or greater than 0.10%, and not more than
0.15%, relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c) Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
greater than 0.05%, and less than 0.15%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical
composition in an greater than 0.06%, and not more than 0.10%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
greater than 0.05%, and not more than 0.10%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method.
51. The pharmaceutical composition of claim 50, wherein methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.12%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or is present in the
pharmaceutical composition in an amount not more than 0.1%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method.
52. The pharmaceutical composition of claims 48 or 49, wherein a)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount greater than 0.02%, and
less than 0.05%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.02%, or greater than 0.03%, and less than
0.1%, relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c) Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical
composition in an greater than 0.02%, and less than 0.06%, relative
to the concentration of laquinimod, based on a determination by an
HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method.
53. The pharmaceutical composition of claim 52, wherein
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.05%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
54. The pharmaceutical composition of claim 41, further comprising
at least one pharmaceutically acceptable carrier.
55. The pharmaceutical composition of claim 54, wherein a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount not more than 1.0%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or b)
N-Ethylaniline (NEA) is present in the pharmaceutical composition
in an amount not more than 0.5%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or c) the
combined amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is not more
than 0.5%, relative to the concentration of laquinimod, based on a
determination by an HPLC method.
56. The pharmaceutical composition of claim 55, wherein
N-Ethylaniline (NEA) is present in the pharmaceutical composition
in an amount not more than 0.1%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
57. The pharmaceutical composition of claims 55 or 56, wherein a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount less than 0.05%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount less than 0.05%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.5%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or d) N-Ethylaniline
(NEA) is present in the pharmaceutical composition in an amount
less than 0.1%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or e) the combined amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is less than
0.1%, relative to the concentration of laquinimod, based on a
determination by an HPLC method.
58. The pharmaceutical composition of claim 57, wherein
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
59. The pharmaceutical composition of claims 57 or 58, wherein a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methy-3-oxopropanamido)ben-
zoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount less than 0.02%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount less than 0.02%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.03%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or d)
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
less than 0.02%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or e) N-Ethylaniline (NEA) is
present in the pharmaceutical composition in an less than 0.03%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or f) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
less than 0.02%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or g) the combined amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is less than
0.03%, relative to the concentration of laquinimod, based on a
determination by an HPLC method.
60. The pharmaceutical composition of claim 59, wherein
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.02%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
61. The pharmaceutical composition of claim 54, wherein a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount greater than 0.02%, and not more than 1.0%, relative
to the concentration of laquinimod, based on a determination by an
HPLC method, or b) N-Ethylaniline (NEA) is present in the
pharmaceutical composition in an greater than 0.03%, and not more
than 0.5%, relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c) the combined amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is greater
than 0.03%, and not more than 0.5%, relative to the concentration
of laquinimod, based on a determination by an HPLC method.
62. The pharmaceutical composition of claim 61, wherein
N-Ethylaniline (NEA) is present in the pharmaceutical composition
in an amount not more than 0.1%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
63. The pharmaceutical composition of claim 54, wherein a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount greater than 0.05%, and not more than 1.0%, relative
to the concentration of laquinimod, based on a determination by an
HPLC method, or b) N-Ethylaniline (NEA) is present in the
pharmaceutical composition in an greater than 0.1%, and not more
than 0.5%, relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c) the combined amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is greater
than 0.1%, and not more than 0.5%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
64. The pharmaceutical composition of claim 54, wherein a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount greater than 0.02%, and less than 0.05%, relative to
the concentration of laquinimod, based on a determination by an
HPLC method, or b) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or c)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.02%, and less than 0.5%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or d) Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or e) N-Ethylaniline (NEA) is present in the pharmaceutical
composition in an greater than 0.03%, and less than 0.1%, relative
to the concentration of laquinimod, based on a determination by an
HPLC method, or f) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or g) the combined amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is greater
than 0.03%, and less than 0.1%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
65. The pharmaceutical composition of claim 64, wherein
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.03%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or is
present in the pharmaceutical composition in an amount less than
0.1%, relative to the concentration of laquinimod, based on a
determination by an HPLC method.
66. The pharmaceutical composition of any one of claims 41-65,
comprising laquinimod sodium salt.
67. The pharmaceutical composition of any one of claims 54-66, in a
oral unit dosage form, or in the form of a capsule, a tablet, or a
liquid suspension.
68. The pharmaceutical composition of claim 67, wherein the oral
unit dosage form comprises more than 0.3 mg laquinimod, more than
0.5 mg laquinimod or more than 0.6 mg laquinimod.
69. A process for preparing
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) comprising the steps of: a) adding sodium
hydroxide solution to a suspension of
5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide in water, b) stirring the mixture of step
a) followed by addition of hydrochloric acid solution, c)
extracting the aqueous solution with ethyl acetate, d) washing the
organic phase with brine, e) drying the organic phase over sodium
sulfate, f) filtering the suspension, g) evaporating the filtrate,
h) purifying the residue by crystallization from isopropyl alcohol,
i) cooling the suspension followed by filtering and washing with
isopropyl alcohol, and j) obtaining and drying the resulting white
solid.
70.
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamid-
o)benzoic acid (BH-3-HLAQ) prepared by the process of claim 69.
71. A process for preparing
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) comprising the
steps of: a) heating a mixture of
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) and dimethylsulfoxide, b) cooling the mixture of step
a), and c) filtering the mixture of step b) and collecting the
resulting filtrate.
72. 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) prepared by
the process of claim 71.
73. A process for preparing
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) comprising the steps of: a) heating a mixture of ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in a solution of hydrochloric acid in acetic acid, b)
cooling the mixture of step a), c) diluting the mixture of step b)
with 2-propanol and further cooling the diluted mixture, and d)
filtering off the crystals resulting from step c).
74.
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) prepared by the process of claim 73.
75. A process for preparing Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) comprising the steps of: a) forming sodium dimethylmalonate
by reaction of dimethylmalonate in dimethlformamide with sodium
methoxide solution, b) reacting the intermediate
5-chloro-1-methyl-h-benzo[D][1,3]oxazine-2,4-dione with sodium
dimethylmalonate to form methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) sodium salt, and c) acidifying methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) sodium salt to methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME).
76. Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) prepared by the process of claim 75.
77. A process for preparing Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) comprising the steps of: a) adding sodium hydride to a
solution of 5-chloro-1-methyl-1h-benzo[D][1,3]oxazine-2,4-dione and
diethyl malonate in dimethylformamide, b) heating the mixture of
step a) while stirring, c) cooling the solution of step b), d)
quenching the reaction mixture of step c), e) acidifying the
mixture of step d), f) filtering then drying the mixture of step
e), and g) crystallizing the crude product of step f) by dissolving
in ethanol following by slow cooling.
78. Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbox-
ylate (MCQEE) prepared by the process of claim 77.
79. A process for testing whether a sample of laquinimod contains
an undesirable impurity which comprises determining whether the
sample contains a compound having the structure: ##STR00039##
80. A process for preparing a validated pharmaceutical composition
comprising laquinimod comprising: a) obtaining a batch of
laquinimod; b) determining the amount of at least one of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the batch using by an HPLC method; and c) preparing the
pharmaceutical composition from the batch only if i) the batch is
determined to have not more than 0.15%
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), relative to
the concentration of laquinimod, or ii) the batch is determined to
have not more than 0.15%
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), relative to the concentration of laquinimod, or iii)
the batch is determined to have not more than 0.15%, 0.12% or 0.1%
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), relative to the concentration of laquinimod, or iv) the
batch is determined to have not more than 0.1% N-Ethylaniline
(NEA), relative to the concentration of laquinimod, or v) the batch
is determined to have not more than 0.1% Ethyl
5-chloro-4-hydroxy-1l-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE), relative to the concentration of laquinimod.
81. The process of claim 80, wherein in step c) the pharmaceutical
composition is prepared from the batch only if the batch is
determined to have not more than 0.12% Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), relative to the concentration of laquinimod, or wherein in
step c) the pharmaceutical composition is prepared from the batch
only if the batch is determined to have not more than 0.1% Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), relative to the concentration of laquinimod.
82. A process for preparing a pharmaceutical composition comprising
laquinimod, or for distributing a validated batch of a
pharmaceutical composition comprising laquinimod, comprising a)
obtaining a batch of laquinimod or of the pharmaceutical
composition; b) performing stability testing with a sample of the
batch; c) determining the total amount of at least one of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the sample of the batch after stability testing by an
HPLC method; and d) validating the batch for distribution or
preparing the pharmaceutical composition from the batch only if the
sample of the batch after stability testing contains i) not more
than a total of 0.15% relative to the concentration of laquinimod
of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), or ii) not
more than a total of 0.15% relative to the concentration of
laquinimod of
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), or iii) not more than a total of 0.15%, 0.12% or 0.1%
relative to the concentration of laquinimod of Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), or iv) not more than a total of 0.1% relative to the
concentration of laquinimod of N-Ethylaniline (NEA), or v) not more
than a total of 0.1% relative to the concentration of laquinimod of
Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE).
83. The process of claim 82, further comprising step e)
distributing the batch if in step d) the batch is validated for
distribution.
84. The process of claims 82 or 83, wherein in step d) the batch is
validated for distribution or the pharmaceutical composition is
prepared from the batch only if the sample of the batch after
stability testing contains not more than 0.12% Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) relative to the concentration of laquinimod, or wherein in
step d) the batch is validated for distribution or the
pharmaceutical composition is prepared from the batch only if the
sample of the batch after stability testing contains not more than
0.1% Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), relative to the concentration of laquinimod.
85. A process for validating a batch of a pharmaceutical product
containing laquinimod or a pharmaceutically acceptable salt thereof
and a pharmaceutically acceptable carrier for distribution
comprising a) subjecting a sample of the batch to stability
testing; b) determining the amount of at least one of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the sample of the batch after stability testing by an
HPLC method; and c) validating the batch for distribution only if
the sample of the batch after stability testing contains i) not
more than a total of 1.0% relative to the concentration of
laquinimod of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ), or ii) not more than a total of 0.5% or
0.1% relative to the concentration of laquinimod of N-Ethylaniline
(NEA), or iii) not more than a total of 0.5% relative to the
concentration of laquinimod of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) combined.
86. The process of claim 85 wherein in step c) the batch is
validated for distribution only if the sample of the batch after
stability testing not more than a total of the 0.1% of
N-Ethylaniline (NEA) relative to the concentration of
laquinimod.
87. A process for preparing a packaged pharmaceutical composition
comprising laquinimod or a pharmaceutically acceptable salt thereof
comprising: a) obtaining a batch of pharmaceutical composition
comprising laquinimod or a pharmaceutically acceptable salt
thereof; b) performing stability testing with a sample from the
batch; c) determining the amount of at least one of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the sample by an HPLC method after stability testing;
and d) packaging the pharmaceutical composition in only if i) the
content of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) in the sample is determined to be not more
than 1.0% to the concentration of laquinimod, or ii) the content of
N-Ethylaniline (NEA) in the sample is determined to be not more
than 0.5% or 0.1% to the concentration of laquinimod. iii) the
combined content of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the sample is determined to be not more than a
total of 0.5% relative to the concentration of laquinimod.
88. The process of claim 87, wherein in step d) the pharmaceutical
composition is packaged only if the content of N-Ethylaniline (NEA)
in the sample is determined to be not more than 0.1% to the
concentration of laquinimod.
89. The process of any one of claims 69-88, wherein the laquinimod
is laquinimod sodium salt.
90. An impurity or a salt thereof for use, as a reference standard
to detect trace amounts of the impurity in a pharmaceutical
composition comprising laquinimod or a pharmaceutically acceptable
salt thereof, wherein the impurity is selected from the group
consisting of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE).
91. A method of determining the concentration of an impurity in a
pharmaceutical composition comprising laquinimod, the method
comprising, a) Preparing a sample solution from the pharmaceutical
composition, b) Preparing a standard solution comprising the
impurity c) Preparing a resolution solution comprising laquinimod
and the impurity, d) Preparing a buffer solution by dissolving
ammonium acetate in water and adjusting to pH of 7.0.+-.0.05 with
aqueous ammonia or glacial acetic acid, e) Preparing a diluent
solution comprising the buffer solution and acetonitrile, f)
Preparing a blank solution comprising the diluent solution and
aqueous acetonitrile, g) Injecting into the HPLC the resolution
solution, the blank solution, the standard solution, and the sample
solution, h) Running the HPLC using ultraviolet absorption at 240
nm and the diluent solution as the mobile phase, i) Determining the
retention time (RT) and the areas of the peaks of the impurity in
the chromatograms of the sample solution, and j) Performing
quantitation of the impurity with respect to the corresponding
peaks in the chromatograms of the standard solutions, wherein the
impurity is 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2
dihydroquinoline-3-carboxamide (5-HLAQ) or Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE).
92. A method of determining the concentration of an impurity in a
pharmaceutical composition comprising laquinimod, the method
comprising, a) Preparing a sample solution from the pharmaceutical
composition, b) Preparing a standard solution comprising the
impurity c) Preparing a resolution solution comprising laquinimod
and the impurity, d) Preparing a buffer solution by dissolving
ammonium acetate in water, and adjusting to pH of 7.0.+-.0.05 with
aqueous ammonia or glacial acetic acid, e) Preparing a blank
solution comprising the buffer solution and acetonitrile, f)
Injecting into the HPLC the resolution solution, the blank
solution, the standard solution, and the sample solution, g)
Running the HPLC using ultraviolet absorption at 240 nm and a
mobile phase of a mixture of the buffer solution, and acetonitrile.
h) Determining the retention time (RT) and the areas of the peaks
of the impurity in the chromatograms of the sample solution, i)
Performing quantitation of the impurity with respect to the
corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is N-Ethylaniline (NEA),
1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3'-[1]ethylindolin-[2]-
-one](SPIRO-LAQ), or
5-Chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide (3-HLAQ).
93. A method of determining the concentration of an impurity in a
pharmaceutical composition comprising laquinimod and a
pharmaceutically acceptable carrier, the method comprising, a)
Preparing a sample solution from the pharmaceutical composition, b)
Preparing a standard solution comprising the impurity c) Preparing
a Quantitation Limit (QL) solution comprising the impurity, d)
Preparing a resolution solution comprising laquinimod and the
impurity, e) Preparing a buffer solution by dissolving ammonium
dihydrogen phosphate in water, and adjusting to pH of 7.0.+-.0.10
with aqueous ammonia or phosphoric acid, f) Preparing a blank
solution comprising the buffer solution and acetonitrile, g)
Injecting into the HPLC the resolution solution, the blank
solution, the QL solution, the standard solution and the sample
solution, h) Running the HPLC using a ultraviolet absorption at 212
nm and a mobile phase of a mixture of the buffer solution,
acetonitrile, and methanol, i) Determining the retention time (RT)
and the areas of the peaks of the impurity in the chromatograms of
the sample solution, and j) Performing quantitation of the impurity
with respect to the corresponding peaks in the chromatograms of the
standard solutions, wherein the impurity is
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) or
N-ethyl-4-hydroxy-1-methyl-5-(2,3,4,5,6-pentahydroxyhexylamino)-2-oxo-N-p-
henyl-1,2-dihydroquinoline-3-carboxamide (MEG-LAQ).
94. A method of determining the concentration of an impurity in a
pharmaceutical composition comprising laquinimod and a
pharmaceutically acceptable carrier, the method comprising, a)
Preparing a sample solution from the pharmaceutical composition, b)
Preparing a standard solution comprising the impurity c) Preparing
a resolution solution comprising laquinimod and the impurity, d)
Preparing a buffer solution by dissolving ammonium acetate in
water, and adjusting to PH or 7.0.+-.0.05 with aqueous ammonia or
glacial acetic acid, e) Preparing a blank solution comprising the
buffer solution and acetonitrile, f) Injecting into the HPLC the
resolution solution, the blank solution, the standard solution and
the sample solution, g) Running the HPLC using a ultraviolet
absorption at 242 nm and the blank solution as the mobile phase, h)
Determining the retention time (RT) and the areas of the peaks of
the impurity in the chromatograms of the sample solution, and i)
Performing quantitation of the impurity with respect to the
corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) or N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2
dihydroquinoline-3-carboxamide (5-HLAQ).
95. A method of determining the concentration of an impurity in a
pharmaceutical composition comprising laquinimod and a
pharmaceutically acceptable carrier, the method comprising, a)
Preparing a sample solution from the pharmaceutical composition, b)
Preparing a standard solution comprising the impurity c) Preparing
a resolution solution comprising laquinimod and the impurity, d)
Preparing a blank solution comprising methanol and acetonitrile, e)
Preparing a buffer solution by dissolving ammonium acetate in
water, and adjusting to pH of 7.0.+-.0.05 with aqueous ammonia or
glacial acetic acid, f) Injecting into the HPLC the resolution
solution, the blank solution, and the sample solution, g) Running
the HPLC using a ultraviolet absorption at 240 nm and a mobile
phase comprising acetonitrile and the buffer solution, h)
Determining the retention time (RT) and the areas of the peaks of
the impurity in the chromatograms of the sample solution, and i)
Performing quantitation of the impurity with respect to the
corresponding peaks in the chromatograms of the standard solutions,
wherein the impurity is N-Ethylaniline (NEA),
5-Chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide (3-HLAQ) or
1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3'-[1]ethylindolin-[2]-
-one](SPIRO-LAQ).
Description
[0001] This application claims priority of U.S. Provisional
Application No. 61/785,575, filed Mar. 14, 2013, the entire content
of which is hereby incorporated by reference herein.
[0002] Throughout this application, various publications are
referred to by first author and year of publication. Full citations
for these publications are presented in a References section
immediately before the claims. Disclosures of the publications
cited in the References section in their entireties are hereby
incorporated by reference into this application in order to more
fully describe the state of the art as of the date of the invention
described herein.
BACKGROUND OF THE INVENTION
[0003] Laquinimod is a compound which has been shown to be
effective in the acute experimental autoimmune encephalomyelitis
(aEAE) model (U.S. Pat. No. 6,077,851). Its chemical name is
N-ethyl-N-phenyl-1,2,-dihydro-4-hydroxy-5-chloro-1-methyl-2-oxoquinoline--
3-carboxamide, and its Chemical Registry number is 248281-84-7.
[0004] Laquinimod is a small molecule having the following chemical
structure:
##STR00001##
[0005] Laquinimod sodium has high oral bioavailability and has been
suggested as an oral formulation for the treatment of Multiple
Sclerosis (MS). (Polman, C. et al., (2005) "Treatment with
laquinimod reduces development of active MRI lesions in relapsing
MS", Neurology. 64:987-991; Sandberg-Wollheim M, et al. (2005)
"48-week open safety study with high-dose oral laquinimod in
patients", Mult Scler. 11:S154). Studies have also shown that
laquinimod can reduce development of active MRI lesions in
relapsing MS. (Polman, C. et al., (2005) "Treatment with laquinimod
reduces development of active MRI lesions in relapsing MS",
Neurology. 64:987-991).
[0006] In order to prepare laquinimod as a pharmaceutical drug
product, manufacturing processes need to take into consideration
the possibility of the impurities disclosed herein being present in
the product. Early syntheses of laquinimod produced product
contaminated with impurities such as methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
and 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline (K.
Jansson et al., (2006) "Synthesis and Reactivity of Laquinimod, a
Quinoline-3-carboxamide: Intramolecular Transfer of the Enol Proton
to a Nitrogen Atom as a Plausible Mechanism for Ketene Formation,
J. Org. Chem., 71, 1658-1667). Several other impurities have also
been disclosed (U.S. Pat. No. 6,077,851; U.S. Pat. No. 6,875,869;
and U.S. Pat. No. 7,884,208). U.S. Patent Application Publication
No. 2012/0010239, the contents of which are incorporated by
reference into this application, discloses the impurity
5-chloro-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxa-
mide. U.S. Patent Application Publication No. 2013/0217724, the
contents of which are incorporated by reference into this
application, discloses the impurity
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide. U.S. Patent Application Publication No. 2013/00345256, the
contents of which are incorporated by reference into this
application, discloses the impurity
N-ethyl-4-hydroxy-1-methyl-5-(methyl(2,3,4,5,6-pentahydroxyhexyl)amino)-2-
-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide. U.S. Pat. No.
8,178,127, the contents of which are incorporated by reference into
this application, discloses the impurities
2-chloro-6-(1-ethyl-N-methyl-2-oxoindoline-3-carboxamido)benzoic
acid,
1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3'-[1]ethylindolin-[2]-
-one and
5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl,1,2,3,4-te-
trahydroquinoline-3-carboxamide.
[0007] The processes of synthesis of laquinimod and the preparation
of its sodium salt are disclosed in U.S. Pat. No. 6,077,851. An
additional process of synthesis of laquinimod acid, but not
laquinimod sodium, having low levels of certain impurities is
disclosed in U.S. Pat. No. 6,875,869. U.S. Pat. No. 7,884,208
teaches a process for the preparation of laquinimod sodium which
removes certain impurities present after the salt formation step,
thus resulting in a crystalline mixture of higher purity as well as
a crystalline mixture having large particles, and improved tapped
and bulk density. Pharmaceutical compositions comprising laquinimod
sodium are disclosed in PCT International Application Publication
No. WO 2005/074899.
[0008] In the preparation of laquinimod sodium disclosed in U.S.
Pat. No. 6,077,851, laquinimod acid was so suspended in ethanol,
and 5M sodium hydroxide solution was added. After stirring, the
resulting precipitate was filtered, washed with ethanol, and dried.
The method used to make laquinimod sodium in U.S. Pat. No.
6,077,851 is commonly referred to as a slurry-to-slurry salt
formation.
[0009] In the slurry-to-slurry salt formation method of U.S. Pat.
No. 6,077,851, the laquinimod sodium is not dissolved in solution.
Any solid impurities, if present in the laquinimod sodium
suspension, are therefore not removed by filtration.
[0010] U.S. Pat. No. 6,875,869 discloses a process of preparing the
base compound laquinimod in high yield and low level of impurities.
However, the process in U.S. Pat. No. 6,875,869 is only for
synthesis of the base compound (laquinimod acid) and not the salt.
As such, the slurry-to-slurry salt formation process would still be
needed to form the sodium salt.
[0011] U.S. Pat. No. 7,884,208 teaches an improved process for
preparing laquinimod sodium resulting in crystals of higher purity
as well as crystals having improved crystalline characteristics,
e.g., comprising no more than 2 ppm of a heavy metal and having
higher tapped density. In the processes disclosed in Examples 13-17
of U.S. Pat. No. 7,884,208, laquinimod sodium is dissolved in water
to form an aqueous solution; the aqueous solution is concentrated;
and then a water-miscible anti-solvent is added to the concentrated
solution to form laquinimod sodium crystals. The process of U.S.
Pat. No. 7,884,208 removes the impurities after salt formation,
thus resulting in laquinimod sodium of higher purity than the
laquinimod sodium produced directly from the "slurry to slurry"
process of U.S. Pat. No. 6,077,851.
SUMMARY OF THE INVENTION
[0012] The subject invention provides a mixture of crystalline
laquinimod sodium particles, wherein (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
40 microns or less or (ii) 50% or more of the total amount by
volume of the laquinimod sodium particles have a size of 15 microns
or less, and wherein: [0013] a) the mixture has a bulk density of
0.2 g/mL to 0.4 g/mL; [0014] b) the mixture has a tapped density of
0.40 g/mL to 0.7 g/mL; [0015] c) an amount of heavy metal which is
present in the mixture is no more than 20 ppm of heavy metal
relative to the amount by weight of laquinimod sodium; [0016] d) an
amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) which
is present in the mixture is no more than 0.15% relative to the
amount of laquinimod sodium as measured by HPLC; [0017] e) an
amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the mixture is no more than 0.15% relative to the
amount of laquinimod sodium as measured by HPLC; [0018] f) an
amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) which is present in the mixture is no more than 0.12%
relative to the amount of laquinimod sodium as measured by HPLC,
[0019] g) an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbox-
ylate (MCQME) which is present in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC;
[0020] h) an amount of N-ethylaniline (NEA) which is present in the
mixture is no more than 0.10% relative to the amount of laquinimod
sodium as measured by HPLC; or [0021] i) an amount of
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide (5-HLAQ) which is present in the mixture is no more than
0.10% relative to the amount of laquinimod sodium as measured by
HPLC.
[0022] The subject invention provides a mixture of crystalline
laquinimod sodium particles, wherein (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
less than 40 microns, (ii) 50% or more of the total amount by
volume of the laquinimod sodium particles have a size of less than
15 microns, and (iii) 10% or more of the total amount by volume of
the laquinimod sodium panicles have a size of less than 5 microns
and wherein: [0023] a) the mixture has a bulk density of 0.2 g/mL
to 0.4 g/mL; [0024] b) the mixture has a tapped density of 0.40
g/mL to 0.7 g/mL; [0025] c) an amount of heavy metal which is
present in the mixture is no more than 20 ppm of heavy metal
relative to the amount by weight of laquinimod sodium; [0026] d) an
amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) which
is present in the mixture is no more than 0.15% relative to the
amount of laquinimod sodium as measured by HPLC; [0027] e) an
amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) which is present in the mixture is no more than 0.15%
relative to the amount of laquinimod sodium as measured by HPLC;
[0028] f) an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbox-
ylate (MCQME) which is present in the mixture is no more than 0.12%
relative to the amount of laquinimod sodium as measured by HPLC;
[0029] g) an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) which is present in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC;
[0030] h) an amount of N-ethylaniline (NEA) which is present in the
mixture is no more than 0.10% relative to the amount of laquinimod
sodium as measured by HPLC; or [0031] i) an amount of
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide (5-HLAQ) which is present in the mixture is no more than
0.10% relative to the amount of laquinimod sodium as measured by
HPLC.
[0032] The subject invention provides a pharmaceutical composition
comprising the mixture as described herein and a pharmaceutically
acceptable carrier.
[0033] The subject invention provides a process of
recrystallization of laquinimod sodium comprising: [0034] a)
dissolving an amount of laquinimod sodium in water to form an
aqueous solution [0035] b) concentrating the aqueous solution to
form a concentrated solution comprising approximately 1.7-1.8 mL of
water per gram of laquinimod sodium; [0036] c) adding acetone to
the concentrated solution of step b); and [0037] d) isolating
recrystallized laquinimod sodium.
[0038] The subject invention also provides for a mixture of
crystalline laquinimod sodium particles prepared by the process as
described herein, and a pharmaceutical composition comprising said
mixture.
[0039] The subject invention provides a mixture of crystalline
laquinimod sodium particles, wherein (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
40 microns or less or (ii) 50% or more of the total amount by
volume of the laquinimod sodium particles have size of 15 microns
or less, and wherein [0040] a) an amount of aluminium in the
mixture is less than 5 ppm relative to the amount by weight of
laquinimod sodium; [0041] b) an amount of calcium in the mixture is
less than 60 ppm relative to the amount by weight of laquinimod
sodium; [0042] c) an amount of copper in the mixture is less 1 ppm
relative to the amount by weight of laquinimod sodium; or [0043] d)
an amount of zinc in the mixture is less than 7 ppm relative to the
amount by weight of laquinimod sodium.
[0044] The subject invention also provides a pharmaceutical
composition comprising the mixture as described herein and a
pharmaceutically acceptable carrier.
[0045] The subject invention also provides an isolated compound
having the structure:
##STR00002##
or a salt thereof.
[0046] The subject invention also provides an composition
comprising a compound having the structure:
##STR00003## [0047] or a salt thereof, wherein the composition is
free of laquinimod or a salt thereof.
[0048] The subject invention also provides a pharmaceutical
composition comprising an amount of laquinimod and at least one of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE), wherein a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount not more than 10%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount not more than 10%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount not more than 10%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or d)
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 10%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or e) N-Ethylaniline
(NEA) is present in the pharmaceutical composition in an amount not
more than 10%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or f) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
not more than 10%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
[0049] The subject invention also provides a process for preparing
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) comprising the steps of: a) adding sodium
hydroxide solution to a suspension of
5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide in water, b) stirring the mixture of step
a) followed by addition of hydrochloric acid solution, c)
extracting the aqueous solution with ethyl acetate, d) washing the
organic phase with brine, e) drying the organic phase over sodium
sulfate, f) filtering the suspension, g) evaporating the filtrate,
h) purifying the residue by crystallization from isopropyl alcohol,
i) cooling the suspension followed by filtering and washing with
isopropyl alcohol, and j) obtaining and drying the resulting white
solid.
[0050] The subject invention also provides
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-1-3-HLAQ) prepared by the process described
above.
[0051] The subject invention also provides a process for preparing
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) comprising the
steps of: a) heating a mixture of
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) and dimethylsulfoxide, b) cooling the mixture of step
a), and c) filtering the mixture of step b) and collecting the
resulting filtrate.
[0052] The subject invention also provides
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) prepared by the
process described above.
[0053] The subject invention also provides a process for preparing
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) comprising the steps of: a) heating a mixture of ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in a solution of hydrochloric acid in acetic acid, b)
cooling the mixture of step a), c) diluting the mixture of step b)
with 2-propanol and further cooling the diluted mixture, and d)
filtering off the crystals resulting from step c).
[0054] The subject invention also provides
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) prepared by the process described above.
[0055] The subject invention also provides a process for preparing
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) comprising the steps of: a) forming sodium dimethylmalonate
by reaction of dimethylmalonate in dimethlformamide with sodium
methoxide solution, b) reacting the intermediate
5-chloro-1-methyl-1h-benzo[D][1,3]oxazine-2,4-dione with sodium
dimethylmalonate to form methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) sodium salt, and c) acidifying methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) sodium salt to methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME).
[0056] The subject invention also provides Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) prepared by the process described above.
[0057] The subject invention also provides a process for preparing
Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) comprising the steps of: a) adding sodium hydride to a
solution of 5-chloro-1-methyl-1h-benzo[D][1,3]oxazine-2,4-dione and
diethyl malonate in dimethylformamide, b) heating the mixture of
step a) while stirring, c) cooling the solution of step b), d)
quenching the reaction mixture of step c), e) acidifying the
mixture of step d), f) filtering then drying the mixture of step
e), and g) crystallizing the crude product of step f) by dissolving
in ethanol following by slow cooling.
[0058] The subject invention also provides Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) prepared by the process described above.
[0059] The subject invention also provides a process for testing
whether a sample of laquinimod contains an undesirable impurity
which comprises determining whether the sample contains a compound
having the structure:
##STR00004##
[0060] The subject invention also provides a process for preparing
a validated pharmaceutical composition comprising laquinimod
comprising: a) obtaining a batch of laquinimod; b) determining the
amount of at least one of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the batch using by an HPLC method; and c) preparing the
pharmaceutical composition from the batch only if [0061] i) the
batch is determined to have not more than 0.15%
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), relative to
the concentration of laquinimod, or [0062] ii) the batch is
determined to have not more than 0.15%
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), relative to the concentration of laquinimod, or
[0063] iii) the batch is determined to have not more than 0.15%,
0.12% or 0.1% Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), relative to the concentration of laquinimod, or [0064] iv)
the batch is determined to have not more than 0.1% N-Ethylaniline
(NEA), relative to the concentration of laquinimod, or [0065] v)
the batch is determined to have not more than 0.1% Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE), relative to the concentration of laquinimod.
[0066] The subject invention also provides a process for preparing
a pharmaceutical composition comprising laquinimod, or for
distributing a validated batch of a pharmaceutical composition
comprising laquinimod, comprising a) obtaining a batch of
laquinimod or of the pharmaceutical composition; b) performing
stability testing with a sample of the batch; c) determining the
total amount of at least one of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the sample of the batch after stability testing by an
HPLC method; and d) validating the batch for distribution or
preparing the pharmaceutical composition from the batch only if the
sample of the batch after stability testing contains [0067] i) not
more than a total of 0.15% relative to the concentration of
laquinimod of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
or [0068] ii) not more than a total of 0.15% relative to the
concentration of laquinimod of
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), or [0069] iii) not more than a total of 0.15%, 0.12%
or 0.1% relative to the concentration of laquinimod of Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), or [0070] iv) not more than a total of 0.1% relative to
the concentration of laquinimod of N-Ethylaniline (NEA), or [0071]
v) not more than a total of 0.1% relative to the concentration of
laquinimod of Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylat-
e (MCQEE).
[0072] The subject invention also provides a process for validating
a batch of a pharmaceutical product containing laquinimod or a
pharmaceutically acceptable salt thereof and a pharmaceutically
acceptable carrier for distribution comprising a) subjecting a
sample of the batch to stability testing; b) determining the amount
of at least one of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido-
)benzoic acid (BH-3-1-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the sample of the batch after stability testing by an
HPLC method; and c) validating the batch for distribution only if
the sample of the batch after stability testing contains [0073] i)
not more than a total of 1.0% relative to the concentration of
laquinimod of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ), or [0074] ii) not more than a total of 0.5%
or 0.1% relative to the concentration of laquinimod of
N-Ethylaniline (NEA), or [0075] iii) not more than a total of 0.5%
relative to the concentration of laquinimod of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) combined.
[0076] The subject invention also provides a process for preparing
a packaged pharmaceutical composition comprising laquinimod or a
pharmaceutically acceptable salt thereof comprising: a) obtaining a
batch of pharmaceutical composition comprising laquinimod or a
pharmaceutically acceptable salt thereof; b) performing stability
testing with a sample from the batch; c) determining the amount of
at least one of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the sample by an HPLC method after stability testing;
and d) packaging the pharmaceutical composition in only if [0077]
i) the content of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido-
)benzoic acid (BH-3-HLAQ) in the sample is determined to be not
more than 1.0% to the concentration of laquinimod, or [0078] ii)
the content of N-Ethylaniline (NEA) in the sample is determined to
be not more than 0.5% or 0.1% to the concentration of laquinimod.
[0079] iii) the combined content of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the sample is determined to be not more than a
total of 0.5% relative to the concentration of laquinimod.
[0080] The subject invention also provides an impurity or a salt
thereof for use, as a reference standard to detect trace amounts of
the impurity in a pharmaceutical composition comprising laquinimod
or a pharmaceutically acceptable salt thereof, wherein the impurity
is selected from the group consisting of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE).
[0081] The subject invention also provides a method of determining
the concentration of an impurity in a pharmaceutical composition
comprising laquinimod, the method comprising, a) Preparing a sample
solution from the pharmaceutical composition, b) Preparing a
standard solution comprising the impurity, c) Preparing a
resolution solution comprising laquinimod and the impurity, d)
Preparing a buffer solution by dissolving ammonium acetate in water
and adjusting to pH of 7.0.+-.0.05 with aqueous ammonia or glacial
acetic acid, e) Preparing a diluent solution comprising the buffer
solution and acetonitrile, f) Preparing a blank solution comprising
the diluent solution and aqueous acetonitrile, g) Injecting into
the HPLC the resolution solution, the blank solution, the standard
solution, and the sample solution, h) Running the HPLC using
ultraviolet absorption at 240 nm and the diluent solution as the
mobile phase, i) Determining the retention time (RT) and the areas
of the peaks of the impurity in the chromatograms of the sample
solution, and j) Performing quantitation of the impurity with
respect to the corresponding peaks in the chromatograms of the
standard solutions,
wherein the impurity is
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2
dihydroquinoline-3-carboxamide (5-HLAQ) or Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE).
[0082] The subject invention also provides a method of determining
the concentration of an impurity in a pharmaceutical composition
comprising laquinimod, the method comprising, a) Preparing a sample
solution from the pharmaceutical composition, b) Preparing a
standard solution comprising the impurity, c) Preparing a
resolution solution comprising laquinimod and the impurity, d)
Preparing a buffer solution by dissolving ammonium acetate in
water, and adjusting to pH of 7.0.+-.0.05 with aqueous ammonia or
glacial acetic acid, e) Preparing a blank solution comprising the
buffer solution and acetonitrile, f) Injecting into the HPLC the
resolution solution, the blank solution, the standard solution, and
the sample solution, g) Running the HPLC using ultraviolet
absorption at 240 nm and a mobile phase of a mixture of the buffer
solution, and acetonitrile, h) Determining the retention time (RT)
and the areas of the peaks of the impurity in the chromatograms of
the sample solution, and i) Performing quantitation of the impurity
with respect to the corresponding peaks in the chromatograms of the
standard solutions,
wherein the impurity is N-Ethylaniline (NEA),
1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3'-[1]ethylindolin-[2]-
-one] (SPIRO-LAQ), or
5-Chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide (3-HLAQ).
[0083] The subject invention also provides a method of determining
the concentration of an impurity in a pharmaceutical composition
comprising laquinimod and a pharmaceutically acceptable carrier,
the method comprising, a) Preparing a sample solution from the
pharmaceutical composition, b) Preparing a standard solution
comprising the impurity, c) Preparing a Quantitation Limit (QL)
solution comprising the impurity, d) Preparing a resolution
solution comprising laquinimod and the impurity, e) Preparing a
buffer solution by dissolving ammonium dihydrogen phosphate in
water, and adjusting to pH of 7.0.+-.0.10 with aqueous ammonia or
phosphoric acid, f) Preparing a blank solution comprising the
buffer solution and acetonitrile, g) Injecting into the HPLC the
resolution solution, the blank solution, the QL solution, the
standard solution and the sample solution, h) Running the HPLC
using a ultraviolet absorption at 212 nm and a mobile phase of a
mixture of the buffer solution, acetonitrile, and methanol, i)
Determining the retention time (RT) and the areas of the peaks of
the impurity in the chromatograms of the sample solution, and j)
Performing quantitation of the impurity with respect to the
corresponding peaks in the chromatograms of the standard
solutions,
wherein the impurity is
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) or
N-ethyl-4-hydroxy-1-methyl-5-(2,3,4,5,6-pentahydroxyhexylamino)-2-oxo-N-p-
henyl-1,2-dihydroquinoline-3-carboxamide (MEG-LAQ).
[0084] The subject invention also provides a method of determining
the concentration of an impurity in a pharmaceutical composition
comprising laquinimod and a pharmaceutically acceptable carrier,
the method comprising, a) Preparing a sample solution from the
pharmaceutical composition, b) Preparing a standard solution
comprising the impurity, c) Preparing a resolution solution
comprising laquinimod and the impurity, d) Preparing a buffer
solution by dissolving ammonium acetate in water, and adjusting to
PH or 7.0.+-.0.05 with aqueous ammonia or glacial acetic acid, e)
Preparing a blank solution comprising the buffer solution and
acetonitrile, f) Injecting into the HPLC the resolution solution,
the blank solution, the standard solution and the sample solution,
g) Running the HPLC using a ultraviolet absorption at 242 nm and
the blank solution as the mobile phase, h) Determining the
retention time (RT) and the areas of the peaks of the impurity in
the chromatograms of the sample solution, and i) Performing
quantitation of the impurity with respect to the corresponding
peaks in the chromatograms of the standard solutions,
wherein the impurity is
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) or N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2
dihydroquinoline-3-carboxamide (5-HLAQ).
[0085] The subject invention also provides a method of determining
the concentration of an impurity in a pharmaceutical composition
comprising laquinimod and a pharmaceutically acceptable carrier,
the method comprising, a) Preparing a sample solution from the
pharmaceutical composition, b) Preparing a standard solution
comprising the impurity, c) Preparing a resolution solution
comprising laquinimod and the impurity, d) Preparing a blank
solution comprising methanol and acetonitrile, e) Preparing a
buffer solution by dissolving ammonium acetate in water, and
adjusting to pH of 7.0.+-.0.05 with aqueous ammonia or glacial
acetic acid, f) Injecting into the HPLC the resolution solution,
the blank solution, and the sample solution, g) Running the HPLC
using a ultraviolet absorption at 240 nm and a mobile phase
comprising acetonitrile and the buffer solution, h) Determining the
retention time (RT) and the areas of the peaks of the impurity in
the chromatograms of the sample solution, and i) Performing
quantitation of the impurity with respect to the corresponding
peaks in the chromatograms of the standard solutions,
wherein the impurity is N-Ethylaniline (NEA),
5-Chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide (3-HLAQ) or
1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3'-[1]ethylindolin-[2]-
-one] (SPIRO-LAQ).
