U.S. patent application number 15/711502 was filed with the patent office on 2018-04-19 for salts and polymorphs of a tetracycline compound.
The applicant listed for this patent is Paratek Pharmaceuticals, Inc.. Invention is credited to Raymond Cvetovich, Tadeusz Warchol.
Application Number | 20180104262 15/711502 |
Document ID | / |
Family ID | 41340587 |
Filed Date | 2018-04-19 |
United States Patent
Application |
20180104262 |
Kind Code |
A1 |
Cvetovich; Raymond ; et
al. |
April 19, 2018 |
SALTS AND POLYMORPHS OF A TETRACYCLINE COMPOUND
Abstract
Crystalline forms, including salts and polymorphs, of a compound
useful in the treatment of tetracycline compound-responsive states
are provided herein. The crystalline compounds are useful for the
treatment or prevention of conditions and disorders such as
bacterial infections and neoplasms, as well as other known
applications for tetracycline compounds in general.
Inventors: |
Cvetovich; Raymond; (Scotch
Plains, NJ) ; Warchol; Tadeusz; (Northborough,
MA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Paratek Pharmaceuticals, Inc. |
Boston |
MA |
US |
|
|
Family ID: |
41340587 |
Appl. No.: |
15/711502 |
Filed: |
September 21, 2017 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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14962326 |
Dec 8, 2015 |
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15711502 |
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13749101 |
Jan 24, 2013 |
9227921 |
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14962326 |
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12471758 |
May 26, 2009 |
8383610 |
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13749101 |
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61128712 |
May 23, 2008 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61P 31/12 20180101;
A61P 35/00 20180101; A61P 31/10 20180101; C07C 237/26 20130101;
A61K 31/65 20130101; A61P 35/02 20180101; C07B 2200/13 20130101;
A61P 19/10 20180101; A61P 31/00 20180101; A61P 17/00 20180101; Y02A
50/30 20180101; A61P 29/00 20180101; A61P 31/04 20180101; C07C
231/12 20130101; C07C 2603/46 20170501; A61P 3/10 20180101; A61P
19/02 20180101; A61P 1/12 20180101; C07C 303/32 20130101; C07C
309/30 20130101 |
International
Class: |
A61K 31/65 20060101
A61K031/65; C07C 303/32 20060101 C07C303/32; C07C 309/30 20060101
C07C309/30; C07C 237/26 20060101 C07C237/26; C07C 231/12 20060101
C07C231/12 |
Claims
1. A crystalline form of Compound 1: ##STR00006##
2. A tosylate salt of Compound 1: ##STR00007##
3. A crystalline form of the salt of claim 2.
4. (canceled)
5. A polymorph of the crystalline form of claim 3, characterized by
an X-ray powder diffraction pattern including peaks at
approximately 8.06, 13.02, and 18.83 .degree. 20 using Cu K.alpha.
radiation.
6. The polymorph of claim 5, characterized by an X-ray powder
diffraction pattern including peaks at approximately 8.06, 11.41,
13.02, 18.83, 20.54 and 24.53 .degree. 2.theta. using Cu K.alpha.
radiation.
7. The polymorph of claim 5, characterized by an X-ray powder
diffraction pattern including peaks at approximately 5.60, 8.06,
8.57, 11.41, 13.02, 15.58, 18.83, 20.54 and 24.53 .degree. 2.theta.
using Cu K.alpha. radiation.
8. (canceled)
9. A method of preparing a stable crystalline tosylate salt of
Compound 1: ##STR00008## wherein the method comprises: dissolving a
freebase of Compound 1 in a first solvent or combination of
solvents to form a first solution; dissolving p-toluenesulfonic
acid in a second solvent or combination of solvents to form a
second solution; and combining said first and second solution to
form a third solution.
10-29. (canceled)
30. A polymorph of the crystalline form of claim 3, characterized
by an X-ray powder diffraction pattern including peaks at
approximately 11.88 and 16.12 .degree. 2.theta. using Cu K.alpha.
radiation.
31. A polymorph of the crystalline form of claim 3, characterized
by an X-ray powder diffraction pattern including peaks at
approximately 7.82, 11.88, 16.12 and 21.46 .degree. 2.theta. using
Cu K.alpha. radiation.
32. A polymorph of the crystalline form of claim 3, characterized
by an X-ray powder diffraction pattern including peaks at
approximately 5.11 and 15.60 .degree. 2.theta. using Cu K.alpha.
radiation.
33. A polymorph of the crystalline form of claim 3, characterized
by an X-ray powder diffraction pattern including peaks at
approximately 5.11, 8.89, 10.34, 11.76 and 15.60 .degree. 2.theta.
using Cu K.alpha. radiation.
34. A method of treating a tetracycline responsive state in a
subject in need thereof, the method comprising administering to
said subject an effect amount of a crystalline form of a tosylate
salt of Compound 1: ##STR00009##
35. The method of claim 34, wherein the crystalline form of a
tosylate salt of Compound 1 is a polymorph selected from the group
consisting of: a polymorph characterized by an X-ray powder
diffraction pattern including peaks at approximately 8.06, 13.02,
and 18.83 .degree. 2.theta. using Cu K.alpha. radiation, a
polymorph characterized by an X-ray powder diffraction pattern
including peaks at approximately 5.11 and 15.60 .degree. 2.theta.
using Cu K.alpha. radiation, and a polymorph characterized by an
X-ray powder diffraction pattern including peaks at approximately
11.88 and 16.12 .degree. 2.theta. using Cu K.alpha. radiation.
36. The method of claim 34, wherein said crystalline form of a
tosylate salt of Compound 1 is administered as a pharmaceutical
composition comprising said crystalline form of a tosylate salt of
Compound 1 and a pharmaceutically acceptable diluent, excipient or
carrier.
37. The method of claim 34, wherein said tetracycline responsive
state is a bacterial infection.
38. The method of claim 37, wherein said bacterial infection is
associated with gram positive bacteria.