BRIEF DESCRIPTION OF FIGURES
[0086] FIG. 1: FIG. 1 from U.S. Pat. No. 7,884,208.
[0087] FIG. 2: FIG. 2 from U.S. Pat. No. 7,884,208.
[0088] FIG. 3: HPLC Data--Example chromatogram of mixture of
laquinimod sodium.
[0089] FIG. 4: HPLC Data--Pure chromatogram of Laquinimod
Sodium.
[0090] FIG. 5: Microscopic photograph of typical batch of crude
Laquinimod Sodium at a first magnification level.
[0091] FIG. 6: Microscopic photograph of typical batch of crude
Laquinimod Sodium at a second magnification level.
[0092] FIG. 7: Microscopic photograph of Batch C at a first
magnification level.
[0093] FIG. 8: Microscopic photograph of Batch Cat a second
magnification level.
[0094] FIG. 9: 1H-NMR Spectrum of MCQ in DMSO.
[0095] FIG. 10: 13C-NMR Spectrum of MCQ in DMSO.
[0096] FIG. 11: Mass Spectrum of MCQ (ES+ mode).
[0097] FIG. 12: Fourier Transform-Infrared Spectrum of MCQ in
KBr.
[0098] FIG. 13: 1H-NMR Spectrum of MCQCA in D.sub.2O+KOH.
[0099] FIG. 14: 13C-NMR Spectrum of MCQCA in D.sub.2O+KOH.
[0100] FIG. 15: Mass Spectrum of MCQCA (ES+ mode).
[0101] FIG. 16: Fourier Transform Infrared Spectrum of MCQCA.
[0102] FIG. 17: 1H-NMR Spectrum of MCQEE in CDCl.sub.3.
[0103] FIG. 18: 13C-NMR Spectrum of MCQEE in CDCl.sub.3.
[0104] FIG. 19: Mass Spectrum of MCQEE (ES.sup.+ mode).
[0105] FIG. 20: Fourier Transform Infrared Spectrum of MCQEE.
[0106] FIG. 21: 1H-NMR Spectrum of 5-HLAQ in DMSO.
[0107] FIG. 22: 13C-NMR Spectrum of 5-HLAQ in DMSO.
[0108] FIG. 23: Mass Spectrum of 5-HLAQ (ES+ mode).
[0109] FIG. 24: Fourier Transform Infrared Spectrum of 5-HLAQ.
[0110] FIG. 25: 1H-NMR Spectrum of DELAQ in CDCl.sub.3.
[0111] FIG. 26: 13C-NMR Spectrum of DELAQ in CDCl.sub.3.
[0112] FIG. 27: Mass Spectrum of DELAQ (ES+ mode).
[0113] FIG. 28: Fourier Transform Infrared Spectrum of DELAQ in
KBr.
[0114] FIG. 29: 1H-NMR Spectrum of 3-HLAQ in CDCl.sub.3.
[0115] FIG. 30: 13C-NMR Spectrum of 3-HLAQ in CDCl.sub.3.
[0116] FIG. 31: Mass Spectrum of 3-HLAQ (ES+ mode).
[0117] FIG. 32: Fourier Transform Infrared Spectrum of 3-HLAQ.
[0118] FIG. 33: 1H-NMR Spectrum of SPIRO-LAQ in DMSO.
[0119] FIG. 34: 13C-NMR Spectrum of SPIRO-LAQ in DMSO.
[0120] FIG. 35: Mass Spectrum of SPIRO-LAQ (ES+ mode).
[0121] FIG. 36: Fourier Transform Infrared Spectrum of
SPIRO-LAQ.
[0122] FIG. 37: 1H-NMR of BH-3-HLAQ in DMSO.
[0123] FIG. 38: 13C-NMR of BH-3-HLAQ in DMSO.
[0124] FIG. 39: Mass Spectrum of BH-3-HLAQ (ES+ mode).
[0125] FIG. 40: FT-IR Spectrum of BH-3-HLAQ.
[0126] FIG. 41: Resolution test chromatogram (Laquinimod Sodium
Before Milling (Cryst) and Drug Substance)--Identification By
Retention Time And Determination Of Assay And Polar
Impurities/Degradation Products (Polar IDD) By HPLC.
[0127] FIG. 42: Diluent chromatogram (Laquinimod Sodium Before
Milling (Cryst) and Drug Substance)--Identification By Retention
Time And Determination Of Assay And Polar Impurities/Degradation
Products (Polar IDD) By HPLC.
[0128] FIG. 43: Typical chromatogram for assay (Laquinimod Sodium
Before Milling (Cryst) and Drug Substance)--Identification By
Retention Time And Determination Of Assay And Polar
Impurities/Degradation Products (Polar IDD) By HPLC.
[0129] FIG. 44: Typical chromatogram for Impurities testing
(Laquinimod Sodium Before Milling (Cryst) and Drug
Substance)--Identification By Retention Time And Determination Of
Assay And Polar Impurities/Degradation Products (Polar IDD) By
HPLC.
[0130] FIG. 45: Resolution test chromatogram (Laquinimod Sodium
Before Milling (Cryst) and Drug Substance)--Determination Of
N-Ethylaniline And Nonpolar Impurities (Non-Polar IDD) By HPLC.
[0131] FIG. 46: Diluent chromatogram (Laquinimod sodium before
milling (Cryst) and Drug Substance) Determination Of N-Ethylaniline
And Nonpolar Impurities (Non-Polar IDD) By HPLC.
[0132] FIG. 47: Typical sample chromatogram (Laquinimod sodium
before milling (Cryst) and Drug Substance)--Determination Of
N-Ethylaniline And Nonpolar Impurities (Non-Polar IDD) by HPLC.
[0133] FIG. 48: Resolution Solution 1 (Resolution Test)
Chromatogram (Laquinimod Capsules) Identification And Determination
Of Assay And Polar Impurities/Degradation Products By HPLC.
[0134] FIG. 49: Resolution Solution 2 Chromatogram (Laquinimod
Capsules) Identification And Determination Of Assay And Polar
Impurities/Degradation Products By HPLC.
[0135] FIG. 50: Diluent (Blank) chromatogram (Laquinimod Capsules)
Identification and Determination of Assay and Polar
Impurities/Degradation Products By HPLC.
[0136] FIG. 51: Sample chromatogram for Assay (Laquinimod Capsules)
Identification and Determination of Assay and Polar
Impurities/Degradation Products By HPLC.
[0137] FIG. 52: Sample chromatogram for determination of
Impurities/Degradation products (Laquinimod Capsules)
Identification and Determination of Assay and Polar
Impurities/Degradation Products by HPLC.
[0138] FIG. 53: Resolution test chromatogram (Laquinimod Capsules)
Determination of Non-Polar Impurities/Degradation Products.
[0139] FIG. 54: Blank (Diluent) chromatogram (Laquinimod Capsules)
Determination of Non-Polar Impurities/Degradation Products.
[0140] FIG. 55: Sample chromatogram (Laquinimod Capsules)
Determination of Non-Polar Impurities/Degradation Products.
[0141] FIG. 56: Typical Chromatogram of Resolution Solution
(Laquinimod Capsules)--Determination of MEG-LAQ and BH-3-HLAQ by
HPLC.
[0142] FIG. 57: Blank (Diluent 2) chromatogram (Laquinimod
Capsules) Determination of MEG-LAQ and BH-3-HLAQ by HPLC.
[0143] FIG. 58: Chromatogram of QL Solution (Laquinimod
Capsules)--Determination of MEG-LAQ and BH-3-HLAQ by HPLC.
[0144] FIG. 59: Chromatogram of Standard Solution (Laquinimod
Capsules)--Determination of MEG-LAQ and BH-3-HLAQ by HPLC.
[0145] FIG. 60: Typical Sample Chromatogram (Laquinimod
Capsules)--Determination of MEG-LAQ and BH-3-HLAQ by HPLC.
DETAILED DESCRIPTION OF THE INVENTION
[0146] Laquinimod is a small molecule having the following chemical
structure:
##STR00005##
Laquinimod
[0147] It is an oral immunomodulator which has demonstrated
therapeutic effect in various experimental inflammatory/autoimmune
animal models, such as Experimental Autoimmune Encephalomyelitis
(EAE), an animal model for Multiple Sclerosis (MS), Dextran Sodium
Solphate (DSS) induced colitis for Inflammatory Bowel Disease,
Non-Obese Diabetic (NOD) mice for Type I Diabetes (IDDM),
Experimental Autoimmune Neuritis (EAN) for Guillain-Barre Syndrome,
Systemic Lupus Erythematosus (SLE), lupus nephritis, lupus
arthritis, Crohn's Disease and Rheumatoid arthritis. The
therapeutic activity of laquinimod in these models results from a
variety of mechanistic effects, including reduction of leukocyte
infiltration into target tissues by modulation of
chemokine-mediated T-cell adhesion, modulation of cytokine balance,
down regulation of MHC class II resulting in alteration of antigen
presentation, and effects on dendritic cells subpopulations (PCT
International Application Publication No. WO2013/169746).
[0148] A pharmaceutically acceptable salt of laquinimod includes
lithium, sodium, potassium, magnesium, calcium, manganese, copper,
zinc, aluminum and iron. Salt formulations of laquinimod and the
process for preparing the same are described, e.g., in U.S. Patent
Application Publication No. 2005/0192315 and PCT International
Application Publication No. WO 2005/074899, which are hereby
incorporated by reference into this application.
[0149] The subject invention provides a mixture of crystalline
laquinimod sodium particles, wherein (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
40 microns or less or (ii) 50% or more of the total amount by
volume of the laquinimod sodium particles have a size of 15 microns
or less, and wherein: [0150] a) the mixture has a bulk density of
0.2 g/mL to 0.4 g/mL; [0151] b) the mixture has a tapped density of
0.40 g/mL to 0.7 g/mL; [0152] c) an amount of heavy metal which is
present in the mixture is no more than 20 ppm of heavy metal
relative to the amount by weight of laquinimod sodium; [0153] d) an
amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) which
is present in the mixture is no more than 0.15% relative to the
amount of laquinimod sodium as measured by HPLC; [0154] e) an
amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) which is present in the mixture is no more than 0.15%
relative to the amount of laquinimod sodium as measured by HPLC;
[0155] f) an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbox-
ylate (MCQME) in the mixture is no more than 0.12% relative to the
amount of laquinimod sodium as measured by HPLC; [0156] g) an
amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) which is present in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC;
[0157] h) an amount of N-ethylaniline (NEA) which is present in the
mixture is no more than 0.10% relative to the amount of laquinimod
sodium as measured by HPLC; or [0158] i) an amount of
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide (5-HLAQ) which is present in the mixture is no more than
0.10% relative to the amount of laquinimod sodium as measured by
HPLC.
[0159] In an embodiment of the mixture, (i) 90% or more of the
total amount by volume of the laquinimod sodium particles have a
size of less than 40 microns or (ii) 50% or more of the total
amount by volume of the laquinimod sodium particles have a size of
less than 15 microns.
[0160] In an embodiment of the mixture, 10% or more of the total
amount by volume of the laquinimod sodium particles have a size of
5 microns or less and wherein: [0161] a) the mixture has a tapped
density of 0.40 g/mL to 0.7 g/mL; or [0162] b) an amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) which is
present in the mixture is no more than 0.15% relative to the amount
of laquinimod sodium as measured by HPLC.
[0163] The subject invention provides a mixture of crystalline
laquinimod sodium particles, wherein (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
less than 40 microns, (ii) 50% or more of the total amount by
volume of the laquinimod sodium particles have a size of less than
15 microns, and (iii) 10% or more of the total amount by volume of
the laquinimod sodium particles have a size of less than 5 microns
and wherein: [0164] a) the mixture has a bulk density of 0.2 g/mL
to 0.4 g/mL; [0165] b) the mixture has a tapped density of 040 g/mL
to 0.7 g/mL; [0166] c) an amount of heavy metal which is present in
the mixture is no more than 20 ppm of heavy metal relative to the
amount by weight of laquinimod sodium [0167] d) an amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) which is
present in the mixture is no more than 0.15% relative to the amount
of laquinimod sodium as measured by HPLC; [0168] e) an amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) which is present in the mixture is no more than 0.15%
relative to the amount of laquinimod sodium as measured by HPLC;
[0169] f) an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbox-
ylate (MCQME) which is present in the mixture is no more than 0.12%
relative to the amount of laquinimod sodium as measured by HPLC;
[0170] g) an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylase
(MCQME) which is present in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC;
[0171] h) an amount of N-ethylaniline (NEA) which is present in the
mixture is no more than 0.10% relative to the amount of laquinimod
sodium as measured by HPLC; or [0172] i) an amount of
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide (5-HLAQ) which is present in the mixture is no more than
0.10% relative to the amount of laquinimod sodium as measured by
HPLC.
[0173] In an embodiment of the mixture, the mixture is prepared in
a single batch comprising 2.5 kg or more of laquinimod sodium. In
an embodiment of the mixture, the laquinimod sodium particles are
determined based on an unmilled sample of the mixture. In an
embodiment of the mixture, the size and amount by volume of
laquinimod sodium particles are determined based on a milled sample
of the mixture.
[0174] In an embodiment of the mixture, the mixture has a bulk
density of 0.2 g/mL to 0.4 mL. In an embodiment of the mixture, the
mixture has a tapped density of 0.40 g/mL to 0.7 g/mL.
[0175] In an embodiment of the mixture, an amount of aluminium in
the mixture is less than 5 ppm relative to the amount by weight of
laquinimod sodium. In an embodiment of the mixture, an amount of
aluminium in the mixture is less than 2 ppm relative to the amount
by weight of laquinimod sodium. In an embodiment of the mixture, an
amount of calcium in the mixture is less than 60 ppm relative to
the amount by weight of laquinimod sodium. In an embodiment of the
mixture, an amount of calcium in the mixture is less than 25 ppm
relative to the amount by weight of laquinimod sodium, hi an
embodiment of the mixture, an amount of copper in the mixture is
less than 1 ppm relative to the amount by weight of laquinimod
sodium. In an embodiment of the mixture, an amount of copper in the
mixture is less than 0.6 ppm relative to the amount by weight of
laquinimod sodium. In an embodiment of the mixture, an amount of
zinc in the mixture is less than 7 ppm relative to the amount by
weight of laquinimod sodium. In an embodiment of the mixture, an
amount of zinc in the mixture is less than 4 ppm relative to the
amount by weight of laquinimod sodium. In an embodiment of the
mixture, an amount of heavy metal in the mixture is no more than 20
ppm relative to the amount by weight of laquinimod sodium.
[0176] In an embodiment of the mixture, a total amount of polar
impurities in the mixture is no more than 1.00% relative to the
amount of laquinimod sodium as measured by HPLC.
[0177] In an embodiment of the mixture, an amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) in the mixture
is no more than 0.15% relative to the amount of laquinimod sodium
as measured by HPLC. In an embodiment of the mixture, an amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the mixture is no more than 0.15% relative to the
amount of laquinimod sodium as measured by HPLC. In an embodiment
of the mixture, an amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) in the mixture is no more than 0.12% relative to the amount
of laquinimod sodium as measured by HPLC. In another embodiment,
the amount of MCQME in the mixture is no more than 0.10% relative
to the amount of laquinimod sodium as measured by HPLC. In an
embodiment of the mixture, an amount of ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the mixture is no more than 0.10% relative to the amount
of laquinimod sodium as measured by HPLC. In an embodiment of the
mixture, a total amount of non-polar impurities in the mixture is
no more than 0.50% relative to the amount of laquinimod sodium as
measured by HPLC. In an embodiment of the mixture, an amount of
N-ethylaniline (NEA) in the mixture is no more than 0.10% relative
to the amount of laquinimod sodium as measured by HPLC. In an
embodiment of the mixture, an amount of
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide (5-HLAQ) in the mixture is no more than 0.10% relative to
the amount of laquinimod sodium as measured by HPLC. In an
embodiment of the mixture, an amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxa-
mide (DELAQ) in the mixture is no more than 0.10% relative to the
amount of laquinimod sodium as measured by HPLC.
[0178] In an embodiment of the mixture, an amount of laquinimod
acid in the mixture is no more than 1.00% relative to the amount of
laquinimod sodium as measured by HPLC. In an embodiment of the
mixture, an amount of dimethyl malonate in the mixture is no more
than 0.10% relative to the amount of laquinimod sodium as measured
by HPLC. In an embodiment of the mixture, an amount of diethyl
malonate in the mixture is no more than 0.10% relative to the
amount of laquinimod sodium as measured by HPLC. In an embodiment
of the mixture, an amount of dimethyl sulfate in the mixture is no
more than 1 ppm relative to the amount by weight of laquinimod
sodium. In an embodiment of the mixture, an amount of water in the
mixture is no more than 1.5% by weight relative to the amount of
laquinimod sodium as measured by K.F. coulometric titration. In an
embodiment of the mixture, an amount of sodium from 5.8% to 6.4%
relative to the amount by weight of laquinimod sodium.
[0179] In an embodiment of the mixture, an amount of ethanol in the
mixture is no more than 5000 ppm relative to the amount by weight
of laquinimod sodium. In an embodiment of the mixture, an amount of
n-heptane in the mixture is no more than 5000 ppm relative to the
amount by weight of laquinimod sodium. In an embodiment of the
mixture, an amount of n-octane in the mixture is no more than 2000
ppm relative to the amount by weight of laquinimod sodium. In
another embodiment of the mixture, an amount of n-octane in the
mixture is no more than 200 ppm relative to the amount by weight of
laquinimod sodium. In another embodiment of the mixture, an amount
of n-octane in the mixture is no more than 20 ppm relative to the
amount by weight of laquinimod sodium. In an embodiment of the
mixture, an amount of methanol in the mixture is no more than 3000
ppm relative to the amount by weight of laquinimod sodium. In an
embodiment of the mixture, an amount of acetone in the mixture is
no more than 5000 ppm relative to the amount by weight of
laquinimod sodium. In an embodiment of the mixture, an amount of
dioxane in the mixture is no more than 380 ppm relative to the
amount by weight of laquinimod sodium. In an embodiment of the
mixture, an amount of dimethyl formamide in the mixture is no more
than 880 ppm relative to the amount by weight of laquinimod
sodium.
[0180] In an embodiment of the mixture, the recited compound is
present in the mixture. In another embodiment, the recited compound
is present in at least trace amounts.
[0181] The subject invention provides a pharmaceutical composition
comprising the mixture of the subject invention and a
pharmaceutically acceptable carrier.
[0182] In an embodiment of the pharmaceutical composition, a total
amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the pharmaceutical composition is no more than
0.50% relative to the amount of laquinimod sodium as measured by
HPLC. In an embodiment of the pharmaceutical composition, a total
amount of polar impurities in the pharmaceutical composition is no
more than 2.00% relative to the amount of laquinimod sodium as
measured by HPLC. In an embodiment of the pharmaceutical
composition, an amount of N-ethylaniline (NEA) in the
pharmaceutical composition is no more than 0.50% relative to the
amount of laquinimod sodium as measured by HPLC. In an embodiment
of the pharmaceutical composition, an amount of
5-chloro-N-ethyl-3-hydroxy-1-methyl-5-2,4-dioxo-N-phenyl-1,2,3,4-tetrahyd-
roquinoline-3-carboxamide (3-HLAQ) in the pharmaceutical
composition is no more than 0.50% relative to the amount of
laquinimod sodium as measured by HPLC. In an embodiment of the
pharmaceutical composition, a total amount of non-polar impurities
in the pharmaceutical composition is no more than 1.00% relative to
the amount of laquinimod sodium as measured by HPLC. In an
embodiment of the pharmaceutical composition, an amount of
N-ethyl-4-hydroxy-1-methyl-5-(methyl(2,3,4,5,6-pentahydroxyhexyl)amino)-2-
-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide (MEG-LAQ) in the
pharmaceutical composition is no more than 1.00% relative to the
amount of laquinimod sodium as measured by HPLC.
[0183] In an embodiment of the pharmaceutical composition, an
amount of water in the pharmaceutical composition is no more than
1.50% relative to the amount of laquinimod sodium as measured by
K.F. coulometric titration. In an embodiment of the pharmaceutical
composition, an amount of water in the pharmaceutical composition
is no more than 0.80% relative to the amount of laquinimod sodium
as measured by K.F. coulometric titration. In an embodiment of the
pharmaceutical composition, an amount of sodium from 5.8% to 6.4%
relative to the amount by weight of laquinimod sodium.
[0184] In an embodiment of the pharmaceutical composition, an
amount of ethanol in the pharmaceutical composition is no more than
5000 ppm relative to the amount by weight of laquinimod sodium. In
an embodiment of the pharmaceutical composition, an amount of
n-heptane in the pharmaceutical composition is no more than 5000
ppm relative to the amount by weight of laquinimod sodium. In an
embodiment of the pharmaceutical composition, an amount of n-octane
in the pharmaceutical composition is no more than 2000 ppm relative
to the amount by weight of laquinimod sodium. In another embodiment
of the pharmaceutical composition, an amount of n-octane in the
pharmaceutical composition is no more than 200 ppm relative to the
amount by weight of laquinimod sodium. In another embodiment of the
pharmaceutical composition, an amount of n-octane in the
pharmaceutical composition is no more than 20 ppm relative to the
amount by weight of laquinimod sodium. In an embodiment of the
pharmaceutical composition, an amount of methanol in the
pharmaceutical composition is no more than 380 ppm relative to the
amount by weight of laquinimod sodium. In an embodiment of the
pharmaceutical composition, an amount of acetone in the
pharmaceutical composition is no more than 880 ppm relative to the
amount by weight of laquinimod sodium. In an embodiment of the
pharmaceutical composition, an amount of dioxane in the
pharmaceutical composition is no more than 380 ppm relative to the
amount by weight of laquinimod sodium. In an embodiment of the
pharmaceutical composition, an amount of dimethyl formamide in the
pharmaceutical composition is no more than 880 ppm relative to the
amount by weight of laquinimod sodium.
[0185] In an embodiment of the pharmaceutical composition, the
recited compound is present in the pharmaceutical composition. In
another embodiment, the recited compound is present in at least
trace amounts.
[0186] The subject invention provides a method of treating a
subject afflicted with a form of multiple sclerosis, lupus
nephritis, lupus arthritis, rheumatoid arthritis, a BDNF-related
disorder, Crohn's disease, a GABA-related disorder, a cannabinoid
receptor type 1 (CB1) mediated disorder, or an ocular inflammatory
disorder comprising administering to the subject the mixture of the
subject invention or the pharmaceutical composition of the subject
invention so as to thereby treat the subject.
[0187] The subject invention provides a method for alleviating a
symptom of multiple sclerosis, lupus nephritis, lupus arthritis,
rheumatoid arthritis, a BDNF-related disorder, Crohn's disease, a
GABA-related disorder, a cannabinoid receptor type 1 (CB1) mediated
disorder, or an ocular inflammatory disorder in a subject afflicted
with a form of multiple sclerosis comprising administering to the
subject the mixture of any one of the subject invention or the
pharmaceutical composition of the subject invention thereby
alleviating the symptom of multiple sclerosis in the subject.
[0188] In one embodiment, the mixture or the pharmaceutical
composition for use in the treatment of, or alleviation of symptoms
of, a form of multiple sclerosis, lupus nephritis, lupus arthritis,
rheumatoid arthritis, a BDNF-related disorder, Crohn's disease, a
GABA-related disorder, a cannabinoid receptor type 1 (CB1) mediated
disorder, or an ocular inflammatory disorder.
[0189] The subject invention provides a use of the mixture or the
pharmaceutical composition for the manufacture of a medicament for
treating, or alleviating a symptom of, a form of multiple
sclerosis, a GABA-related disorder, a cannabinoid receptor type 1
(CB1) mediated disorder, lupus nephritis, lupus arthritis,
rheumatoid arthritis, a BDNF-related disorder, Crohn's disease, or
an ocular inflammatory disorder.
[0190] The subject invention provides a process of
recrystallization of laquinimod sodium comprising: a) dissolving an
amount of laquinimod sodium in water to form an aqueous solution;
b) concentrating the aqueous solution to form a concentrated
solution comprising approximately 1.7-1.8 mL of water per gram of
laquinimod sodium; c) adding acetone to the concentrated solution
of step b); and d) isolating recrystallized laquinimod sodium.
[0191] In an embodiment of the process, the amount of laquinimod
sodium in step a) is 2.5 kg or greater. In an embodiment of the
process, step a) is performed with 10-12 mL of water per gram of
laquinimod sodium. In an embodiment of the process, step a) is
performed with approximately 11 mL of water per gram of laquinimod
sodium. In an embodiment of the process, step a) is performed by
heating the aqueous solution to a temperature of 58-75.degree. C.
In an embodiment of the process, step a) is performed by heating
the aqueous solution to a temperature of 60-73.degree. C.
[0192] In an embodiment of the process, crystallization occurs
after step a) and before step c).
[0193] In an embodiment of the process, crystallization is induced
by rapid stirring during or after the concentrating step b). In an
embodiment of the process, crystallization is induced by addition
of a seed crystal during or after the concentrating step b). In an
embodiment of the process, crystallization occurs without addition
of a seed crystal.
[0194] In an embodiment of the process, step b) is performed under
conditions appropriate to induce crystallization at the
concentration of 1.7-1.8 mL of water per gram of laquinimod sodium.
In an embodiment of the process, step b) is performed at
28-45.degree. C. hi an embodiment of the process, step b) is
performed at 30-40.degree. C. In an embodiment of the process, step
c) is performed with the concentrated solution at 40-55.degree. C.
In an embodiment of the process, step c) is performed with the
concentrated solution at 45-50.degree. C. In an embodiment of the
process, step c) is performed with 6-12 mL of acetone per gram of
laquinimod sodium. In an embodiment of the process, step c) is
performed with approximately 10 mL of acetone per gram of
laquinimod sodium. In an embodiment of the process, step c) is
performed over a period of 1-4 hours. In an embodiment of the
process, step c) is performed over a period of 1.2-2.5 hours. In an
embodiment of the process, step c) is followed by cooling the
solution to a temperature no less than -14.degree. C. and no more
than 6.degree. C. In an embodiment of the process, step c) is
followed by cooling the solution to a temperature no less than
-4.degree. C. and no more than 4.degree. C.
[0195] In an embodiment of the process, the solution is cooled over
a period of 3-5 hours. In an embodiment of the process, the
solution is cooled over a period of 3.5-4.5 hours.
[0196] In an embodiment of the process, step d) further comprises
washing the recrystallized laquinimod sodium with 1-4 mL of acetone
per gram of crude laquinimod sodium used in step a). In an
embodiment of the process, step d) further comprises washing the
recrystallized laquinimod sodium with approximately 3 mL of acetone
per gram of crude laquinimod sodium used in step a). In an
embodiment of the process, step d) further comprises drying the
recrystallized laquinimod sodium for no less than one hour at
30-40.degree. C. under a vacuum of no more than 50 mmHg.
[0197] In an embodiment of the process, the isolated recrystallized
laquinimod sodium in step d) is a mixture of crystalline laquinimod
sodium particles having a particle size distribution such that (i)
90% or more of the total amount by volume of the laquinimod sodium
particles have a size of 40 microns or less, (ii) 50% or more of
the total amount by volume of the laquinimod sodium particles have
a size of 15 microns or less, and (iii) 10% or more of the total
amount by volume of the laquinimod sodium particles have a size of
less than 5 microns or less.
[0198] The subject invention provides a mixture of crystalline
laquinimod sodium particles prepared by the process of the subject
invention.
[0199] In an embodiment of the mixture of crystalline laquinimod
sodium particles prepared by the process of the subject invention,
(i) 90% or more of the total amount by volume of the laquinimod
sodium particles have a size of 40 microns or less, (ii) 50% or
more of the total amount by volume of the laquinimod sodium
particles have a size of 15 microns or less, and (iii) 10% or more
of the total amount by volume of the laquinimod sodium particles
have a size of 5 microns or less.
[0200] In an embodiment of the mixture, [0201] a) the mixture has a
bulk density of 0.2 out to 0.4 OIL; [0202] b) the mixture has a
tapped density of 0.40 g/mL to 0.7 g/mL; [0203] c) an amount of
aluminium in the mixture is less than 5 ppm relative to the amount
by weight of laquinimod sodium; [0204] d) an amount of calcium in
the mixture is less than 60 ppm relative to the amount by weight of
laquinimod sodium; [0205] e) an amount of copper in the mixture is
less than 1 ppm relative to the amount by weight of laquinimod
sodium; [0206] f) an amount of zinc in the mixture is less than 7
ppm relative to the amount by weight of laquinimod sodium; [0207]
g) an amount of heavy metal in the 20 ppm relative to the amount by
weight of laquinimod sodium; [0208] h) a total amount of polar
impurities in the mixture is no more than 1.00% relative to the
amount of laquinimod sodium as measured by HPLC; [0209] i) an
amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) in
the mixture is no more than 0.15% relative to the amount of
laquinimod sodium as measured by HPLC; [0210] j) an amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the mixture is no more than 0.15% relative to the
amount of laquinimod sodium as measured by HPLC; [0211] k) an
amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) in the mixture is no more than 0.12% relative to the amount
of laquinimod sodium as measured by HPLC; [0212] l) an amount of
methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) in the mixture is no more than 0.10% relative to the amount
of laquinimod sodium as measured by HPLC; [0213] m) an amount of
ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the mixture is no more than 0.10% relative to the amount
of laquinimod sodium as measured by HPLC; [0214] n) a total amount
of non-polar impurities in the mixture is no more than 0.50%
relative to the amount of laquinimod sodium as measured by HPLC;
[0215] o) an amount of N-ethylaniline (NEA) in the mixture is no
more than 0.10% relative to the amount of laquinimod sodium as
measured by HPLC; [0216] p) an amount of laquinimod acid in the
mixture is no more than 1.00% relative to the amount of laquinimod
sodium as measured by HPLC; [0217] q) an amount of dimethyl
malonate in the mixture is no more than 0.10% relative to the
amount of laquinimod sodium as measured by HPLC; [0218] r) an
amount of diethyl malonate in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC;
[0219] s) an amount of dimethyl sulfate in the mixture is no more
than 1 ppm relative to the amount by weight of laquinimod sodium;
[0220] t) an amount of water in the mixture is no more than 1.5% by
weight relative to the amount of laquinimod sodium as measured by
K.F. coulometric titration; [0221] u) the mixture comprises an
amount of sodium 5.8% to 64% relative to the amount by weight of
laquinimod sodium; [0222] v) an amount of ethanol in the mixture is
no more than 5000 ppm relative to the amount by weight of
laquinimod sodium; [0223] w) an amount of n-heptane in the mixture
is no more than 5000 ppm relative to the amount by weight of
laquinimod sodium; [0224] x) an amount of n-octane in the mixture
is no more than 2000 ppm relative to the amount by weight of
laquinimod sodium; [0225] y) an amount of n-octane in the mixture
is no more than 200 ppm relative to the amount by weight of
laquinimod sodium; [0226] z) an amount of n-octane in the mixture
is no more than 20 ppm relative to the amount by weight of
laquinimod sodium; [0227] aa) an amount of methanol in the mixture
is no more than 3000 ppm relative to the amount by weight of
laquinimod sodium; [0228] bb) an amount of acetone in the mixture
is no more than 5000 ppm relative to the amount by weight of
laquinimod sodium; [0229] cc) an amount of dioxane in the mixture
is no more than 380 ppm relative to the amount by weight of
laquinimod sodium; or [0230] dd) an amount of di methyl formamide
in the mixture is no more than 880 ppm relative to the amount by
weight of laquinimod sodium.
[0231] In an embodiment of the mixture, [0232] a) the mixture has a
bulk density of 0.2 g/mL to 0.4 g/mL; [0233] b) the mixture has a
tapped density of 0.40 g/mL to 0.7 g/mL; [0234] c) an amount of
aluminium relative to the amount by weight of laquinimod sodium;
[0235] d) an amount of calcium in the mixture is less than 60 ppm
relative to by weight of laquinimod sodium; [0236] e) an amount of
copper in the mixture is less than 1 ppm relative to the amount by
weight of laquinimod sodium; [0237] f) an amount of zinc in the
mixture is less than 7 ppm relative to the amount by weight of
laquinimod sodium; [0238] g) an amount of heavy metal in the
mixture is no more than 0.002% relative to the amount by weight of
laquinimod sodium; [0239] h) a total amount of polar impurities in
the mixture is no more than 1.00% relative to the amount of
laquinimod sodium as measured by HPLC; [0240] i) an amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) in the mixture
is no more than 0.15% relative to the amount of laquinimod sodium
as measured by HPLC; [0241] j) an amount of
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the mixture is no more than 0.15% relative to the
amount of laquinimod sodium as measured by HPLC; [0242] k) an
amount of methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) in the mixture is no more than 0.12% relative to the amount
of laquinimod sodium as measured by HPLC; [0243] l) an amount of
methyl
5-.chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) in the mixture is no more than 0.10% relative to the amount
of laquinimod sodium as measured by HPLC; [0244] m) an amount of
ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the mixture is no more than 0.10% relative to the amount
of laquinimod sodium as measured by HPLC; [0245] n) a total amount
of non-polar impurities in the mixture is no more than 0.50%
relative to the amount of laquinimod sodium as measured by HPLC;
[0246] o) an amount of N-ethylaniline (NEA) in the mixture is no
more than 0.10% relative to the amount of laquinimod sodium as
measured by HPLC; [0247] p) an amount of laquinimod acid in the
mixture is no more than 1.00% relative to the amount of laquinimod
sodium as measured by HPLC; [0248] q) an amount of dimethyl
malonate in the mixture is no more than 0.10% relative to the
amount of laquinimod sodium as measured by HPLC; [0249] r) an
amount of diethyl malonate in the mixture is no more than 0.10%
relative to the amount of laquinimod sodium as measured by HPLC;
[0250] s) an amount of dimethyl sulfate in the mixture is no more
than 1 ppm relative to the amount by weight of laquinimod sodium;
[0251] t) an amount of water in the mixture is no more than 1.5% by
weight relative to the amount of laquinimod sodium as measured by
K.F. coulometric titration; [0252] u) the mixture comprises an
amount of sodium from 5.8% to 6.4% relative to the amount by weight
of laquinimod sodium; [0253] v) an amount of ethanol in the mixture
is no more than 5000 ppm relative to the amount by weight of
laquinimod sodium; [0254] w) an amount of n-heptane in the mixture
is no more than 5000 ppm relative to the amount by weight of
laquinimod sodium; [0255] x) an amount of n-octane in the mixture
is no more than 2000 ppm relative to the amount by weight of
laquinimod sodium; [0256] y) an amount of methanol in the mixture
is no more than 3000 ppm relative to the amount by weight of
laquinimod sodium; [0257] z) an amount of acetone in the mixture is
no more than 5000 ppm relative to the amount by weight of
laquinimod sodium; [0258] aa) an amount of dioxane in the mixture
is no more than 380 ppm relative to the amount by weight of
laquinimod sodium; and [0259] bb) an amount of dimethyl formamide
in the mixture is no more than 880 ppm relative to the amount by
weight of laquinimod sodium.
[0260] In one embodiment, the amount of n-octane in the mixture is
no more than 200 ppm. In another embodiment, the amount of n-octane
in the mixture is no more than 20 ppm.
[0261] In an embodiment of the mixture, the recited compound is
present in the mixture. In another embodiment, the recited compound
is present in at least trace amounts.
[0262] The subject invention provides a pharmaceutical composition
comprising the mixture of crystalline laquinimod sodium particles
prepared by the process of the subject invention, and a
pharmaceutically acceptable carrier.
[0263] In an embodiment of the pharmaceutical composition, [0264]
a) a total amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) and
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the pharmaceutical composition is no more than
0.50% relative to the amount of laquinimod sodium as measured by
HPLC; [0265] b) a total amount of polar impurities in the
pharmaceutical composition is no more than 2.00% relative to the
amount of laquinimod sodium as measured by HPLC; [0266] c) an
amount of N-ethylaniline (NEA) in the pharmaceutical composition is
no more than 0.50% relative to the amount of laquinimod sodium as
measured by HPLC; [0267] d) a total amount of non-polar impurities
in the pharmaceutical composition is no more than 1.00% relative to
the amount of laquinimod sodium as measured by HPLC; [0268] e) an
amount of water in the pharmaceutical composition is no more than
0.80% relative to the amount of laquinimod sodium as measured by
K.F. coulometric titration; [0269] f) the pharmaceutical
composition comprises an amount of sodium from 5.8% to 6.4%
relative to the amount by weight of laquinimod sodium; [0270] g) an
amount of ethanol in the pharmaceutical composition is no more than
5000 ppm relative to the amount by weight of laquinimod sodium;
[0271] h) an amount of n-heptane in the pharmaceutical composition
is no more than 5000 ppm relative to the amount by weight of
laquinimod sodium; [0272] i) an amount of n-octane in the
pharmaceutical composition is no more than 2000 ppm relative to the
amount by weight of laquinimod sodium; [0273] j) an amount of
n-octane in the pharmaceutical composition is no more than 200 ppm
relative to the amount by weight of laquinimod sodium; [0274] k) an
amount of n-octane in the pharmaceutical composition is no more
than 20 ppm relative to the amount by weight of laquinimod sodium;
[0275] l) an amount of methanol in the pharmaceutical composition
is no more than 380 ppm relative to the amount by weight of
laquinimod sodium; [0276] m) an amount of acetone in the
pharmaceutical composition is no more than 880 ppm relative to the
amount by weight of laquinimod sodium; [0277] n) an amount of
dioxane in the pharmaceutical composition is no more than 380 ppm
relative to the amount by weight of laquinimod sodium; or [0278] o)
an amount of dimethyl formamide in the pharmaceutical composition
is no more than 880 ppm relative to the amount by weight of
laquinimod sodium.
[0279] In an embodiment of the pharmaceutical composition, [0280]
a) a total amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) and
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the pharmaceutical composition is no more than
0.50% relative to the amount of laquinimod sodium as measured by
HPLC; [0281] b) a total amount of polar impurities in the
pharmaceutical composition is no more than 2.00% relative to the
amount of laquinimod sodium as measured by HPLC; [0282] c) an
amount of N-ethylaniline (NEA) in the pharmaceutical composition is
no more than 0.50% relative to the amount of laquinimod sodium as
measured by HPLC; [0283] d) a total amount of non-polar impurities
in the pharmaceutical composition is no more than 1.00% relative to
the amount of laquinimod sodium as measured by HPLC; [0284] e) an
amount of water in the pharmaceutical composition is no more than
0.80% relative to the amount of laquinimod sodium as measured by
K.F. coulometric titration; [0285] f) the pharmaceutical
composition comprises an amount of sodium from 5.8% to 6.4%
relative to the amount by weight of laquinimod sodium; [0286] g) an
amount of ethanol in the pharmaceutical composition is no more than
5000 ppm relative to the amount by weight of laquinimod sodium;
[0287] h) an amount of n-heptane in the pharmaceutical composition
is no more than 5000 ppm relative to the amount by weight of
laquinimod sodium; [0288] i) an amount of n-octane in the
pharmaceutical composition is no more than 2000 ppm relative to the
amount by weight of laquinimod sodium; [0289] j) an amount of
methanol in the pharmaceutical composition is no more than 380 ppm
relative to the amount by weight of laquinimod sodium; [0290] k) an
amount of acetone in the pharmaceutical composition is no more than
880 ppm relative to the amount by weight of laquinimod sodium;
[0291] l) an amount of dioxane in the pharmaceutical composition is
no more than 380 ppm relative to the amount by weight of laquinimod
sodium; and [0292] m) an amount of dimethyl formamide in the
pharmaceutical composition is no more than 880 ppm relative to the
amount by weight of laquinimod sodium.