39. The method of claim 37, wherein the bacterial infection is
associated with gram negative bacteria.
40. The method of claim 37, wherein said bacterial infection is
associated with a bacterial species selected from the group
consisting of K. pneumoniae, Salmonella, E. hirae, A. baumanii, B.
catarrhalis, H influenzae, P. aeruginosa, E. faecium, E. coli, S.
aureus and E. faecalis.
41. The method of claim 37, wherein the bacterial infection is
resistant to other tetracycline antibiotics.
42. The method of claim 41, wherein the other tetracycline
antibiotics are selected from one or more of tetracycline,
minocycline, doxycycline, sancycline, chlortetracycline,
demeclocyclin, oxytetracycline, chelocardin, rolitetracycline,
lymecycline, methacycline, apicycline, clomocycline, pipacycline,
mepylcycline, meglucycline, guamecycline, penimocycline, or
etamocycline.
Description
RELATED APPLICATION
[0001] This application is a continuation application of U.S.
patent application Ser. No. 14/962,326, filed on Dec. 8, 2015;
which is a continuation application of U.S. patent application Ser.
No. 13/749,101, filed on Jan. 24, 2013, now U.S. Pat. No.
9,227,921; which is a division of U.S. patent application Ser. No.
12/471,758, filed on May 26, 2009, now U.S. Pat. No. 8,383,610;
which claims the benefit of the filing date under 35 U.S.C. 119(e)
of U.S. Provisional Application No. 61/128,712, filed on May 23,
2008. The entire contents of each of the foregoing applications are
incorporated herein by reference.
BACKGROUND OF THE INVENTION
[0002] The development of the tetracycline antibiotics was the
direct result of a systematic screening of soil specimens collected
from many parts of the world for evidence of microorganisms capable
of producing bactericidal and/or bacteriostatic compositions. The
first of these novel compounds was introduced in 1948 under the
name chlortetracycline. Two years later, oxytetracycline became
available. The elucidation of the chemical structure of these
compounds confirmed their similarity and furnished the analytical
basis for the production of a third member of this group in 1952,
tetracycline. A new family of tetracycline compounds, without the
ring-attached methyl group present in earlier tetracyclines, was
prepared in 1957 and became publicly available in 1967; and
minocycline was in use by 1972.
[0003] Recently, research efforts have focused on developing new
tetracycline antibiotic compositions effective under varying
therapeutic conditions and routes of administration. New
tetracycline analogues have also been investigated which may prove
to be equal to or more effective than the originally introduced
tetracycline compounds. Examples include U.S. Pat. Nos. 2,980,584;
2,990,331; 3,062,717; 3,165,531; 3,454,697; 3,557,280; 3,674,859;
3,957,980; 4,018,889; 4,024,272; and 4,126,680. These patents are
representative of the range of pharmaceutically active tetracycline
and tetracycline analogue compositions.
[0004] Historically, soon after their initial development and
introduction, the tetracyclines were found to be highly effective
pharmacologically against rickettsiae; a number of gram-positive
and gram-negative bacteria; and the agents responsible for
lymphogranuloma venereum, inclusion conjunctivitis, and
psittacosis. Hence, tetracyclines became known as "broad spectrum"
antibiotics. With the subsequent establishment of their in vitro
antimicrobial activity, effectiveness in experimental infections,
and pharmacological properties, the tetracyclines as a class
rapidly became widely used for therapeutic purposes. However, this
widespread use of tetracyclines for both major and minor illnesses
and diseases led directly to the emergence of resistance to these
antibiotics even among highly susceptible bacterial species both
commensal and pathogenic (e.g., pneumococci and Salmonella). The
rise of tetracycline-resistant organisms has resulted in a general
decline in use of tetracyclines and tetracycline analogue
compositions as the antibiotic of choice.
[0005] Each pharmaceutical compound has an optimal therapeutic
blood concentration and a lethal concentration. The bioavailability
of the compound determines the dosage strength in the drug
formulation necessary to obtain the ideal blood level. If the drug
can crystallize as two or more polymorphs differing in
bioavailability, the optimal dose will depend on the polymorph
present in the formulation. Some drugs show a narrow margin between
therapeutic and lethal concentrations. Chloramphenicol-3-palmitate
(CAPP), for example, is a broad-spectrum antibiotic known to
crystallize in at least three polymorphic forms and one amorphous
form. The most stable form, A, is marketed. The difference in
bioactivity between this polymorph and another form, B, is a factor
of eight, thus creating the possibility of fatal overdosages of the
compound if unwittingly administered as form B due to alterations
during processing and/or storage. Therefore, regulatory agencies,
such as the United States Food and Drug Administration, have begun
to place tight controls on the polymorphic content of the active
component in solid dosage forms. In general, for drugs that exist
in polymorphic forms, if anything other than the pure,
thermodynamically preferred polymorph is to be marketed, the
regulatory agency may require batch-by-batch monitoring. Thus, it
becomes important for both medical and commercial reasons to
produce and market the pure drug in its most thermodynamically
stable polymorph, substantially free of other kinetically favored
polymorphs.
[0006] For instance, salt forms of a compound, and polymorphic
forms of the free compound or salt, are known in the pharmaceutical
art to affect, for example, the solubility, dissolution rate,
bioavailability, chemical and physical stability, flowability,
fractability, and compressibility of the compound as well as the
safety and efficacy of drug products based on the compound (see,
e.g., Knapman, Modern Drug Discovery, 2000, 3(2): 53).
[0007] Accordingly, identification of a salt form or free base of a
compound with optimal physical and chemical properties will advance
the development of tetracycline compounds as pharmaceuticals. The
most useful of such physical and chemical properties include: easy
and reproducible preparation, crystallinity, non-hygroscopicity,
aqueous solubility, stability to visible and ultraviolet light, low
rate of degradation under accelerated stability conditions of
temperature and humidity, low rate of isomerization between
isomeric forms, and safety for long-term administration to
humans.