[0293] In one embodiment, the amount of n-octane in the
pharmaceutical composition is no more than 200 ppm relative to the
amount by weight of laquinimod sodium. In another embodiment, the
amount of n-octane in the pharmaceutical composition is no more
than 20 ppm relative to the amount by weight of laquinimod
sodium.
[0294] The subject invention provides a mixture of crystalline
laquinimod sodium particles wherein (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
40 microns or less or (ii) 50% or more of the total amount by
volume of the laquinimod sodium particles have a size of 15 microns
or less, and wherein a) an amount of aluminium in the mixture is
less than 5 ppm relative to the amount by weight of laquinimod
sodium; b) an amount of calcium in the mixture is less than 60 ppm
relative to the amount by weight of laquinimod sodium; c) an amount
of copper in the mixture is less than 1 ppm relative to the amount
by weight of laquinimod sodium; or d) an amount of zinc in the
mixture is less than 7 ppm relative to the amount by weight of
laquinimod sodium.
[0295] In an embodiment of the pharmaceutical composition, the
recited compound is present in the pharmaceutical composition. In
another embodiment, the recited compound is present in at least
trace amounts.
[0296] The subject invention provides a pharmaceutical composition
comprising the mixture of the subject invention and a
pharmaceutically acceptable carrier.
[0297] In an embodiment of the mixture, 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
1 micron or greater, 2 microns or greater, 3 microns or greater or
4 microns or greater. In an embodiment of the mixture, 90% of the
total amount by volume of the laquinimod sodium particles have a
size of greater than 1 micron, greater than 2 microns, greater than
3 microns, greater than 4 microns or greater than 5 microns.
[0298] In a further embodiment of the mixture, the mixture has a
tapped density of 0.4 g/mL to 0.7 g/mL, 0.44 g/mL to 0.7 g/mL, 0.45
g/mL to 0.7 g/mL, 0.46 g/mL to 0.7 g/mL or 0.5 g/mL to 0.7 g/mL. In
a further embodiment of the mixture, the mixture has a bulk density
of 0.2 g/mL to 0.4 g/mL, 0.2 g/mL to 0.33 g/mL, 0.2 g/mL to 0.31
g/mL.
[0299] In a further embodiment of the pharmaceutical composition,
the pharmaceutical composition comprises mannitol. In a further
embodiment of the pharmaceutical composition, the pharmaceutical
composition comprises meglumine. In a further embodiment of the
pharmaceutical composition, the pharmaceutical composition
comprises sodium stearyl fumarate.
[0300] In a further embodiment of the pharmaceutical composition,
not less than 70% of the labeled amount of laquinimod is dissolved
in 30 minutes. In a further embodiment of the pharmaceutical
composition, capsules of the pharmaceutical composition contain
90-110% of the labeled amount of laquinimod. In a further
embodiment of the pharmaceutical composition, capsules of the
pharmaceutical composition contain 95-105% of the labeled amount of
laquinimod. In a further embodiment of the pharmaceutical
composition, capsules of the pharmaceutical composition contain
98.0-102.0% of the labeled amount of laquinimod.
[0301] In a further embodiment of the pharmaceutical composition,
the pharmaceutical composition has content uniformity conforming to
the U.S. Pharmacopeia. In a further embodiment of the
pharmaceutical composition, the pharmaceutical composition has
content uniformity conforming to European Pharmacopeia.
[0302] The subject invention also provides an isolated compound
having the structure:
##STR00006##
or a salt thereof.
[0303] The subject invention also provides a composition comprising
a compound having the structure:
##STR00007## [0304] or a salt thereof, wherein the composition is
free of laquinimod or a salt thereof.
[0305] The subject invention also provides a pharmaceutical
composition comprising an amount of laquinimod and at least one of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE), wherein a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount not more than 10%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount not more than 10%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount not more than 10%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or d)
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 10%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or e) N-Ethylaniline
(NEA) is present in the pharmaceutical composition in an amount not
more than 10%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or f) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
not more than 10%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
[0306] In an embodiment of the pharmaceutical composition as
described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) is present in the pharmaceutical composition in an amount not
more than 0.15%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount not more than 0.15%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or c)
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.15%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or d) N-Ethylaniline
(NEA) is present in the pharmaceutical composition in an amount not
more than 0.1%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
not more than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
[0307] In one embodiment, methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.12%, relative to the concentration of laquinimod,
based on a determination by an HPLC method. In another embodiment,
methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
[0308] In an embodiment of the pharmaceutical composition as
described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) is present in the pharmaceutical composition in an amount
less than 0.05%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or c) Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
less than 0.05%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is
present in the pharmaceutical composition in an less than 0.06%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
less than 0.05%, relative to the concentration of laquinimod, based
on a determination by an HPLC method.
[0309] In one embodiment,
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.05%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
[0310] In an embodiment of the pharmaceutical composition as
described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) is present in the pharmaceutical composition in an amount
less than 0.02%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.03%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or c)
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
less than 0.02%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or d) N-Ethylaniline (NEA) is
present in the pharmaceutical composition in an less than 0.02%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
less than 0.02%, relative to the concentration of laquinimod, based
on a determination by an HPLC method.
[0311] In one embodiment,
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.02%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
[0312] In an embodiment of the pharmaceutical composition as
described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) is present in the pharmaceutical composition in an amount
greater than 0.02%, and not more than 0.15%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.02%, and not more than 0.15%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or c) Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
greater than 0.02%, and not more than 0.15%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical
composition in an greater than 0.02%, and not more than 0.1%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
greater than 0.02%, and not more than 0.1%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method.
[0313] In one embodiment, methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.12%, relative to the concentration of laquinimod,
based on a determination by an HPLC method. In another embodiment,
methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method. In yet another
embodiment,
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.03%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
[0314] In an embodiment of the pharmaceutical composition as
described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) is present in the pharmaceutical composition in an amount
greater than 0.05%, and not more than 0.15%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.05%, or greater than 0.10%, and not more than
0.15%, relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c) Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
greater than 0.05%, and less than 0.15%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical
composition in an greater than 0.06%, and not more than 0.10%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
greater than 0.05%, and not more than 0.10%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method.
[0315] In one embodiment, methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.12%, relative to the concentration of laquinimod,
based on a determination by an HPLC method. In another embodiment,
methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
not more than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
[0316] In an embodiment of the pharmaceutical composition as
described herein, a) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or b)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.02%, or greater than 0.03%, and less than
0.1%, relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c) Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or d) N-Ethylaniline (NEA) is present in the pharmaceutical
composition in an greater than 0.02%, and less than 0.06%, relative
to the concentration of laquinimod, based on a determination by an
HPLC method, or e) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method.
[0317] In an embodiment,
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.05%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
[0318] In an embodiment of the pharmaceutical composition as
described herein, the composition further comprises at least one
pharmaceutically acceptable carrier.
[0319] In an embodiment of the pharmaceutical composition as
described herein, a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount not more than 1.0%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or b)
N-Ethylaniline (NEA) is present in the pharmaceutical composition
in an amount not more than 0.5%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or c) the
combined amount of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is not more
than 0.5%, relative to the concentration of laquinimod, based on a
determination by an HPLC method.
[0320] In one embodiment, N-Ethylaniline (NEA) is present in the
pharmaceutical composition in an amount not more than 0.1%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method.
[0321] In an embodiment of the pharmaceutical composition as
described herein, a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount less than 0.05%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount less than 0.05%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.5%, relative to the concentration of laquinimod,
based on a determination by an HPLC method, or d) N-Ethylaniline
(NEA) is present in the pharmaceutical composition in an amount
less than 0.1%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or e) the combined amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is less than
0.1%, relative to the concentration of laquinimod, based on a
determination by an HPLC method.
[0322] In one embodiment,
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
[0323] In an embodiment of the pharmaceutical composition as
described herein, a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount less than 0.02%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or b)
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) is present in
the pharmaceutical composition in an amount less than 0.02%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.03%, relative to the concentration of
laquinimod, based on a determination by an HPLC method, or d)
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
less than 0.02%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or e) N-Ethylaniline (NEA) is
present in the pharmaceutical composition in an less than 0.03%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method, or f) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
less than 0.02%, relative to the concentration of laquinimod, based
on a determination by an HPLC method, or g) the combined amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is less than
0.03%, relative to the concentration of laquinimod, based on a
determination by an HPLC method.
[0324] In one embodiment,
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.02%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
[0325] In an embodiment of the pharmaceutical composition as
described herein, a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount greater than 0.02%, and not more than 1.0%, relative
to the concentration of laquinimod, based on a determination by an
HPLC method, or b) N-Ethylaniline (NEA) is present in the
pharmaceutical composition in an greater than 0.03%, and not more
than 0.5%, relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c) the combined amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is greater
than 0.03%, and not more than 0.5%, relative to the concentration
of laquinimod, based on a determination by an HPLC method.
[0326] In one embodiment, N-Ethylaniline (NEA) is present in the
pharmaceutical composition in an amount not more than 0.1%,
relative to the concentration of laquinimod, based on a
determination by an HPLC method.
[0327] In an embodiment of the pharmaceutical composition as
described herein, a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount greater than 0.05%, and not more than 1.0%, relative
to the concentration of laquinimod, based on a determination by an
HPLC method, or b) N-Ethylaniline (NEA) is present in the
pharmaceutical composition in an greater than 0.1%, and not more
than 0.5%, relative to the concentration of laquinimod, based on a
determination by an HPLC method, or c) the combined amount of
5-Chloro-4-hydroxy-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carbox-
ylic acid (MCQCA) present in the pharmaceutical composition is
greater than 0.1%, and not more than 0.5%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method.
[0328] In an embodiment of the pharmaceutical composition as
described herein, a)
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) is present in the pharmaceutical composition
in an amount greater than 0.02%, and less than 0.05%, relative to
the concentration of laquinimod, based on a determination by an
HPLC method, or b) 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
(MCQ) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or c)
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.02%, and less than 0.5%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or d) Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or e) N-Ethylaniline (NEA) is present in the pharmaceutical
composition in an greater than 0.03%, and less than 0.1%, relative
to the concentration of laquinimod, based on a determination by an
HPLC method, or f) Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) is present in the pharmaceutical composition in an amount
greater than 0.02%, and less than 0.05%, relative to the
concentration of laquinimod, based on a determination by an HPLC
method, or g) the combined amount of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) present in the pharmaceutical composition is greater
than 0.03%, and less than 0.1%, relative to the concentration of
laquinimod, based on a determination by an HPLC method.
[0329] In one embodiment,
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount greater than 0.03%, relative to the concentration of
laquinimod, based on a determination by an HPLC method. In another
embodiment,
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is present in the pharmaceutical composition in an
amount less than 0.1%, relative to the concentration of laquinimod,
based on a determination by an HPLC method.
[0330] In one embodiment, the pharmaceutical composition as
described herein further comprises laquinimod sodium salt. In
another embodiment, the pharmaceutical composition is in an oral
unit dosage form. In another embodiments, it is in the form of a
capsule, a tablet, or a liquid suspension. In another embodiment,
the oral unit dosage form comprises more than 0.3 mg laquinimod. In
another embodiments, the oral unit dosage form comprises more than
0.5 mg laquinimod. In yet another embodiment, the oral unit dosage
form comprises more than 0.6 mg laquinimod.
[0331] The subject invention also provides a process for preparing
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) comprising the steps of: a) adding sodium
hydroxide solution to a suspension of
5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide in water, b) stirring the mixture of step
a) followed by addition of hydrochloric acid solution, c)
extracting the aqueous solution with ethyl acetate, d) washing the
organic phase with brine, e) drying the organic phase over sodium
sulfate, f) filtering the suspension, g) evaporating the filtrate,
h) purifying the residue by crystallization from isopropyl alcohol,
i) cooling the suspension followed by filtering and washing with
isopropyl alcohol, and j) obtaining and drying the resulting white
solid.
[0332] The subject invention also provides
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) prepared by the process described above.
[0333] The subject invention also provides a process for preparing
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) comprising the
steps of: a) heating a mixture of
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) and dimethylsulfoxide, b) cooling the mixture of step
a), and c) filtering the mixture of step b) and collecting the
resulting filtrate.
[0334] The subject invention also provides
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) prepared by the
process described above.
[0335] The subject invention also provides a process for preparing
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) comprising the steps of: a) heating a mixture of ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in a solution of hydrochloric acid in acetic acid, b)
cooling the mixture of step a), c) diluting the mixture of step b)
with 2-propanol and further cooling the diluted mixture, and d)
filtering off the crystals resulting from step c).
[0336] The subject invention also provides
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) prepared by the process described above.
[0337] The subject invention also provides a process for preparing
Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) comprising the steps of: a) forming sodium dimethylmalonate
by reaction of dimethylmalonate in dimethlformamide with sodium
methoxide solution, b) reacting the intermediate
5-chloro-1-methyl-1h-benzo[D][1,3]oxazine-2,4-dione with sodium
dimethylmalonate to form methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) sodium salt, and c) acidifying methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) sodium salt to methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME).
[0338] The subject invention also provides Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) prepared by the process described above.
[0339] The subject invention also provides a process for preparing
Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) comprising the steps of: a) adding sodium hydride to a
solution of 5-chloro-1-methyl-1h-benzo[D][1,3]oxazine-2,4-dione and
diethyl malonate in dimethylformamide, b) heating the mixture of
step a) while stirring, c) cooling the solution of step b), d)
quenching the reaction mixture of step c), e) acidifying the
mixture of step d), f) filtering then drying the mixture of step
e), and g) crystallizing the crude product of step f) by dissolving
in ethanol following by slow cooling.
[0340] The subject invention also provides Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) prepared by the process described above.
[0341] The subject invention also provides a process for testing
whether a sample of laquinimod contains an undesirable impurity
which comprises determining whether the sample contains a compound
having the structure:
##STR00008##
[0342] The subject invention also provides a process for preparing
a validated pharmaceutical composition comprising laquinimod
comprising: a) obtaining a batch of laquinimod; b) determining the
amount of at least one of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the batch using by an HPLC method; and c) preparing the
pharmaceutical composition from the batch only if [0343] i) the
batch is determined to have not more than 0.15%
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ), relative to
the concentration of laquinimod, or [0344] ii) the batch is
determined to have not more than 0.15%
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), relative to the concentration of laquinimod, or
[0345] iii) the batch is determined to have not more than 0.15%,
0.12% or 0.1% Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), relative to the concentration of laquinimod, or [0346] iv)
the batch is determined to have not more than 0.1% N-Ethylaniline
(NEA), relative to the concentration of laquinimod, or [0347] v)
the batch is determined to have not more than 0.1% Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE), relative to the concentration of laquinimod.
[0348] In one embodiment, in step c) the pharmaceutical composition
is prepared from the batch only if the batch is determined to have
not more than 0.12% Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), relative to the concentration of laquinimod. In another
embodiment, in step c) the pharmaceutical composition is prepared
from the batch only if the batch is determined to have not more
than 0.1% Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyla-
te (MCQME), relative to the concentration of laquinimod.
[0349] The subject invention also provides a process for preparing
a pharmaceutical composition comprising laquinimod, or for
distributing a validated batch of a pharmaceutical composition
comprising laquinimod, comprising a) obtaining a batch of
laquinimod or of the pharmaceutical composition; b) performing
stability testing with a sample of the batch; c) determining the
total amount of at least one of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the sample of the batch after stability testing by an
HPLC method; and d) validating the batch for distribution or
preparing the pharmaceutical composition from the batch only if the
sample of the batch after stability testing contains [0350] i) not
more than a total of 0.15% relative to the concentration of
laquinimod of 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
or [0351] ii) not more than a total of 0.15% relative to the
concentration of laquinimod of
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA), or [0352] iii) not more than a total of 0.15%, 0.12%
or 0.1% relative to the concentration of laquinimod of Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), or [0353] iv) not more than a total of 0.1% relative to
the concentration of laquinimod of N-Ethylaniline (NEA), or [0354]
v) not more than a total of 0.1% relative to the concentration of
laquinimod of Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylat-
e (MCQEE).
[0355] In one embodiment, the process further comprises step e)
distributing the batch if in step d) the batch is validated for
distribution. In another embodiments, in step d) the batch is
validated for distribution or the pharmaceutical composition is
prepared from the batch only if the sample of the batch after
stability testing contains not more than 0.12% Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) relative to the concentration of laquinimod. In another
embodiment, in step d) the batch is validated for distribution or
the pharmaceutical composition is prepared from the batch only if
the sample of the batch after stability testing contains not more
than 0.1% Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), relative to the concentration of laquinimod.
[0356] The subject invention also provides a process for validating
a batch of a pharmaceutical product containing laquinimod or a
pharmaceutically acceptable salt thereof and a pharmaceutically
acceptable carrier for distribution comprising a) subjecting a
sample of the batch to stability testing; b) determining the amount
of at least one of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido-
)benzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the sale of the batch after stability testing by an HPLC
method; and c) validating the batch for distribution only if the
sample of the batch after stability testing contains [0357] i) not
more than a total of 1.0% relative to the concentration of
laquinimod of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ), or [0358] ii) not more than a total of 0.5%
or 0.1% relative to the concentration of laquinimod of
N-Ethylaniline (NEA), or [0359] iii) not more than a total of 0.5%
relative to the concentration of laquinimod of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) combined.
[0360] In an embodiment, in step c) the batch is validated for
distribution only if the sample of the batch after stability
testing not more than a total of the 0.1% of N-Ethylaniline (NEA)
relative to the concentration of laquinimod.
[0361] The subject invention also provides a process for preparing
a packaged pharmaceutical composition comprising laquinimod or a
pharmaceutically acceptable salt thereof comprising; a) a)
obtaining a batch of pharmaceutical composition comprising
laquinimod or a pharmaceutically acceptable salt thereof; b)
performing stability testing with a sample from the batch; c)
determining the amount of at least one of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido-
)benzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) in the sample by an HPLC method after stability testing;
and d) d) packaging the pharmaceutical composition in only if
[0362] i) the content of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) in the sample is determined to be not more
than 1.0% to the concentration of laquinimod, or [0363] ii) the
content of N-Ethylaniline (NEA) in the sample is determined to be
not more than 0.5% or 0.1% to the concentration of laquinimod.
[0364] iii) the combined content of
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ) and
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) in the sample is determined to be not more than a
total of 0.5% relative to the concentration of laquinimod.
[0365] In one embodiment, in step d) the pharmaceutical composition
is packaged only if the content of N-Ethylaniline (NEA) in the
sample is determined to be not more than 0.1% to the concentration
of laquinimod.
[0366] In one embodiments, the laquinimod is laquinimod sodium
salt.
[0367] The subject invention also provides an impurity or a salt
thereof for use, as a reference standard to detect trace amounts of
the impurity in a pharmaceutical composition comprising laquinimod
or a pharmaceutically acceptable salt thereof, wherein the impurity
is selected from the group consisting of
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ),
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME), N-Ethylaniline (NEA), and Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE).
[0368] The subject invention also provides a method of determining
the concentration of an impurity in a pharmaceutical composition
comprising laquinimod, the method comprising, a) Preparing a sample
solution from the pharmaceutical composition, b) Preparing a
standard solution comprising the impurity, c) Preparing a
resolution solution comprising laquinimod and the impurity, d)
Preparing a buffer solution by dissolving ammonium acetate in water
and adjusting to pH of 7.0.+-.0.05 with aqueous ammonia or glacial
acetic acid, e) Preparing a diluent solution comprising the buffer
solution and acetonitrile, f) Preparing a blank solution comprising
the diluent solution and aqueous acetonitrile, g) Injecting into
the HPLC the resolution solution, the blank solution, the standard
solution, and the sample solution, h) Running the HPLC using
ultraviolet absorption at 240 nm and the diluent solution as the
mobile phase, i) Determining the retention time (RT) and the areas
of the peaks of the impurity in the chromatograms of the sample
solution, and j) Performing quantitation of the impurity with
respect to the corresponding peaks in the chromatograms of the
standard solutions,
wherein the impurity is
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME). N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2
dihydroquinoline-3-carboxamide (5-HLAQ) or Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE).
[0369] The subject invention also provides a method of determining
the concentration of an impurity in a pharmaceutical composition
comprising laquinimod, the method comprising, a) Preparing a sample
solution from the pharmaceutical composition, b) Preparing a
standard solution comprising the impurity, c) Preparing a
resolution solution comprising laquinimod and the impurity, d)
Preparing a buffer solution by dissolving ammonium acetate in
water, and adjusting to pH of 7.0.+-.0.05 with aqueous ammonia or
glacial acetic acid, e) Preparing a blank solution comprising the
buffer solution and acetonitrile, f) Injecting into the HPLC the
resolution solution, the blank solution, the standard solution, and
the sample solution, g) Running the HPLC using ultraviolet
absorption at 240 nm and a mobile phase of a mixture of the buffer
solution, and acetonitrile, h) Determining the retention time (RT)
and the areas of the peaks of the impurity in the chromatograms of
the sample solution, and i) Performing quantitation of the impurity
with respect to the corresponding peaks in the chromatograms of the
standard solutions,
wherein the impurity is N-Ethylaniline (NEA),
1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3'-[l]ethylindolin-[2]-
-one](SPIRO-LAQ), or
5-Chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide (3-HLAQ).
[0370] The subject invention also provides a method of determining
the concentration of an impurity in a pharmaceutical composition
comprising laquinimod and a pharmaceutically acceptable carrier,
the method comprising, a) Preparing a sample solution from the
pharmaceutical composition, b) Preparing a standard solution
comprising the impurity, c) Preparing a Quantitation Limit (QL)
solution comprising the impurity, d) Preparing a resolution
solution comprising laquinimod and the impurity, e) Preparing a
buffer solution by dissolving ammonium dihydrogen phosphate in
water, and adjusting to pH of 7.0.+-.0.10 with aqueous ammonia or
phosphoric acid, f) Preparing a blank solution comprising the
buffer solution and acetonitrile, g) Injecting into the HPLC the
resolution solution, the blank solution, the QL solution, the
standard solution and the sample solution, h) Running the HPLC
using a ultraviolet absorption at 212 nm and a mobile phase of a
mixture of the buffer solution, acetonitrile, and methanol, i)
Determining the retention time (RT) and the areas of the peaks of
the impurity in the chromatograms of the sample solution, and j)
Performing quantitation of the impurity with respect to the
corresponding peaks in the chromatograms of the standard
solutions,
wherein the impurity is
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid (BH-3-HLAQ) or
N-ethyl-4-hydroxy-1-methyl-5-(2,3,4,5,6-pentahydroxyhexylamino)-2-oxo-N-p-
henyl-1,2-dihydroquinoline-3-carboxamide (MEG-LAQ).
[0371] The subject invention also provides a method of determining
the concentration of an impurity in a pharmaceutical composition
comprising laquinimod and a pharmaceutically acceptable carrier,
the method comprising, a) Preparing a sample solution from the
pharmaceutical composition, b) Preparing a standard solution
comprising the impurity, c) Preparing a resolution solution
comprising laquinimod and the impurity, d) Preparing a buffer
solution by dissolving ammonium acetate in water, and adjusting to
PH or 7.0.+-.0.05 with aqueous ammonia or glacial acetic acid, e)
Preparing a blank solution comprising the buffer solution and
acetonitrile, f) Injecting into the HPLC the resolution solution,
the blank solution, the standard solution and the sample solution,
g) Running the HPLC using a ultraviolet absorption at 242 nm and
the blank solution as the mobile phase, h) Determining the
retention time (RT) and the areas of the peaks of the impurity in
the chromatograms of the sample solution, and i) Performing
quantitation of the impurity with respect to the corresponding
peaks in the chromatograms of the standard solutions,
wherein the impurity is
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one (MCQ),
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyli-
c acid (MCQCA), Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQME) or N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2
dihydroquinoline-3-carboxamide (5-HLAQ).
[0372] The subject invention also provides a method of determining
the concentration of an impurity in a pharmaceutical composition
comprising laquinimod and a pharmaceutically acceptable carrier,
the method comprising, a) Preparing a sample solution from the
pharmaceutical composition, b) Preparing a standard solution
comprising the impurity, c) Preparing a resolution solution
comprising laquinimod and the impurity, d) Preparing a blank
solution comprising methanol and acetonitrile, e) Preparing a
buffer solution by dissolving ammonium acetate in water, and
adjusting to pH of 7.0.+-.0.05 with aqueous ammonia or glacial
acetic acid, f) Injecting into the HPLC the resolution solution,
the blank solution, and the sample solution, g) Running the HPLC
using a ultraviolet absorption at 240 nm and a mobile phase
comprising acetonitrile and the buffer solution, h) Determining the
retention time (RT) and the areas of the peaks of the impurity in
the chromatograms of the sample solution, and i) Performing
quantitation of the impurity with respect to the corresponding
peaks in the chromatograms of the standard solutions,
wherein the impurity is N-Ethylaniline (NEA),
5-Chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide (3-HLAQ) or
1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3'-[l]ethylindolin-[2]-
-one](SPIRO-LAQ).
[0373] Each embodiment disclosed herein is contemplated as being
applicable to each of the other disclosed embodiments. Thus, all
combinations of the various elements described herein are within
the scope of the invention. For example, any embodiment for an
element of a mixture a contemplated as being applicable to a
pharmaceutical composition. As another example, an embodiment to a
tapped density of 0.46 g/mL to 0.7 g/mL is contemplated as being
applicable to a mixture having an amount of aluminium in the
mixture less than 5 ppm relative to the amount by weight of
laquinimod sodium.
Comparison to the Prior Art
[0374] Applicants have unexpectedly achieved a mixture of
crystalline laquinimod sodium that is improved over the laquinimod
sodium mixture of the prior art.
[0375] U.S. Pat. No. 7,884,208 teaches a process for
recrystallizing laquinimod sodium which produces a mixture of
crystalline laquinimod sodium having larger crystals, lowered
impurity levels and certain improved crystalline characteristics as
compared to the laquinimod sodium crystals known at the time.
[0376] Specifically, the process of U.S. Pat. No. 7,884,208
achieves a mixture of crystalline laquinimod sodium having (i) a
mixture wherein 10% or more of the total amount by volume of the
laquinimod sodium particles has a size of greater than 40 microns
and wherein 50% or more of the total amount by volume of the
laquinimod sodium particles has a size of greater than 15 microns,
(ii) high density (tapped and bulk), (iii) low heavy metal content,
and (iv) low content of certain polar impurities.
[0377] However, the recrystallization process of U.S. Pat. No.
7,884,208 (Examples 13-17, Tables 1-4) does not produce a mixture
of recystallized laquinimod sodium particles wherein 90% or more of
the total amount by volume of the laquinimod sodium have a size of
40 microns or less, or 50% or more of the total amount by volume of
the laquinimod sodium has a size of 15 microns or less.
[0378] Likewise, the recrystallization process of U.S. Pat. No.
7,884,208 does not produce a mixture of recystallized laquinimod
sodium particles wherein 90% or more of the total amount by volume
of the laquinimod sodium have a size of less than 40 microns,
wherein 50% or more of the total amount by volume of the laquinimod
sodium has a size of less than 15 microns, and wherein 10% or more
of the total amount by volume of the laquinimod sodium has a size
of less than 5 microns.
[0379] Example 14 of U.S. Pat. No. 7,884,208 produced a mixture of
recystallized laquinimod sodium particles wherein 10% or more of
the total amount by volume of the laquinimod sodium have a size of
less than 5 microns. However, this Example also shows reduced
quality of crystalline characteristics, specifically Tapped
Density. The mixture produced by Example 14 has an acceptable
D(0.1) value but an undesired Tapped Density.
[0380] Conversely, Example 13 of U.S. Pat. No. 7,884,208 produced a
mixture having high Tapped Density, but did not produce crystals
wherein 90% or more of the total amount by volume of the laquinimod
sodium particles have a size of 40 microns or less or 50% or more
of the total amount by volume of the laquinimod sodium particles
have a size of 15 microns or less.
[0381] Importantly, U.S. Pat. No. 7,884,208 is unable to achieve
the advantages of recrystallization, i.e., better density and
impurity profiles, while also producing laquinimod sodium crystals
wherein 90% or more of the total amount by volume of the laquinimod
sodium particles have a size of 40 microns or less, or 50% or more
of the total amount by volume of the laquinimod sodium particles
have a size of 15 microns or less.
[0382] U.S. Pat. No. 7,884,208, by way of Example 1 (batches A, B
and C) and Tables 1-3, also teaches that the process disclosed in
U.S. Pat. No. 6,077,851 results in a mixture of crystalline
laquinimod sodium having all four of (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
40 microns or less, or 50% or more of the total amount by volume of
the laquinimod sodium particles have a size of 15 microns or less,
(ii) poor density (tapped and bulk), (iii) high heavy metal
content, and (iv) high polar impurity content. Importantly,
although U.S. Pat. No. 6,077,851 achieves a mixture wherein 90% or
more of the total amount by volume of the laquinimod sodium
particles have a size of 40 microns or less or, 50% or more of the
total amount by volume of the laquinimod sodium particles have a
size of 15 microns or less, but it does not achieve crystals having
acceptable density or low levels of impurities.
[0383] The prior art contains no teaching of a process for
preparing laquinimod sodium wherein 90% or more of the total amount
by volume of the laquinimod sodium particles have a size of 40
microns or less, or 50% or more of the total amount by volume of
the laquinimod sodium particles have a size of 15 microns or less,
and having desirable density and purity profiles.
[0384] The present invention provides a process which is capable of
producing a mixture of recystallized laquinimod sodium crystals
wherein 90% or more of the total amount by volume of the laquinimod
sodium particles have a size of 40 microns or less or (ii) 50% or
more of the total amount by volume of the laquinimod sodium
particles have a size of 15 microns or less, and having desirable
density and purity profiles, specifically, (i) high density (tapped
or bulk), (ii) low heavy metal content, or (iii) low polar impurity
content.
[0385] The mixture of laquinimod sodium provided by the present
invention is achieved by an improved recrystallization process.
Improved Recrystallization Process
[0386] The laquinimod sodium manufactured by the recrystallization
processes of the present invention has improved purity and density
profiles over the laquinimod sodium disclosed in U.S. Pat. No.
6,077,851 and improved crystalline characteristics, especially
smaller particles, over U.S. Pat. No. 7,884,208.
[0387] The modified recrystallization process of the present
invention unexpectedly results in different recrystallization
conditions than achieved by the process disclosed in U.S. Pat. No.
7,884,208 and, thusly, results in different products. Specifically,
concentrating the aqueous solution to 1.7-1.8 unexpectedly results
in crystalline laquinimod sodium particles having reduced levels of
impurities, improved crystalline characteristics, and wherein 90%
or more of the total amount by volume of the laquinimod sodium
particles have a size of 40 microns or less, 50% or more of the
total amount by volume of the laquinimod sodium particles have a
size of 15 microns or less, and 10% or more of the total amount by
volume of the laquinimod sodium particles have a size of 5 microns
or less.
[0388] Without being limited to any one particular theory, an
important factor affecting crystallization is initial concentration
of crystallizing solution. Careful control of concentration of the
solution can facilitate initiation of spontaneous crystallization
prior to the end of the concentration step. The process of the
present invention can initiate spontaneous crystallization prior to
addition of acetone.
[0389] Concentrating the aqueous solution to form a concentrated
solution comprising approximately 1.7-1.8 ml of water per gram of
laquinimod sodium is an important aspect of the present
invention.
Improved Mixture of Crystalline Laquinimod Sodium
[0390] U.S. Pat. No. 7,884,208 teaches advantages associated with
large particles. Specifically, U.S. Pat. No. 7,884,208 teaches that
larger particles of laquinimod sodium are more "processable" when
making pharmaceutical compositions and that smaller particles are
often associated with dust-like properties which may interfere with
processing, and sometimes associated with flowability problems
which may interfere with manufacturing. Further, U.S. Pat. No.
7,884,208 teaches that chemical stability has been shown to be
decreased by the increase in surface area that results from smaller
particle size. (Felmeister, A. Chpt 88, Remington's Pharmaceutical
Sciences, 15.sup.th Edition, Mack Publishing Company, Easton, Pa.
(1975)).
[0391] However, the subject invention has unexpectedly achieved an
improved mixture of laquinimod sodium wherein (i) 90% or more of
the total amount by volume of the laquinimod sodium particles have
a size of 40 microns or less, (ii) 50% or more of the total amount
by volume of the laquinimod sodium particles have a size of 15
microns or less, and (iii) 10% or more of the total amount by
volume of the laquinimod sodium particles have a size of 5 microns
or less.
[0392] Laquinimod has been graded as a highly potent API, demanding
special caution and avoiding material contact with workers and
environment. Laquinimod has been graded as having the highest
potency rate, corresponding to a recommended acceptable daily
intake (ADI) during operations and manufacturing of less than 0.01
mg/day or <1 .mu.g/m.sup.3 as an 8-hour TWA. High potency
compounds are associated with controls, whether engineering,
administrative or procedure-related, that afford the desired level
of worker protection. For example, high potency compounds may
require no human intervention or manual operations. (Bruce D.
Naumann, Control Banding In The Pharmaceutical Industry,
http://www.aioh.org.au/downloads/documents/ControlBandingBNaumaun.pdf)
[0393] Importantly, although the mixture of the subject invention
may be milled or unmilled, the present invention is achieved
without the need of a milling operation.
[0394] The mixture of laquinimod sodium of the present invention
overcomes the potential problems associated with mixtures having
large particles. With respect to processing and manufacturing, the
small size of the laquinimod sodium particles of the present
invention may obviate the need for milling and comminution steps.
U.S. Pat. No. 7,884,208 reflected the understanding in the art that
there are problems associated with small particles in
pharmaceutical compositions, but the laquinimod sodium of the
present invention has no problems associated with stability,
processing or manufacturing.
[0395] Comminution introduces its own set of problems to a drug
substance beyond the disadvantages of introducing an additional
drug processing step. For example, milling can introduce
impurities, new polymorphs, amorphous sections in the crystalline
structure of the API, other changes to particle morphology,
differences in agglomeration, increased solubility, changes in
moisture levels, and changes in compressibility (Hausner, "The Role
of Particle Size in the Development of Generic Products" 2003). As
a result, comminution may affect the efficacy and safety of a drug
substance. Some of the disadvantages of comminution are illustrated
by the side effects resulting from microcrystalline Nitrofurantoin
compared to macrocrystalline Nitrofurantoin. (Brumfitt, W. and J.
M. T. Hamilton-Miller, J. Antimicrobial Chemotherapy 42:363:371
(1998)).
[0396] Accordingly, it is advantageous to produce a drug substance
which does not contain large particles to avoid the inefficiencies
of additional process steps such as milling or sieving. The
laquinimod sodium of the present invention provides a mixture of
laquinimod sodium particles having small particle sizes which
avoids safety problems and additional problems related to
milling.
[0397] Another concern during formulation processes is maintaining
uniformity of content of the drug product. In the case of
laquinimod, the unit dose of laquinimod is quite low relative to
the total weight of the drug product, e.g., tablet or capsule. A
typical formulation, for example, may comprise only a small amount
of laquinimod, e.g., 0.3, 0.6 or 1.2 mg, in a capsule with total
weight of over 200 mg. As such, small fluctuation in the amount of
laquinimod due to problems of flowability, segregation, uniformity,
or poor homogenous distribution could result in a large percent
deviation from the desired amount, e.g., 0.3, 0.6 or 1.2 mg. The
mixture of laquinimod sodium of the present invention provides high
uniformity of content and minimal fluctuations in the amount of
laquinimod in the capsules.
TABLE-US-00001 TABLE 1 Uniformity of Laquinimod Blend (UoB) and
Uniformity of the content of filled capsules (UoC) Batch UoB AVG
UoB RSD UoC UoC AV a 99.5 1.67 99.3 4.5 b 100.6 0.61 98.7 2.7 c
102.6 1.10 99.9 2.2 d 97.3 0.90 99.4 4.7 e 101.3 1.02 100.6 2.4 f
101.3 1.02 100.8 3.6 g 97.4 0.85 96.4 5.9 h 101.6 1.08 98.6 2.8 i
98.5 0.71 94.2 8.6 j (0.3 mg) 101.2 0.66 96.9 8.7 k 101.2 0.66 94.6
7.1 1 97.1 1.35 96.8 5 m 95.3 0.88 98.6 3.5 n 98.6 0.91 101.4 5.2 o
97.1 0.59 96.6 5.7 p 98.8 0.58 98.4 5.3 q 98.2 0.54 98.5 4.3 r 97.2
0.96 100.7 2.4 s 102.2 1.20 100.2 4.6 t 102.9 1.82 98.4 3.3 u 103.6
1.01 99.7 4.4 v 97.9 0.5 98.5 1.4 w 98.7 0.5 99.2 1.6 xA 95.3 0.7
94.2 12.1 xB y 97.1 1.2 98 z1 (0.6 mg) 99.8 1.8 97.4 2.2 z2 (0.6
mg) 99.8 1.8 97.4 2.2 UoB: Uniformity of Blend, RSD is the
parameter to describe the uniformity before filling into capsules;
UoC: Uniformity of the content of the filled capsule, AV is the
acceptance value which is related to RSD to uniformity.
[0398] Uniformity of the shape of laquinimod particles is also an
important concern during formulation as a lack of uniformity of
shape can cause variation in density of drug substance and cause
problems during drug product formation, e.g., capsule or tablet
formation. Crystalline laquinimod sodium particles are rod-shaped
particles. It is known that milling operations may result in
changes to particle shape.
[0399] Decreased particle size is known to result in faster
dissolution profiles. The rate of dissolution of small particles is
usually faster than that of large particles because a greater
surface area of the drug substance is in contact with the liquid
medium. When formulating a drug with a low dissolution rate, it is
desirable to decrease particle size in order to increase
dissolution and thus facilitate rapid gastrointestinal or oral
absorption.
[0400] In such cases where drug substances have no recognized
problems associated with dissolution rate, particle size reduction
may be inadvisable and even deleterious. Increasing surface area
can increase degradation rates of the drug substance. As discussed,
for example, in U.S. Pat. Nos. 8,178,127 and 7,989,473, laquinimod
sodium is susceptible to degradation.
[0401] Unexpectedly, in spite of known disadvantages associated
with small particle sizes, it was found that an improved drug
substance and drug product resulted from a mixture of crystalline
laquinimod sodium particles wherein (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
40 microns or less, (ii) 50% or more of the total amount by volume
of the laquinimod sodium particles have a size of 15 microns or
less, and (iii) 10% or more of the total amount by volume of the
laquinimod sodium particles have a size of 5 microns or less.
[0402] Accordingly, an advantage of the recrystallization process
of the present invention is that the resulting mixture of
crystalline laquinimod sodium has particles having small particle
sizes, which is associated with high uniformity and homogeneity
with respect to distribution of the API into capsules, tablets and
other drug products. Laquinimod sodium crystals having small
particle sizes can obviate or reduce the need for additional
milling steps. The small particle sizes of the laquinimod sodium of
the present invention are achieved without sacrificing desirable
purity or density profiles and without the need for prior milling
operations.
[0403] Another advantage of the present invention is that
laquinimod sodium crystals have a higher density than the
laquinimod sodium crystals produced by the slurry-to-slurry process
of U.S. Pat. No. 6,077,851. Low tapped density is anathema to
certain prized qualities in a drug substance or drug product such
as compressibility, the ability of a powder to decrease in volume
under pressure, and compactability, the ability of a powder to be
compressed into a tablet of certain strength or hardness. Crystals
with low tapped density are also known to have poor flowability,
which results in a lack of uniformity of content in finished dosage
forms, especially in tablets. (Rudnic et al. Chpt. 45, Remington's
Pharmaceutical Sciences, 20.sup.th Edition, Lippincott Williams
& Wilkins, Baltimore, Md. (2000)) Uniformity of content is
especially important for pharmaceutical compositions comprising a
potent drug substance, e.g., Laquinimod sodium.