SUMMARY OF THE INVENTION
[0008] In one embodiment, the invention pertains, at least in part,
to a stable solid state form, such as a crystalline form, of the
aminoalkyl tetracycline compound, Compound 1:
##STR00001##
(4S,4AS,5AR,12AS)-4-7-Bis(dimethylamino)-9{[(2,2-dimethylpropyl)amino]met-
hyl}-3,10,12,12A-tetrahydroxy-1,11-dioxo-1,4,4A,5,5A,6,11,12A-octahydrotet-
racene-2-carboxamide (9-(2,2-dimethy 1-propy
1-aminomethyl)-minocycline)
[0009] In another embodiment, the invention pertains, at least in
part, to an HCl salt of Compound 1. In another embodiment, the
invention pertains, at least in part, to a tosylate
(p-toluenesulfonate) salt of Compound 1. In another embodiment, the
invention pertains, at least in part, to a mesylate salt of
Compound 1.
[0010] In another embodiment, the invention pertains, at least in
part, to a stable crystalline form of Compound 1.
[0011] In another embodiment, the invention pertains, at least in
part, to a stable crystalline form of a salt of Compound 1. For
example, the stable crystalline form of a salt is a stable
crystalline form of a tosylate, HCl or mesylate salt of Compound
1.
[0012] In another embodiment, the invention pertains, at least in
part, to a polymorph of Compound 1.
[0013] In another embodiment, the invention pertains, at least in
part, to a polymorph of a salt of Compound 1.
[0014] For example, the invention relates to a polymorph of a
tosylate salt of Compound 1. The invention relates, in part to a
form 1 polymorph of Compound 1. The invention relates, in part to a
form 2 polymorph of Compound 1. The invention relates, in part to a
form 3 polymorph of Compound 1.
[0015] For example, a form 1 polymorph of a tosylate salt of
Compound 1 has X-ray powder diffraction peaks at approximately
8.06, 13.02, and 18.83 .degree. 2.theta. using Cu K.alpha.
radiation. In some embodiments, the form 1 polymorph of a tosylate
salt of Compound 1 has X-ray powder diffraction peaks at
approximately 8.06, 11.41, 13.02, 18.83, 20.54, and 24.53 .degree.
2.theta. using Cu K.alpha. radiation. In some embodiments, the form
1 polymorph of a tosylate salt of Compound 1 has X-ray powder
diffraction peaks at approximately 5.60, 8.06, 8.57, 11.41, 13.02,
15.58, 18.83, 20.54, and 24.53 .degree. 2.theta. using Cu K.alpha.
radiation.
[0016] For example, a form 1 polymorph of a tosylate salt of
Compound 1 is stable at temperature in a range from about 0.degree.
C. to about 70.degree. C. In some embodiments, the form 1 polymorph
of a tosylate salt of Compound 1 is stable at temperature in a
range from about 5.degree. C. to about 50.degree. C. In some
embodiments, the form 1 polymorph of a tosylate salt of Compound 1
is stable at temperature in a range from about 20.degree. C. to
about 30.degree. C.
[0017] The form 1 polymorph of a tosylate salt of Compound 1 can be
obtained by crystallizing the tosylate salt of said Compound 1 from
isopropanol.
[0018] For example, a form 2 polymorph of a tosylate salt of
Compound 1 has X-ray powder diffraction pattern peaks at 7.82,
11.88, 16.12 and 21.46 .degree. 28 using Cu Ka radiation.
[0019] For example, a form 3 polymorph of a tosylate salt of
Compound 1 has X-ray powder diffraction pattern peaks at 5.11,
8.89, 10.34, 11.76 and 15.60 .degree. 28 using Cu K.alpha.
radiation.
[0020] In yet another embodiment, the invention includes
pharmaceutical compositions comprising a crystalline form of
Compound 1 and a pharmaceutically acceptable diluent, excipient or
carrier.
[0021] For example, the pharmaceutical composition of the invention
includes a composition comprising a polymorph of Compound 1 and a
pharmaceutically acceptable diluent, excipient or carrier.
[0022] In another embodiment, the pharmaceutical composition of the
invention includes a salt of Compound 1 and a pharmaceutically
acceptable diluent, excipient or carrier. For example, the salt can
be an HCl salt, a tosylate salt, or a mesylate salt.
[0023] In one embodiment, the pharmaceutical composition of the
invention includes a polymorph of a salt of Compound 1 and a
pharmaceutically acceptable diluent, excipient or carrier. For
example, the polymorph can be a polymorph of tosylate salt, HCl
salt, or mesylate salt of Compound 1.
[0024] In some embodiments, the pharmaceutical composition
comprises a polymorph of Compound 1, or salt thereof in a pure
form.
[0025] In another embodiment, the pharmaceutical composition of the
invention includes a polymorph of the tosylate salt of Compound 1
and a pharmaceutically acceptable diluent, excipient or carrier.
For example, the polymorph can be a form 1, a form 2, or a form 3
polymorph of tosylate salt of Compound 1.
[0026] In some embodiments, the pharmaceutical composition
comprises a polymorph of tosylate salt, HCl salt, or mesylate salt
of Compound 1 in a pure form.
[0027] In another aspect of the invention, the salt of Compound 1
is more stable than the free base of Compound 1.
[0028] In another embodiment, the invention includes a method for
preparing a stable crystalline form of Compound 1.
[0029] In another embodiment, the invention includes a method for
preparing a stable crystalline form of a salt of Compound 1. For
example, the stable crystalline can be a crystalline of a tosylate,
HCl, or mesylate salt of Compound 1.
[0030] In another embodiment, the invention includes a method for
preparing a polymorph of a salt of Compound 1. For example, the
polymorph can be a polymorph of a tosylate, HCl, or mesylate salt
of Compound 1.
[0031] In another embodiment, the invention includes a method for
preparing a polymorph of a tosylate salt of Compound 1. For
example, the polymorph can be a form 1, a form 2, or a form 3
polymorph of a tosylate salt of Compound 1.