[0404] Compared to the slurry-to-slurry process of U.S. Pat. No.
6,077,851, the present invention also shows low aggregation of the
particles and, additionally, provides particles with acceptable
density and lower levels of impurities. As shown in FIGS. 5-8, the
crude laquinimod resulting from the process described in U.S. Pat.
No. 6,077,851 has a high rate of aggregates (FIGS. 5 and 6),
compared to a low rate of aggregates of the present invention
(FIGS. 7 and 8).
[0405] Another advantage of the present invention is that the
process of the present invention is environmentally friendly
without sacrificing desirable crystalline characteristics.
Specifically, by use of water as the primary solvent, the present
invention achieves both environmental friendliness and improved
crystalline characteristics, specifically with respect to particle
size distribution over U.S. Pat. No. 7,884,208.
Terms
[0406] As used herein, and unless stated otherwise, each of the
following terms shall have the definition set forth below.
[0407] As used herein, "laquinimod" means laquinimod acid or a
pharmaceutically acceptable salt thereof, including laquinimod
sodium.
[0408] As used herein, "laquinimod acid" is
N-ethyl-N-phenyl-1,2,-dihydro-4-hydroxy-5-chloro-1-methyl-2-oxoquinoline--
3-carboxamide, and its Chemical Registry number is 248281-84-7.
"Laquinimod sodium" is the sodium salt of laquinimod acid.
[0409] As used herein, D(0.1) is the particle size, in microns,
below which 10% by volume distribution of the population is
found.
[0410] As used herein, D(0.5) is the particle size, in microns,
below which 50% by volume distribution of the population is
found.
[0411] As used herein, D(0.9) is the particle size, in microns,
below which 90% by volume distribution of the population is
found.
[0412] As used herein, "crystalline characteristics" includes
particle size distribution, bulk density and tapped density.
[0413] As used herein, "drug substance" refers to the active
ingredient in a drug product or for use in a drug product, which
provides pharmacological activity or other direct effect in the
diagnosis, cure, mitigation, treatment, or prevention of disease,
or to affect the structure or any function of the body of man or
animals.
[0414] As used herein, "drug product" refers to the formulated or
finished dosage form containing the drug substance as well as at
least one pharmaceutically acceptable carrier.
[0415] As used herein, a composition that is "free" of a chemical
entity means that the composition contains, if at all, an amount of
the chemical entity which cannot be avoided following an
affirmative act intended to purify the composition by separating
the chemical entity from the composition.
[0416] For example, a composition which is "free" of an alkalizing
agent means that the alkalizing agent, if present at all, is a
minority component of the composition by weight. Preferably, when a
composition is "free" of a component, the composition comprises
less than 0.1 wt %, 0.05 wt %, 0.02 wt %, or 0.01 wt % of the
component.
[0417] As used herein, an "isolated" compound is a compound
isolated from the crude reaction mixture following an affirmative
act of isolation. The act of isolation involves separating the
compound from the other known components of the crude reaction
mixture, with some impurities, unknown side products and residual
amounts of the other known components of the crude reaction mixture
permitted to remain. Purification is an example of an affirmative
act of isolation.
[0418] As used herein, "stability testing" refers to tests
conducted at specific time intervals and various environmental
conditions (e.g., temperature and humidity) to see if and to what
extent a drug product degrades over its designated shelf life time.
The specific conditions and time of the tests are such that they
accelerate the conditions the drug product is expected to encounter
over its shelf life. For example, detailed requirements of
stability testing for finished pharmaceuticals are codified in 21
C.F.R .sctn.211.166, the entire content of which is hereby
incorporated by reference.
[0419] As used herein, "dissolution rate" is determined based on
the amount of drug substance dissolved in 30 min. as indicated in
the U.S. Pharmacopeia <711>.
[0420] As used herein, "atmospheric pressure" refers to a pressure
of about 1 atm.
[0421] As used herein, "ambient temperature" refers to a
temperature of about 20.degree. C. to about 30.degree. C.
[0422] As used herein, "about" in the context of a numerical value
or range means .+-.10% of the numerical value or range recited or
claimed.
[0423] As used herein, "approximately" in the context of a
numerical value or range means .+-.5% of the numerical value or
range recited or claimed.
[0424] The term "stable pharmaceutical composition" as used herein
in connection with the composition according to the invention
denotes a composition, which preserves the physical
stability/integrity and/or chemical stability/integrity of the
active pharmaceutical ingredient during storage.
[0425] Furthermore, "stable pharmaceutical composition" is
characterized by its level of degradation products not exceeding 5%
at 40.degree. C./75% RH after 6 months or 3% at 55.degree. C./75%
RH after two weeks, compared to their level in time zero.
[0426] As used herein, "treating" and "treatment" encompasses,
e.g., inducing inhibition, regression, or stasis of a disease,
disorder or condition, or ameliorating or alleviating a symptom of
a disease, disorder or condition. "Ameliorating" or "alleviating" a
condition or state as used herein shall mean to relieve or lessen
the symptoms of that condition or state. "Inhibition" of disease
progression or disease complication in a subject as used herein
means preventing or reducing the disease progression and/or disease
complication in the subject.
[0427] "Administering to the subject" means the giving of,
dispensing of, or application of medicines, drugs, or remedies to a
subject to relieve, cure, or reduce the symptoms associated with a
condition, e.g., a pathological condition.
[0428] The drug substance of the present invention, e.g.,
laquinimod sodium, may be administered in admixture with suitable
pharmaceutical diluents, extenders, excipients, or carriers
(collectively referred to herein as a pharmaceutically acceptable
carrier) suitably selected with respect to the intended form of
administration and as consistent with conventional pharmaceutical
practices. The unit will be in a form suitable for oral, rectal,
topical, intravenous or direct injection or parenteral
administration. The compounds can be administered alone or mixed
with a pharmaceutically acceptable carrier.
[0429] This carrier can be a solid or liquid, and the type of
carrier is generally chosen based on the type of administration
being used. The active agent can be co-administered in the form of
a tablet or capsule, liposome, as an agglomerated powder or in a
liquid form. Examples of suitable solid carriers include lactose,
sucrose, gelatin and agar. Capsules or tablets can be easily
formulated and can be made easy to swallow or chew; other solid
forms include granules, and bulk powders.
[0430] Capsules or tablets may contain suitable binders,
lubricants, disintegrating agents, diluents, coloring agents,
flavoring agents, flow-inducing agents, and melting agents. For
instance, for oral administration in the dosage unit form of a
tablet or capsule, the active drug component can be combined with
an oral, non-toxic, pharmaceutically acceptable, inert carrier such
as lactose, gelatin, agar, starch, sucrose, glucose, methyl
cellulose, magnesium stearate, dicalcium phosphate, calcium
sulfate, mannitol, sorbitol and the like.
[0431] Suitable binders include starch, gelatin, natural sugars
such as glucose or beta-lactose, corn sweeteners, natural and
synthetic gums such as acacia, tragacanth, or sodium alginate,
povidone, carboxymethylcellulose, polyethylene glycol, waxes, and
the like.
[0432] Lubricants used in these dosage forms include sodium oleate,
sodium stearate, magnesium stearate, sodium benzoate, sodium
acetate, sodium chloride, stearic acid, sodium stearyl fumarate,
talc and the like.
[0433] Disintegrators include, without limitation, starch, methyl
cellulose, agar, bentonite, xanthan gum, croscarmellose sodium,
sodium starch glycolate and the like.
[0434] Specific examples of the techniques, pharmaceutically
acceptable carriers and excipients that may be used to formulate
oral dosage forms of the present invention are described, e.g., in
U.S. Patent Application Publication No. 2005/0192315, PCT
International Application Publication Nos. WO 2005/074899, WO
2007/047863, and WO 2007/146248, and in U.S. Pat. No. 7,589,208.
For example, the oral dosage form of the present invention may
comprise an alkaline-reacting component, said component preferably
amounting from about 1 to 20% by weight of the formulation in order
to keep the pH above 8.
[0435] Techniques and compositions for making dosage forms useful
in the present invention are described in the following references:
7 Modern Pharmaceutics, Chapters 9 and 10 (Banker & Rhodes,
Editors, 1979); Pharmaceutical Dosage Forms: Tablets (Lieberman et
al., 1981); Ansel, Introduction to Pharmaceutical Dosage Forms 2nd
Edition (1976); Remington's Pharmaceutical Sciences, 17th ed. (Mack
Publishing Company, Easton, Pa., 1985); Advances in Pharmaceutical
Sciences (David Ganderton, Trevor Jones, Eds., 1992); Advances in
Pharmaceutical Sciences Vol. 7. (David Ganderton, Trevor Jones,
James McGinity, Eds., 1995); Aqueous Polymeric Coatings for
Pharmaceutical Dosage Forms (Drugs and the Pharmaceutical Sciences,
Series 36 (James McGinity, Ed., 1989); Pharmaceutical Particulate
Carriers: Therapeutic Applications: Drugs and the Pharmaceutical
Sciences, Vol 61 (Alain Rolland, Ed., 1993); Drug Delivery to the
Gastrointestinal Tract (Ellis Horwood Books in the Biological
Sciences. Series in Pharmaceutical Technology; J. G. Hardy, S. S.
Davis, Clive G. Wilson, Eds.); Modem Pharmaceutics Drugs and the
Pharmaceutical Sciences, Vol 40 (Gilbert S. Banker, Christopher T.
Rhodes, Eds.). All of the aforementioned publications are
incorporated by reference herein. Examples of suitable liquid
dosage forms include solutions or suspensions in water,
pharmaceutically acceptable fats and oils, alcohols or other
organic solvents, including esters, emulsions, syrups or elixirs,
suspensions, solutions and/or suspensions reconstituted from
non-effervescent granules and effervescent preparations
reconstituted from effervescent granules. Such liquid dosage forms
may contain, for example, suitable solvents, preservatives,
emulsifying agents, suspending agents, diluents, sweeteners,
thickeners, and melting agents.
[0436] Oral dosage forms optionally contain flavorants and coloring
agents. Parenteral and intravenous forms may also include minerals
and other materials to make them compatible with the type of
injection or delivery system chosen.
[0437] The compounds used in the method of the present invention
may also be administered in the form of liposome delivery systems,
such as small unilamellar vesicles, large unilamallar vesicles, and
multilamellar vesicles. Liposomes can be formed from a variety of
phospholipids, such as cholesterol, stearylamine, or
phosphatidylcholines. The compounds may be administered as
components of tissue-targeted emulsions.
[0438] The compounds used in the method of the present invention
may also be coupled to soluble polymers as targetable drug carriers
or as a prodrug. Such polymers include polyvinylpyrrolidone, pyran
copolymer, polyhydroxylpropylmethacrylamide-phenol,
polyhydroxyethylaspartamidephenol, or polyethyleneoxide-polylysine
substituted with palmitoyl residues. Furthermore, the compounds may
be coupled to a class of biodegradable polymers useful in achieving
controlled release of a drug, for example, polylactic acid,
polyglycolic acid, copolymers of polylactic and polyglycolic acid,
polyepsilon caprolactone, polyhydroxy butyric acid,
polyorthoesters, polyacetals, polydihydropyrans, polycyanoacylates,
and crosslinked or amphipathic block copolymers of hydrogels.
[0439] For oral administration in liquid dosage form, the oral drug
components are combined with any oral, non-toxic, pharmaceutically
acceptable inert carrier such as ethanol, glycerol, water, and the
like. Examples of suitable liquid dosage forms include solutions or
suspensions in water, pharmaceutically acceptable fats and oils,
alcohols or other organic solvents, including esters, emulsions,
syrups or elixirs, suspensions, solutions and/or suspensions
reconstituted from non-effervescent granules and effervescent
preparations reconstituted from effervescent granules. Such liquid
dosage forms may contain, for example, suitable solvents,
preservatives, emulsifying agents, suspending agents, diluents,
sweeteners, thickeners, and melting agents.
[0440] Liquid dosage forms for oral administration can contain
coloring and flavoring to increase patient acceptance. In general,
water, a suitable oil, saline, aqueous dextrose (glucose), and
related sugar solutions and glycols such as propylene glycol or
polyethylene glycols are suitable carriers for parenteral
solutions. Solutions for parenteral administration preferably
contain a water soluble salt of the active ingredient, suitable
stabilizing agents, and if necessary, buffer substances.
Antioxidizing agents such as sodium bisulfite, sodium sulfite, or
ascorbic acid, either alone or combined, are suitable stabilizing
agents. Also used are citric acid and its salts and sodium EDTA. In
addition, parenteral solutions can contain preservatives, such as
benzalkonium chloride, methyl- or propyl-paraben, and
chlorobutanol. Suitable pharmaceutical carriers are described in
Remington's Pharmaceutical Sciences, Mack Publishing Company, a
standard reference text in this field.
[0441] The drug substance of the present invention, e.g.,
laquinimod sodium, may be administered in various forms, including
those detailed herein. The treatment with the compound may be a
component of a combination therapy or an adjunct therapy, i.e. the
subject or patient in need of the drug is treated or given another
drug for the disease in conjunction with one or more of the instant
compounds. This combination therapy can be sequential therapy where
the patient is treated first with one drug and then the other or
the two drugs are given simultaneously. These can be administered
independently by the same route or by two or more different routes
of administration depending on the dosage forms employed.
[0442] Gelatin capsules may contain the active ingredient compounds
and powdered carriers, such as lactose, starch, cellulose
derivatives, magnesium stearate, stearic acid, and the like.
Similar diluents can be used to make compressed tablets. Both
tablets and capsules can be manufactured as immediate release
products or as sustained release products to provide for continuous
release of medication over a period of hours. Compressed tablets
can be sugar coated or film coated to mask any unpleasant taste and
protect the tablet from the atmosphere, or enteric coated for
selective disintegration in the gastrointestinal tract.
[0443] The compounds used in the method of the present invention
may also be administered in intranasal form via use of suitable
intranasal vehicles, or via transdermal routes, using those forms
of transdermal skin patches well known to those of ordinary skill
in that art. To be administered in the form of a transdermal
delivery system, the dosage administration will generally be
continuous rather than intermittent throughout the dosage
regimen.
[0444] Parenteral and intravenous forms may also include minerals
and other materials to make them compatible with the type of
injection or delivery system chosen.
[0445] A dosage unit of the compounds used in the method of the
present invention may comprise a single compound or mixtures
thereof with additional antibacterial agents. The compounds can be
administered in oral dosage forms as tablets, capsules, pills,
powders, granules, elixirs, tinctures, suspensions, syrups, and
emulsions. The compounds may also be administered in intravenous
(bolus or infusion), intraperitoneal, subcutaneous, or
intramuscular form, or introduced directly, e.g. by injection,
topical application, or other methods, into or onto a site of
infection, all using dosage forms well known to those of ordinary
skill in the pharmaceutical arts.
[0446] A "dose" or "dosage unit" of laquinimod as measured in
milligrams refers to the milligrams of laquinimod acid present in a
preparation, regardless of the form of the preparation. A dosage
unit may comprise a single compound or mixtures of compounds
thereof. A dosage unit can be prepared for oral dosage forms, such
as tablets, capsules, pills, powders, and granules. For example,
the "dose" or "dosage unit" of laquinimod may be 0.3, 0.6, or 1.2
mg.
[0447] As used herein, a "pharmaceutically acceptable" component is
one that is suitable for use with humans and/or animals without
undue adverse side effects (such as toxicity, irritation, and
allergic response) commensurate with a reasonable benefit/risk
ratio.
[0448] As used herein, "pharmaceutically acceptable carrier" refers
to a carrier or excipient that is suitable for use with humans
and/or animals without undue adverse side effects (such as
toxicity, irritation, and allergic response) commensurate with a
reasonable benefit/risk ratio. It can be a pharmaceutically
acceptable solvent, suspending agent or vehicle, for delivering the
instant compounds to the subject. The carrier may be liquid or
solid and is selected with the planned manner of administration in
mind. "Pharmaceutically acceptable carrier" includes "fillers",
which fill out the size of a tablet or capsule, making it practical
to produce and convenient for the consumer to use. By increasing
the bulk volume, the fillers make it possible for the final product
to have the proper volume for patient handling. "Pharmaceutically
acceptable carrier" also includes "lubricants", which prevent
ingredients from clumping together and from sticking to the tablet
punches or capsule filling machine. Lubricants also ensure that
tablet formation and ejection can occur with low friction between
the solid and die wall. "Pharmaceutically acceptable carrier" also
includes inert carriers such as lactose, gelatin, agar, starch,
sucrose, glucose, methyl cellulose, dicalcium phosphate, calcium
sulfate, mannitol, sorbitol, microcrystalline cellulose and the
like. Liposomes are also a pharmaceutically acceptable carrier.
[0449] It is understood that where a parameter range is provided,
all integers within that range, and tenths and hundredth thereof,
are also provided by the invention. For example, "0.15-0.35%"
includes 0.15%, 0.16%, 0.17% etc. up to 0.35%.
[0450] The subject invention is also intended to include all
isotopes of atoms occurring on the compounds disclosed herein,
including impurities. Isotopes include those atoms having the same
atomic number but different mass numbers. By way of general example
and without limitation, isotopes of hydrogen include tritium and
deuterium. Isotopes of carbon including C-13 and C-14.
[0451] As used herein, "detection limit" for an analytical method
used in screening or testing for the presence of a compound in a
sample is a threshold under which the compound in a sample cannot
be detected by the analytical method used. The detection limits of
a given HPLC method for detecting an impurity in a sample
containing laquinimod may vary based on the method and the impurity
or impurities being detected. For example, the detection limit of
the typical HPLC method for detecting
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid (MCQCA) is 0.03% and the detecting limit of a given method for
detecting methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxyla-
te (MCQME), 5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline
(MCQ),
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
(MCQEE) and unknown impurities is 0.02%.
[0452] As used herein, "quantitation limit" for an analytical
method used in screening or testing for the presence of a compound
in a sample is a threshold under which the compound in a sample
cannot be quantified by the analytical method used. The
quantitation limits of a given HPLC method for detecting an
impurity in a sample containing laquinimod may vary based on the
impurity or impurities being detected.
[0453] As used herein, "density" is a measurement defined as the
mass of a substance per unit volume.
[0454] As used herein, "bulk density" or "BD" refers to a density
measurement of a loose, uncompacted substance, wherein the volume
of the substance includes the air trapped between particles.
[0455] As used herein, "tapped density" or "TD" refers to a density
measurement of a substance that has been tapped or vibrated, thus
minimizing the volume of the substance by eliminating or minimizing
the air trapped between particles.
[0456] As used herein, "rapid stirring" refers to stirring which
splashes solvent onto the walls of the vessel.
[0457] As used herein, "blend uniformity" refers to the homogeneity
of blend or granulate including laquinimod sodium particles prior
to encapsulation, tableting or otherwise finalizing the drug
product beyond formation of the final blend, and can represent
either one sample or the average of more than one sample. Blend
uniformity may be measured, for example, by taking 10 samples that
represent the upper, middle and lower layer of each batch of the
final blend, performing an HPLC assay to measure the amount of
active ingredient in the samples, and comparing the amount of
active ingredient in each sample to the labeled amount of active
ingredient. The standard deviation and relative standard deviation
can be determined based on the individual amounts of the tested
samples expressed as percentages of the labeled amount of drug
substance in each sample.
[0458] As used herein, "content uniformity" refers to the
homogeneity of the laquinimod sodium content among dosage forms,
e.g., capsules or tablets, after formulation. The uniformity of
dosage units by content uniformity of the pharmaceutical
composition described herein meets the U.S. Pharmacopeia
<905> Acceptance Value and range (as specified); L1=15.0 and
L2=25.0. Content uniformity may be measured, for example, as
indicated by the United States Pharmacopoeia which includes 1)
assaying ten tablets (or other dosage form of the drug product) to
ensure that the relative standard deviation (RSD) of active content
is less than or equal to 6.0% and no value is outside 85-115%; and
2) assaying twenty more tablets (or other dosage form of the drug
product) to ensure that the RSD for all thirty is less than or
equal to 7.8%, no more than one value is outside 85-115% and no
value is outside 75-125% of stated content.
[0459] As used herein, "residual solvents" include ethanol,
n-heptane, n-octane, methanol, acetone, dioxane, and dimethyl
formamide. Residual solvents may be determined, for example, based
on the manufacturer's statements of residual solvent levels in the
active ingredients/excipients and calculation as per U.S.
pharmacopeia <467> Option 2, product meets the USP
<467> Residual Solvents limit criteria. Testing is not
necessarily required.
[0460] As used herein, "NMT" means no more than.
[0461] As used herein, "LT" means less than.
[0462] As used herein, "MCQME" means methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate.
MCQME is disclosed in U.S. Pat. No. 7,560,557 and J. Org. Chem.,
2006, 71, 1658-1667, the contents of which are incorporated by
reference into this application. MCQME has the structure:
##STR00009##
[0463] As used herein, "MCQEE" means ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate.
MCQEE is disclosed in U.S. Pat. No. 7,560,557, the contents of
which are incorporated by reference into this application. MCQEE
has the structure:
##STR00010##
[0464] As used herein, "MCQ" means
5-chloro-4-hydroxy-1-methylquinolin-2(1H)-one. MCQ is disclosed in
U.S. Pat. No. 7,560,557 and J. Org. Chem., 2006, 71, 1658-1667, the
contents of which are incorporated by reference into this
application. MCQ has the structure:
##STR00011##
[0465] As used herein, "MCQCA" means
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid. MCQCA is disclosed in U.S. Pat. No. 7,560,557 and J. Org.
Chem., 2006, 71, 1658-1667, the contents of which are incorporated
by reference into this application. MCQCA has the structure:
##STR00012##
[0466] As used herein, "NEA" means N-ethyl aniline. NEA is
disclosed in U.S. Pat. No. 7,560,557, the contents of which are
incorporated by reference into this application. NEA has the
structure:
##STR00013##
[0467] As used herein, "5-HLAQ" means
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide. 5-HLAQ is disclosed in PCT International Application No.
PCT/US13/26476 and U.S. Application Publication No. US 2013/0217724
A1, the contents of which are incorporated by reference into this
application. 5-HLAQ has the structure:
##STR00014##
[0468] As used herein, "3-HLAQ" means
5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-1,2,3,4-tetrahydro-
quinoline-3-carboxamide. 3-HLAQ is disclosed in PCT International
Application No. PCT/US2008/013890 and U.S. Pat. No. 8,178,127 B2,
the contents of which are incorporated by reference into this
application. 3-HLAQ has the structure:
##STR00015##
[0469] As used herein, "MEG-LAQ" means
N-ethyl-4-hydroxy-1-methyl-5-(methyl(2,3,4,5,6-pentahydroxyhexyl)amino)-2-
-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide. MEG-LAQ is
disclosed in U.S. Application Publication No. US 2013/0345256 A1,
the contents of which are incorporated by reference into this
application. MEG-LAQ has the structure:
##STR00016##
[0470] As used herein, "DELAQ" means
5-chloro-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxa-
mide. DELAQ is disclosed in PCT International Application No.
PCT/US2011/043391 and U.S. Application Publication No. US
2012/0010239 A1, the contents of which are incorporated by
reference into this application. DELAQ has the structure:
##STR00017##
[0471] As used herein, "SPIRO-LAQ" means
1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3'-[l]ethylindolin-[2]-
-one]. SPIRO-LAQ is disclosed in PCT International Application No.
PCT/US2008/013890 and U.S. Pat. No. 8,178,127 B2, the contents of
which are incorporated by reference into this application.
SPIRO-LAQ has the structure:
##STR00018##
[0472] As used herein, "BH-3-HLAQ" means
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid. BH-3-HLAQ has the structure:
##STR00019##
[0473] As used herein, "DMM" means dimethylmalonate. DMM is a
synthetic reagent, and has the structure:
##STR00020##
[0474] As used herein, "DMS" means Dimethyl sulfate. DMS is a
synthetic reagent.
[0475] As used herein, "MCIA" means
5-chloro-1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione. MCIA has the
structure
##STR00021##
[0476] As used herein, "DMF" means N,N-dimethylformamide.
[0477] As used herein, "LOD" means loss on drying.
[0478] The following commercially available solvents of appropriate
quality are also used as analytical standards for testing of drug
substance: Ethanol, n-heptane, n-octane, methanol, acetone,
1,4-dioxane and N,N-dimethylformamide (DMF). The ID number and
source of analytical standards for dimethyl malonate is S4695599
(Merck), sodium methyl sulfate is A0294777 (Arcos), and
13C2-dimethyl sulfate is P-5052 (Holland Moran).
[0479] By any range disclosed herein, it is meant that all
hundredth, tenth and integer unit amounts within the range are
specifically disclosed as part of the invention. Thus, for example,
0.01 mg to 50 mg means that 0.02, 0.03 . . . 0.09; 0.1, 0.2 . . .
0.9; and 1, 2 . . . 49 mg unit amounts are included as embodiments
of this invention.
[0480] Impurities are measured by common pharmacopeial methods
unless otherwise specified.
[0481] As used herein, an "anti-solvent" is a solvent in which
laquinimod sodium is slightly soluble, very slightly soluble,
practically insoluble, or insoluble at room temperature
(20-25.degree. C.). The solubility terms are defined below, in
accordance with the United States Pharmacopoeia XXV.
TABLE-US-00002 Term Parts of solvent required for 1 part solute
Slightly soluble From 100 to 1000 Very slightly soluble From 1000
to 10,000 Practically insoluble 10,000 and over Insoluble 10,000
and over
[0482] The purification of impure crystalline compounds is usually
attained by recrystallization from a suitable solvent or mixture of
solvents. (Vogel's Textbook of Practical Organic Chemistry. 5'
edition. Longman Scientific & Technical, 1989.) The
recrystallization process generally comprises the following steps:
a) dissolving the impure crystalline substance in a suitable
solvent near the boiling point; b) filtering the hot solution from
particles of insoluble material and dust; c) allowing the hot
solution to cool to cause the dissolved substance to crystallize
out; and d) separating the crystals from the supernatant solution.
(Id.) However, standard recrystallization techniques were
accompanied by low or no yields when applied to laquinimod sodium
as taught in U.S. Pat. No. 7,884,208. As shown in Examples 2-7 of
U.S. Pat. No. 7,884,208, attempts to recrystallize laquinimod
sodium using standard recrystallization procedures resulted in poor
yields, if any. The process of U.S. Pat. No. 7,884,208 overcomes
the difficulties associated with recrystallizing laquinimod sodium
by use of an anti-solvent in which laquinimod sodium is practically
insoluble. In addition, the process of U.S. Pat. No. 7,884,208
concentrates the laquinimod sodium aqueous solution before the
addition of the anti-solvent. The process of the present invention
is an improvement over the process of U.S. Pat. No. 7,884,208.
[0483] This invention will be better understood by reference to the
Experimental Details and Examples which follow, but those skilled
in the art will readily appreciate that the specific experiments
detailed are only illustrative of the invention as described more
fully in the claims which follow thereafter.
EXPERIMENTAL DETAILS
Determination of Powder Density
Bulk Density
[0484] 1. Mix powder; [0485] 2. Tare a 50 ml empty cylinder on a
0.01 g sensitivity balance; [0486] 3. Transfer the powder, without
compacting, to the cylinder being held at approximately a 45 degree
angle to achieve an untapped apparent volume of 40 to 50 ml. [0487]
4. Bring the cylinder containing the sample to a vertical position
by a sharp move in order to level the volume for reading. [0488] 5.
Read the apparent volume (Va) to the nearest graduated unit; [0489]
6. Weigh the cylinder with sample (the balance gives sample weight
M); [0490] 7. Calculate bulk density in g/ml according to the
following equation: BD=M/Va; [0491] 8. Perform steps 1-7 again and
report the average data of duplicates.
Tapped Density
[0491] [0492] 1. Put the same cylinder used to calculate Bulk
Density in a Quantachrome Dual Autotap instrument; [0493] 2.
Perform 1250 taps; [0494] 3. Read the tapped volume (Vf) to the
nearest graduated unit; [0495] 4. Calculate the tapped density in
g/ml according to the following equation: TD=M/Vf; [0496] 5.
Perform steps 1-4 again and report the average data of
duplicates.
Determination of Particle Size
[0497] The particle size distributions were measured by Malvern
Laser Diffraction, using the Mastersizer S model. Laser diffraction
relies on the fact that diffraction angle of light is inversely
proportional to particle size. Properties of particles are measured
and interpreted as measurements of a sphere (a sphere being the
only shape that can be described by one unique number). In
addition, laser diffraction calculates a particle size distribution
based around volume terms, thus eliminating particle count from the
determination of particle size. The Mastersizer S model measures
particles using a single technique and a single range setting.
[0498] D(0.1) is the particle size, in microns, below which 10% by
volume distribution of the population is found. D(0.5) is the
particle size, in microns, below which 50% by volume distribution
of the population is found. D(0.9) is the particle size, in
microns, below which 90% by volume distribution of the population
is found.
Determination of Heavy Metals
[0499] Metal content was measured using inductively coupled plasma
atomic emission spectrometry using an inductively coupled plasma
atomic emission spectrometry ("ICP-AES") system manufactured by
Spectro (Kleve, Germany). Sample digestion was performed in 65%
nitric acid, and the internal standard used was scandium.
[0500] Note: In the following examples the volumes of solvents used
are calculated relative to starting weight of laquinimod sodium.
The yields are calculated in weight percent.
Determination of Purity--an Exemplary HPLC Procedure for
Identification and Determination of Assay and Polar
Impurities/Degradation Products in Laquinimod Capsules
[0501] Laquinimod sodium and polar impurity/degradation products
were determined by isocratic reversed phase high performance liquid
chromatography (RP-HPLC), using an ODS-3V column and a mobile phase
comprised of a mixture of ammonium acetate buffer at pH 7.0 (80%)
and acetonitrile (20%). The detection technique was ultraviolet
absorption at 240 nm.
Specific HPLC Conditions:
[0502] Column & Packing: Inertsil ODS-3V, 5 .mu.m,
4.6.times.250 mm, GL Sciences Inc. [0503] Guard column: Opti-Guard
C 18, 1.times.10 mm [0504] Mobile phase: Acetonitrile:Buffer pH
7.0-20:80 (v/v). Mix and degas [0505] Buffer pH 7.0 preparation:
Dissolve 7.7 g Ammonium acetate in 2000 mL water and adjust to pH
7.0.+-.0.05 with aqueous ammonia or glacial acetic acid. Filter
through a 0.45 .mu.m membrane filter. [0506] Flowrate: 15 mL/min
[0507] Detection: UV at 240 nm [0508] Injection volume: 50 .mu.L
[0509] Diluent A: Acetonitrile/Water--50:50 (v/v) [0510] Diluent B
(and blank): Mobile Phase [0511] Column temperature: 40.degree. C.
[0512] Autosampler temperature: 5.degree. C. [0513] Run time: 40
minutes
Typical HPLC Procedure:
1. Standard Solutions Preparation
1.1 Laquinimod Standard Stock Solution (Solution S)
[0514] Weigh accurately in duplicate about 15 mg of laquinimod
sodium standard into a 50 mL volumetric flask. Dilute with diluents
A up to 2/3 of the volume, sonicate for 2 minutes in a cold
sonication bath and dilute to volume with diluents A.
[0515] Concentration of standard stock solution is about 300
.mu.g/mL laquinimod sodium. Standard stock solution may be used for
one month when stored in a refrigerator 2.degree. C.-8.degree.
C.
1.2 Laquinimod Standard Working Solution for Assay (Solution A)
[0516] Dilute 3 mL of the Standard Stock Solution to 10 mL with
diluents B (dilution factor 3.33).
[0517] Concentration of Laquinimod sodium is about 90 .mu.g/mL.
Concentration expressed as laquinimod (acid) is about 85
.mu.g/mL.
[0518] Standard working solution A may be used for 7 days when
stored in a refrigerator (2.degree. C.-8.degree. C.).
1.3 MCQCA Standard Stock Solution
[0519] Weigh accurately about 18 mg of MCQCA standard into a 100 mL
volumetric flask. Dilute to volume with acetonitrile, sonicate (in
a cold sonication bath) until the substance is completely
dissolved--stock MCQCA solution.
[0520] Concentration of MCQCA is about 180 .mu.g/mL.
[0521] MCQCA Stock standard solution should be freshly
prepared.
1.4 Standard Solution for Determination of Impurities (Solution
1)
[0522] Prepare a solution in diluents B, containing Laquinimod in a
concentration of 0.2% and MCQCA--in a concentration of 0.1%, with
respect to the working concentration of Laquinimod in Standard
solution A. As an example, apply the following procedure.
[0523] Transfer 4.0 mL of laquinimod sodium standard solution for
assay (Solution A) and 1.0 mL of MCQCA stock standard solution to a
100 mL volumetric flask and dilute to volume with the diluents B
(intermediate dilution).
[0524] Place 2.5 mL of this intermediate dilution into a 50 mL
volumetric flask and make up to volume with diluents B.
[0525] Total dilution factor for laquinimod standard is 1666.67,
for MCQCA 2000.
[0526] Concentration of laquinimod sodium is about 0.18 .mu.g/mL
(0.2%)
[0527] Concentration of MCQCA is about 0.09 .mu.g/mL (0.1%, QL
level).
[0528] Standard solution I may be used for 24 hours when stored in
a refrigerator.
2. Resolution Solutions Preparation
2.1 Mixed Solution
[0529] Prepare solution containing the following potential
impurities standards (markers) using the Diluent A as a
solvent:
Mixed Solution:
[0530] MCQ: 5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one
[0531] MCQCA:
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid
[0532] MCQMA: Methyl
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate
[0533] 5-HLAQ:
N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide
[0534] The Mixed Solution may be prepared as follows:
[0535] Weigh about 3 mg of each impurity standard/marker into a 100
mL volumetric flask, dissolve (sonication is acceptable) and dilute
to volume with the Diluent A.
[0536] Concentration of each impurity in the Mixed Solution is
about 30 .mu.g/mL. Mixed Solution may be used for up to 4 months
when stored frozen at about -20.degree. C. For this purpose, the
freshly prepared Mixed Solution should be divided into aliquots,
immediately frozen and stored at -20.degree. C. After thawing, the
aliquots should be mixed well and should not be refrozen.
2.2 Stock Solutions of Additional Impurities
[0537] Weigh about 3 mg of MCQEE (Ethyl
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate)
into a 100 mL volumetric flask, dissolve (sonication is acceptable)
and dilute to volume with the Diluent A. This is MCQEE Stock
solution.
[0538] This solution may be used for up to 4 months when stored
frozen at about -20.degree. C.
[0539] For this purpose, the freshly prepared MCQEE Stock Solution
should be divided into aliquot, immediately frozen and stored at
-20.degree. C. After thawing, the aliquots should be mixed well and
should not be refrozen.
[0540] Weigh about 3 mg of MEG-LAQ (Meglumine Adduct of Laquinimod)
into a 100 mL volumetric flask, dissolve (sonication is acceptable)
and dilute to volume with the Diluent A. This is MEG-LAQ Stock
solution.
[0541] This solution may be used for one week when stored in
refrigerator (2.degree. C.-8.degree. C.).
2.3 Resolution Solutions
[0542] Prepare two Resolution Solutions separately as follows,
using the Mixed Solution.
2.3.1 Resolution Solution 1
[0543] Transfer 3 mL of laquinimod standard stock solution
(solution S), 0.3 mL of the Mixed Solution and 0.3 mL of the MCQEE
Stock solution to a 10 mL volumetric flask and dilute to volume
with the Diluent B. This is Resolution Solution 1.
[0544] Concentration of laquinimod sodium in it is about 90
.mu.g/mL. Concentration of each impurity--is about 0.9 .mu.g/mL
(about 1% with respect to the working concentration of
laquinimod).
[0545] Resolution Solution 1 is used for resolution test (for
system suitability) and for determination of retention times
(RT)/relative retention times (RRT) of five impurities: MCQ, MCQCA,
MCQME, MCQEE, and 5-HLAQ.
[0546] Resolution Solution 1 may be used for 9 days if stored in a
refrigerator 2.degree. C.-8.degree. C.).
2.3.2 Resolution Solution 2
[0547] Transfer 3 mL of laquinimod standard stock solution
(solution S), 0.3 mL of the Mixed Solution and 0.3 mL of the
MEG-LAQ Stock solution to a 10 mL volumetric flask and dilute to
volume with the Diluent B. This is Resolution Solution 2.
[0548] Concentration of laquinimod sodium in it is about 90
.mu.g/mL. Concentration of each impurity--is about 0.9 .mu.g/mL
(about 1% with respect to the working concentration of
laquinimod).
[0549] Resolution Solution 2 is used for determination of retention
time of MEG-LAQ.
[0550] Resolution Solution 2 may be used for 9 days if stored in a
refrigerator 2.degree. C.-8.degree. C.).
3. Sample Solution Preparation
[0551] Weigh accurately 20 capsules and completely empty their
contents into a mortar. Pay attention to complete emptying of
capsule contents into the mortar, using spatula when necessary.
Weigh the empty capsules. Calculate the average weight of the
capsule contents. Mix and grind the capsule contents in a mortar
and keep the powder in a tightly closed container protected from
light.
[0552] Weigh accurately, in duplicate, the amount of powder
corresponding to 7 capsules, into a 50 mL volumetric flask.
[0553] Add diluents B up to 2/3 of the volume, shake for 30 minutes
at 200 mot/min. Dilute to volume with Diluent B. Mix well. Filter
before use through a 0.45 .mu.m GHP ACRODISC GF filter or
equivalent, discarding the first 0.5-1 mL.
[0554] Working concentration of laquinimod (acid) is about 84
ug/mL. Immediately after preparation place sample solutions into a
refrigerator or in a cooled to 5.degree. C. autosampler. The sample
solutions may be used for 24 hours when kept at the temperature
2.degree. C.-8.degree. C.
4. Procedure
[0555] Inject the Resolution Solutions, Diluent B (Blank), Standard
Solutions for assay and IDD and Sample solutions, according to
standard operating procedures.
[0556] Determine the retention time (RT) and the area of the
laquinimod peak in the chromatograms of Sample and Standard
Solutions for identification and assay.
[0557] Determine the RT, the relative retention time (RRT) and peak
areas of all known impurities and any other impurities in the
chromatograms of Sample Solutions, for calculation of the content
of impurities/degradation products.
[0558] Ignore early elution peaks of excipients and system peaks
(See chromatogram for determination of impurities/degradation
products). For example. Use integration inhibition between 0 and
RRT 0.15 (about 2.5 minutes).
[0559] Adjust integration parameters to reject peaks with area less
than 10% of the average response of Laquinimod peak in the
injections of Standard Solution I (for determination of
impurities/degradation products).
[0560] Disregard peak of MEG-LAQ in sample injections (identified
following Resolution Solution 2). The content of MEG-LAQ is tested
by Example 23.
5. System Suitability Test
5.1 Resolution Test
[0561] Typical retention time of laquinimod peak is 15.5.+-.2.0
minutes.
[0562] Tailing factor (USP) for laquinimod peak should be not more
than 2.0.
[0563] Resolution factor for all the pairs of peaks should be not
less than (NLT) 2.
[0564] RRT of the peaks of known impurities/degradation products
should be as follows:
[0565] MCQME: 0.33-0.38
[0566] MCQ: 0.49-0.58
[0567] MCQEE: 0.56-0.65
[0568] MCQCA: 0.71-0.85
[0569] 5-HLAQ: 1.2-1.4 (Should not be more than 23 minutes)
[0570] MEG-LAQ peak is substantially broadened in comparison with
neighboring peaks. Retention time of MEG-LAQ is variable, being
very sensitive to slightest changes in chromatographic conditions
(pH, % acetonitrile, Temperature, etc.) and therefore should be
defined using its peak in the chromatogram of the Resolution
Solution 2. Typically, its RRT is about 0.66.