[0032] In one embodiment, the invention includes a method for
preparing a form 1 of polymorph of a tosylate salt of Compound 1,
wherein the method comprises: combining Compound 1 with a solvent
to produce a slurry; and adding p-toluenesulfonic acid. For
example, the solvent can be an alcoholic solvent, such as
isopropanol. The p-toluenesulfonic acid is provided in an amount of
from 25 to 75 wt % relative to the amount of said Compound 1, for
example, from 25 to 50 wt %, from 30 to 40 wt %, or 33 wt %
relative to the amount of said Compound 1. For example, the
p-toluenesulfonic acid is provided in a form of p-toluenesulfonic
acid monohydrate.
[0033] For example, the slurry is warmed prior to the addition of
p-toluenesulfonic acid.
[0034] For example, the slurry is stirred after the addition of
p-toluenesulfonic acid. For example, the stirring is conducted at a
temperature in a range from 20 to 25.degree. C. For example, the
stirring is conducted for 10 to 24 hours.
[0035] For example, the slurry is dried. For example, the water
content of the supernatant of said slurry is in a range from 0.2 to
1.0 mg/mL, or in a range from 0.4 to 0.8 mg/mL.
[0036] In yet another embodiment, the invention includes a method
for preparing a form 1 polymorph of a tosylate salt of Compound 1,
wherein the method comprises: preparing a solution of Compound 1 in
a solvent or a combination of solvents; and adding a solution of
p-toluenesulfonic acid in a solvent or a combination of
solvents.
[0037] For example, the solvent is an alcoholic solvent, such as
methanol, ethanol, or isopropanol. For example, the combination of
solvents includes an alcoholic solvent. For example, the
combination of solvents further comprises a second alcoholic
solvent. For example, the combination of solvents includes ethanol
and isopropanol. For example, the combination of solvents further
includes an anti-solvent, such as ketone, ether, and ester. For
example, the ether includes, but is not limited to, methyl-t-butyl
ether. For example, the combination of solvents includes an
alcoholic solvent and an anti-solvent. For example, the combination
of solvents includes methanol and methyl-t-butyl ether.
[0038] For example, p-toluenesulfonic acid is provided in an amount
of from 25 to 75 wt %, from 30 to 50 wt %, from 35 to 45 wt %, or
40 wt % relative to the amount of said Compound 1. For example,
p-toluenesulfonic acid is provided in a form of a p-toluenesulfonic
acid monohydrate.
[0039] For example, the solution is prepared at a temperature in a
range from 0 to 60.degree. C., at a temperature in a range from 15
to 45.degree. C., or at a temperature in a range from 20 to
25.degree. C.
[0040] For example, the solution is warmed after it is prepared.
For example, the solution is maintained at a temperature in a range
from 20 to 50.degree. C., or at about 45.degree. C.
[0041] For example, the method further comprises adding a seed
crystal of monotosylate salt of Compound 1 to produce a slurry. The
slurry may be stirred for 10 to 24 hours or for about 22 hours. The
slurry may be stirred at a temperature in a range from 15 to
45.degree. C. or at about 20.degree. C. The slurry may be dried.
For example, the water content of the slurry is in a range of 1 to
10 wt %, or in a range of 2 to 6 wt %, or about 3 wt %.
[0042] In another embodiment, the invention includes a method for
preparing a form 1 polymorph of a tosylate salt of Compound 1,
wherein the method comprises: dissolving a freebase of Compound 1
in a first solvent or combination of solvents to form a first
solution; dissolving p-toluenesulfonic acid in a second solvent or
combination of solvents to form a second solution; and combining
said first and second solution to form a third solution.
[0043] In one embodiment, the first and second solvent or
combination of solvents can be the same or different. In another
embodiment, the solvent can be an alcoholic solvent, such as
methanol, ethanol, and isopropanol. In another embodiment, the
combination of solvents is a combination of two alcoholic solvents,
including, but not limited to ethanol and isopropanol. In a
preferred example, the volume-to-volume ratio of ethanol and
isopropanol is 2 to 1. In yet another embodiment, the combination
of solvents is a combination that includes, but is not limited to,
an alcoholic solvent and an anti-solvent (e.g., a ketone, an ether,
an ester, etc.). For example, the combination of solvents is a
combination that includes, but is not limited to, methanol and
methyl-t-butyl ether. In a preferred example, the volume-to-volume
ratio of methanol and methyl-t-butyl ether is 1 to 1.2.
[0044] In another embodiment, the method further comprises adding a
form 1 polymorph tosylate salt of Compound 1 to the third solution
to form a fourth solution. For example, the form 1 polymorph
tosylate salt is a seed crystal. In some embodiments, the fourth
solution forms a slurry upon stirring. The slurry may be washed
with a solvent or a combination of solvents, which may be the same
or different from the first solvent or combination of solvent, or
the second solvent or combination of solvents. The slurry may be
dried.
[0045] In another embodiment, the invention relates to a pure
composition comprising Compound 1, where in the composition is
about 90-100%, preferably 95-100%, more preferably 98-100%
(wt./wt.) or 99-100% (wt./wt.) pure; e.g., less than about 10%,
less than about 5%, less than about 2% or less than about 1%
impurity is present. Such impurities include, e.g., degradation
products, oxidized products, epimers, solvents, and/or other
undesirable impurities.
[0046] In yet another embodiment, the invention includes a method
for treating a tetracycline responsive state in a subject, by
administering to the subject an effective amount of a crystalline
form of Compound 1. For example, the subject is a human
subject.
[0047] In yet another embodiment, the invention includes a method
for treating a tetracycline responsive state in a subject, by
administering to the subject an effective amount of a stable salt
of Compound 1. For example, the stable salt is a tosylate, an HCl,
or a mesylate salt of Compound 1.
[0048] In yet another embodiment, the invention includes a method
for treating a tetracycline responsive state in a subject, by
administering to the subject an effective amount of a polymorph of
Compound 1.
[0049] In yet another embodiment, the invention includes a method
for treating a tetracycline responsive state in a subject, by
administering to the subject an effective amount of a polymorph of
salt of Compound 1. For example, the polymorph can be a polymorph
of a tosylate, an HCl, or a mesylate salt of Compound 1.
[0050] In yet another embodiment, the invention includes a method
for treating a tetracycline responsive state in a subject, by
administering to the subject an effective amount of a polymorph of
tosylate salt of Compound 1. For example, the polymorph of tosylate
can be a form 1, a form 2, or a form 3 polymorph of tosylate salt
of Compound 1.
[0051] For example, the tetracycline responsive state is a
bacterial infection. The bacterial infection can be associated with
gram positive bacteria, or gram negative bacteria. In some
embodiments, the bacterial infection is associated with E. coli, S.
aureus, or E. faecalis.
[0052] In some embodiments, the bacterial infection is resistant to
other tetracycline antibiotics, which include, but are not limited
to, tetracycline, minocycline, doxycycline, sancycline,
chlortetracycline, demeclocyclin, oxytetracycline, chelocardin,
rolitetracycline, lymecycline, methacycline, apicycline,
clomocycline, pipacycline, mepylcycline, meglucycline,
guamecycline, penimocycline, and etamocycline.
BRIEF DESCRIPTION OF THE DRAWINGS
[0053] FIG. 1 provides an X-ray powder diffraction pattern of a
sample comprising crystalline Compound 1 at 25.degree. C.;
[0054] FIG. 2 provides an X-ray powder diffraction pattern at
25.degree. C. of the starting material (E00285), form 1 tosylate
salt, form 2 tosylate salt, form 3 tosylate salt, and the amorphous
tosylate salt form of Compound 1;
[0055] FIG. 3 provides a comparison of X-ray powder diffraction
patterns at 25.degree. C. for crystalline Compound 1 (E00285) and a
sample comprising form 1 tosylate salt, obtained from
recrystallization of amorphous tosylate salt of Compound 1 in
IPA;
[0056] FIG. 4 provides a comparison of X-ray powder diffraction
patterns at 25.degree. C. for samples comprising form 1 tosylate
salt, form 2 tosylate salt, and form 3 tosylate salt that were
dried overnight in a vacuum;
[0057] FIG. 5 provides a variable temperature X-ray powder
diffraction analysis of samples comprising form 2 tosylate
salt;
[0058] FIG. 6 provides a variable temperature X-ray powder
diffraction analysis of samples comprising form 3 tosylate
salt;
[0059] FIG. 7 provides a variable temperature X-ray powder
diffraction analysis of samples comprising a slurry of a 50:50
mixture of form 1 tosylate salt and form 3 tosylate salt in
IPA;
[0060] FIG. 8 provides a high resolution X-ray powder diffraction
pattern of a sample comprising form 1 tosylate salt of Compound
1;
[0061] FIG. 9 provides a high resolution X-ray powder diffraction
pattern of a sample comprising form 2 tosylate salt of Compound 1
(93.2% HPLC purity);
[0062] FIG. 10 provides a high resolution X-ray powder diffraction
pattern of a sample comprising form 3 tosylate salt of Compound 1
(96.7% HPLC purity).
[0063] FIG. 11 provides a summary of the polymorphism of the
tosylate salt of Compound 1.
DETAILED DESCRIPTION OF THE INVENTION
[0064] Tetracycline-type antibiotic compounds have long been known
to have limited stability in the solid phase freebase form. One
such non-crystalline tetracycline analog compound,
(4S,4AS,5AR,12AS)-4-7-Bis(dimethylamino)-9-{[(2,2-dimethylpropyl)amino]me-
thyl}-3,10,12,12A-tetrahydroxy-1,11-dioxo-1,4,4A,5,5A,6,11,12A-octahydrote-
tracene-2-carboxamide (Compound 1; MW=556.66,
MF=C.sub.29H.sub.40N.sub.4O.sub.7), has limited stability in the
solid phase upon exposure to air, light and/or moisture.
##STR00002##
(4S,4AS,5AR,12AS)-4-7-Bis(dimethylamino)-9{[(2,2-dimethylpropyl)amino]met-
hyl}-3,10,12,12A-tetrahydroxy-1,11-dioxo-1,4,4A,5,5A,6,11,12A-octahydrotet-
racene-2-carboxamide
[0065] Specifically, Compound 1 is a yellow amorphous solid that is
unstable at temperatures higher than 0.degree. C. and when exposed
to air. Compound 1 must be stored at temperatures below 0.degree.
C. with limited exposure in the solid phase to air, light and
moisture. Outside of these limited exposure conditions, Compound 1
degrades to produce degradation products including air degradation
products 2, 3 and 4, as well as the 4-epi-isomer 5.
##STR00003##
[0066] Prior to this disclosure, no stable crystalline forms or
stable crystalline acid salts of Compound 1 were known.
[0067] The present invention relates to crystalline Compound 1,
salt forms of Compound 1, polymorphic forms of Compound 1 or
polymorphic forms of salts of Compound 1; pharmaceutical
compositions comprising the crystalline forms, salt forms,
polymorphic forms, or polymorphic forms of salts of Compound 1;
methods of making the crystalline forms, salt forms, polymorphic
forms, or polymorphic forms of salts of Compound 1; and methods of
their use for the treatment of tetracycline-responsive states.
1. SOLID FORM COMPOUNDS
[0068] Compound 1 is a tetracycline compound. The term
"tetracycline compound" includes many compounds with a similar ring
structure to tetracycline. Examples of tetracycline compounds
include: tetracycline, chlortetracycline, oxytetracycline,
demeclocycline, methacycline, sancycline, doxycycline, and
minocycline.
[0069] The free base and certain pharmaceutically acceptable salts
of Compound 1 are described in U.S. application Ser. No.