5.2 System Precision Test
[0571] Evaluate laquinimod standards for assay and IDD in order to
test the system precision according to the standard operating
procedures.
[0572] MCQCA in Solution I is used to test the sensitivity of the
system. RSD of the area of six injections of Std 1 as well as the
difference between Std 1 and Std 2 should be no more than (NMT)
20%.
5.3 Blank
[0573] Injection diluents B to detect system peaks.
6. Identification by Retention Time (RT)
[0574] The RT of the main peak obtained in the sample chromatogram
should correspond to that obtained for the laquinimod peak in the
injection of Standard Solution.
7. Calculation and Report
7.1 Assay Calculation
[0575] % Assay ( to Label Claim ) = Area Smp .times. Conc Std
.times. 0.94 .times. V Smp .times. AvgWt CapsContent Area Std
.times. W Smp .times. Label Claim ##EQU00001##
[0576] Where 0.94 is the conversion factor of laquinimod sodium
salt to laquinimod (acid).
7.2 Calculation and Evaluation of Impurities/Degradation
Products
7.2.1 Calculation of Relative Retention Time (RRT)
[0577] R R T Impurity = R T Impurity R T Laquinimod
##EQU00002##
7.2.2 Calculation of Content of Impurities/Degradation Products
[0578] % Impurity = Area impurity .times. Conc Std .times. V Smp
.times. AvgWT CapsContent .times. 0.94 .times. RRF Area Std .times.
W Smp .times. Label Claim ##EQU00003##
Area.sub.impurity is the area of an impurity/degradation product
(known or unknown) peak in the Sample Solution.
[0579] Area.sub.std is the laquinimod peak in chromatogram of
Standard Solution I.
[0580] 0.94 is the conversion factor of laquinimod sodium salt to
laquinimod (acid).
[0581] RRF is the relative response factors of
impurities/degradation products calculated as the following ratio:
slope of Laquinimod regression line/slope of impurity regression
line.
[0582] The values for relative response factors with respect to
laquinimod are: MCQME: 0.74; MCQ: 0.65; MCQEE: 0.85; MCQCA: 0.62;
and 5-HLAQ: 1.0.
[0583] RRF for unknown impurities/degradation products is taken as
1.0.
7.2.3 Evaluation and Report of Impurities/Degradation products
[0584] Quantitation level (QL) MCQME, MCQ, MCQEE, 5-HLAQ, and
unknown impurities is 005%. Detection level (DL) of MCQME, MCQ,
MCQEE, 5-HLAQ and unknown impurities is 0.02%. QL of MCQCA is 0.1%.
Detection level DL of MCQCA is 0.03%.
[0585] Correlate all the peaks in sample chromatogram with those in
the system suitability chromatogram, with .+-.5% of the actual
corresponding retention times. Report data as shown in Table 2.
TABLE-US-00003 TABLE 2 Reporting Guidelines for HPLC data Result
Report Specified impurities 5-HLAQ .gtoreq.0.05% The calculated
result <0.05% <0.05% <0.02% <0.02% (or ND) MCQ and
MCQCA .gtoreq.0.01% The calculated result sum <0.01% <0.01%
<0.03% <0.03% (or ND) Other impurities MCQME, MCQEE
.gtoreq.0.05% The calculated result <0.05% <0.05% <0.02%
<0.02% (or ND) Unknown peaks .gtoreq.0.05% The calculated result
(by RRT to <0.05% <0.05% laquinimod) <0.02% Not to be
reported* (or ND) Total .gtoreq.0.05% The sum of calculated results
<0.05% <0.05% (or <0.02% or ND) *If no impurities were
detected, report: any other <0.2%.
TABLE-US-00004 TABLE 3 Example System Suitability Results 1 RT USP
USP Int Name RT Ratio Area Resolution Trailing Type 1 MCQME 5.460
0.36 35801 1.07 BB 2 MCQ 8.260 0.54 60687 8.2 1.16 BV 3 MCQEE 9.249
0.6 24029 2.6 1.05 VB 4 MCQCA 12.031 0.78 18609 5.7 1.34 BB 5
Laquinimod 15.332 12332469 5.2 1.84 BB 6 5-HLAQ 20.451 1.33 89463
7.3 1.04 BB
Example 1
Modified Recrystallization of Laquinimod Sodium-- Pilot Production,
100 Fold Scale-Up, (Pilot Scale Batches A and B)
[0586] Recrystallization of laquinimod sodium was performed on
pilot scale (Batches A and B) as follows.
[0587] Re-crystallization of Laquinimod Na on pilot scale is
performed in two glass-lined reactors (Reactor A, 30 liter volume
and Reactor B 60 liter). Solid product is filtered and dried in
Hastelloy C agitated filter-dryer with 20 micron mesh.
[0588] Batch size is 2.5 kg of starting crude Laquinimod Na.
[0589] Batch of crude Laquinimod Na (2.5 kg) is introduced to
Reactor A with 10 volumes of process water. The batch is heated to
60-73.degree. C. at stirring until complete dissolution of
solid.
[0590] The hot solution in Reactor A is transferred to Reactor B
through 0.2 .mu.m filtration system. Reactor A and filters washed
with 1.2 volumes of process water and the wash is transferred to
the Reactor B.
[0591] Vacuum is built-up and the solution in the Reactor B is
evaporated at P<45 mmHg and jacket temperature T<65.degree.
C. until volume of the residue reaches 5.4 liters (2.16 volumes).
Then atmospheric pressure is built-up and jacket temperature
40-50.degree. C. is adjusted. The batch is stirred for not less
than 10 minutes and then seeded with Laquinimod Na crystals to
initiate crystallization.
[0592] The batch is stirred at 45.degree. C. for additional 90
minutes and 7.9 volumes of acetone are added to the reactor in
1.5-2.5 hrs. Reactor temperature during the addition maintained
between 40 and 50.degree. C.
[0593] Resulting slurry is cooled to 0.+-.4.degree. C. during
3.5-4.5 hrs and stirred at this temperature for 10-15 hrs. Then the
slurry is transferred to filter-dryer and solid is filtered under
pressure of nitrogen.
[0594] The cake is washed twice (22 kg) with acetone, purged with
nitrogen and then dried under vacuum (P<50 mmHg) and elevated
temperature (T=40.degree. C.) at agitation.
[0595] Dry product is discharged, sampled for analysis and
packed.
Discussion of Example 1
Example 1
[0596] The pilot scale process of recrystallization of laquinimod
sodium was based on Example 15 of U.S. Pat. No. 7,884,208. The
starting material was crude laquinimod sodium having low particle
size (d(0.1)=1-2.mu., d(0.5)=6-11.mu.; d(0.9)=20-35.mu.) and
appears as aggregated solid. Example 15 of U.S. Pat. No. 7,884,208
involves 25.0 g of laquinimod sodium (laboratory scale) prepared
according to the method disclosed in U.S. Pat. No. 6,875,869. In
Example 15, the 25.0 g of laquinimod sodium is dissolved in an
aqueous solution of laquinimod sodium and then evaporated under
vacuum at stirring to a concentrated solution having a volume ratio
of 2.14 v/w, the resulting residue is seeded to induce
crystallization then treated with an anti-solvent (acetone).
[0597] The modified pilot scale process was performed with 2.5 kg
of laquinimod sodium which is a 100-fold scale up from Example 15.
In addition, the modified pilot scale process had significant
differences from the laboratory scale process of Example 15 of U.S.
Pat. No. 7,884,208. Specifically, evaporation on the laboratory
scale was performed in a round-bottom flask in a rotary evaporator
without stirring, while evaporation on the pilot scale was
performed in a reactor with stirring. On the pilot scale, the
evaporation residue is stirred aggressively, liquid splashes on the
reactor walls, solid depositions form, and crystallization was
spontaneous. On the laboratory scale, a metastable solution could
be concentrated to a volume ratio of 2.1-2.2 v/w at which point
crystallization did not take place and nucleation was controlled by
seeding. On the pilot scale, conditions and concentration were such
that spontaneous crystallization took place, i.e., crystallization
was induced without seeding.
[0598] Surprisingly, the pilot batches did not result in laquinimod
sodium particles having a particle size distribution expected based
on Example 15 of U.S. Pat. No. 7,884,208. Instead, applicants
unexpected found that the pilot batches resulted in a mixture of
recrystallized laquinimod sodium particles wherein (i) 90% or more
of the total amount by volume of the laquinimod sodium particles
have a size of less than 40 microns, (ii) 50% or more of the total
amount by volume of the laquinimod sodium particles have a size of
less than 15 microns, and (iii) 10% or more of the total amount by
volume of the laquinimod sodium particles have a size of less than
5 microns. A comparison of particle size distributions resulting
from the two different processes is shown in Table 4.
TABLE-US-00005 TABLE 4 Laboratory Scale Modified Pilot Scale 25 g
batch 2.5 kg batch PSD by Example 15 from U.S. Pilot Scale Pilot
Scale Malvern Pat. No. 7,884,208 Batch A Batch B d(0.1) um 6.1 3.3
4.1 d(0.5) um 21.2 12.9 13.5 D(0.9) um 51.8 33.8 33.9 Appearance
White free- White free- White free- flowing powder flowing powder
flowing powder
[0599] The process of U.S. Pat. No. 7,884,208 and the process of
Example 1, above, each produce different products and are not
equivalent processes. Applicants' pilot scale version of the
process of U.S. Pat. No. 7,884,208 resulted in substantially
different conditions from the prior art and resulted in a
substantially different product having smaller particle sizes as
shown in Table 4.
[0600] Since laquinimod Na is a potent drug substance, small
particle size is advantageous for this API. Formation of
non-aggregated laquinimod sodium crystals with reduced particle
size could provide better uniformity of drug product and avoid
milling or de-lumping operations. The starting material, crude
laquinimod sodium, appears as aggregated solid. The re-crystallized
product is free flowing powder. Powders with smaller particles have
a stronger trend to aggregate. Crude laquinimod sodium prepared by
slurry-to-slurry recrystallization (i.e., the process of U.S. Pat.
No. 6,077,851) produces particles having a low particle size and
are highly aggregated. The modified process produces particles
having a low particle size and are free flowing.
[0601] The aim was scalable crystallization procedure giving
smaller crystal size, PSD and low aggregation on laboratory, pilot
and commercial scale. The desirable PSD profile was the following:
d(0.1)<5 .mu.m, d(0.5)<15 .mu.m and d(0.9)<40 .mu.m. The
method is based on spontaneous crystallization initiated in aqueous
phase prior to acetone addition. The important factor affecting
crystallization is initial concentration of crystallizing solution.
In the new crystallization procedure, reduced water volume ratio in
the end of evaporation from 2.14 v/w to 1.7-1.8 v/w. Higher
concentration of the solution ensures initiation of spontaneous
crystallization in the end of evaporation operation and provides
higher supersaturation level and lower crystal size.
Example 2
Recrystallization of Laquinimod Sodium--Laboratory Scale
(Laboratory Scale Batch A)
[0602] Recrystallization of laquinimod sodium was performed on
laboratory scale (Batch A) as follows.
[0603] All operations of Laquinimod Na re-crystallization step
including evaporation were performed on laboratory scale in
transparent agitated glass reactors equipped with stirrer,
thermometers and circulating bath for heating and cooling.
[0604] 25 g crude Na Laquinimod and 275 ml deionized water
introduced into 250 ml stirred jacketed glass reactor. The mixture
is stirred and heated to 70.degree. C., after complete dissolution
of the solid the solution is filtered through paper filter.
Resulting clear filtrate introduced to 250 ml jacketed glass
reactor equipped with circulating bath, stirrer, thermometer and
vacuum distillation system.
[0605] Vacuum is applied and water is distilled at stirring,
pressure during the evaporation is 38-40 mbar and jacket
temperature is 55.degree. C.
[0606] After distillation of ca. 2/3 volume spontaneous
crystallization on the reactor wall above liquid level is
observed.
[0607] The distillation is continued until the residue volume
reaches 45 ml then atmospheric pressure is build up and the batch
is stirred at 50.degree. C. for one hour. On this step intensive
crystallization takes place.
[0608] 200 ml acetone is added to the resulting slurry in one hour
and the batch is stirred for one additional hour at 50.degree.
C.
[0609] The batch is cooled to 0-5.degree. C. during one hour and
filtered on Buchner filter. The solid cake is washed with 75 ml of
acetone.
[0610] Collected wet product (28.0 g) is dried in oven under vacuum
at 50.degree. C. to constant weight.
[0611] Dry product--23.8 g; Crystallization yield--95.2%
Analysis:
[0612] Microscopic observation--rod-shape particles
[0613] Particle Size Distribution by Malvern:
[0614] D(0.1).sup.=2.3 .mu.m; D(0.5)=10.8 .mu.m; D(0.9)=32.7
.mu.m
Example 3
Recrystallization of Laquinimod Sodium--Laboratory Scale
(Laboratory Scale Batch B)
[0615] Recrystallization of laquinimod sodium was performed on
laboratory scale (Batch B) as follows.
[0616] All operations of Laquinimod Na re-crystallization step
including evaporation were performed on laboratory scale in
transparent agitated glass reactors equipped with stirrer,
thermometers and circulating bath for heating and cooling.
[0617] 25 g Na Laquinimod crude and 275 ml deionized water
introduced into 250 ml stirred jacketed glass reactor. The mixture
is stirred and heated to 70.degree. C., after complete dissolution
of the solid the solution is filtered through paper filter.
Resulting clear filtrate introduced to 250 ml jacketed glass
reactor equipped with circulating bath, stirrer, thermometer and
vacuum distillation system. Vacuum is applied and water is
distilled at stirring, pressure during the evaporation is 38-40
mbar and jacket temperature is 55.degree. C. During the
distillation spontaneous crystallization on the reactor wall is
observed when the residue volume reached ca. 120 ml. The
distillation is continued until the residue volume reaches 45 ml
then atmospheric pressure is build up and the batch is stirred at
50.degree. C. for one hour. On this step intensive crystallization
takes place.
[0618] 200 ml acetone is added to the resulting slurry in one hour
and the batch is stirred for one additional hour at 50.degree.
C.
[0619] The batch is cooled to 0-5.degree. C. during one hour,
stirred at this temperature for one additional hour and filtered on
Buchner filter. The solid cake is washed with 75 ml of acetone.
[0620] Collected wet product (27.5 g) is dried in oven under vacuum
at 50.degree. C. to constant weight.
[0621] Dry product--23.65 g; Crystallization yield--94.6%
Analysis:
[0622] Microscopic observation--rod-shape particles
[0623] Particle Size Distribution by Malvern:
[0624] D(0.1)=2.6 .mu.m; D(0.5)=12.4 .mu.m; D(0.9)=34.3 .mu.m
TABLE-US-00006 TABLE 5 Laboratory scale Laquinimod Na
crystallization results Laboratory Scale Laboratory Scale
Laboratory Scale Batch No. Batch A Batch B Starting material, crude
.sup. 25 g 25 g Laquinimod Na, g Dry product, g 23.8 g 23.65 g
Crystallization yield, % 95.2% 94.6% PSD D(0.1) .mu.m 2.3 2.6
D(0.5) .mu.m 10.8 12.4 D(0.9) .mu.m 32.7 34.3
Discussion of Example 2 and Example 3
[0625] The results of Example 2 and Example 3 are summarized in
Table 5. Table 5 shows that the process reliably produced a mixture
of laquinimod sodium crystals a mixture of recrystallized
laquinimod sodium particles wherein (i) 90% or more of the total
amount by volume of the laquinimod sodium particles have a size of
less than 40 microns, (ii) 50% or more of the total amount by
volume of the laquinimod sodium particles have a size of less than
15 microns, and (iii) 10% or more of the total amount by volume of
the laquinimod sodium particles have a size of less than 5
microns.
Example 4
Recrystallization of Laquinimod Sodium--Production Scale
(Production Scale Batches C, D and E)
[0626] Recrystallization of laquinimod sodium was performed on a
production scale (Batches C, D and E) as follows.
[0627] Re-crystallization of Laquinimod Na on the production scale
is performed in two 250 liter glass-lined reactors (Reactor I and
Reactor II). Solid product is filtered and dried in Hastelloy C-22
agitated filter-dryer with 20 micron mesh.
[0628] Batch size is 6.5-7.5 kg of dry API.
[0629] Batch of crude Laquinimod Na is introduced to Reactor I with
11 volumes of process water. The batch is heated to 60-73.degree.
C. at stirring until complete dissolution of solid.
[0630] The hot solution in Reactor I is circulated through 0.2
.mu.m filtration system at heating and stirring during 15-20
minutes. After the circulation completion filtered solution is
transferred to Reactor II through 0.2 .mu.m filter. Reactor I and
filters washed with 1.75 volumes of process water and the wash is
transferred to the Reactor II.
[0631] Vacuum is build-up and the solution in the Reactor II is
evaporated at P<45 mmHg and jacket temperature T<65.degree.
C. until volume of the residue reaches 14-16 liter (ca. 1.7-1.8 v/w
water/weight crude laquinimod Na starting material). On this step
spontaneous crystallization is initiated on the reactor walls. Then
atmospheric pressure is build-up and jacket temperature
40-50.degree. C. is adjusted.
[0632] The batch is stirred for not less than 10 minutes.
[0633] The batch is stirred at 45.degree. C. for additional 90
minutes and 7.9 volumes of acetone are added to the reactor in
1.5-2.5 hrs. Reactor temperature during the addition maintained
between 40 and 50.degree. C.
[0634] Resulting slurry is cooled to 0.+-.4.degree. C. during 2-5
hrs and stirred at this temperature for 10-15 hrs. Then the slurry
is transferred to filter-dryer and solid is filtered under pressure
of nitrogen.
[0635] The cake is washed twice (2.times.10 liter) with acetone,
purged with nitrogen and then dried under vacuum (P<50 mmHg) and
elevated temperature (T=35.+-.5.degree. C.) at agitation.
[0636] Dry product is discharged, sampled for analysis and
packed.
[0637] Data for 3 typical production GMP batches is summarized in
Table 6. The PSD values presented in Table 6 are in a good
accordance to the results of laboratory scale experiments presented
in Example 2 and Example 3.
TABLE-US-00007 TABLE 6 Production scale Laquinimod Na
crystallization results Production Scale Batch No. Batch C Batch D
Batch E Starting material, crude 8.50 8.55 9.45 Laquinimod Na, kg
Dry product, kg 7.40 7.00 6.80 Crystallization yield, % 87 82 72
Product Apperance White free- White free- White free- flowing
flowing flowing powder powder powder Bulk Density, g/ml 0.308 0.288
0.245 Tapped Density, g/ml 0.609 0.609 0.460 PSD D(0.1) .mu.m 4.1
4.1 2.5 D(0.5) .mu.m 14.0 14.7 9.5 D(0.9) .mu.m 32.0 33.6 21.0
Batch C and D had reduced levels of impurities (Table 7) and good
bulk and tapped density (Table 6).
[0638] Analysis of powder density of laquinimod sodium produced on
the production scale (22 crystallization batches) shows that the
bulk density varies in range between 0.237 and 0.364 g/ml. Tapped
density is with 0.432 and 0.609 g/ml.
[0639] The results of analyzing Batches C and D are shown in Table
7 and Table 8.
TABLE-US-00008 TABLE 7 Analytical Results Test Specifications Batch
C Batch D Assay 98.0-102.0% 99.3% 99.9% (by HPLC) Related Polar MCQ
NMT 0.15% LT 0.02% LT 0.02% Substances (by HPLC) MCQCA NMT 0.15% LT
0.03% LT 0.03% MCQEE NMT 0.10% LT 0.02% LT 0.02% MCQME NMT 0.12% LT
0.02% LT 0.02% 5-HLAQ NMT 0.10% LT 0.02% LT 0.02% Any other NMT
0.10% LT 0.02% LT 0.02% impurities Total impurities NMT 1.00% LT
0.05% LT 0.05% Related Non-Polar N-Ethyl NMT 0.10% LT 0.02% LT
0.02% Substances (by HPLC) aniline Any other NMT 0.10% LT 0.02% LT
0.02% impurities Total impurities NMT 0.50% LT 0.02% LT 0.02%
Des-ethyl laquinimod NMT 0.10% LT 0.1% LT 0.1% (DELAQ) content (by
HPLC) LAQ content by HPLC) NMT 1.00% LT 0. 2% LT 0. 2%
Dimethylmalonate NMT 0.10% LT 0.05% LT 0.05% content (by HPLC)
Dimethyl sulfate NMT 1 ppm LT 1 ppm LT 1 ppm content (by LC-MS)
Water (by K.F coulometric) NMT 1.5% (w/w 0.3% (w/w) 0.2% (w/w)
Heavy metals (by IPC-ABS) NMT 20 ppm LT 20 ppm LT 20 ppm Sodium
content 5.8-6.4% 6.1% 6.1% Residual solvents Ethanol NMT 5000 ppm
LT 5 ppm LT 5 ppm n-Heptane NMT 5000 ppm LT 10 ppm LT 10 ppm
n-octane NMT 2000 ppm LT 10 ppm LT 10 ppm Methanol NMT 3000 ppm LT
30 ppm LT 30 ppm Acetone NMT 5000 ppm LT 250 ppm LT 250 ppm Dioxane
NMT 380 ppm LT 10 ppm LT 10 ppm DMF NMT 880 ppm LT 40 ppm LT 40 ppm
Microbiological tests Total viable NMT 1000 CFU/g LT 10 CFU/g LT 10
CFU/g aerobic count Fungi/yeasts NMT 100 CFU/g LT 10 CFU/g LT 10
CFU/g and moulds Escherica Coli Absence of E-Coli Absence
Absence
TABLE-US-00009 TABLE 8 Metal impurities in PPM of Laquinimod Sodium
Example 1, (Batch D) of Example 17 of Example Example U.S. Pat. No.
U.S. Pat. No. 4 New 4 New Impurity 7,884,208 7,884,208 Batch C
Batch D Al 14.0 5.6 1.63 1.13 Ca 165 65 6.3 22 Cr 2.6 <0.5 0.55
<0.26 Cu 2.8 1.3 0.325 <0.26 Fe 31.5 5.8 3.44 3.55 Ni 5.5
<0.5 0.79 0.61 S 466 <1 6.7 3.9 Zn 20.5 7.5 <1 3.15
TABLE-US-00010 TABLE 9 Purification of Laquinimod Sodium from MCQME
Starting material, Re-crystallized Crude product Batch No. Crude C
Appearance Aggregated white Free-flowing white solid powder MCQME
by HPLC, % 0.10 N.D.* *N.D.--Not Detected (<0.02%)
[0640] Microscopic photographs of typical batches of Crude and
re-crystallized Laquinimod Na at different magnification are
presented on FIGS. 5-8.
Discussion of Example 4
[0641] The modified crystallization procedure demonstrated good
reproducibility of particle size distribution on the production
scale. Reduction of the evaporation residue volume to a ratio of
1.7-1.8 v/w and initiation of spontaneous crystallization provided
desirable crystal size. Improved crystallization procedure
demonstrates reduction in crystal size to a level of d(0.9)<40
.mu.m and good reproducibility of Particle Size Distribution on the
production scale.
[0642] The product with reduced crystal size has no trend to
aggregation and does not need milling or de-lumping for
homogenization. The product with similar PSD prepared by
slurry-to-slurry procedure is aggregated and thus problematic in
formulation.
[0643] The modified crystallization procedure also results in
laquinimod sodium having desirable density and purity profiles.
[0644] The improved crystallization procedure also provides
effective purification from organic impurities, e.g., MCQME.
[0645] The data shown in Table 9 demonstrates complete removal of
MCQME impurity by re-crystallization of commercial scale batch of
Laquinimod sodium. Since this intermediate has genotoxic potential
it should be purified to undetectable level. The crystallization
process also provides purification of all other known organic
impurities to the level below the limit of detection.
[0646] Microscopic photographs of typical batches of crude and
re-crystallized Laquinimod sodium at different magnification are
presented on FIGS. 5-8 and show rod-shape morphology of the both
products. At the same time, the Crude presented on FIGS. 5 and 6
are much more aggregated than the re-crystallized "Cryst" product
of FIGS. 7 and 8.
Example 5
Laquinimod Capsules of Pharmaceutical composition of Laquinimod
Sodium
[0647] Laquinimod capsules are manufactured according to the method
as described in Example 2 of PCT International Application
Publication No. WO 2007/146248, the entire content of which is
hereby incorporated by reference. Steps of Example 2 of WO
2007/146248 are performed. Each capsule contains 064 mg of
laquinimod sodium equivalent to 0.6 mg laquinimod.
[0648] The capsules had quantities of impurities within
specification based on HPLC relative to the amount of
laquinimod.
[0649] The capsules have a water content of no more than 1.5%.
[0650] The dissolution profiles, content uniformity, and residual
solvents of the encapsulated pharmaceutical composition conforms to
U.S. Pharmacopeia <711> (dissolution), U.S. Pharmacopeia
<905> (uniformity), and U.S. Pharmacopeia <467>.
[0651] Each capsule contains 90.0-110.0% of the labeled amount.
[0652] The capsules contain a total aerobic microbial count (TAMC)
of NMT 10.sup.3 cfulg, a total combined yeasts/moulds count (TYMC)
of NMT 10.sup.2 cfu/g, and an absence of Escherichia Coli in 1
g.
Discussion of Example 5
[0653] Example 5 demonstrates that, in a commercial-scale
production, pharmaceutical compositions of laquinimod can be
prepared with non-detectable levels or a low level of polar
impurities and non-polar impurities.
Example 6
Preparation Of Laquinimod Sodium Reference Standard
[0654] Laquinimod sodium primary reference standard batch was
prepared by crystallization of laquinimod sodium batch, followed by
purification by crystallization from a water/acetone mixture (1:4.5
w/w). The crystallization was performed by dissolving the drug
substance batch in water while heating. The clear solution was
filtered and concentrated to a known residue volume under reduced
pressure. Acetone was added and the solution was cooled. The
obtained crystals were filtered, washed and dried. The
chromatographic purity of this batch was found to be 100.0%. The
total of both polar and non-polar impurities was LT 0.05%, content
of DELAQ was LT 0.1%, and laquinimod acid content was LT 0.2%.
Example 7
Preparation of MCQ
[0655] MCQ is a white solid. The molecular structure, chemical
formula and molecular weight of MCQ are provided below.
##STR00022##
[0656] MCQ was prepared in the following manner: MCQCA and DMSO
were heated at 75.degree. C. for 2 hours and were then cooled to
room temperature. Water was added and the precipitate was collected
by filtration, washed with water and dried in a vacuum oven at
50.degree. C. Material of sufficient purity to be suitable for use
as a reference standard was obtained.
[0657] MCQ was characterized by NMR, MS, elemental analysis and
FT-IR as detailed hereafter.
Nuclear Magnetic Resonance (NMR) Spectroscopy
[0658] The .sup.1H-NMR and .sup.13C-NMR characterization of MCQ was
performed in DMSO on a Bruker 300 MHz instrument. The peak
assignments are summarized in Table 10. The .sup.1H-NMR and
.sup.13C-NMR spectra are presented in FIG. 9 and FIG. 10,
respectively.
TABLE-US-00011 TABLE 10 .sup.13C and .sup.1H-NMR Peak Assignments
for MCQ .sup.13C (ppm) .sup.1H (ppm) Multiplicity (Hz) *
N--CH.sub.3 29.30 3H 3.55 brs C # 2 161.53 -- -- C # 3 99.57 1H
5.94 brs C # 4 161.53 -- -- C # 4a ca. 113.2 -- -- C # 5 ca. 130.5
-- -- C # 6 124.78 1H 7.47 d (8.0) C # 7 130.93 1H 7.53 t (8.0) C #
8 114.22 1H 7.29 d (8.0) C # 8a 142.41 -- -- *brs = broad singlet,
d = doublet, t = triplet
Mass Spectroscopy (MS)
[0659] The mass spectrum of MCQ was obtained on a Q-TOF
Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray
ionization in positive ion mode (ES.sup.+). The ratio between MS
signals containing .sup.35Cl and .sup.37Cl corresponds to the
natural abundances of these isotopes (about 3:1). The spectrum
shown in FIG. 11 is in agreement with the molecular weight of MCQ.
The attribution of the main signals in ES.sup.+ mass spectrum of
MCQ is presented in Table 1.
TABLE-US-00012 TABLE 11 Attribution of Main Peaks of ES.sup.+
Spectrum of MCQ m/z Attribution 210 [M(.sup.35Cl) + H].sup.+ 212
[M(.sup.37Cl) + H].sup.+ 232 [M(.sup.35Cl) + Na].sup.+ 234
[M(.sup.37Cl) + Na].sup.+
[0660] The test for elemental analysis was performed on a
Perkin-Elmer 2400 Series II C H N Analyzer. The results for MCQ are
presented in Table 12. C, H and N weight percentages are accurate
and reproducible to within .+-.0.3%. The accuracy for halogens is
.+-.0.5%.
TABLE-US-00013 TABLE 12 Elemental Analysis Results for MCQ Element
% C % H % N % Cl Theoretical 57.30 3.85 6.68 16.91 Experimental
57.31 3.64 6.54 17.41
Infrared Spectroscopy
[0661] The FT-IR spectrum of MCQ was measured with a Nicolet 6700
"Thermo Scientific" FT-IR apparatus. A representative spectrum is
presented in FIG. 12. A summary of the band assignments is provided
in Table 13.
TABLE-US-00014 TABLE 13 Summary of IR Band Assignments of MCQ in
KBr Transition Energy (cm.sup.-1) Band Assignment 702-1280 Aromatic
C--H deformation vibration 1093 and 1135 Aromatic C--Cl stretching
1164-1280 Aromatic C--OH stretching 1164-1413 C--N stretching
1554-1613 Aromatic C--C stretching 1449-1613 Aromatic C.dbd.C
stretching 1637 C.dbd.O stretching 2890 N--CH.sub.3 stretching 3450
OH stretching
Example 8
Preparation Of MCQCA
[0662] MCQCA is a white solid. The molecular structure, chemical
formula and molecular weight of MCQCA are provided below.
##STR00023##
[0663] MCQCA was prepared in the following manner:
[0664] A mixture of MCQEE in a 2.8M solution of HCl in acetic acid
was heated for 6 hours at 65.degree. C. using a reflux condenser.
The mixture was cooled to room temperature, diluted with 2-propanol
and was further cooled to 8.degree. C. The crystals were filtered
off, washed with 2-propanol and dried in a vacuum oven at
50.degree. C.
[0665] Material of sufficient purity to be suitable for use as a
reference standard was obtained.
[0666] MCQCA was characterized by NMR, MS, elemental analysis and
FT-IR as detailed below.
Nuclear Magnetic Resonance (NMR) Spectroscopy
[0667] The 1H-NMR and 13C-NMR characterization of MCQCA was
performed in D2O+KOH on a Bruker 300 MHz instrument. The peak
assignments are summarized in Table 14. The 1H-NMR and 13C-NMR
spectra are presented in FIG. 13 and FIG. 14, respectively.
TABLE-US-00015 TABLE 14 .sup.13C and .sup.1H-NMR Peak Assignments
for MCQCA .sup.13C (ppm) .sup.1H (ppm) Multiplicity (Hz) *
N--CH.sub.3 29.78 3H 3.43 s C # 2 162.31 -- -- C # 3 113.46 -- -- C
# 4 176.93 -- -- C # 4a 119.28 -- -- C # 5 131.56 -- -- C # 6
124.88 1H 7.05 d (8.0) C # 7 129.38 1H 7.21 t (8.0) C # 8 114.03 1H
7.14 d (8.0) C # 8a 141.32 -- -- C # 9 169.62 -- -- *d = doublet, t
= triplet, s = singlet
Mass Spectroscopy (MS)
[0668] The mass spectrum of MCQCA was obtained on a Q-TOF
Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray
ionization in positive ion mode (ES.sup.+). The ratio between MS
signals containing .sup.35Cl and .sup.37Cl corresponds to the
natural abundances of these isotopes (about 3:1). The spectrum
shown in FIG. 15 is in agreement with the molecular weight of
MCQCA. The attribution of the main signals in ES.sup.+ mass
spectrum of MCQCA is presented in Table 15.
TABLE-US-00016 TABLE 15 Attribution of Main Peaks of ES.sup.+ Mass
Spectrum of MCQCA m/z Attribution 236 [M(.sup.35Cl) +
H--H.sub.2O].sup.+ 238 [M(.sup.37Cl) + H--H.sub.2O].sup.+ 254
[M(.sup.35Cl) + H].sup.+ 256 [M(.sup.37Cl) + H].sup.+ 276
[M(.sup.35Cl) + Na].sup.+ 278 [M(.sup.37Cl) + Na].sup.+
Elemental Analysis
[0669] The test for elemental analysis was performed on a
Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight
percentages are accurate and reproducible to within .+-.0.3%. The
accuracy for halogens is .+-.0.5%.
[0670] The results for MCQCA are presented in Table 16.
TABLE-US-00017 TABLE 16 Elemental Analysis Results for MCQCA
Element % C % H % N % Cl Theoretical 52.09 3.18 5.52 13.98
Experimental 52.11 3.16 5.56 13.68
Infrared Spectroscopy
[0671] The attenuated total reflectance (ATR) FT-IR spectrum of
MCQCA was measured with a Nicolet 6700 "Thermo Scientific" FT-IR
apparatus. A representative spectrum is presented in FIG. 16. A
summary of the band assignments is provided in Table 17.
TABLE-US-00018 TABLE 17 Summary of IR Band Assignments of MCQCA
Transition Energy (cm.sup.-1) Band Assignment 706-1206 Aromatic
C--H deformation vibration 1008-1102 Aromatic C--Cl stretching
1148-1284 Aromatic C--OH stretching 1008-1434 C--N stretching
1571-1678 Aromatic C--C stretching 1571-1678 Aromatic C.dbd.C
stretching 1678 C.dbd.O stretching 2520 COO--H stretching 3090
Aromatic C--H stretching
Example 9
Preparation Of MCQME
[0672] MCQME is a white to yellowish solid. The molecular
structure, chemical formula and molecular weight of MCQME are
provided below.
##STR00024##
[0673] The preparation of MCQME started with the formation of
sodium dimethylmalonate by reaction of dimethylmalonate in DMF with
sodium methoxide solution. Then, the intermediate MCIA was reacted
with sodium dimethylmalonate to form the MCQME sodium salt. Finally
the MCQME sodium salt was acidified to MCQME, which was washed with
water and dried. The following is a detailed description of the
preparation of MCQME.
##STR00025##
[0674] DMF (66.5 kg [70.4 liter]), dimethyl malonate (9.6 kg [72.7
mole, 8.3 liter]) and a 30% methanolic solution of sodium methoxide
(1.5 kg [63.9 mole]) were charged to a reactor. The reaction
mixture was stirred and heated to 82.degree. C. to 88.degree. C.
and kept at this temperature for 11/2 to 21/2 hours in order to
form sodium dimethyl malonate. Then the mixture was cooled to
30-40.degree. C. Vacuum was applied and the reactor jacket
temperature was heated gradually in order to evaporate out
methanol. After evaporation of methanol (not less than 12 kg [14.5
liter] distillate) the vacuum was broken with nitrogen and the
reaction mixture was heated to 82.degree. C. to 88.degree. C. C.
Then MCIA (8.5 kg [40.1 mole]) was added gradually to the reaction
mixture and washed with DMF (10 kg [10.6 liter]). The reactor
content was cooled to less than 75.degree. C., vacuum was applied,
the reactor jacket temperature was gradually raised and methanol
was evaporated (12 kg [15 liter] distillate). The vacuum was broken
with nitrogen and the reaction mixture was cooled to 12.degree. C.
to 35.degree. C. Process water (119 Kg) was added gradually,
keeping the reaction mixture temperature at 12.degree. C. to
35.degree. C., then the reaction mixture was heated to
42-48.degree. C. 32% Aqueous HCl solution (11.1 kg [97.3 mole]) was
added during 2 to 3 hours and remains of hydrochloric solution in
feeding line were washed out with DMF. The formed slurry is cooled
to 20.degree. C. to 30.degree. C. and was maintained at that
temperature for not less than 30 minutes.
[0675] The slurry was transferred gradually to a filter-dryer.
Mother liquor was removed by pressing the slurry. Process water
(29.8 kg for each cycle) was added to the filter-dryer and pressed
out. Washing of the filter-dryer content with water was performed
until the pH of outflow washing liquor was NLT 4. The product was
dried under vacuum at 40.degree. C. to 50.degree. C. (jacket
temperature of filterdryer).
[0676] Vacuum drying was performed until the LOD of a sample from
the filter-dryer was less than 2.0% and the water content (by
Karl-Fischer) was not more than 0.5%. Then the jacket was cooled to
20.degree. C. to 30.degree. C. and dry MCQME was discharged.
[0677] The yield is 62% to 87%.
[0678] The dry MCQME was delumped, analyzed and released.
Example 10
Preparation Of MCQEE
[0679] MCQEE is a light yellow solid. The molecular structure,
chemical formula and molecular weight of MCQEE are provided
below.
##STR00026##
[0680] MCQEE was prepared in the following manner:
[0681] Sodium hydride was added to a solution of MCIA and diethyl
malonate in DMF. The mixture was gradually heated to 95.degree. C.
and stirred for 3.5 hours, then cooled to 35.degree. C. The
reaction mixture was quenched with water and then acidified with
37% HCl. After 1 hour at 8.degree. C. the mixture was filtered and
washed with water (until the pH of the mother liquor was 4.5) and
dried in a vacuum oven at 40.degree. C. The crude product was
crystallized by dissolving in ethanol at 85.degree. C. followed by
slow cooling (in an ice water bath). The obtained crystals were
filtered, washed with ethanol and dried in vacuum oven at
50.degree. C.
[0682] Material of sufficient purity to be suitable for use as a
reference standard was obtained.
[0683] MCQEE was characterized by NMR, MS, elemental analysis and
FT-IR as detailed below.
Nuclear Magnetic Resonance (NMR) Spectroscopy
[0684] The .sup.1H-NMR and .sup.13C-NMR characterization of MCQEE
was performed in CDCl.sub.3 on a Bruker 300 MHz instrument. The
peak assignments are summarized in Table 18. The .sup.1H-NMR and
.sup.13C-NMR spectra are presented in FIG. 17 and FIG. 18,
respectively.
TABLE-US-00019 TABLE 18 .sup.13C and .sup.1H-NMR Peak Assignments
for MCQEE .sup.13C (ppm) .sup.1H (ppm) Multiplicity (Hz) *
N--H.sub.3 30.08 3H 3.63 s C # 2 158.76 -- -- C # 3 98.18 -- -- C #
4 172.99 -- -- C # 4a 112.48 -- -- C # 5 134.50 -- -- C # 6 125.96
1H 7.23 d (8.0) C # 7 133.28 1H 7.48 t (8.0) C # 8 113.29 1H 7.24 d
(8.0) C # 8a 143.53 -- -- C # 9 172.67 -- -- C # 1{grave over ( )}
62.56 2H 4.51 q (7.0) C # 2{grave over ( )} 14.20 3H 1.48 t (7.0) *
d = doublet, t = triplet, s = singlet, q = quartet
Mass Spectroscopy (MS)
[0685] The mass spectrum of MCQEE was obtained on a Q-TOF
Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray
ionization in positive ion mode (ES.sup.+). The ratio between MS
signals containing .sup.35Cl and .sup.37Cl corresponds to the
natural abundances of these isotopes (about 3:1). The spectrum
shown in FIG. 19 is in agreement with the molecular weight of
MCQEE. The attribution of the main signals in ES.sup.+ mass
spectrum of MCQEE is presented in Table 19.