10/786,881, corresponding to U.S. Publication. No. 2005/0026876 A1.
There is no teaching or suggestion of crystalline forms of Compound
1, or that any of the described salt forms are superior to the
others, as judged by the list of properties described above.
[0070] Thus, the present invention addresses the need for improved
tetracycline compounds and the need for improved solid state forms
of tetracycline compounds for manufacturing and
bioavailability.
[0071] The solid state form of the tetracycline compound, Compound
1, can be a crystalline form. The crystalline form of the compound
can be a free base. Crystalline forms of different salts of the
free base compound can be formed. Examples of acids which can be
used to convert the free base to a salt include, but are not
limited to, HCl, p-toluenesulfonic acid, trifluoroacetic acid,
methylsulfonic acid, benzenelsulfonic acid, and acetic acid.
[0072] The neutral forms of the compounds may be regenerated by
contacting the salt with a base or acid and isolating the parent
compound in the conventional manner. The parent form of the
compound may differ from the various salt forms in certain physical
properties, such as solubility in polar solvents.
[0073] As described herein, a process through which different
crystalline forms of Compound 1 can be generated has been
developed. More specifically, the inventors have shown that the
crystalline form obtained mainly depends on the nature of the
solvent used in the process. For the purposes of this description
the term "crystalline form" refers to either a polymorphic form or
a non-amorphous form, without distinction. "Polymorphic form"
refers to an organized structure involving only molecules of the
solute and having a characteristic crystalline signature.
[0074] The terms "polymorphs" and "polymorphic forms" and related
terms herein refer to crystalline forms of the same molecule, and
different polymorphs may have different physical properties such
as, for example, melting temperatures, heats of fusion,
solubilities, dissolution rates and/or vibrational spectra as a
result of the arrangement or conformation of the molecules in the
crystal lattice. The differences in physical properties exhibited
by polymorphs affect pharmaceutical parameters such as storage
stability, compressibility and density (important in formulation
and product manufacturing), and dissolution rates (an important
factor in bioavailability). Differences in stability can also
result from changes in chemical reactivity (e.g., differential
oxidation, such that a dosage form discolors more rapidly when
comprised of one polymorph than when comprised of another
polymorph) or mechanical property (e.g., tablets crumble on storage
as a kinetically favored polymorph converts to thermodynamically
more stable polymorph) or both (e.g., tablets of one polymorph are
more susceptible to breakdown at high humidity). As a result of
solubility/dissolution differences, in the extreme case, some
polymorphic transitions may result in lack of potency or, at the
other extreme, toxicity. In addition, the physical properties of
the crystal may be important in processing, for example, one
polymorph might be more likely to form solvates or might be
difficult to filter and wash free of impurities (i.e., particle
shape and size distribution might be different between
polymorphs).
[0075] Polymorphs of a molecule can be obtained by a number of
methods, as known in the art. Such methods include, but are not
limited to, melt recrystallization, melt cooling, solvent
recrystallization, de solvation, rapid evaporation, rapid cooling,
slow cooling, vapor diffusion and sublimation.
[0076] Techniques for characterizing polymorphs include, but are
not limited to, differential scanning calorimetry (DSC), X-ray
powder diffractometry (XRPD), single crystal X-ray diffractometry,
vibrational spectroscopy, e.g., IR and Raman spectroscopy, solid
state NMR, hot stage optical microscopy, scanning electron
microscopy (SEM), electron crystallography and quantitative
analysis, particle size analysis (PSA), surface area analysis,
solubility studies and dissolution studies.
[0077] The term, "solvate", as used herein, refers to a crystal
form of a substance which contains solvent. The term "hydrate"
refers to a solvate wherein the solvent is water.
[0078] A desolvated solvate is a crystal form of a substance which
can only be made by removing the solvent from a solvate.
[0079] The term, "amorphous form", as used herein, refers to a
noncrystalline form of a substance.
[0080] As used herein, the term "pure" means about 90-100%,
preferably 95-100%, more preferably 98-100% (wt./wt.) or 99-100%
(wt./wt.) pure compound; e.g. less than about 10%, less than about
5%, less than about 2% or less than about 1% impurity is present.
Such impurities include, e.g., degradation products, oxidized
products, epimers, solvents, and/or other undesirable
impurities.
[0081] As used herein, a compound is "stable" where significant
amounts of degradation products are not observed under constant
conditions of humidity, light exposure and at temperatures higher
than 0.degree. C. over a period of four weeks. A compound is not
considered to be stable at a certain condition when degradation
impurities appear or an area percentage of existing impurities
begins to grow. The amount of degradation growth as a function of
time is important in determining compound stability.
[0082] All ranges set forth herein are intended to encompass the
indicated endpoints of the range as well as all included values and
ranges, including those not specifically set forth.
[0083] The present invention is directed to crystalline forms, salt
forms and polymorphs of Compound 1; compositions comprising the
crystalline forms, salts and polymorphs alone or in combination
with other active ingredients; methods of preparing the
crystalline, salts and polymorphs; and methods of their use in the
modulation of tetracycline compound receptive states. While not
intending to be bound by any particular theory of operation, the
storage stability, compressibility, density or dissolution
properties of the crystalline forms, salts and polymorphs are
beneficial for manufacturing, formulation and bio-availability of
the tetracycline compound.
[0084] Preferred salts and polymorphs of the invention are those
that are characterized by physical properties, e.g., stability,
solubility, hygroscopicity and dissolution rate, appropriate for
clinical and therapeutic dosage forms. Preferred polymorphs of the
invention are those that are characterized by physical properties,
e.g., crystal morphology, compressibility and hardness, suitable
for manufacture of a solid dosage form. Such properties can be
determined using techniques such as X-ray diffraction, microscopy,
IR spectroscopy, thermal analysis and hygroscopicity analysis, as
described herein and known in the art.