TABLE-US-00020 TABLE 19 Attribution of Main Peaks of ES.sup.+ Mass
Spectrum of MCQEE m/z Attribution 236 [M(.sup.35Cl) +
H--(CH.sub.3--CH.sub.2--OH)].sup.+ 238 [M(.sup.37Cl) +
H--(CH.sub.3--CH.sub.2--OH)].sup.+ 282 [M(.sup.35Cl) + H].sup.+ 284
[M(.sup.37Cl) + H].sup.+ 304 [M(.sup.35Cl) + Na].sup.+ 306
[M(.sup.37Cl) + Na].sup.+
Elemental Analysis
[0686] The test for elemental analysis was performed on a
Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight
percentages are accurate and reproducible to within .+-.0.3%. The
accuracy for halogens is .+-.0.5%. The results for MCQEE are
presented in Table 20.
TABLE-US-00021 TABLE 20 Elemental Analysis Results for MCQEE
Element % C % H % N % Cl Theoretical 55.43 4.29 4.97 12.59
Experimental 55.44 4.25 4.87 12.57
Infrared Spectroscopy
[0687] The attenuated total reflectance (ATR) FT-IR spectrum of
MCQEE was measured with a Nicolet 6700 "Thermo Scientific" FT-IR
apparatus, A representative spectrum is presented in FIG. 20. A
summary of the band assignments is provided in Table 21.
TABLE-US-00022 TABLE 21 Summary of IR Band Assignments of MCQEE
Transition Energy (cm.sup.-1) Band Assignment 676-1213 Aromatic
C--H deformation vibration 1019-1092 C--Cl stretching 1145-1438
C--N stretching 1145-1270 Aromatic C--OH stretching 1438-1603
Aromatic C--C stretching, Aromatic C.dbd.C stretching 1656 C.dbd.O
stretching
Example 11
Preparation Of 5-HLAQ
[0688] 5-HLAQ is an off-white solid. The molecular structure,
chemical formula and molecular weight of 5-HLAQ are provided
below.
##STR00027##
[0689] 5-HLAQ is unstable in most organic solvents as well as in
aqueous acidic medium. Therefore purification by the commonly used
methods is extremely difficult. Consequently, purification of the
material was performed by formation of the di-acetate derivative
followed by basic hydrolysis, acidic precipitation and rapid
filtration.
[0690] 5-HLAQ was prepared by a multi-step procedure, depicted
below.
##STR00028##
[0691] The synthesis involved: 1) preparation of 5-MeO-laquinimod
(compound 2) from 2-amino-6-methoxybenzoic acid (compound 1); 2)
preparation of 5-HLAQ crude (compound 3); 3) acetylation of the
phenolic groups to produce the di-acetate-laquinimod derivative
(compound 4); 4) hydrolysis of the di-acetate derivative to produce
the purified 5-HLAQ (compound 5).
[0692] The procedure for the preparation of 5-HLAQ is further
described step-wise in the following paragraphs:
5-MeO-laquinimod (Compound 2)
[0693] 5-MeO-laquinimod was obtained by a four-step synthetic
procedure starting from 2-amino-6-methoxybenzoic acid (compound 1).
These four steps are identical to the manufacturing process of
laquinimod starting from ACBA.
5-HLAQ Crude (Compound 3)
[0694] 5-MeO-laquinimod was dissolved in dichloromethane and the
solution was cooled to 0-5.degree. C. AlCl.sub.3 was added and the
reaction mixture was stirred for 0.5 hour at 0-5.degree. C.
followed by 7 hours' stirring at ambient temperature. The solution
was evaporated to dryness at 30.degree. C. (in a water bath) and
water was added. The obtained grey solid was filtered, washed with
1N HCl and dried at 30.degree. C. in a vacuum oven.
Di-acetate-laquinimod (Compound 4)
[0695] Acetic anhydride was added to a solution of 5-HLAQ crude in
pyridine and the reaction mixture was stirred for 1 hour at room
temperature. The pyridine was evaporated to dryness and the oily
residue was dissolved in dichloromethane. The organic solution was
washed with 1N HCl followed by aqueous washings. The crude 2 was
purified by flash chromatography on silica gel (mobile phase: 1%
methanol in dichloromethane).
5-HLAQ (Compound 5)
[0696] 1N NaOH solution was added to a suspension of 4 in ethanol.
The reaction mixture was stirred for 40 min and the ethanol was
evaporated. The residue was acidified with 5N HCl down to pH 1-2
and the white solid was filtered, washed with water and dried.
5-HLAQ was obtained as a white solid with a purity of >99% by
HPLC.
[0697] Material of sufficient purity to be suitable for use as a
reference standard was obtained.
[0698] 5-HLAQ was characterized by NMR, MS, elemental analysis and
FT-IR as detailed below.
Nuclear Magnetic Resonance (NMR) Spectroscopy
[0699] The .sup.1H-NMR and .sup.13C-NMR characterization of 5-HLAQ
was performed in DMSO on a Bruker 300 MHz instrument. The peak
assignments are summarized in Table 22. The .sup.1H-NMR and
.sup.13C-NMR spectra are presented in FIG. 21 and FIG. 22,
respectively.
TABLE-US-00023 TABLE 22 .sup.13C and .sup.1H-NMR Peak Assignments
for 5-HLAQ .sup.13C (ppm) .sup.1H (ppm) Multiplicity (Hz) *
N--CH.sub.3 29.09 3H 3.39 brs C # 2 157.95 -- -- C # 3 102.85 -- --
C # 4 159.65 -- -- C # 4a -- -- -- C # 5 155.20 -- -- C # 6 107.79
1H 6.84 d (8.0) C # 7 131.91 1H 7.36 t (8.0) C # 8 106.35 1H 6.61 d
(8.0) C # 8a 140.47 -- -- C # 9 158.79 -- -- N--CH.sub.2--CH.sub.3
42.81 2H 3.76 dq (7) N--CH.sub.2--CH.sub.3 12.91 3H 1.08 t (7) C #
1{grave over ( )} 141.59 -- -- C # 2{grave over ( )}/6{grave over (
)} 126.84 1H 7.3 m C # 3{grave over ( )}/5{grave over ( )} 128.46
1H 7.3 m C # 4{grave over ( )} 127.31 1H 7.18 m * brs = broad
singlet, d = doublet, t = triplet, s = singlet, m = multiplet, dq =
double quartet
Mass Spectroscopy (MS)
[0700] The mass spectrum of 5-HLAQ was obtained on a Q-TOF
Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray
ionization in positive ion mode (ES.sup.+).
[0701] The spectrum shown in FIG. 23 is in agreement with the
molecular weight of 5-HLAQ. The attribution of the main signals in
ES.sup.+ mass spectrum of 5-HLAQ is presented in Table 23
below.
TABLE-US-00024 TABLE 23 Attribution of Main Peaks of ES.sup.+ Mass
Spectrum of 5-HLAQ m/z Attribution 339 [M + H].sup.+ 361 [M +
Na].sup.+
Elemental Analysis
[0702] The test for elemental analysis was performed on a
Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight
percentages are accurate and reproducible to within .+-.0.3%. The
accuracy for halogens is .+-.0.5%. The results for 5-HLAQ are
presented in Table 24.
TABLE-US-00025 TABLE 24 Elemental Analysis Results for 5-HLAQ
Element % C % H % N Theoretical 67.44 5.36 8.28 Experimental 67.05
5.06 8.07
[0703] The attenuated total reflectance (ATR) FT-IR spectrum of
5-HLAQ was measured with a Nicolet 6700 "Thermo Scientific" FT-IR
apparatus. A representative spectrum is presented in FIG. 24. A
summary of the band assignments is provided in Table 25.
TABLE-US-00026 TABLE 25 Summary of IR Band Assignments of 5-HLAQ
Transition Energy (cm.sup.-1) Band Assignment 686-1257 Aromatic
C--H deformation vibration 1019-1416 C--N stretching 1157-1249
Aromatic C--OH stretching 1586-1631 Aromatic C--C stretching,
Aromatic C.dbd.C stretching 1659 C.dbd.O stretching
Example 12
Preparation Of DELAQ
[0704] DELAQ is an off-white to pale yellow solid. The molecular
structure, chemical formula and molecular weight of DELAQ are
provided below.
##STR00029##
[0705] DELAQ was prepared in the following manner:
[0706] A mixture of MCQME, n-heptane, n-octane and aniline was
heated in a jacketed reactor to Tj=135.degree. C. and the volatiles
were distilled for 3 hours. The distillation was stopped and
additional portions of n-heptane and aniline were added to the
mixture. The distillation was continued for an additional 2 hours
at Tj=135.degree. C. The reaction mixture was cooled to room
temperature and n-heptane was added. The crystalline solid formed
was collected by filtration and washed with n-heptane. The wet
material was dried in a vacuum oven at 50.degree. C.
[0707] Material of sufficient purity to be suitable for use as a
reference standard was obtained.
[0708] DELAQ was characterized by NMR, MS, elemental analysis and
FT-IR, as detailed below.
Nuclear Magnetic Resonance (NMR) Spectroscopy
[0709] The .sup.1H-NMR and .sup.13C-NMR characterization of DELAQ
was performed in CDCl.sub.3 on a Bruker 300 MHz instrument. The
peak assignments are summarized in Table 26. The .sup.1H-NMR and
.sup.13C-NMR spectra are presented in FIG. 25 and FIG. 26,
respectively.
TABLE-US-00027 TABLE 26 .sup.13C and .sup.1H-NMR Peak Assignments
for DELAQ .sup.13C (ppm) .sup.1H (ppm) Multiplicity (Hz) *
N--CH.sub.3 30.23 3H 3.68 s NH -- 12.57 brs C # 2 162.09 -- -- C #
3 96.84 -- -- C # 4 173.81 -- -- C # 4a 113.50 -- -- C # 5 134.51
-- -- C # 6 126.61 1H 7.27 m C # 7 132.89 1H 7.47 m C # 8 113.81 1H
7.27 m C # 8a 142.08 -- -- C # 9 169.61 -- -- C # 1{grave over ( )}
137.01 -- -- C # 2{grave over ( )}/6{grave over ( )} 121.29 1H 7.65
m C # 3{grave over ( )}/5{grave over ( )} 129.02 1H 7.36 m C #
4{grave over ( )} 124.95 1H 7.17 m * brs = broad singlet, s =
singlet, m = multiplet,
Mass Spectroscopy (MS)
[0710] The mass spectrum of DELAQ was obtained on a Q-TOF
Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray
ionization in positive ion mode (ES.sup.+). The ratio between MS
signals containing .sup.35Cl and .sup.37Cl corresponds to the
natural abundances of these isotopes (about 3:1). The spectrum
shown in FIG. 27 is in agreement with the molecular weight of
DELAQ. The attribution of the main signals in ES.sup.+ mass
spectrum of DELAQ is presented in Table 27.
TABLE-US-00028 TABLE 27 Attribution of Main Peaks of ES.sup.+ Mass
Spectrum of DELAQ m/z Attribution 236 [M(.sup.35Cl) +
H--(CH.sub.3)--(Ph)].sup.+ 238 [M(.sup.37Cl) +
H--(CH.sub.3)--(Ph)].sup.+ 329 [M(.sup.35Cl) + H].sup.+ 331
[M(.sup.37Cl) + H].sup.+
Elemental Analysis
[0711] The test for elemental analysis was performed on a
Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight
percentages are accurate and reproducible to within .+-.0.3%. The
accuracy for halogens is .+-.0.5%. The results for DELAQ are
presented in Table 28.
TABLE-US-00029 TABLE 28 Elemental Analysis Results for DELAQ
Element % C % H % N % Cl Theoretical 62.11 3.99 8.52 10.78
Experimental 61.90 3.83 8.38 10.96
Infrared Spectroscopy
[0712] The FT-IR spectrum of DELAQ was measured with a Nicolet 6700
"Thermo Scientific" FT-IR apparatus. A representative spectrum is
presented in FIG. 28. A summary of the band assignments is provided
in Table 29.
TABLE-US-00030 TABLE 29 Summary of IR Band Assignments of DELAQ in
KBr Transition Energy (cm.sup.-1) Band Assignment 686-1203 Aromatic
C--H deformation vibration 735-833 N--H deformation vibration
1077-1089 C--Cl stretching 1203-1449 C--N stretching 1203 Aromatic
C--OH stretching 1489-1620 Aromatic C--C stretching, Aromatic
C.dbd.C stretching 1659 C.dbd.O stretching 2951 COO--H
stretching
Example 13
Preparation Of 3-HLAQ
[0713] 3-HLAQ is a yellow solid. The molecular structure, chemical
formula and molecular weight of 3-HLAQ are provided below.
##STR00030##
[0714] 3-HLAQ was prepared in the following manner:
[0715] A solution of laquinimod sodium in water was added drop-wise
to a solution of disodium hydrogenphosphate dihydrate and oxone
(2KHSO.sub.5.KHSO.sub.4.K.sub.2SO.sub.4) in water. 10% NaOH was
added until the pH was 8.0. The solution was mixed for 30 minutes
at room temperature. The obtained solid was filtered, washed with
water and dried in a vacuum oven at 50.degree. C. The crude product
was recrystallized twice from ethyl acetate and n-heptane mixture.
Each time the product was isolated by cooling the mixture to
10.degree. C. and stirring for 1 hour. The obtained crystals were
filtered off, washed with n-heptane and dried in a vacuum oven at
50.degree. C.
[0716] Material of sufficient purity to be suitable for use as a
reference standard was obtained.
[0717] 3-HLAQ was characterized by NMR, MS, elemental analysis and
FT-IR, as detailed below.
Nuclear Magnetic Resonance (NMR) Spectroscopy
[0718] The .sup.1H-NMR and .sup.13C-NMR characterization of 3-HLAQ
was performed in CDCl.sub.3 on a Bruker 300 MHz instrument. The
peak assignments are summarized in Table 30. The .sup.1H-NMR and
.sup.13C-NMR spectra are presented in FIG. 29 and FIG. 30,
respectively.
TABLE-US-00031 TABLE 30 .sup.1H and .sup.13C -NMR Peak Assignments
for 3-HLAQ .sup.13C (ppm) .sup.1H (ppm) Multiplicity (Hz) *
N--CH.sub.3 30.56 3H 3.21 s C # 2 165.45 -- -- C # 3 77.92 -- -- C
# 4 188.90 -- -- C # 4a 136.16 -- -- C # 5 ca. 131.6 -- -- C # 6
126.75 1H 7.14 d (8.0) C # 7 134.90 1H 7.35 t (8.0) C # 8 113.78 1H
6.84 d (8.0) C # 8a 144.51 -- -- C # 9 167.87 -- --
N--CH.sub.2--CH.sub.3 46.96 2H 3.79 q (6.0) N--CH.sub.2--CH.sub.3
12.55 3H 1.16 bt (6.0) C # 1{grave over ( )} 137.19 -- -- C #
2{grave over ( )}/6{grave over ( )} 131.62 1H 7.07-7.15 m C #
3{grave over ( )}/5{grave over ( )} 128.91 1H 7.07-7.15 m C #
4{grave over ( )} 129.11 1H 7.07-7.15 m * d = doublet, t = triplet,
s = singlet, m = multiplet, q = quartet, bt = broad triplet
Mass Spectroscopy (MS)
[0719] The mass spectrum of 3-HLAQ was obtained on a Q-TOF
Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray
ionization in positive ion mode (ES.sup.+). The ratio between MS
signals containing .sup.35Cl and .sup.37Cl corresponds to the
natural abundances of these isotopes (about 3:1). The spectrum
shown in FIG. 31 is in agreement with the molecular weight of
3-HLAQ. The attribution of the main signals in ES.sup.+ mass
spectrum of 3-HLAQ is presented in Table 31.
TABLE-US-00032 TABLE 31 Attribution of Main Peaks of ES.sup.+ Mass
Spectrum of 3-HLAQ m/z Attribution 120 ##STR00031## 148
##STR00032## 373 [M(.sup.35Cl) + H].sup.+ 375 [M(.sup.37Cl) +
H].sup.+ 395 [M(.sup.35Cl) + Na].sup.+ 397 [M(.sup.37Cl) +
Na].sup.+
Elemental Analysis
[0720] The test for elemental analysis was performed on a
Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight
percentages are accurate and reproducible to within .+-.0.3%. The
accuracy for halogens is .+-.0.5%. The results for 3-HLAQ are
presented in Table 32.
TABLE-US-00033 TABLE 32 Elemental Analysis Results for 3-HLAQ
Element % C % H % N % Cl Theoretical 61.21 4.60 7.51 9.51
Experimental 61.09 4.68 7.38 9.30
Infrared Spectroscopy
[0721] The attenuated total reflectance (ATR) FT-IR spectrum of
3-HLAQ was measured with a Nicolet 6700 "Thermo Scientific" FT-IR
apparatus. A representative spectrum is presented in FIG. 32. A
summary of the band assignments is provided in Table 33.
TABLE-US-00034 TABLE 33 Summary of IR Band Assignments of 3-HLAQ
Transition Energy (cm.sup.-1) Band Assignment 794-1244 Aromatic
C--H deformation vibration 1035-1116 C--Cl stretching 1061-1417
C--N stretching 1458-1480 CH.sub.2 deformation vibration 1583
Aromatic C--C stretching, Aromatic C.dbd.C stretching 1663-1693
C.dbd.O stretching 2979 C--H stretching in N--CH.sub.2
Example 14
Preparation Of SPIRO-LAQ
[0722] SPIRO-LAQ is a white powder. The molecular structure,
chemical formula and molecular weight of SPIRO-LAQ are provided
below.
##STR00033##
[0723] SPIRO-LAQ was prepared in the following manner:
[0724] A mixture of laquinimod (free acid), ammonium cerium (IV)
nitrate, ethanol and acetic acid in water was mixed for 1 hour.
Then the precipitate was washed and filtered with water followed by
ethanol, and dried in a vacuum oven at 50.degree. C.
[0725] Material of sufficient purity to be suitable for use as a
reference standard was obtained.
[0726] SPIRO-LAQ was characterized by NMR, MS, elemental analysis
and FT-IR as detailed below.
Nuclear Magnetic Resonance (NMR) Spectroscopy
[0727] The .sup.1H-NMR and .sup.13C-NMR characterization of
SPIRO-LAQ was performed in DMSO on a Bruker 300 MHz instrument. The
peak assignments are summarized in Table 34. The .sup.1H-NMR and
.sup.13C-NMR spectra are presented in FIG. 33 and FIG. 34,
respectively.
TABLE-US-00035 TABLE 34 .sup.13C and .sup.1H-NMR Peak Assignments
for SPIRO-LAQ .sup.13C (ppm) .sup.1H (ppm) Multiplicity (Hz) *
N--CH.sub.3 30.95 3H 3.44 brs C # 2 164.10 -- -- C # 3 71.76 -- --
C # 4 185.60 -- -- C # 4a 124.66 -- -- C # 5 133.49 -- -- C # 6
126.13 1H 7.54 d (8.0) C # 7 136.03 1H 7.74 t (8.0) C # 8 115.77 1H
7.20 d (8.0) C # 8a 145.62 -- -- N--CH.sub.2--CH.sub.3 34.90 2H
3.75 q (8.0) N--CH.sub.2--CH.sub.3 12.40 3H 1.16 t (8.0) C #
2{grave over ( )} 167.40 -- -- C # 3{grave over ( )}a 117.83 -- --
C # 4{grave over ( )} 124.87 1H 7.20 d (8.0) C # 5{grave over ( )}
122.84 1H 7.38 ddd (8.0, 6.5, 2.5) C # 6{grave over ( )} 129.84 1H
7.0-7.1 m C # 7{grave over ( )} 109.64 1H 7.0-7.1 m C # 7{grave
over ( )}a 143.17 -- -- * brs = broad singlet, d = doublet, t =
triplet, s = singlet, m = multiplet, dd = double doublet, ddd =
double double doublet, q = quartet
Mass Spectroscopy (MS)
[0728] The mass spectrum of SPIRO-LAQ was obtained on a Q-TOF
Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray
ionization in positive ion mode (ES.sup.+). The ratio between MS
signals containing .sup.35Cl and .sup.37Cl corresponds to the
natural abundances of these isotopes (about 3:1). The spectrum
shown in FIG. 35 is in agreement with the molecular weight of
SPIRO-LAQ. The attribution of the main signals in ES.sup.+ mass
spectrum of SPIRO-LAQ is presented in Table 35.
TABLE-US-00036 TABLE 35 Attribution of Main Peaks of ES.sup.+ Mass
Spectrum of SPIRO-LAQ m/z Attribution 311 [M(.sup.35Cl) +
H--(CH.sub.2CH.sub.3)--(CH.sub.3)].sup.+ 313 [M(.sup.37Cl) +
H--(CH.sub.2CH.sub.3)--(CH.sub.3)].sup.+ 355 [M(.sup.35Cl) +
H].sup.+ 357 [M(.sup.37Cl) + H].sup.+
Elemental Analysis
[0729] The test for elemental analysis was performed on a
Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight
percentages are accurate and reproducible to within .+-.0.3%. The
accuracy for halogens is .+-.0.5%. The results for SPIRO-LAQ are
presented in Table 36.
TABLE-US-00037 TABLE 36 Elemental Analysis Results for SPIRO-LAQ
Element % C % H % N % Cl Theoretical 64.32 4.26 7.90 9.99
Experimental 64.08 4.22 7.75 10.12
Infrared Spectroscopy
[0730] The attenuated total reflectance (ATR) FT-IR spectrum of
SPIRO-LAQ was measured with a Nicolet 6700 "Thermo Scientific"
FT-IR apparatus. A representative spectrum is presented in FIG. 36.
A summary of the band assignments is provided in Table 37.
TABLE-US-00038 TABLE 37 Summary of IR Band Assignments of SPIRO-LAQ
Transition Energy (cm.sup.-1) Band Assignment 687-1244 Aromatic
C--H deformation vibration 1020-1091 C--Cl stretching 1020-1454
C--N stretching 1153-1278 Aromatic C--OH stretching 1582 Aromatic
C--C stretching, Aromatic C.dbd.C stretching 1667-1738 C.dbd.O
stretching
Example 15
Preparation of MEG-LAQ
[0731] The molecular structure, chemical formula and molecular
weight of MEG-LAQ is presented below.
##STR00034##
An aqueous solution of laquinimod sodium and meglumine was refluxed
and bubbled with air for about 1 month. The obtained solution was
diluted twice with water and acidified with concentrated
hydrochloric acid to pH 1-2. The aqueous solution was filtered
followed by extraction with chloroform. Then, a concentrated
ammonium hydroxide solution was added to the aqueous solution, up
to neutralization. The solution was evaporated and the obtained
brown syrup was washed with methanol. Meglumine was solidified and
filtered followed by silica gel addition to the methanolic
solution. The solvent was evaporated and the obtained mixture was
purified by silica-gel column chromatography (mobile phase: 20%
methanol in dichloromethane).
[0732] This synthesis and purification affords a material of
sufficient purity to be suitable for use as a standard.
Example 16
Preparation of BH-3-HLAQ
[0733] The molecular structure, chemical formula and molecular
weight of BH-3-HLAQ is presented below.
##STR00035##
[0734] A 1N NaOH solution was added to a suspension of 3-HLAQ in
water. The yellow solution was stirred for 0.5 hr followed by the
addition of 1N HCl solution. The aqueous solution that contained
white solid was extracted with ethyl acetate. The organic phase was
washed with brine and dried over sodium sulfate. The suspension was
filtered, evaporated to dryness and the solid residue was purified
by crystallization from IPA:water mixture (1:3.75 v/v). The
suspension was cooled to 0-5.degree. C. and was kept for 1 hr,
filtered and washed with IPA:water mixture (1:10 v/v). The obtained
white solid was dried in a vacuum oven at 50.degree. C.
[0735] At the time of initial qualification, the BH-3-HLAQ standard
was demonstrated to conform to the molecular structure by IR,
elemental analysis, MS, and NMR. Additional testing included
chromatographic purity and loss on drying.
[0736] This synthesis and purification affords a material of
sufficient purity to be suitable for use as a standard.
[0737] For structure elucidation purposes, a sample of BH-3-HLAQ
was characterized by NMR, MS, elemental analysis and FT-IR as
depicted below.
NMR Spectroscopy
[0738] The .sup.1H-NMR and .sup.13C-NMR characterization of
BH-3-HLAQ was performed in DMSO on a Bruker 300 MHz instrument. The
peak assignments are summarized in Table 38. The .sup.1H-NMR and
.sup.13C-NMR spectra are presented in FIG. 37 and FIG. 38,
respectively.
TABLE-US-00039 TABLE 38 .sup.13C and .sup.1H-NMR Peak Assignments
for BH-3-HLAQ .sup.13C (ppm) .sup.1H (ppm) Multiplicity (Hz) *
CO.sub.2H 165.62 -- -- C # 1 130.08 -- -- C # 2 135.13 -- -- C # 3
130.20 1H 7.77 d (8.0) C # 4 131.66 1H 7.66 t (8.0) C # 5 129.38 1H
7.19 d (8.0) C # 6 140.13 -- -- N--CH.sub.3 37.43 3H 3.09 -- C # 1'
167.55 -- -- C # 2' 68.67 1H 4.25 s C # 3' 168.93 -- --
NCH.sub.2CH.sub.3 43.43 2H 3.43, 3.38 ABq of q (14.0, 7.0, 2.0)
NCH.sub.2CH.sub.3 12.24 3H 0.86 t (7.0) C # 7 137.34 -- -- C # 8/12
127.86 2H 6.46 br C # 9/11 128.93 2H 7.27 m C # 10 127.80 1H 7.27 m
* s = singlet, br = broad, m = multiplet, d = doublet, t = triplet,
q = quartet
Mass Spectrometry
[0739] The mass spectrum of BH-3-HLAQ was obtained on a Q-TOF
Micro-TM-MICROMASS (TOF) mass spectrometer, using electrospray
ionization in positive ion mode (ES.sup.+). The spectrum shown in
FIG. 39 is in agreement with the molecular weight of BH-3-HLAQ. The
attribution of the main signals in ES.sup.+ mass spectrum of
BH-3-HLAQ is presented in Table 39.
TABLE-US-00040 TABLE 39 Attribution of Main Peaks of ES.sup.+ Mass
Spectrum of BH-3-HLAQ m/z Attribution 391 [M + H].sup.+ 413 [M +
Na].sup.+
FT-IR
[0740] The attenuated total reflectance (ATR) FT-IR spectrum of
BH-3-HLAQ was measured with a Nicolet 6700 "Thermo Scientific"
FT-IR apparatus. FIG. 40 shows a typical spectrum. A summary of the
band assignments is shown in Table 40.
TABLE-US-00041 TABLE 40 Summary of IR Band Assignments of BH-3-HLAQ
Transition Energy (cm.sup.-1) Band Assignment 703, 737, 769
Aromatic C--H deformation vibration 1090 Aromatic C--Cl stretching
1492 Aromatic C.dbd.C stretching 1586 C--N stretching 1652, 1728
C.dbd.O stretching 3081 Aromatic C--H stretching 3442 O--H
stretching
Elemental Analysis
[0741] The test for elemental analysis was performed on a
Perkin-Elmer 2400 Series II C H N Analyzer. C, H and N weight
percentages are accurate and reproducible to within .+-.0.3%. The
accuracy for halogens is .+-.0.5%. The results for BH-3-HLAQ are
presented in Table 41.
TABLE-US-00042 TABLE 41 Elemental Analysis Results for BH-3-HLAQ
Element % C % H % N % Cl Theoretical 58.39 4.90 7.17 9.07
Experimental 58.31 4.79 7.15 9.56
Example 17
Specification of Laquinimod Sodium Drug Substance
##STR00036##
[0743] Laquinimod sodium is a white to off-white powder, freely
soluble in water which should be stored in a well-closed container,
protected from light, at room temperature.
[0744] The specifications of Laquinimod sodium are as follows:
TABLE-US-00043 TABLE 42 TEST SPECIFICATIONS Description White to
off-white powder Identification: a) HPLC (refer to The retention
time of the main peak Example 19) in the sample chromatogram
corresponds to that of the Standard. (See, Example 19) b) Infrared
Spectrum The transmission spectrum exhibits minima at the same
wavenumbers as the reference standard. c) Ultraviolet spectrum The
absorption spectrum exhibits maxima at the same wavelengths as the
reference standard. d) sodium identification Conforms. Assay of
LAQ-Na (by HPLC, 98.0-102.0% refer to Example 19) Related
Substances a) Polar impurities (refer MCQ - NMT 0.15% to Table 43)
MCQCA - NMT 0.15% MCQME - NMT 0.12% 5-HLAQ - NMT 0.10% MCQEE - NMT
0.10% Any other impurity - NMT 0.10% Total - NMT 1.00% b)
N-ethylaniline and N-ethylaniline - NMT 0.10% non-polar impurities
Any other impurity - NMT 0.10% (refer to Table 43) Total - NMT
0.50% c) DELAQ content (by DELAQ - NMT 0.1% HPLC) LAQ content (by
HPLC) NMT 1.0% Dimethyl Malonate NMT 0.1% content (by HPLC)
Dimethyl sulfate NMT 1 ppm content (by LC-MS) Water content (by KF
NMT 1.5% (w/w) coulometric) Heavy metals NMT 20 ppm Sodium Content
5.8-6.4% Color of solution Not more intensely colored than
reference solution BY.sub.7 Free Chloride NMT 0.1% Residual
solvents Ethanol - NMT 5000 ppm (by GC) n-Heptane - NMT 5000 ppm
n-Octane - NMT 2000 ppm Methanol - NMT 5000 ppm Acetone - NMT 5000
ppm Dioxane - NMT 380 ppm Residual solvent - DMF NMT 880 ppm (by HS
GC) Microbial purity test Total viable aerobic count - NMT 10.sup.3
cfu/g Fungi/yeast and mold - NMT 10.sup.2 cfu/g Escherichia Coli -
Absence/g For MCQME, alternative specifications are NMT 10% and LT
10%.
[0745] Table 43 lists the non-pharmacopoeial analytical methods
currently in use for the determination of impurity levels in the
drug substance at the time of release and during stability testing.
The method numbers and the detection and quantitation limits for
each impurity are also listed.
TABLE-US-00044 TABLE 43 Analytical Methods for Determination of
Impurities Method Quantitation Detection Parameter Method.sup.1
Type Limit Limit MCQ Example 19 RP-HPLC 0.05% 0.02% MCQCA Example
19 RP-HPLC 0.05%.sup.2 0.02% MCQME Example 19 RP-HPLC 0.05% 0.02%
5-HLAQ Example 19 RP-HPLC 0.05% 0.02% MCQEE Example 19 RP-HPLC
0.05% 0.02% LAQ Example 19 RP-HPLC 0.2% 0.03% NEA Example 20
RP-HPLC 0.06% 0.02%
Example 18
Specification of Laquinimod Sodium Drug Product
[0746] Laquinimod drug product was prepared as 0.6 mg capsules of
laquinimod sodium. The specifications of laquinimod sodium (0.6 mg)
capsules are as follows:
TABLE-US-00045 TABLE 44 Release and Shelf Life Specifications Test
Use.sup.1 Acceptance Criteria Description R Off-white opaque cap
and body, hard gelatin capsule filed with white to off-white
granulate. Imprinted "0.6 mg" on the body, and "LAQ" on the cap.
Appearance S Off-white opaque cap and body, hard gelatin capsule
filed with white to off-white granulate. Imprinted "0.6 mg" on the
body, and "LAQ" on the cap. No cracks, discoloration and other
appearance defects are Observed Identification R By HPLC (Example
21): The retention time of the major peak in the chromatogram of
sample preparation corresponds to that in the chromatogram of the
standard preparation. R By UV Spectrum: The sample spectrum
exhibits the same maxima as that of the standard Dissolution R + S
Not less than 70% (Q) of the labeled amount is dissolved in 30 min.
Conforms to current Ph. Eur. (2.9.3) Uniformity of R Conforms to
current Ph. Eur. (2.9.40) Dosage Units by Content Uniformity Assay
R + S 95.0-105.0% of the labeled amount (By HPLC, Example 21)
Impurities/ R + S Sum of MCQ.sup.2 and MCQCA.sup.3: NMT 0.5%
Degradation 5-HLAQ.sup.4: NMT 0.5% Product Any Other: NMT 0.5%
Determination MEG - LAQ.sup.5: NMT 1.0% (See, Table 46
BH-3-HLAQ.sup.6: NMT 1.0% for methods) Water Content R NMT 0.8% S
NMT 1.5% Identification R Positive for the correct color of
Color.sup.7 Microbiological R + S Total aerobic microbial count
(TAMC): Examination of NMT 10.sup.3 cfu/g Non-Sterile Total
combined yeasts/moulds count (TYMC): Products.sup.8 NMT 10.sup.2
cfu/g Absence of Escherichia Coli in 1 g .sup.1R = Release; S =
Stability .sup.2MCQ: 5-chloro-4hydroxy-1-methylquinolin-2(1H)-one
.sup.3MCQCA:
5-chloro-4-hydroxy-1methyl-2-oxo-1,2-dihydroquinoline-3-carboxylic
acid .sup.45-HLAQ:
N-ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carb-
oxamide .sup.5MEG-LAQ: N-ethyl-4-hydroxy-1
methyl-5-(methyl(2,3,4,5,6-pentahydroxyhexyl)amino)-2-oxo-N-phenyl
1,2-dihydroquinoline-3-Carboxamide. .sup.6BH-3-HLAQ:
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl-3-oxopropanamido)be-
nzoic acid .sup.7Frequency: Pilot batches, first three production
batches and occasional test upon request thereafter.
.sup.8Frequency: For Release: Pilot batches, first 3 production
batches and every fifth batch thereafter, or at least one batch per
year if less than 5 batches per year are manufactured. For
Stability: a) For long term testing at zero time and annually
and/or expiry date. b) For intermediate testing at zero time and 12
months. c) For accelerated testing at zero time and 6 months.
[0747] Finished product stability protocol package sizes were the
largest and smallest of each marketing container/closure and/or
unidose configuration.
TABLE-US-00046 TABLE 45 Stability Batch Storage Conditions Test
Intervals Frequency Long-term testing 0, 3, 6, 9, 12, 18, 24 Pilot
batches, first 25 .+-. 2.degree. C./ and 36 months and/or three
production 60 .+-. 5% RH at expiry date.sup.9 batches, an annual
batch thereafter and variations as requested. Accelerated testing
0, 1, 2, 3, and 6 Pilot batches and 40 .+-. 2.degree. C. months
variations as requested. 75 .+-. 5% RH Accelerated testing 0, 3,
and 6 months First three 40 .+-. 2.degree. C./ production batches.
75 .+-. 5% RH Intermediate testing.sup.10 0, 3, 6, 9 and 12 Pilot
batches, first 30 .+-. 2.degree. C./ months three production 65
.+-. 5% RH batches, and variations as requested. .sup.9For
validation batches and commercial batches only. .sup.10Only if
accelerated testing fails.
[0748] The following examples describe non-pharmacopoeial
analytical methods for the determination of impurity levels in the
drug product at the time of release and during stability testing.
The method numbers and the detection and quantitation limits for
each impurity are also listed, as well as the corresponding
validation numbers.
TABLE-US-00047 TABLE 46 Analytical Methods for Determination of
Impurities in the Drug Product Example Method Quantitation
Detection Parameter Number Type Limit Limit MCQ & MCQCA Example
21 RP-HPLC 0.1% 0.03% 5-HLAQ Example 21 RP-HPLC 0.05% 0.02%
NEA.sup.1 Example 22 RP-HPLC 0.1% 0.03% 3-HLAQ.sup.1 Example 22
RP-HPLC 0.1% 0.03% MEG-LAQ Example 23 RP-HPLC 0.2% 0.1% BH-3-HLAQ
Example 23 RP-HPLC 0.05% 0.02% .sup.1NEA and 3-HLAQ were omitted
from the commercial specifications since they were never observed.
The monitoring of these impurities in the primary stability batches
will continue until completion of the stability programs, using the
listed method.
Example 19
Method of Determining Amount of MCQ, MCQCA, 5-HLAQ, MCQME and MCQEE
in a Sample of Laquinimod Drug Substance
[0749] Laquinimod sodium drug substance was analyzed by HPLC. The
amount of MCQ, MCQCA, MCQME, MCQEE and 5-HLAQ was determined using
the following HPLC method.
1. HPLC Conditions
TABLE-US-00048 [0750] Column & Packing Inertsil ODS-3V, 5
.mu.m, 4.6 .times. 250 mm, GL Sciences Guard column Opti-Guard C
18, 1 mm Detection UV at 240 nm Flow rate 1.5 mL/min Injection
volume 50 .mu.L Column temperature 40.degree. C. Autosampler
5.degree. C. temperature Run time 30 minutes (Run time for
standards may be shortened to at least 25 minutes.) Mobile phase
800 mL of Ammonium acetate buffer 1 solution, pH 7.0; and 200 mL
Acetonitrile. Mix well and degas. Buffer pH 7.0 Dissolve 7.7 g of
Ammonium acetate in 2000 mL water and adjust to pH 7.0 .+-. 0.05
with aqueous ammonia or glacial acetic acid. Filter using 0.45
.mu.m membrane filter. Diluent A Acetonitrile/Water (1:1) Diluent B
Mobile phase Blank solution Diluent A/Diluent B (3:7) Note: Adjust
flow rate and mobile phase composition to achieve the required
system suitability parameters.
[0751] All preparations are performed in amber flasks with non
transparent caps. Immediately after preparation place the solutions
in a refrigerator or in a cooled to 5.degree. C. autosampler
rack.
2. Standard Solutions Preparation
2.1 Stock Standard Solution (Solution S)
[0752] Weigh accurately about 15 mg of Laquinimod sodium standard
into a 50 mL volumetric flask. Dilute with diluent A up to 2/3 of
the volume, sonicate for about 2 minutes in cold sonication bath
and dilute to volume with the Diluent A. Concentration of Stock
standard solution is about 300 .mu.g/mL. Stock standard solution
may be stored for one month in refrigerator.
2.2 Standard Solution for Assay (Solution A)
[0753] Dilute 3 mL of the Stock standard solution to 10 mL with
diluent B (dilution factor 3.33). Concentration of Laquinimod
sodium is about 90 .mu.g/mL. Standard solution A may be used for 7
days when stored in a refrigerator.
2.3 MCQCA Stock Standard Solution
[0754] Weigh accurately about 18 mg of MCQCA standard into a 100 mL
volumetric flask. Dilute to volume with acetonitrile, sonicate
until the substance is completely dissolved--stock MCQCA
solution.
[0755] Concentration of MCQCA is about 180 .mu.g/mL.
[0756] MCQCA Stock standard solution should be freshly
prepared.
2.4 Standard Solution for Impurities Determination (Solution I)
[0757] Prepare a solution in diluent B, containing Laquinimod in a
concentration of 0.2% and MCQCA--in a concentration of 0.1%, with
respect to the working concentration of Laquinimod in Standard
solution A. As an example, apply the following procedure.
[0758] Transfer 4.0 mL of Laquinimod sodium standard solution for
assay (Solution A) and 1.0 mL of MCQCA stock standard solution to a
100 mL volumetric flask and dilute to volume with the diluent B
(Intermediate dilution solution).
[0759] Place 2.5 mL of this intermediate dilution into a 50 mL
volumetric flask and make up to volume with the diluent B.
[0760] Total dilution factor for Laquinimod standard is 1666.67,
for MCQCA 2000.
[0761] Concentration of Laquinimod sodium is about 0.18 .mu.g/mL
(0.2%).
[0762] Concentration of MCQCA is about 0.09 .mu.g/mL (0.1%, QL
level).
[0763] Standard solution I may be used for 24 hours when stored in
a refrigerator.
3. Resolution Solution Preparation
3.1 Mixed Solution
[0764] Prepare a solution of the following impurities in the
Diluent A:
TABLE-US-00049 Abbreviation Full name MCQ
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one MCQCA
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-
3-carboxylic acid MCQME Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-
dihydroquinoline-3-carboxylate 5-HLAQ
N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-
dihydroquinoline-3-carboxamide MCQEE Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-
dihydroquinoline-3-carboxylate
[0765] Mixed solution may be prepared as follows:
[0766] Weigh about 3 mg of each impurity to a 100 mL volumetric
flask, dissolve and dilute to volume with the Diluent A--Mixed
solution. Concentration of each impurity in the Mixed solution is
about 30 .mu.g/mL. The Mixed solution is used only for the
determination of the exact retention times of the known impurities
and may be used up to four months when stored at about -20.degree.
C. For this purpose, the freshly prepared Mixed solution is divided
into aliquots and stored immediately at about -20.degree. C. After
thawing, the aliquots should not be refrozen.
3.2 Resolution Solution
[0767] Transfer 3 mL of the stock standard solution (Solution S)
and 0.3 mL of the Mixed solution to a 10 mL volumetric flask and
dilute to volume with the Diluent B. Concentration of Laquinimod
sodium form is about 90 .mu.g/mL. Concentration of each impurity is
about 0.9 .mu.g/mL (1% with respect to Laquinimod sodium).
Resolution solution may be used for 9 days if stored in
refrigerator.
4. Sample Preparation
[0768] Weigh accurately about 15 mg of Laquinimod sodium Cryst or
DS into a 50 mL volumetric flask. Dilute with the Diluent A up to
2/3 of the volume, sonicate for about 2 minutes in cold sonication
bath and dilute to volume with the Diluent A--Stock sample
solution.
[0769] Dilute 3 mL of the Stock sample solution to 10 mL with the
Diluent B--Working sample solution (dilution factor 3.33).
[0770] Concentration of Laquinimod sodium is about 90 .mu.g/mL.
[0771] The sample must be analyzed not later than within 24 hours
after preparation.
5. Procedure
[0772] Inject the Resolution solution, Blank, Standard solutions
for assay and IDD, Laquinimod sample solutions according to the
relevant SOP's.
6. System Suitability Test
6.1 Resolution Test
[0773] 1. The typical retention time for the Laquinimod peak is
15.5.+-.2.0 minutes.
[0774] 2. The tailing factor for the Laquinimod peak should be not
more than 2.0.
[0775] 3. Resolution factor of NLT 2 between the peaks should be
achieved.
[0776] 4. The typical RRT of the specified impurity peaks should be
as the following:
TABLE-US-00050 IMPURITY Typical RRT MCQME 0.33-0.38 MCQ 0.49-0.58
MCQEE 0.56-0.65 MCQCA 0.72-0.85 5-HLAQ 1.2-1.4* *Retention time of
5-HLAQ should be not more than 23 minutes.
6.2 System Precision Test
[0777] Evaluate Laquinimod standards for assay and IDD in order to
test the system precision according to the relevant SOP's.
[0778] MCQCA in Solution I is used to test the sensitivity of the
system. The RSD of the area of the six injections of Std 1 as well
as the difference between Std1 and Std2 should be NMT 20%.
7. Identification by Retention Time (RT)
[0779] The RT of the main (Laquinimod) peak obtained in the sample
chromatogram should correspond to that obtained for the Standard
solution.
8. Calculation and Report
8.1 Assay Calculation for Laquinimod Sodium
[0780] Area of smp .times. Conc . of std .times. 100 Area of std (
sol A ) .times. Conc . of smp = % Assay of Laquinimod sodium
##EQU00004##
[0781] Report the calculated result of the Assay on dry basis after
subtraction of Laquinimod acid content.
8.2 Evaluation and Calculation of Impurities
[0782] Calculations are performed with respect to the diluted
Laquinimod sodium standard (Solution I).
[0783] Quantitation level of MCQME, MCQ, MCQEE, 5-HLAQ and unknown
impurities is 0.05%.
[0784] Detection level of MCQME, MCQ, MCQEE, 5-HLAQ and unknown
impurities is 0.02%.
[0785] Reporting level of MCQCA is 0.10%.
[0786] Detection level of MCQCA is 0.03%
[0787] Adjust integration parameters to reject peaks with the area
less than 10% of Laquinimod sodium standard (solution I) average
area.
[0788] Correlate the known impurity peaks in the sample
chromatogram with those in the system suitability chromatogram
within .+-.5% of the actual corresponding retention times.
[0789] Calculation of impurities in Laquinimod sodium:
Area of imp .times. R R F .times. Conc . of std .times. 100 Area of
Laq Na std ( Sol I ) .times. Conc . of smp = % Impurityof
Laquinimodsodium ##EQU00005##
[0790] RRF--relative response factors of impurities calculated as
the following ratio: slope of Laquinimod sodium regression
line/slope of impurity regression line
[0791] Values for Relative Response Factors (RRF) with respect to
Laquinimod sodium are:
TABLE-US-00051 Unknown impurities/ degradation Impurity MCQME MCQ
MCQEE MCQCA products RRF 0.70 0.60 0.80 0.58 1.0
9. Reporting of Impurities
[0792] Report impurities as follows:
TABLE-US-00052 Result Report MCQME, MCQ, MCQEE, .gtoreq.0.05% The
calculated result 5-HLAQ <0.05% <0.05% <0.02% (or ND)
<0.02% MCQCA .gtoreq.0.10% The calculated result <0.10%
<0.10% <0.03% (or ND) <0.03% Unknown impurities
.gtoreq.0.05% The calculated result <0.05% <0.05% <0.02%
(or ND) Not to be reported* Total Variant 1 The sum of calculated
results Variant 2 <0.10% Variant 3 <0.05% Variant 4 <0.03%
*If no unknown impurities were detected, report: Any other
<0.02%. Variant 1: At least one of the impurities .gtoreq.0.05%
or MCQCA .gtoreq.0.10% Variant 2: All impurities <0.05% and
MCQCA <0.10% Variant 3: All impurities <0.05% and MCQCA
<0.03% Variant 4: All impurities <0.02% and MCQCA
<0.03%
[0793] FIG. 41 presents a resolution test chromatogram according to
the above method.
System Suitability Results for FIG. 41.
TABLE-US-00053 [0794] USP USP Int Name RT RRT Area Resolution
Tailing Type 1 MCQME 5.682 0.37 167799 1.1 BB 2 MCQ 8.863 0.57
199156 8.8 1.1 BB 3 MCQEE 9.710 0.63 158206 2.1 1.1 BB 4 MCQCA
11.572 0.75 228929 3.8 1.6 BB 5 Laq 15.481 13270800 6.1 1.6 BB 6
5-HLAQ 20.482 1.32 129109 6.6 1.0 BB
[0795] FIG. 42 presents a diluent chromatogram according to the
above method.
[0796] FIG. 43 presents a typical chromatogram for assay according
to the above method.
Peak Results for FIG. 43.
TABLE-US-00054 [0797] Name RT Area % Area Int Type 1 Laq 15.598
12787417 100.00 BB
[0798] FIG. 44 presents a typical chromatogram for Impurities
testing according to the above method.
Peak Results for FIG. 44.
TABLE-US-00055 [0799] Name RT Area % Area Int Type 1 Laq 15.598
12790697 100.00 BB
Example 20
Method of Determining Amount of NEA, 3-HLAQ and SPIRO-LAQ in a
Sample of Laquinimod Drug Substance
[0800] Laquinimod sodium drug substance was analyzed by HPLC. The
amount of NEA, 3-HLAQ And SPIRO-LAQ was determined using the
following HPLC method.
1. HPLC Conditions
TABLE-US-00056 [0801] Column & Packing Inertsil ODS-3V, 5
.mu.m, 4.6 .times. 250 mm, GL Sciences Guard column Opti-Guard C
18, 1 mm (recommended) Detection UV at 240 nm Flow rate 1.5 mL/min
Injection volume 50 .mu.L Column temperature 40.degree. C.
Autosampler 5.degree. C. temperature Run time 30 minutes (Run time
for standards may be reduced to at least 18 minutes.) Mobile phase
50% Solution A-50% Solution B Solution A 800 mL of Ammonium acetate
buffer solution, pH 7.0; and 200 mL Acetonitrile, degas Solution B
350 mL of Ammonium acetate buffer solution, pH 7.0; and 650 mL
Acetonitrile. Mix Solution B on a magnetic stirrer for about 0.5
hour and then degas. Buffer pH 7.0 Dissolve 7.7 g of Ammonium
acetate in 2000 mL water and adjust to pH 7.0 .+-. 0.05 with
aqueous ammonia or glacial acetic acid. Filter through a 0.45 .mu.m
membrane filter. Diluent A Acetonitrile/Water (1:1) Diluent B
Solution B Note: Adjust flow rate and mobile phase composition to
achieve the required system suitability parameters.
[0802] All the preparations should be performed in amber flasks. It
is recommended to wash volumetric flasks and pipettes with
acetonitrile before use. The use of plastic Pasteur pipettes should
be excluded.
2. Standard Solution Preparation
2.1 NEA Standard Stock Solution
[0803] Weigh accurately about 25 .mu.L (about 25 mg) of
N-Ethylaniline (NEA) standard into a 50 mL volumetric flask. Dilute
to volume with Diluent A.
[0804] Concentration of stock standard solution is about 500
.mu.g/mL.
[0805] NEA stock standard solution may be used for three weeks when
stored in a refrigerator.
2.2 NEA Standard Primary Dilution Solution
[0806] Dilute 0.5 mL of NEA standard stock solution to 50 mL with
Diluent B.
[0807] Concentration of N-Ethylaniline is about 5 .mu.g/mL.
2.3 NEA Working Standard Solution
[0808] Dilute 0.5 mL of NEA standard primary dilution solution to
25 mL with Diluent B (total dilution factor--5000).
[0809] Concentration of N-Ethylaniline is about 0.1 g/mL (about
0.1% of the working concentration).
[0810] NEA working standard solution may be used for four days when
stored in a refrigerator.
3 Resolution Solution Preparation
3.1 Laquinimod Sodium Stock Solution
[0811] Weigh accurately about 15 mg of Laquinimod sodium standard
into a 50 mL volumetric flask. Dilute with Diluent A up to 2/3 of
the volume, sonicate for about 2 minutes in cold sonication bath
and dilute to volume with Diluent A.
[0812] Laquinimod stock solution may be used for one month when
stored in refrigerator.
3.2 SPIRO-LAQ Stock Solution
[0813] Weight about 2.5 mg of SPIRO-LAQ into a 50 mL volumetric
flask. Dissolve and dilute to volume with a mixture of 95%
acetonitrile--5% water.
3.3 3-HLAQ Stock Solution
[0814] Weight about 2.5 mg of 3-HLAQ into a 50 mL volumetric flask.
Dissolve and dilute to volume with 95% acetonitrile--5% water.
3.4 Resolution Solution
[0815] Transfer the following aliquots to 50 mL volumetric flask
and dilute to volume with Diluent B:
[0816] 15 mL of Laquinimod sodium stock solution, 1.0 mL of
SPIRO-LAQ stock solution, 1.0 mL of 3-HLAQ stock solution and 0.1
mL of NEA standard stock solution.
[0817] Concentration of Laquinimod sodium is about 90 .mu.g/mL.
[0818] Concentration of NEA, SPIRO-LAQ and 3-HLAQ is about 1
.mu.g/mL (1%).
[0819] The Resolution Solution is used only for the determination
of the retention times of Laquinimod, NEA, 3-HLAQ and SPIRO-LAQ and
may be used for 38 days when stored at -20.degree. C. For this
purpose, the freshly prepared resolution solution should be divided
into aliquots and stored immediately at -20.degree. C. After
thawing, the solution aliquots should not be refrozen.
[0820] Mix vigorously resolution solution after thawing before
injection.
[0821] After thawing, the Resolution Solution may be used for 4
days when stored in refrigerator.
4. Sample Solution Preparation
[0822] Weigh accurately about 15 mg of Laquinimod sodium Cryst or
DS into a 50 mL volumetric flask. Dilute with Diluent B up to 2/3
of the volume, sonicate until the substance is completely dissolved
and dilute to volume with Diluent B--sample stock solution.
[0823] Dilute 3 mL of sample stock solution to 10 mL with Diluent B
(dilution factor 3.33).
[0824] Concentration of Laquinimod sodium is about 90 .mu.g/mL.
[0825] Note: Immediately after preparation put the sample solution
to refrigerator or at cooled to 5.degree. C. auto sampler. The
sample must be analyzed not later than 12 hours after
preparation.
5. Procedure
[0826] Inject resolution solution, Diluent B (two or three times),
Standard and Sample solutions according to relevant SOP.
6. System Suitability Test
6.1 Resolution Test
[0827] The typical retention time is 2.2.+-.0.5 minutes for
Laquinimod peak.
[0828] The typical retention time is 12.2.+-.1.5 minutes for NEA
peak.
[0829] The chromatographic mapping of the specified impurity peaks
should be in the following order: 3-HLAQ, NEA and SPIRO-LAQ.
[0830] Resolution of NLT 2 for both pairs of peaks should be
achieved.
6.2 System Precision Test
[0831] Evaluate standards in order to test system precision
according to the relevant SOP.
6.3 Blank
[0832] Inject Diluent B to detect system peaks.
7. Evaluation and Calculation
[0833] Quantitation limit for NEA and any other impurities is
0.06%
[0834] Detection limit for NEA and any other impurities is
0.02%.
[0835] Adjust integration parameters to reject peaks with the area
less than 15% of standard area.
[0836] Disregard all the impurity peaks eluting between 0 and the
RT of Laquinimod plus one minute.
[0837] Calculate the content of non-polar impurities following the
equation:
Area of imp in smp .times. Conc . of std .times. 100 .times. R R F
Area of std .times. Conc . of smp = % Impurity ##EQU00006##
[0838] Where RRF is the Relative Response Factor of NEA to
impurities as per the Table:
TABLE-US-00057 Impurity NEA 3-HLAQ SPIRO-LAQ Unknown impurities RRF
1.00 1.37 0.92 1.23
8. Report Mode
TABLE-US-00058 [0839] Result Report NEA, 3-HLAQ or SPIRO-
.gtoreq.0.06% The calculated result LAQ <0.06% <0.06%
<0.02% (or ND) <0.02% Unknown impurities (by .gtoreq.0.06%
The calculated result RRT to Laquinimod) <0.06% <0.06%
<0.02% (or ND) Not to be reported* Total .gtoreq.0.06% The sum
of calculated results <0.06% <0.06% <0.02, or ND <0.02%
*If no unknown impurities were detected, report: Any other
<0.02%.
[0840] FIG. 45 presents a resolution test chromatogram according to
the above method.
System Suitability Separation Results for FIG. 45.
TABLE-US-00059 [0841] Int Height USP Name Type RT (.mu.V) Area
Resolution 1 Lag bb 2.156 1098982 12220719 2 System BB 3.442 173
1293 5.2 peak 3 3-HLAQ BB 10.067 7917 119088 21.9 4 NEA BB 11.034
12544 165542 2.5 5 Spiro- BB 12.496 8797 146324 3.6 LAQ
[0842] FIG. 46 presents a diluent chromatogram in according to the
above method.
Peak Results for FIG. 46.
TABLE-US-00060 [0843] Name Int Type RT Height (.mu.V) Area % Area 1
System peak BB 3.423 313 2191 100.00
[0844] FIG. 47 presents a typical sample chromatogram according to
the above method.
Peak Results for FIG. 47.
TABLE-US-00061 [0845] Name Int Type RT Height (.mu.V) Area % Area 1
Laquinimod bb 2.145 96369 11564891 99.99 2 System peak BB 3.421 166
1216 0.01
Example 21
Method of Determining Amount of MCQ, MCQCA, 5-HLAQ, MCQME and MCQEE
in a Sample of Laquinimod Drug Product
[0846] Laquinimod capsules containing 0.6 mg Laquinimod were
analyzed by HPLC. The amount of MCQ, MCQCA, 5-HLAQ, MCQME and MCQEE
was determined using the following HPLC method.
1. HPLC Conditions
TABLE-US-00062 [0847] Column & Packing Inertsil ODS-3V, 5
.mu.m, 4.6 .times. 250 mm, GL Sciences Inc. Guard column Opti-Guard
C 18, 1 .times. 10 mm Mobile phase Acetonitrile: Buffer pH
7.0-20:80 (v/v). Mix and degas Buffer pH 7.0 Dissolve 7.7 g
Ammonium acetate in 2000 mL preparation water and adjust to pH 7.0
.+-. 0.05 with aqueous ammonia or glacial acetic acid. Filter
through a 0.45 .mu.m membrane filter. Flow rate 1.5 mL/min
Detection UV at 240 nm Injection volume 50 .mu.L Diluent A
Acetonitrile/Water-50:50 (v/v) Diluent B (and Mobile phase Blank)
Column temperature 40.degree. C. Autosampler 5.degree. C.
temperature Run time 40 minutes Run time for standards May be
shortened to 23 minutes Note: Flow rate, mobile phase composition
and column temperature may be varied in order to achieve the
required system suitability parameters.
[0848] All the preparations should be protected from light. For
liquid preparations use amber flasks with non transparent caps. Use
only glass Pasteur pipettes. Standard and sample solutions should
be kept refrigerated at 2-8.degree. C. after preparation.
2. Standard Solutions Preparation
2.1 Laquinimod Standard Stock Solution (Solution S)
[0849] Weigh accurately in duplicate about 15 mg of Laquinimod
sodium standard into a 50 mL volumetric flask. Dilute with diluent
A up to 2/3 of the volume, sonicate for 2 minutes in a cold
sonication bath and dilute to volume with diluent A.
[0850] Concentration of standard stock solution is about 300
.mu.g/mL Laquinimod sodium.
[0851] Standard stock solution may be used for one month when
stored in a refrigerator (2.degree. C.-8.degree. C.).
2.2 Laquinimod Standard Working Solution for Assay (Solution A)
[0852] Dilute 3 mL of the Standard Stock Solution to 10 mL with
diluent B (Dilution factor 3.33).
[0853] Concentration of Laquinimod sodium is about 90 .mu.g/mL.
Concentration expressed as Laquinimod (acid) is about 85
.mu.g/mL.
[0854] Standard working solution A may be used for 7 days when
stored in a refrigerator (2.degree. C.-8.degree. C.).
2.3 MCQCA Standard Stock Solution
[0855] Weigh accurately about 18 mg of MCQCA standard into a 100 mL
volumetric flask. Dilute to volume with acetonitrile, sonicate (in
a cold sonication bath) until the substance is completely
dissolved--stock MCQCA solution.
[0856] Concentration of MCQCA is about 180 .mu.g/mL.
[0857] MCQCA Stock standard solution should be freshly
prepared.
2.4 Standard Solution for Determination of Impurities (Solution
I)
[0858] Prepare a solution in diluent B, containing Laquinimod in a
concentration of 0.2% and MCQCA--in a concentration of 0.1%, with
respect to the working concentration of Laquinimod in Standard
solution A. As an example, apply the following procedure.
[0859] Transfer 4.0 mL of Laquinimod sodium standard solution for
assay (Solution A) and 1.0 mL of MCQCA stock standard solution to a
100 mL volumetric flask and dilute to volume with the diluent B
(Intermediate dilution).
[0860] Place 2.5 mL of this intermediate dilution into a 50 mL
volumetric flask and make up to volume with the diluent B.
[0861] Total dilution factor for Laquinimod standard is 1666.67,
for MCQCA 2000.
[0862] Concentration of Laquinimod sodium is about 0.18 .mu.g/mL
(0.2%).
[0863] Concentration of MCQCA is about 0.09 .mu.g/mL (0.1%, QL
level).
[0864] Standard solution I may be used for 24 hours when stored in
a refrigerator.
3. Resolution Solutions Preparation
3.1 Mixed Solution
[0865] Prepare solution containing the following potential
impurities standards (markers) using the Diluent A as a
solvent:
Mixed Solution:
TABLE-US-00063 [0866] Impurity Abbreviation Full name MCQ
5-Chloro-4-hydroxy-1-methylquinolin-2(1H)-one MCQCA
5-Chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-
3-carboxylic acid MCQME Methyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-
dihydroquinoline-3-carboxylate 5-HLAQ
N-Ethyl-4,5-dihydroxy-1-methyl-2-oxo-N-phenyl-1,2-
dihydroquinolin-3-carboxamide
[0867] The Mixed Solution may be prepared as follows:
[0868] Weigh about 3 mg of each impurity standard/marker into a 100
mL volumetric flask, dissolve (sonication is acceptable) and dilute
to volume with the Diluent A.
[0869] Concentration of each impurity in the Mixed Solutions is
about 30 .mu.g/mL.
[0870] Mixed Solution may be used for up to 4 months when stored
frozen at about -20.degree. C.
[0871] For this purpose, the freshly prepared Mixed Solution should
be divided into aliquots, immediately frozen and stored at
-20.degree. C. After thawing, the aliquots should be mixed well and
should not be refrozen.
3.2 Stock Solutions of Additional Impurities
[0872] Weigh about 3 mg of MCQEE (Ethyl
5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate)
into a 100 mL volumetric flask, dissolve (sonication is acceptable)
and dilute to volume with the Diluent A. This is MCQEE Stock
solution.
[0873] This Solution may be used for up to 4 months when stored
frozen at about -20.degree. C.
[0874] For this purpose, the freshly prepared MCQEE Stock Solution
should be divided into aliquots, immediately frozen and stored at
-20.degree. C. After thawing, the aliquots should be mixed well and
should not be refrozen.
[0875] Weigh about 3 mg of MEG-LAQ (Meglumine Adduct of Laquinimod)
into a 100 mL volumetric flask, dissolve (sonication is acceptable)
and dilute to volume with the Diluent A. This is MEG-LAQ Stock
solution.
[0876] This Solution may be used for one week when stored in a
refrigerator (2.degree. C.-8.degree. C.).
3.3 Resolution Solutions
[0877] Prepare two Resolution Solutions separately as follows,
using the Mixed solution.
3.3.1 Resolution Solution 1
[0878] Transfer 3 mL of Laquinimod standard stock solution
(Solution S), 0.3 mL of the Mixed solution and 0.3 mL of the MCQEE
Stock solution to a 10 mL volumetric flask and dilute to volume
with the Diluent B. This is Resolution Solution 1.
[0879] Concentration of Laquinimod sodium in it is about 90
.mu.g/mL. Concentration of each impurity--is about 0.9 g/mL (about
1% with respect to the working concentration of Laquinimod).
[0880] Resolution Solution 1 is used for resolution test (for
system suitability) and for determination of retention times
(RT)/relative retention times (RRT) of five known impurities: MCQ,
MCQCA, MCQME, MCQEE and 5-HLAQ.
[0881] Resolution Solution 1 may be used for 9 days if stored in a
refrigerator (2.degree. C.-8.degree. C.).
[0882] Note: Instead of Resolution Solution i, a Resolution
Solution prepared as per the Method of Example 19 may be used.
3.3.2 Resolution Solution 2
[0883] Transfer 3 mL of Laquinimod standard stock solution
(Solution S), 0.3 mL of the Mixed solution and 0.3 mL of the
MEG-LAQ Stock solution to a 10 mL volumetric flask and dilute to
volume with the Diluent B. This is Resolution Solution 2.
[0884] Concentration of Laquinimod sodium in it is about 90 g/mL.
Concentration of each impurity--is about 0.9 .mu.g/mL (about 1%
with respect to the working concentration of Laquinimod).
[0885] Resolution Solution 2 is used for determination of retention
time of MEG-LAQ.
[0886] Resolution Solution 2 may be used for one week if stored in
a refrigerator (2.degree. C.-8.degree. C.).
4. Sample Solution Preparation
[0887] Weigh accurately 20 capsules and completely empty their
contents into a mortar. Pay attention to complete emptying of
capsule contents into the mortar, using spatula when necessary.
Weigh the empty capsules. Calculate the average weight of the
capsule contents.
[0888] Mix and grind the capsule contents in a mortar and keep the
powder in a tightly closed container protected from light.
[0889] Weigh accurately, in duplicate, the amount of powder
corresponding to 7 capsules, into a 50 mL volumetric flask.
[0890] Add diluent B up to 2/3 of the volume, shake for 30 minutes
at 200 mot/min. Dilute to volume with diluent B. Mix well. Filter
before use through a 0.45 .mu.m GHP ACRODISC GF filter or
equivalent, discarding the first 0.5-1 mL.
[0891] Working concentration of Laquinimod (acid) is about 84
.mu.g/mL.
[0892] Note: Immediately after preparation place sample solutions
into a refrigerator or in a cooled to 5.degree. C. autosampler. The
sample solutions may be used for 24 hours when kept at the
temperature 2.degree. C.-8.degree. C.
5. Procedure
[0893] Inject the Resolution Solutions, Diluent B (Blank), Standard
Solutions for assay and IDD and Sample solutions, according to the
relevant SOP's.
[0894] Determine the retention time (RT) and the area of the
Laquinimod peak in the chromatograms of Sample and Standard
Solutions for identification and assay.
[0895] Determine the RT, the relate rive retention time (RRT) and
peak areas of all known impurities and any other impurities in the
chromatograms of Sample Solutions, for calculation of the content
of impurities/degradation products.
[0896] Ignore early eluting peaks of excipients and system peaks
(See chromatogram for determination of impurities/degradation
products). For example, use integration inhibition between 0 and
RRT 0.15 (about 2.5 minutes).
[0897] Adjust integration parameters to reject peaks with area less
than 10% of the average response of Laquinimod peak in the
injections of Standard Solution I (for determination of
impurities/degradation products).
[0898] Disregard peak of MEG-LAQ in sample injections (identified
following Resolution Solution 2). The content of MEG-LAQ will be
tested by a different method.
6. System Suitability Test
6.1 Resolution Test
[0899] Typical retention time of Laquinimod peak is 15.5.+-.2.0
minutes.
[0900] Tailing factor (USP) for Laquinimod peak should be not more
than 2.0.
[0901] Resolution factor for all the pairs of peaks should be NLT
2.
[0902] RRT of the peaks of known impurities/degradation products
should be as follows:
TABLE-US-00064 Impurity RRT MCQME 0.33-0.38 MCQ 0.49-0.58 MCQEE
0.56-0.65 MCQCA 0.71-0.85 5-HLAQ 1.2-1.4* *Retention time of 5-HLAQ
should not be more than 23 minutes.
[0903] MEG-LAQ peak is substantially broadened in comparison with
neighboring peaks. Retention time of MEG-LAQ is variable, being
very sensitive to slightest changes in chromatographic conditions
(pH, % Acetonitrile, Temperature, etc.) and therefore should be
defined using its peak in the chromatogram of the Resolution
Solution 2. Typically, its RRT is about 0.66.
6.2 System Precision Test
[0904] Evaluate Laquinimod standards for assay and IDD in order to
test the system precision according to the relevant SOP's.
[0905] MCQCA in Solution I is used to test the sensitivity of the
system. RSD of the area of six injections of Std 1 as well as the
difference between Std 1 and Std 2 should be NMT 20%.
6.3 Blank
[0906] Inject diluent B to detect system peaks.
7. Identification by Retention Time (RT)
[0907] The RT of the main peak obtained in the sample chromatogram
should correspond to that obtained for the Laquinimod peak in the
injection of Standard Solution.
8. Calculation and Report
8.1 Assay Calculation
[0908] % Assay ( to Label Claim ) = Area Smp .times. Conc Std
.times. 0.94 .times. V Smp .times. AvgWt CapsContent Area Std
.times. W Smp .times. Label Claim .times. 100 ##EQU00007##
[0909] Where: 0.94: Conversion factor of Laquinimod sodium salt to
Laquinimod (acid).
8.2 Calculation and Evaluation of Impurities/Degradation
Products
8.2.1 Calculation of Relative Retention Time (RRT)
[0910] R R T Impurity = R T Impurity R T Laquinimod
##EQU00008##
8.2.2 Calculation of Content of Impurities/Degradation Products
[0911] % Impurity = Area impurity .times. Conc Std .times. V Smp
.times. AvgWT CapsContent .times. 0.94 .times. R R F Area Std
.times. W Smp .times. Label Claim .times. 100 ##EQU00009##
[0912] Area.sub.Impurity: Area of an impurity/degradation product
(known or unknown) peak in Sample Solution.
[0913] Area.sub.std: Area of Laquinimod peak in chromatogram of
Standard Solution I.
[0914] 0.94: Conversion factor of Laquinimod sodium salt to
Laquinimod (acid).
[0915] RRF: Relative response factors of impurities/degradation
products calculated as the following ratio: slope of Laquinimod
regression line/slope of impurity regression line.
[0916] The values for relative response factors with respect to
Laquinimod are:
TABLE-US-00065 Impurity MCQME MCQ MCQEE MCQCA 5-HLAQ RRF 0.74 0.65
0.85 0.62 1.0
[0917] RRF for unknown impurities/degradation products is taken as
1.0.
8.2.3 Evaluation and Report of Impurities/Degradation Products
[0918] Quantitation level (QL) of MCQME, MCQ, MCQEE, 5-HLAQ and
unknown impurities is 0.05%. Detection level (DL) of MCQME, MCQ,
MCQEE, 5-HLAQ and unknown impurities is 0.02%.
[0919] QL of MCQCA is 0.1%. Detection level DL of MCQCA is
0.03%.
[0920] Correlate all the peaks in sample chromatogram with those in
the system suitability chromatogram, within .+-.5% of the actual
corresponding retention times.
[0921] Report as follows:
TABLE-US-00066 Result Report Specified impurities: 5-HLAQ
.gtoreq.0.05% The calculated result <0.05% <0.05% <0.02%
(or ND) <0.02% MCQ and MCQCA sum .gtoreq.0.1% The calculated
result <0.1% <0.1% <0.03% (or ND) <0.03% Other
impurities: MCQME, MCQEE .gtoreq.0.05% The calculated result
<0.05% <0.05% <0.02% (or ND) <0.02% Unknown peaks (by
RRT to .gtoreq.0.05% The calculated result Laquinimod) <0.05%
<0.05% <0.02% (or ND) Not to be reported* Total .gtoreq.0.05%
The sum of calculated <0.05% (or<0.02 or results ND)
<0.05% *If no impurities were detected, report: any other
<0.02%.
[0922] FIG. 48 presents a resolution test (resolution solution 1)
chromatogram according to the above method.
System Suitability Separation Results for FIG. 48.
TABLE-US-00067 [0923] RT USP USP Int Name RT Ratio Area Resolution
Tailing Type 1 MCQME 5.460 0.36 35801 1.07 BB 2 MCQ 8.260 0.54
60897 8.2 1.16 BV 3 MCQEE 9.249 0.60 24029 2.6 1.05 VB 4 MCQCA
12.031 0.78 18609 5.7 1.34 BB 5 Laq 15.332 12332469 5.2 1.84 BB 6
5-HLAQ 20.451 1.33 89463 7.3 1.04 BB
[0924] FIG. 49 presents a resolution test (resolution solution 2)
chromatogram according to the above method.
System Suitability Separation Results for FIG. 49.
TABLE-US-00068 [0925] RT USP USP Int Name RT Ratio Area Resolution
Tailing Type 1 MCQME 5.457 0.36 151440 1.06 BB 2 MCQ 8.260 0.54
16608 8.1 1.17 BV 3 MCQEE 10.081 0.66 86529 1.7 VB 4 MCQCA 11.856
0.77 183439 1.5 1.9 BB 5 Lag 15.330 12314140 5.8 1.82 BB 6 5-HLAQ
20.446 1.33 120400 7.2 1.04 BB
[0926] FIG. 50 presents a diluent blank chromatogram according to
the above method.
[0927] FIG. 51 presents a sample chromatogram for assay according
to the above method.
[0928] FIG. 52 presents a sample chromatogram for determination of
impurities/degradation products according to the above method.
Example 22
Method of Determining Amount of NEA, 3-HLAQ and SPIRO-LAQ in a
Sample of Laquinimod Drug Product
[0929] Laquinimod capsules containing 0.6 mg Laquinimod were
analyzed by HPLC. The amount of NEA, 3-HLAQ And SPIRO-LAQ was
determined using the following HPLC method.
1. Abbreviations
TABLE-US-00069 [0930] Abbreviation Full name NEA N-Ethylaniline
3-HLAQ 5-chloro-N-ethyl-3-hydroxy-1-methyl-2,4-dioxo-N-phenyl-
1,2,3,4-tetrahydroquinoline-3-carboxamide SPIRO-LAQ
1H,3H-spiro[5-chloro-1-methylquinoline-2,4-dione-3,3'-[1]-
ethylindolin-[2]-one]
2. HPLC Conditions
TABLE-US-00070 [0931] Column & Packing Inertsil ODS-3V, 5
.mu.m, 4.6 .times. 250 mm, GL Sciences Guard column Opti-Guard C
18, 1 .times. 10 mm UV detection 240 nm Mobile phase: 50% Solution
- 50% Solution B (v/v) Solution A: Acetonitrile:Buffer pH 7.0 -
20:80 (v/v) Solution B: Acetonitrile:Buffer pH 7.0 - 65:35 (v/v).
Mix well by magnetic stirring for about 0.5 hour. Buffer Solution
Dissolve 7.7 g Ammonium acetate in 2000 mL pH 7.0 water and adjust
to pH 7.0 .+-. 0.05 with aqueous ammonia or glacial acetic acid.
Filter through a 0.45 .mu.m membrane filter. Flow rate 1.5 mL/min
Injection volume 20 .mu.L Diluent A Methanol/Acetonitrile - 50:50
(v/v) Blank (Final diluent) Methanol/Acetonitrile - 10:90 (v/v)
Column temperature 40.degree. C. Autosampler 5.degree. C.
temperature Run time 20 minutes Needle and seal wash
Acetonitrile/water - 50:50 (v/v) Note: Flow rate, mobile phase
composition and column temperature may be varied to achieve the
required system suitability parameters.
[0932] All the preparations should be protected from light. For
liquid preparations use amber flasks with non transparent caps.
[0933] Use only glass Pasteur pipettes.
[0934] Standard and sample solutions should be kept refrigerated at
2-8.degree. C. after preparation.
3. Standard Solution Preparation
3.1 NEA Standard Stock Solution
[0935] Weigh accurately in duplicate about 18 .mu.L (about 18 mg)
of N-Ethylaniline (NEA) standard into a 50 mL volumetric flask.
Dilute to volume with acetonitrile.
[0936] Concentration of standard stock solution is about 360
.mu.g/mL.
[0937] NEA standard stock solution may be used for three weeks when
stored in a refrigerator (2.degree. C.-8.degree. C.).
3.2 NEA Standard Primary Dilution Solution
[0938] Dilute 0.5 mL NEA standard stock solution to 50 mL with
acetonitrile.
[0939] Concentration of standard primary dilution solution is about
3.6 .mu.g/mL.
3.3 NEA Standard Working Solution
[0940] Transfer 0.5 mL NEA standard primary dilution solution to a
20 mL volumetric flask. Add 4 mL diluent A and dilute to volume
with acetonitrile.
[0941] Concentration of NEA is about 0.09 .mu.g/mL (about 0.1% of
the Laquinimod working concentration).
[0942] NEA working standard solution may be used for one week when
stored in a refrigerator (2.degree. C.-8.degree. C.).
4. Resolution Solution Preparation
[0943] Resolution solution from Example 20 may be used.
4.1 Laquinimod Stock Solution for Resolution Test
[0944] Weigh accurately about 15 mg of Laquinimod sodium standard
into a 50 mL volumetric flask. Dilute with acetonitrile:water
[50:50 (v/v)] up to 2/3 of the volume, sonicate for 2 minutes in a
cold sonication bath and dilute to volume with acetonitrile:water
[50:50 (v/v)].
[0945] Concentration of Laquinimod stock solution is about 300
.mu.g/mL.
[0946] Laquinimod stock solution may be used for one month when
stored in a refrigerator (2.degree. C.-8.degree. C.).
4.2 NEA Stock Solution for Resolution Test
[0947] Weigh accurately about 25 .mu.L (about 25 mg) of
N-Ethylaniline (NEA) standard into a 50 mL volumetric flask. Dilute
to volume with acetonitrile:water [50:50 (v/v)].
[0948] Concentration of NEA stock solution is about 500
.mu.g/mL.
[0949] NEA stock solution may be used for 3 weeks if stored in a
refrigerator (2.degree. C.-8.degree. C.).
4.3 SPIRO-LAQ Stock Solution for Resolution Test
[0950] Weigh about 2.5 mg of SPIRO-LAQ into a 50 mL volumetric
flask. Dissolve and dilute to volume with acetonitrile:water [95:5
(v/v)].
[0951] Concentration of SPIRO-LAQ stock solution is about 50
g/mL.
4.4 3-HLAQ Stock Solution for Resolution Test
[0952] Weigh about 2.5 mg of 3-HLAQ into a 50 mL volumetric flask.
Dissolve and dilute to volume with acetonitrile:water [95:5
(v/v)].
[0953] Concentration of 3-HLAQ stock solution is about 50
.mu.g/mL
4.5 Resolution Solution
[0954] Transfer the following aliquots to a 50 mL volumetric flask
and dilute to volume with mobile phase solution B
[acetonitrile:buffer pH 7.0 (65:35 v/v)]:
[0955] 15 mL of Laquinimod stock solution for resolution test,
[0956] 1.0 mL of SPIRO-LAQ stock solution for resolution test,
[0957] 1.0 mL of 3-HLAQ stock solution for resolution test,
[0958] 0.1 mL of NEA stock solution for resolution test.
[0959] Concentration of Laquinimod sodium is about 90 .mu.g/mL.
[0960] Concentration of NEA, SPIRO-LAQ and 3-HLAQ is about 1
.mu.g/mL (about 1% with respect to Laquinimod working
concentration).
[0961] The resolution solution is used only for the determination
of the retention times of the laquinimod and known potential
impurities/degradation products and may be used for 38 days when
stored at -20.degree. C. For this purpose, the freshly prepared
resolution solution should be divided into aliquots and stored
immediately at -20.degree. C. After thawing, the solution aliquots
should be mixed vigorously and should not be refrozen.
[0962] The resolution solution may be used for 4 days when stored
in a refrigerator (2.degree. C.-8.degree. C.).
5. Sample Solution Preparation
[0963] Weigh accurately at least 10 capsules and empty the contents
in a mortar. Weigh the empty capsules. Calculate the average weight
of the capsule contents.
[0964] Grind the sample in a mortar and keep the powder in a
tightly closed protected from light container.
[0965] Weigh accurately, in duplicate, the amount of powder
corresponding to 3 Laquinimod capsules into a 20 mL volumetric
flask.
[0966] Add 4 to 10 mL of Diluent A and shake for 15 minutes at 200
mot/min to dissolve. Dilute to volume with acetonitrile. Mix well.
Filter before use through a 0.45 .mu.m GHP Acrodisc GF filter or
equivalent, discarding the first 2 mL.
[0967] Concentration of Laquinimod is about 90 .mu.g/mL.
[0968] Note: Immediately after preparation place the sample
solution into a refrigerator or in a cooled to 5.degree. C.
autosampler. The sample should be analyzed not later than within 24
hours after preparation.
6. Procedure
[0969] Inject resolution solution, blank, standard and sample
solutions according to the relevant SOP's for
impurities/degradation products determination.
[0970] Determine the peak area of NEA in standard working
solutions.
[0971] Determine the retention time (RT) of the Laquinimod peak in
the chromatograms from resolution and sample solutions
[0972] Determine the RT, the relative retention time (RRT) to
Laquinimod peak and the peak area of all known impurities and
unknown impurities in the chromatograms from Sample solutions.