1.1 Salts of Compound 1
[0085] In one aspect, the present invention provides crystalline
forms of particular pharmaceutically acceptable salts of Compound
1. This aspect of the invention provides crystalline forms of HCl,
mesylate and tosylate salts of Compound 1:
##STR00004##
(4S,4AS,5AR,12AS)-4-7-Bis(dimethylamino)-9{[(2,2-dimethylpropyl)amino]met-
hyl}-3,10,12,12A-tetrahydroxy-1,11-dioxo-1,4,4A,5,5A,6,11,12A-octahydrotet-
racene-2-carboxamide
[0086] Each salt of the invention can be made from a preparation of
Compound 1. Compound 1 can be synthesized or obtained according to
any method apparent to those of skill in the art. In preferred
embodiments, Compound 1 is prepared according to the methods
described in detail in the examples below. See, e.g., U.S.
Publication No. 2005/0026876 A1, the contents of which are hereby
incorporated by reference in their entirety.
[0087] Alternatively, Compound 1 can be prepared by isolating a
particular salt of Compound 1 and converting such a salt of
Compound 1 to the neutral form by treatment with an appropriate
base. For example, Compound 1 can be prepared by isolating the
hydrochloride salt of Compound 1 by filtration, then converting it
to the neutral form by treatment with monobasic sodium carbonate in
ethyl acetate, or other suitable base.
[0088] Compound 1 prepared by any method can be contacted with an
appropriate acid, either neat (i.e., free from admixture or
dilution) or in a suitable inert solvent or solvents, to yield the
salt forms of the invention. For example, Compound 1 can be
contacted with a p-toluenesulfonic acid to yield the tosylate salt
forms of the invention.
[0089] Stability studies were performed on the free base Compound I
and an amorphous diHCl salt of Compound 1. This salt was formed by
dissolving the compound in aqueous solution, adjusting the pH of
the solution to approximately 4.2, followed by lyophilization. The
free base degraded in less than one month at 40.degree. C., and
approximately three months at 4.degree. C. In contrast, the diHCl
salt of Compound 1 was stable for 6 months at 40.degree. C., and
for two years at room temperature (25.degree. C.).
[0090] As shown in detail in the examples below, the tosylate salt
of Compound 1, and polymorphs thereof, display desirable
properties.
1.2 Polymorphs of Compound 1
[0091] The present invention also provides polymorphs of Compound
1. In certain embodiments, the polymorphs of the invention are
polymorphs of the tosylate salt of Compound 1.
[0092] Each polymorph of the invention can be made from a
preparation of Compound 1. Solid Compound 1 can be dissolved and
then crystallized from the solvent mixtures described below to
yield the polymorphic forms of the invention. In particular
embodiments of the invention, a tosylate salt of Compound 1 can be
dissolved and then crystallized from the solvent mixtures described
below to yield certain polymorphic forms of the invention. In some
embodiments of the invention, free base of Compound 1 can be
dissolved and then acid is added to form a crystalline salt of
Compound 1.
[0093] In one embodiment, the present invention provides a
polymorph of a tosylate salt of Compound 1.
[0094] In a further embodiment, the invention provides a form 1
polymorph of the tosylate salt of Compound 1, having an X-ray
powder diffraction pattern similar to that of FIG. 8, the
characteristics of the diffraction pattern all shown in Table 1.
For example, a particular form 1 polymorph of the invention has
X-ray powder diffraction pattern peaks at 5.60, 8.06, 8.57, 11.41,
13.02, 15.58, 18.83, 20.54 and 24.53 .degree. 2.theta. using Cu
K.alpha. radiation. For example, a particular form 1 polymorph of
the invention has X-ray powder diffraction pattern peaks at 8.06,
11.41, 13.02, 18.83, 20.54 and 24.53 .degree. 2.theta. using Cu
K.alpha. radiation. For example, a particular form 1 polymorph of
the invention has X-ray powder diffraction pattern peaks at 8.06,
13.02, 18.83 and 24.53 .degree. 2.theta.. For example, a particular
form 1 polymorph of the invention has major X-ray powder
diffraction pattern peaks at 8.06 and 18.83 .degree. 2.theta..
TABLE-US-00001 TABLE 1 Angle (2-Theta .degree.) D value (Angstrom)
Intensity (Counts) Intensity (%) 5.60 15.78 347 15.9 8.06 10.97
2184 100.0 8.57 10.30 581 26.6 9.80 9.01 308 14.1 10.89 8.12 233
10.7 11.41 7.75 667 30.5 13.02 6.79 626 28.7 13.78 6.42 261 12.0
14.92 5.93 252 11.5 15.58 5.68 346 15.8 16.10 5.50 262 12.0 17.07
5.19 345 15.8 18.83 4.71 979 44.8 20.54 4.32 838 38.4 21.83 4.07
489 22.4 23.00 3.86 395 18.1 24.53 3.63 661 30.3 25.10 3.55 341
15.6 27.82 3.20 404 18.5 28.48 3.13 357 16.3 30.26 2.95 302 13.8
34.82 2.57 236 10.8 36.19 2.48 254 11.6 37.54 2.39 247 11.3 40.49
2.23 368 16.8
[0095] In another embodiment, the present invention provides form 2
of the tosylate salt of Compound 1. In one embodiment, the form 2
polymorph of the tosylate salt of Compound 1 has an X-ray powder
diffraction pattern similar to that of FIG. 9, the characteristics
of the diffraction pattern all shown in Table 2. For example, a
particular form 2 polymorph of the invention has X-ray powder
diffraction pattern peaks at 7.82, 11.88, 12.68, 16.12, 18.63,
21.46 and 23.74 .degree. 2.theta. using Cu K.alpha. radiation. For
example, a particular form 2 polymorph of the invention has major
X-ray powder diffraction pattern peaks at 7.82, 11.88, 16.12 and
21.46 .degree. 2.theta.. For example, a particular form 2 polymorph
of the invention has X-ray powder diffraction pattern peaks at
11.88 and 16.12 .degree. 2.theta..