[0973] Apply integration inhibition between 0 and the 1.6.times.RT
of Laquinimod peak (RT of system peak).
[0974] Adjust integration parameters to reject peaks with the area
less than 20% of NEA standard area.
7. System Suitability Test
7.1 Resolution Test
[0975] The typical retention time for Laquinimod is 2.2.+-.0.5
minutes.
[0976] The typical retention time for NEA is 12.2.+-.1.5
minutes.
[0977] The RRT of the specified impurity/degradation product peaks
relative to NEA should be as follows:
TABLE-US-00071 Impurity/Degradation product RRT to NEA 3-HLAQ
0.85-0.95 SPIRO-LAQ 1.05-1.20
[0978] Resolution factor of NLT 2 for both known
impurities/degradation products peaks from NEA peak should be
achieved.
7.2 System Precision Test
[0979] Evaluate standards in order to test the system precision
according to the relevant SOP for impurities/degradation products
determination.
7.3 Blank
[0980] Inject blank to detect system peaks.
8 Evaluation, Calculation and Report
8.1 RRT of Impurities/Degradation Products
[0981] R R T Imp = R T Imp R T Laq ##EQU00010##
8.2 Calculation of Content of Impurities/Degradation Products
[0982] Calculate % IDD in Laquinimod capsules as follows:
% Impurity = Area impurity .times. Conc Std NEA .times. V Smp
.times. AvgWT CapsContent .times. R R F Area Std NEA .times. W Smp
.times. Label Claim .times. 100 ##EQU00011##
Area.sub.Imp: Area of the NEA, 3-HLAQ or unknown
impurity/degradation product peak in the sample solution.
[0983] See note in 8.3.
[0984] Area.sub.Std NEA: Area of the NEA in standard working
solution.
[0985] RRF: Relative response factors of impurities calculated as
the following ratio:
[0986] Slope of NEA regression line/slope of impurity regression
line.
[0987] Note: For Unknown impurities/degradation product, the slope
of Laquinimod regression line was used.
[0988] The values for relative response factors with respect to NEA
are:
TABLE-US-00072 Impurity NEA 3-HLAQ Unknown RRF 1.0 1.39 1.15
8.4 Evaluation and Report
[0989] Quantitation limit (QL) for NEA, 3-HLAQ and any other
unknown impurity/degradation product is 0.1% with respect to the
Laquinimod working concentration.
[0990] Detection limit (DL) for NEA, 3-HLAQ and any other unknown
impurity/degradation product is 0.03% with respect to the
Laquinimod working concentration.
[0991] Report content of Impurities/Degradation products as
follows:
TABLE-US-00073 Result Report NEA or 3-HLAQ .gtoreq.0.1% The
calculated result <0.1% <0.1% <0.03% or ND <0.03% Any
other impurity peak .gtoreq.0.1% The calculated result (by RRT to
Laquinimod) <0.1% <0.1% <0.03% or ND Not to be reported**
Total If peak/s .gtoreq.0.1% The sum of calculated If all peaks
<0.1% or results <0.03% <0.1% * Correlate the NEA and
3-HLAQ peaks in the in the sample chromatogram with those in the
system suitability chromatogram within .+-.5% of the actual
retention times. **If no unknown impurities are present, report:
"Any other <0.03%"
[0992] Note: Peaks eluting at RT of the SPIRO-LAQ (.+-.5% of the
actual retention time in system suitability test) report as "any
other impurity" peak.
[0993] FIG. 53 presents a resolution test chromatogram according to
the above method.
System Suitability Separation Results for FIG. 53.
TABLE-US-00074 [0994] Height USP USP Name RT RRT Area (.mu.V)
Tailing Resolution 1 LAQ 2.166 5587169 964422 1.22 2 3-HLAQ 11.228
0.92 81831 4436 1.35 28.23 3 NEA 12.226 63991 4744 1.06 2.34 4
Spiro- 1.14 78255 78255 4725 1.02 4.26 LAQ
[0995] FIG. 54 presents a blank (diluents) chromatogram according
to the above method.
Peak Results for FIG. 54.
TABLE-US-00075 [0996] Name Int Type RT Height (.mu.V) Area % Area 1
System peak BB 3.508 116 858 52.53 2 Sys. peak BB 5.740 41 775
47.47
[0997] FIG. 55 presents a sample chromatogram according to the
above method.
Peak Results for FIG. 55.
TABLE-US-00076 [0998] Name Int Type RT Height (.mu.V) Area % Area 1
Laquinimod BB 2.1459 923247 4717424 99.97 2 System peak BB 3.499 71
666 0.01 3 Sys. peak BB 5.736 38 642 0.01
Example 23
Method of Determining Amount of MEG-LAQ and BH-3-HLAQ in a Sample
of Laquinimod Drug Product
TABLE-US-00077 [0999] Example Method Quantitation Detection
Parameter Number Type Limit Limit MEG-LAQ Example 23 RP-HPLC 0.2%
0.1% BH-3-HLAQ Example 23 RP-HPLC 0.05% 0.02%
[1000] Laquinimod capsules containing 0.6 mg Laquinimod were
analyzed by HPLC. The amount of MEG-LAQ and BH-3-HLAQ was
determined using the following HPLC method.
1. Chromatographic System
TABLE-US-00078 [1001] Column & Packing Inertsil ODS-3 V, 5
.mu.m, 3.0 .times. 150 mm, GL Sciences Inc. Guard column Opti-Guard
C 18, 1 mm, Optimize Technologies (recommended) Column Temperature
43.degree. C. .+-. 4.degree. C. Buffer Solution Dissolve 5.75 g of
Ammonium Dihydrogen Preparation Phosphate in 300 mL water. Filter
through a 0.45 .mu.m membrane filter. Dilute to 2000 mL with water
and adjust to pH 7.0 .+-. 0.10 with aqueous Ammonia or Phosphoric
acid. Apply proportional adjustments for preparation of a different
volume. Solvent A Mixture of 80% (v/v) Buffer Solution and 20%
(v/v) Acetonitrile. Degas Solvent B Mixture of 70% (v/v) Buffer
Solution and 30% (v/v) Methanol. Degas. Mobile Phase Option 1: Work
from 2 reservoirs: 60% (v/v) Solvent A and 40% (v/v) Solvent B;
Adjust to achieve system suitability requirements. Option 2: Work
from 1 reservoir: Mix Solvent A and Solvent B in the ratio 60:40
(v/v); degas after mixing. Flow Rate: 1.0 mL/min Detector UV at 212
nm, 10 mm flow cell path length Autosampler 5.degree. C.
Temperature Injection Volume 35 .mu.L Diluent 1 Acetonitrile/Water
- 50:50 (v/v) Diluent 2/Blank Solvent A Solution Injector Wash
Acetonitrile/Water - 50:50 (v/v) Solution Run Time 30 minutes
[1002] Mobile phase parameters, flow rate and column temperature
may be altered in order to achieve the system suitability
requirements. The Mobile phase parameters may be altered in the
range: 55%-62% (v/v) Solvent A and 45%-38% (v/v) Solvent B. All the
solvents must be of HPLC grade or equivalent. All the preparations
should be kept refrigerated at 2-8.degree. C. and be protected from
light. Use amber flasks (low actinic glassware) with non
transparent caps. Avoid heating of sonication bath.
2. List of Abbreviations
TABLE-US-00079 [1003] Abbeviation Full chemical name MEG-LAQ
N-ethyl-4-hydroxy-1-methyl-5-(methyl(2,3,4,5,6-
pentahydroxyhexyl)amino)-2-oxo-N-phenyl-1,2-
dihydroquinoline-3-carboxamide BH-3-HLAQ
2-chloro-6-(3-(ethyl(phenyl)amino)-2-hydroxy-N-methyl
3-oxopropanamido)benzoic acid
3. Preparation of Standard Solutions
3.1 Stock Solution of BH-3-HLAQ
[1004] Prepare in duplicate. Accurately weigh about 11.5 mg of
BH-3-HLAQ into a 50 mL volumetric flask, sonicate to dissolve and
dilute to volume with the Diluent 1.
[1005] Concentration of BH-3-HLAQ stock solution is about 230
.mu.g/mL.
[1006] BH-3-HLAQ stock solution may be used for 1 week if stored
refrigerated (2.degree. C.-8.degree. C.).
3.2 Stock Solution of MEG-LAQ
[1007] Prepare in duplicate. Accurately weigh about 11.5 mg of
MEG-LAQ into a 50 mL volumetric flask, sonicate to dissolve and
dilute to volume with the Diluent 1. Concentration of MEG-LAQ stock
solution is about 230 .mu.g/mL.
[1008] MEG-LAQ stock solution may be used for 1 week if stored
refrigerated (2.degree. C.-8.degree. C.).
[1009] Note: Due to high hygroscopicity of MEG-LAQ, its exposure to
air should be minimal: close the vial with standard immediately
after weighing; it is recommended to weigh MEG-LAQ under controlled
low humidity conditions.
3.3 Primary Diluted Solution of BH-3-HLAQ
[1010] Transfer 2.0 mL of BH-3-HLAQ Stock solution to a 50 mL
volumetric flask and dilute to volume with Diluent 2. Concentration
of BH-3-HLAQ is about 9.2 .mu.g/mL.
3.4 Primary Diluted Solution of MEG-LAQ
[1011] Transfer 2.0 mL of MEG-LAQ Stock solution to a 50 mL
volumetric flask and dilute to volume with Diluent 2. Concentration
of MEG-LAQ is about 9.2 .mu.g/mL.
3.5 Standard Solution
[1012] Transfer 1.0 mL of the Primary Diluted Solution of BH-3-HLAQ
and 2.5 mL of the Primary Diluted Solution of MEG LAQ to a 50 mL
volumetric flask and dilute to volume with Diluent 2.
[1013] Total dilution factor in this standard solution is: 1250 for
BH-3-HLAQ and 500 for MEG-LAQ (from the stock solution).
[1014] Concentration of BH-3-HLAQ is about 0.18 .mu.g/mL (0.2% of
Laquinimod working concentration), and of MEG-LAQ is about 0.45
.mu.g/mL (0.5% of Laquinimod working concentration).
[1015] Standard solution may be used for 1 week if stored
refrigerated (2.degree. C.-8.degree. C.).
3.6 QL Solution
[1016] Prepare once. Transfer 0.5 mL of Primary Diluted Solution of
BH-3-HLAQ and 2.0 mL of the Primary Diluted Solution of MEG LAQ
(one of the duplicates each) to a 100 mL volumetric flask and
dilute to volume with Diluent 2.
[1017] Concentration of BH-3-HLAQ is about 0.045 .mu.g/mL (0.05% of
Laquinimod working concentration), and of MEG-LAQ is about 0.18
g/mL (0.2% of Laquinimod working concentration).
4. Preparation of Resolution Solution
4.1 Laquinimod Stock Solution
[1018] Prepare once. Weigh about 15 mg of Laquinimod sodium
standard into a 50 mL volumetric flask. Dilute with the Diluent 1
up to 2/3 of the volume, sonicate for 2 minutes in a sonication
bath and dilute to volume with the Diluent 1. Concentration of
Laquinimod stock solution is about 300 .mu.g/mL Laquinimod
sodium.
[1019] Laquinimod Stock solution may be used for 1 month if stored
refrigerated (2.degree. C.-8.degree. C.).
4.2 Resolution Solution
[1020] Transfer 3 mL of Laquinimod stock solution and 1 mL of each
of the Primary Diluted solutions (one of the duplicates of each) to
a 10 mL volumetric flask. Dilute to volume with Diluent 2. Mix
well.
[1021] Concentration of Laquinimod Sodium in the Resolution
Solution is about 90 .mu.g/mL (about 85 g/mL if expressed as
Laquinimod--free acid). Concentration of each impurity is about 0.9
.mu.g/mL (about 1% with respect to the working concentration of
Laquinimod).
[1022] Resolution Solution may be used for one week if stored
refrigerated (2.degree. C. 8.degree. C.).
5. Preparation of Sample Solutions
[1023] Weigh accurately 20 capsules and completely empty their
contents into a mortar. Pay attention to complete emptying of
capsule contents into the mortar, using spatula when necessary.
Weigh the empty capsules. Calculate the average weight of the
capsule contents.
[1024] Mix and grind the capsule contents in a mortar and keep the
powder in a tightly closed container protected from light.
[1025] Weigh accurately, in duplicate, the amount of powder
corresponding to 7 capsules, each into a separate 50 mL volumetric
flask.
[1026] Add diluent 2 up to 2/3 of the volume, shake for 30 minutes
at 200 mot/min. Dilute to volume with diluent 2. Mix well. Filter
before use through a 0.45 .mu.m ACRODISC GHP GF filter or
equivalent, discarding the first 0.5-1 mL.
[1027] Immediately after preparation place sample solutions into a
refrigerator or in a cooled to 5.degree. C. autosampler.
[1028] Working concentration of Laquinimod (acid) is about 84
.mu.g/mL.
[1029] Sample solutions may be used for 24 hours when kept at the
temperature 2.degree. C.-8.degree. C.
6. System Suitability Test
6.1 Resolution Test
[1030] Inject the Resolution solution once.
[1031] Typical retention time of Laquinimod peak* is within the
range 12-18 minutes.
[1032] Order of elution of peaks in the injection of Resolution
Solution and typical RRT of the impurities peaks, with respect to
Laquinimod peak, are as follows:
TABLE-US-00080 Peak MEG-LAQ BH-3-HLAQ Laquinimod Typical RRT*
0.55-0.75 0.80-0.95 1.0 *For information only, may differ for
various lots of columns or as a result of variations in actual
mobile phase composition (Solvent A/Solvent B ratio) and pH.
Acceptance Criteria:
[1033] Tailing factor (USP) for MEG-LAQ and BH-3-HLAQ peaks should
be within 0.8-1.5. [1034] Resolution factor between all the three
peaks (MEG-LAQ, BH-3-HLAQ and Laquinimod) should be not less than
1.5.
[1035] Flow rate, temperature and mobile phase composition (ratio
of Solvent A and Solvent B) may be adjusted to achieve the required
system suitability parameters.
6.2 Blank Injections
[1036] Inject Blank (Diluent 2) twice to detect system peaks.
Disregard system peaks.
6.3 Sensitivity Test
[1037] From 6 replicate injections of the QL Solution, evaluate
areas of BH-3-HLAQ and MEG-LAQ peaks.
[1038] RSD for each of the peaks, in 6 replicates, should be not
more than 20%.
6.4 System Precision Test
[1039] Evaluate areas of BH-3-HLAQ and MEG-LAQ peaks in the
chromatograms of the standard injections in order to test the
system precision according to relevant SOP's.
7. Procedure
7.1 Injections Sequence
[1040] Inject Resolution Solution, Blank--twice, QL solution (6
replicates), Standard solutions and Sample solutions, according to
relevant SOP's.
7.2 Processing of Chromatograms
[1041] Disregard all the peaks other than MEG-LAQ and BH-3-HLAQ in
the injections of standard and sample solutions.
[1042] Determine the retention time (RT) and the areas of the peaks
of MEG-LAQ and BH-3-HLAQ in the chromatograms of Sample solutions,
for identification and quantitation.
[1043] Adjust integration parameters to reject peaks with area less
than 10% of the average response of BH-3-HLAQ peak in the
chromatograms of six successive injections of the first preparation
of Standard Solution.
[1044] This corresponds to 0.02% (DL of BH-3-HLAQ) with respect to
working concentration of Laquinimod.
8. Calculation and Reporting
8.1 Calculation
[1045] Quantitation of each of the impurities (MEG-LAQ and
BH-3-HLAQ) is performed with respect to the corresponding peaks in
the chromatograms of the Standard solutions.
[1046] Use the following equation to calculate the content of
MEG-LAQ and BH-3-HLAQ:
% BH - 3 - HLAQ / MEG - LAQ = Area Smp .times. Conc Std .times. V
Smp .times. AvgWt CapsContent Area Std .times. W Smp .times. Label
Claim .times. 100 ##EQU00012##
[1047] AreaSmp: Area of MEG-LAQ or BH-3-HLAQ peak in the
chromatograms of the Sample solutions
[1048] AreaStd: Area of peak MEG-LAQ (average) or BH-3-HLAQ peak
(average), respectively, in the chromatograms of Standard
solutions
[1049] Label Claim: 0.6 mg
8.2 Evaluation and Reporting
[1050] QL (Quantitation limit) of BH-3-HLAQ is 0.05%; QL of MEG-LAQ
is 0.2%
[1051] DL (Detection limit) of BH-3-HLAQ is 0.02%; DL of MEG-LAQ is
0.1%
[1052] For the values equal or above QL, report the calculated
result.
[1053] For the values below QL but above DL, report: "Less than
0.05% (QL)" for BH-3-HLAQ and "Less than 0.2% (QL)" for
MEG-LAQ.
[1054] For the values below DL, report: "Less than 0.02% (DL)" for
BH-3-HLAQ and "Less than 0.1% (DL)" for MEG-LAQ.
[1055] FIG. 56 presents a typical chromatogram of resolution
solution according to the above method.
Peak Results for FIG. 56.
TABLE-US-00081 [1056] % Int. USP Width USP Height Name RT RTT Area
Area Type Resolution (50%) Tailing (.mu.V) 1 MEG- 9.650 0.66 63177
0.66 BB 1.01 0.56 1322 LAQ 2 BH- 13.203 0.88 145888 1.16 BB 2.61
0.49 0.98 4661 3HLAQ 3 Laq 14962 12369065 86.18 BB 1.76 0.64 2.44
306644
[1057] FIG. 57 presents a bank (diluents 2) chromatogram according
to the above method.
Peak Results for FIG. 57.
TABLE-US-00082 [1058] Name RT Area % Area Int. Type 1 System peak
2.944 12.128 100.00 BB
[1059] FIG. 58 presents a chromatogram of QL solution according to
the above method.
Peak Results for FIG. 58.
TABLE-US-00083 [1060] Name RT Int. Type Height (.mu.V) Area % Area
1 Meg-LAQ 10.080 BB 303 15402 75.11 2 BH-3HLAQ 15.313 BB 201 5777
23.89
[1061] FIG. 59 presents a chromatogram of standard solution
according to the above method.
Peak Results for FIG. 59.
TABLE-US-00084 [1062] Name RT Int. Type Height (.mu.V) Area % Area
1 Meg-LAQ 10.177 BB 760 45319 54.89 2 BH-3HLAQ 15.293 BB 814 25059
35.11
[1063] FIG. 60 presents a typical sample chromatogram according to
the above method.
Peak Results for FIG. 60.
TABLE-US-00085 [1064] Name RT Int. Type Area % Area 1 Meg-LAQ
10.503 BB 20741 68.17 2 BH-3HLAQ 13.952 BB 9683 31.83
Example 24
Computational Toxicology Evaluation of Potentially Genotoxic
Impurities in Laquinimod Sodium Drug Substance and
Intermediates
[1065] DEREK prediction software (DEREK Nexus, version 2.0.0,
2011i, LHASA Ltd, Leeds, UK) was used to evaluate the genotoxic
potential of MCQ and MCQME. Computational toxicology evaluation is
summarized in Table 47.
TABLE-US-00086 TABLE 47 Computational Toxicology Evaluation of MCQ
and MCQME Potential Impurity Origin Toxicophore Endpoint
Probability.sup.1 MCQ By-product Quinoline Mutagenicity Plausible
and potential AMES data negative in the pre-incubation degradation
method (CRL study No. 962723) product MCQME Intermediate in No
structural the synthesis alert and process Weak genotoxic activity
in the in vitro related impurity AMES test (CRL study No. 963892,
but in the DS negative in the in vivo MN (Covance study No.
8246631)
[1066] The DEREK definition of the likelihood level of a structure
being toxic is as follows:
[1067] Plausible: The weight of evidence supports the
proposition.
[1068] Equivocal: There is an equal weight of evidence for and
against the proposition.
Example 25
Batch Analysis Of Laquinimod Sodium Drug Substance
[1069] A number of batches of laquinimod sodium drug substance were
manufactured at various manufacturing facilities and subsequently
analyzed. During manufacture and analysis, sufficient light
protection of sample solution was provided.
[1070] All batches of drug substance were within specification as
provided in Example 17.
[1071] Table 48 and Table 49 shows batch analyses of laquinimod
sodium drug substance batches Table 50 shows batch analyses of
laquinimod acid.
TABLE-US-00087 TABLE 48 Analysis of Laquinimod Sodium Drug
Substance Batches 1-12 Polar Impurities NEA and Non-Polar Imp. Sum
of Any Any MCQ & Other Other MCQ MCQCA MCQCA MCQME MCQEE
Impurity Total NEA Impurity Total Analytical Method Example 19
Example 20 Specifications NMT NMT NMT NMT NMT NMT NMT NMT NMT Batch
No. 0.15 0.15 NA 0.12 0.10 0.10 1.00 0.10 0.10 0.50 Batch 1 LT 0.05
LT 0.05 LT 0.05 LT 0.05 LT 0.05 LT 0.05 LT 0.05 LT 0.05 LT 0.05 LT
0.05 Batch 2 LT 0.05 LT 0.05 LT 0.05 LT 0.05 LT 0.05 LT 0.05 LT
0.05 LT 0.05 LT 0.05 LT 0.05 Batch 3 LT 0.02 LT 0.03 NA LT 0.02 LT
0.02 LT 0.02 LT 0.05 LT 0.02 LT 0.02 LT 0.05 Batch 4 LT 0.02 LT
0.03 NA LT 0.02 LT 0.02 LT 0.02 LT 0.05 LT 0.02 LT 0.02 LT 0.05
Batch 5 LT 0.02 LT 0.03 NA LT 0.02 LT 0.02 LT 0.02 LT 0.05 LT 0.02
LT 0.02 LT 0.06 Batch 6 LT 0.02 LT 0.03 NA LT 0.02 LT 0.02 LT 0.02
LT 0.05 LT 0.02 LT 0.02 LT 0.06 Batch 7 LT 0.02 LT 0.03 NA LT 0.02
LT 0.02 LT 0.02 LT 0.05 LT 0.02 LT 0.02 LT 0.06 Batch 8 LT 0.02 LT
0.03 NA LT 0.02 LT 0.02 LT 0.02 LT 0.05 LT 0.02 LT 0.02 LT 0.06
Batch 9 LT 0.02 LT 0.03 NA LT 0.02 LT 0.02 LT 0.02 LT 0.05 LT 0.02
LT 0.02 LT 0.02 Batch 10 LT 0.02 LT 0.03 NA LT 0.02 LT 0.02 LT 0.02
LT 0.05 LT 0.02 LT 0.02 LT 0.02 Batch 11 LT 0.02 LT 0.03 NA LT 0.02
LT 0.02 LT 0.02 LT 0.05 LT 0.02 LT 0.02 LT 0.02 Batch 12 LT 0.02 LT
0.03 NA LT 0.02 LT 0.02 LT 0.02 LT 0.05 LT 0.02 LT 0.02 LT 0.02 QL
And DL Values Of Impurities Listed In Table 48: Quantitation Limit
Detection Limit MCQ 0.05% 0.02% MCQCA .sup. 0.05%.sup.1 0.02% MCQME
0.05% 0.02% 5-HLAQ 0.05% 0.02% MCQEE 0.05% 0.02% LAQ 0.2% 0.03% NEA
0.06% 0.02% DELAQ Not applicable (limit test) 0.03% DMM Not
applicable (limit test) 0.03% DMS Not applicable (limit test) 0.5
ppm Acetone 250 ppm 25 ppm Dioxane 20 ppm 10 ppm Ethanol 25 ppm 5
ppm n-Heptane 250 ppm 10 ppm Methanol 150 ppm 30 ppm n-Octane 100
ppm 10 ppm DMF 190 ppm 50 ppm .sup.1Reporting limit 0.10%.
TABLE-US-00088 TABLE 49 Analysis of Additional Laquinimod Sodium
Drug Substance Batches 13-18 Related Substances [%] Batch Any
individual impurity Sum of No. .ltoreq.1.0 imp. Specifications MCQ
MCQCA MCQME .ltoreq.2.0 13 Any single impurity <0.05 14 Any
single impurity <0.05 <0.05 15 0.03 0.03 0.03 0.13 16 -- --
0.04 0.17 17 0.01 -- 0.01 0.13 18 -- -- 0.05 0.14
TABLE-US-00089 TABLE 50 Analyses of Laquinimod (Acid Form) Drug
Substance Batches (Batches 19-22) Related Substances [%] Batch NEA
Any No. [%] MCQ MCQCA MCQME Unkown Total 19 -- 0.75 -- -- 1.2 1.95
20 -- -- -- 0.24.sup.4 0.05.sup.4 0.55 0.05.sup.4 0.07.sup.4
0.14.sup.4 21 -- -- -- 0.26.sup.4 0.05.sup.4 0.56 0.05.sup.4
0.07.sup.4 0.13.sup.4 22 <0.05 0.19 <0.5 .sup.4Deviation from
method. Different mobile phase and wavelength used.
Example 26
Observed Impurities at Release and after Long-Term and Accelerated
Storage of Laquinimod Drug Substance
[1072] Impurities were observed in laquinimod sodium batches at
release and after long-term and accelerated storage.
[1073] The ranges of impurity levels observed in the laquinimod
sodium batches, including the primary stability and validation
batches, are summarized in Table 51.
TABLE-US-00090 TABLE 51 Ranges of Impurity Levels Observed in
Laquinimod Drug Substance Batches upon Release and after Long-Term
and Accelerated Storage Range of Observed Parameter Current
Specification Impurity Levels MCQ NMT 0.15% LT 0.02-LT 0.05% MCQCA
NMT 0.15% LT 0.03-LT 0.05% MCQME NMT 0.12% LT 0.02-LT 0.05% 5-HLAQ
NMT 0.10% LT 0.02-LT 0.05% MCQEE NMT 0.10% LT 0.02-LT 0.05% LAQ NMT
1.0% LT 0.025-LT 0.2% NEA NMT 0.10% LT 0.02-LT 0.05% DELAQ NMT 0.1%
LT 0.05-LT 0.1% DMM NMT 0.1% LT 0.05-LT 0.1% DMS NMT 1 ppm LT 1 ppm
Ethanol NMT 5000 ppm LT 5-LT 25 ppm n-Heptane NMT 5000 ppm LT 10-LT
250 ppm Methanol NMT 3000 ppm LT 30-LT 150 ppm Dioxane NMT 380 ppm
LT 10 ppm DMF NMT 880 ppm LT 40-93 ppm Acetone NMT 5000 ppm LT 250
ppm n-Octane NMT 2000 ppm LT 10-LT 20 ppm Heavy Metals NMT 20 ppm
NMT 20 ppm Free chloride NMT 0.1% LT 0.1%
Example 27
Batch Analysis of Laquinimod Sodium Drug Product
[1074] A number of batches of laquinimod sodium drug product were
manufactured at various manufacturing facilities and subsequently
analyzed. During manufacture and analysis, sufficient light
protection of sample solution was provided.
[1075] All batches of drug product were within specification.
[1076] Evaluation of the release data shows that all batches have
been within the specifications in effect at the time of their
release. Furthermore it can be seen that all manufactured batches
of the product proposed for commercialization, e.g., laquinimod 0.6
mg capsules, have shown compliance with the proposed commercial
specifications. Batch-to-batch consistency over time was found in
the results for all the measured parameters. The batch analysis
results demonstrate that the quality of all batches does not differ
significantly from each other and that production at different
manufacturing sites affords material of similar quality.
[1077] Table 52, Table 53, Table 54, Table 55, Table 55, Table 56
and Table 57 relate to batch analyses of laquinimod sodium drug
product batches.
TABLE-US-00091 TABLE 52 Batch Analysis for Laquinimod 0.6 mg
Capsules Batches 23-31 Polar Impurities/ Degradation Products [%]
Non Polar Impurities/ Sum of Degradation Products [%] MCQ & Any
Total Any Total BH-3-HLAQ MCQCA Other Imp. NEA Other Imp. [%]
Current Method Example 21 Example 22 Example 23 Current
Specifications Batch NMT NMT NMT NMT Number NMT 0.5 0.5 2.0 0.5 NMT
0.5 1.0 NMT 1.0 Batch 23 <0.03 <0.02 <0.05 <0.03
<0.03 <0.1 <0.02.sup.5 Batch 24 <0.03 <0.02 <0.05
<0.03 <0.03 <0.1 <0.02.sup.5 Batch 25 <0.03 <0.02
<0.05 <0.03 <0.03 <0.1 <0.02.sup.5 Batch 26 <0.03
<0.02 <0.05 <0.03 <0.03 <0.1 <0.02.sup.5 Batch 27
<0.03 <0.02 <0.05 <0.03 <0.03 <0.1 <0.02.sup.5
0.06.sup.5,6 Batch 28 <0.03 <0.05 <0.05 <0.03 <0.03
<0.1 <0.02.sup.5 Batch 29 <0.03 <0.02 <0.05 <0.03
<0.03 <0.1 <0.02.sup.5 Batch 30 <0.03 <0.05 <0.05
<0.03 <0.03 <0.1 <0.02.sup.5 Batch 31 <0.03 <0.05
<0.05 <0.03 <0.03 <0.1 <0.02.sup.5 .sup.4The
analysis was performed using the previous method for determination
of MEG-LAQ. .sup.5The test for BH-3-HLAQ was introduced after the
release of these batches. The listed results were obtained for
BH-3-HLAQ at the last available stability timepoint at 25.degree.
C./60% RH (18 months for batch 23 and batches 27-30, 9 months for
batches 24-26 and 3 months for batch 31) for all packaging
configurations, with one exception (see footnote 6). .sup.6For
batch 27 in Alu/Alu blisters a result of 0.06% was obtained.
Current QL and DL values for Table 52: Quantitation Limit Detection
Limit Sum of MCQ and 0.1% 0.03% MCQCA 5-HLAQ 0.05% 0.02% NEA.sup.1
0.1% 0.03% 3-HLAQ.sup.1 0.1% 0.03% MEG-LAQ 0.2% 0.1% BH-3-HLAQ
0.05% 0.02% .sup.1NEA and 3-HLAQ were omitted from the commercial
specifications since they were never observed upon batch release
and stability testing completed to date at all tested storage
conditions. The monitoring of these impurities in the primary
stability batches will continue until the end of the stability
program, using the listed method.
TABLE-US-00092 TABLE 53 Batch Analyses for Laquinimod 0.6 mg
Capsules (Batches 32-38) Non Polar Impurities/ Polar Impurities/
Degradation Products [%] Degradation Sum of Products [%] MCQ &
Total Any Total BH-3-HLAQ MCQCA Any Other Impurities NEA Other Imp.
[%] Current Specifications Batch NMT NMT NMT Number NMT 0.5 NMT 0.5
NMT 2.0 0.5 0.5 1.0 NMT 1.0 Batch 32 <0.03 MCQME: <0.02
<0.05 <0.03 <0.03 <0.1 NA.sup.6 MCQEE: <0.02 Batch
33 <0.03 MCQME: <0.02 <0.05 <0.03 <0.03 <0.1
NA.sup.6 MCQEE: <0.02 Batch 34 <0.03 MCQME: <0.02 <0.05
<0.03 <0.03 <0.1 NA.sup.6 MCQEE: <0.02 Batch 35
<0.03 MCQME: <0.02 <0.05 <0.03 <0.03 <0.1
NA.sup.6 MCQEE: <0.02 Batch 36 <0.03 MCQME: <0.02 <0.05
<0.03 <0.03 <0.1 NA.sup.6 MCQEE: <0.02 Batch 37
<0.03 MCQME: <0.02 <0.05 <0.03 <0.03 <0.1
NA.sup.6 MCQEE: <0.02 RRT: 0.23: 0.05 Batch 38 <0.03 MCQME:
<0.02 <0.05 <0.03 <0.03 <0.1 NA.sup.6 MCQEE:
<0.02 .sup.6Determination of BH-3-HLAQ and water content were
introduced in the specifications after the release of this batch.
Current QL and DL values of Table 53: Quantitation Limit Detection
Limit Sum of MCQ 0.1% 0.03% and MCQCA 5-HLAQ 0.05% 0.02% NEA.sup.1
0.1% 0.03% 3-HLAQ.sup.1 0.1% 0.03% MEG-LAQ 0.2% 0.1% BH-3-HLAQ
0.05% 0.02% .sup.1NEA and 3-HLAQ were omitted from the commercial
specifications since they were never observed upon release and
during the stability testing completed to date at all tested
storage conditions. The monitoring of these impurities in the
primary stability batches will continue until the end of the
stability program, using the listed method.
TABLE-US-00093 TABLE 54 Batch Analyses for Laquinimod Capsules 0.3
mg (Batches 39-42) Non Polar Impurities/Degradation Polar
Impurities/Degradation Products [%] Products [%] Sum of MCQ &
Total Any Total MCQCA MCQEE Any Other Impurities NEA Other
Impurities Current Specifications Batch NMT NMT Number NMT 0.5 NA
NMT 0.5 NMT 2.0 0.5 0.5 NMT 1.0 Batch 39 MCQ: <0.02 <0.02 RRT
0.35: <0.05 <0.05 <0.02 <0.02 <0.05 MCQCA: <0.03
Batch 40 <0.03 NA MCQME: <0.02 <0.05 <0.03 <0.03
<0.1 MCQEE: <0.02 Batch 41 <0.03 NA MCQME: <0.02
<0.05 <0.03 <0.03 <0.1 MCQEE: <0.02 Batch 42
<0.03 NA MCQME:< 0.02 <0.05 <0.03 <0.03 <0.1
MCQEE: <0.02
TABLE-US-00094 TABLE 55 Batch Analyses for Laquinimod Capsules 0.3
mg (Batch 43) Non Polar Impurities/ Polar Impurities/Degradation
Products [%] Degradation Products [%] Total Total MCQ MCQEE MCQCA
Any Other Impurities NEA Any Other Impurities Specifications Batch
Strength NMT NMT NMT Number (Formula) 0.5 0.5 NMT 0.5 NMT 0.5 NMT
2.0 0.5 NMT 0.5 NMT 1.0 Batch 0.3 mg <0.02 <0.02 <0.03 RRT
0.35: 0.3 <0.02 RRT 2.30: 0.1 43 (III) <0.05 0.09 RRT 0.18:
0.09 RRT 0.24: 0.08
TABLE-US-00095 TABLE 56 Batch Analyses for Laquinimod Tablets 0.05
mg, 0.25 mg and 0.3 mg (Batches 44-49) Related Substances [%] MCQ
& Any Sum of NEA MCQCA MCQEE unknown Imp. Last Effective
Specifications Batch NMT Number Strength NMT 0.1 NMT 0.5 NMT 0.1
NMT 0.2 1.5 Batch 44 0.05 mg <0.0080 Each <1.0 Each <1.0
Each <1.0 <0.10 Batch 45 0.25 mg 0.009 Each <1.0 Each
<1.0 Each <1.0 <0.10 Batch 46 0.3 mg 0.031 0.22 <0.02
Largest 0.22.sup.4 <0.03 Batch 47 0.3 mg 0.04 0.05 <0.020
Conforms 0.05 Batch 48 0.3 mg 0.03 0.05 <0.020 Conforms 0.05
Batch 49 0.3 mg 0.016 0.17 <0.020 <0.030 0.20 .sup.4An
additional specified impurities were reported: 0.03% of MCQME.
TABLE-US-00096 TABLE 57 Batch Analysis for Laquinimod Oral Solution
1 mg/mL (Batch 50) Batch Number Strength Related Substances [%]
N-Ethylaniline [%] Specifications NMT 4.0 NMT 0.1 Batch 50 1 mg/mL
<4.0 <0.01
REFERENCES
[1078] 1. U.S. Pat. No. 6,077,851; [1079] 2. U.S. Pat. No.
6,875,869; [1080] 3. U.S. Pat. No. 7,560,557; [1081] 4. U.S. Pat.
No. 7,589,208; [1082] 5. U.S. Pat. No. 7,884,208; [1083] 6. U.S.
Pat. No. 7,989,473; [1084] 7. U.S. Pat. No. 8,178,127; [1085] 8.
U.S. Patent Application Publication No. 2006/0188581 A1; [1086] 9.
U.S. Patent Application Publication No. 2012/0010239 A1; [1087] 10.
U.S. Patent Application Publication No. 2013/0217724 A1; [1088] 11.
U.S. Patent Application Publication No. 2013/00345256; [1089] 12.
PCT International Application Publication No. WO 2005/074899;
[1090] 13. PCT International Application Publication No. WO
2003/106424 A1; [1091] 14. PCT International Application
Publication No. WO 2007/047863 A22; [1092] 15. 21 C.F.R.
.sctn.211.166; [1093] 16. Felmeister, A. Chpt 88, Remington's
Pharmaceutical Sciences, 15.sup.th Edition, Mack Publishing
Company, Easton, Pa. (1975); [1094] 17. Hausner, "The Role of
Particle Size in the Development of Generic Products" 2003; [1095]
18. Brumfitt, W. and J. M. T. Hamilton-Miller, J. Antimicrobial
Chemotherapy 42:363:371 (1998); [1096] 19. Rudnic et al. Chpt. 45,
Remington's Pharmaceutical Sciences, 20.sup.th Edition, Lippincott
Williams & Wilkins, Baltimore, Md. (2000); [1097] 20. United
States Pharmacopeia XXV (2000); [1098] 21. United States
Pharmacopeia 34/National Formulary 29, (2011); [1099] 22. 7 Modern
Pharmaceutics, Chapters 9 and 10 (Banker & Rhodes, Editors,
1979); [1100] 23. Pharmaceutical Dosage Forms: Tablets (Lieberman
et al., 1981); [1101] 24. Ansel, Introduction to Pharmaceutical
Dosage Forms 2nd Edition (1976); [1102] 25. Remington's
Pharmaceutical Sciences, 17th ed, (Mack Publishing Company, Easton,
Pa., 1985); [1103] 26. Advances in Pharmaceutical Sciences (David
Ganderton, Trevor Jones, Eds., 1992); [1104] 27. Advances in
Pharmaceutical Sciences Vol. 7. (David Ganderton, Trevor Jones,
James McGinity, Eds., 1995); [1105] 28. Aqueous Polymeric Coatings
for Pharmaceutical Dosage Forms (Drugs and the Pharmaceutical
Sciences, Series 36 (James McGinity, Ed., 1989); [1106] 29.
Pharmaceutical Particulate Carriers: Therapeutic Applications:
Drugs and the Pharmaceutical Sciences, Vol 61 (Alain Rolland, Ed.,
1993); [1107] 30. Drug Delivery to the Gastrointestinal Tract
(Ellis Horwood Books in the Biological Sciences. Series in
Pharmaceutical Technology; J. G. Hardy, S. S. Davis, Clive G.
Wilson, Eds.) (1989); [1108] 31. Modem Pharmaceutics Drugs and the
Pharmaceutical Sciences, Vol 40 (Gilbert S. Banker, Christopher T.
Rhodes, Eds.)(1996); [1109] 32. Vogel's Textbook of Practical
Organic Chemistry. 5.degree. edition. Longman Scientific &
Technical, 1989; [1110] 33. Polman, C. et al., (2005) "Treatment
with laquinimod reduces development of active MRI lesions in
relapsing MS", Neurology. 64:987-991; [1111] 34. Sandberg-Wollheim
M, et al. (2005) "48-week open safety study with high-dose oral
laquinimod in patients", Mult Scler. 11:S154); and [1112] 35. K.
Jansson et al., "Synthesis and Reactivity of Laquinimod, a
Quinoline-3-carboxamide: Intramolecular Transfer of the Enol Proton
to a Nitrogen Atom as a Plausible Mechanism for Ketene Formation
(2006) J. Org. Chem., 71, 1658-1667.
* * * * *
References