TABLE-US-00002 TABLE 2 Angle (2-Theta .degree.) D value (Angstrom)
Intensity (Counts) Intensity (%) 5.317 16.62224 135 10.6 6.272
14.09246 275 21.5 6.511 13.57561 274 21.5 7.108 12.43554 137 10.8
7.821 11.30413 827 64.8 9.712 9.10741 173 13.5 10.783 8.20461 340
26.6 11.875 7.4528 1258 98.6 12.682 6.97995 904 70.8 13.182 6.7162
611 47.9 13.985 6.33261 299 23.4 15.578 5.68838 512 40.1 16.122
5.49766 1100 86.2 16.635 5.32915 467 36.6 17.397 5.09763 697 54.7
18.63 4.76275 967 75.8 20.235 4.38856 647 50.7 20.666 4.298 636
49.8 21.456 4.14147 1276 100 22.51 3.9499 747 58.6 23.744 3.7473
1076 84.3 24.448 3.64103 749 58.7 25.651 3.47293 652 51.1 26.303
3.38824 638 50 27.225 3.27554 678 53.2 27.4 3.25505 747 58.5 27.823
3.20648 763 59.8 28.193 3.16532 588 46.1
[0096] In yet another embodiment, the present invention provides
form 3 of Compound 1. In further embodiments, the form 3 polymorph
of the tosylate salt of Compound 1 has an X-ray powder diffraction
pattern similar to that of FIG. 10, the characteristics of the
diffraction pattern all shown in Table 3. For example, a particular
form 3 polymorph of the invention has X-ray powder diffraction
pattern peaks at 5.11, 8.89, 10.34, 11.76, 13.70, 14.81 and 15.60
.degree. 2.theta. using Cu K.alpha. radiation. For example, a
particular form 3 polymorph of the invention has major X-ray powder
diffraction pattern peaks at 5.11, 8.89, 10.34, 11.76 and 15.60
.degree. 2.theta.. For example, a particular form 3 polymorph of
the invention has major X-ray powder diffraction pattern peaks at
5.11 and 15.60 .degree. 2.theta..
TABLE-US-00003 TABLE 3 Angle (2-Theta .degree.) D value (Angstrom)
Intensity (Counts) Intensity (%) 5.11 17.29 1184 66.4 8.89 9.95 475
26.6 10.34 8.56 431 24.2 11.76 7.53 404 22.7 13.70 6.46 524 29.4
14.81 5.98 552 31 15.60 5.68 1783 100 17.23 5.15 661 37.1 17.93
4.95 1014 56.9 18.35 4.83 832 46.6 18.74 4.74 914 51.3 19.00 4.67
874 49 20.15 4.41 889 49.9 20.36 4.36 913 51.2 20.65 4.30 940 52.7
21.65 4.10 681 38.2 22.59 3.94 923 51.8 23.25 3.83 1206 67.7 23.71
3.75 872 48.9 24.94 3.57 718 40.3 25.43 3.50 551 30.9 26.12 3.41
745 41.8 26.64 3.35 709 39.8 27.15 3.28 689 38.7 27.55 3.24 754
42.3
[0097] The tosylate salt of Compound 1 crystallized as very small,
irregular particles, typically 5-8 microns in size. FIG. 1 depicts
X-ray powder diffraction (XRPD) of a crystalline solid of the
tosylate salt of Compound 1. This compound was shown to melt at
190.degree. C., followed by decomposition.
[0098] Gravimetric vapor sorption was performed on Compound 1 or
its tosylate salt. It was determined that there were 2.5 molecules
of water per molecule of Compound 1. XRPD was performed to compare
the starting material (E00285) with the dehydrated material. The
data indicated no change in form.
2. SYNTHESIS OF COMPOUND 1
[0099] 9-(aminomethyl)-minocycline dihydrochloride (200 mg, 1 eq.),
DMF and trimethylacetaldehyde (45 .mu.l, 1 eq.) were combined in 40
mL flasks and stirred. Triethylamine (150 .mu.L, 3 eq.) was then
added. After stirring at room temperature for several minutes,
NaBH(OAc).sub.3 (175 mg, 2 eq.) and InCl.sub.3 (9 mg, 0.1 eq.) was
added. After one hour, the reaction was clear and red. Liquid
chromatography showed a single product for the reaction. The
reaction was quenched with methanol, the solvent was removed, and
the product was purified using column chromatography.
Purification
[0100] Compound 1 was purified by chromatography by injecting an
aqueous low pH solution of the compound into an HPLC in a polar
organic solvent gradient, and combining the product fractions, such
that the compound was purified. Selection of suitable acidic mobile
phases enhanced process stability and selectivity. Organic and
mineral acid mobile phases were effective at separating by-products
including epimer impurities and closely-eluting by products through
pH control or choice of acid. Acidic mobile phases also protected
against oxidative degradation of the compound.
[0101] For example, the low pH solution had a pH of about 2-3.
Examples of solutions that were used include 0.1% aqueous solutions
of methane sulfonic acid and 0.1% aqueous solutions of
trifluoroacetic acid. In certain embodiments, an isocratic gradient
of 94% of the aqueous solution and 6% acetonitrile or another polar
organic solvent were used to purify the compound from epimeric and
closely eluting by-products.
[0102] The resulting aqueous product fractions may be combined and
the pH may be adjusted to about 4.0-4.5 using a base (e.g., NaOH).
Hydrophobic impurities and oxidative degradents of the compound may
be removed by washing the aqueous solution with a non-polar organic
solvent (e.g., CH.sub.2Cl.sub.2). The organic layers were discarded
and the aqueous layers were combined and retained.
[0103] It should be noted that the organic solvents, such as
methylene chloride, can be used to selectively remove late-eluting
hydrophobic impurities such as 4-carbonyl by products and other
oxidative degradents from the acidic aqueous solution of the
compound.
[0104] The pH of the combined aqueous layers may then be adjusted
to neutral pH, e.g., about 7.5 to about 8.5. The pH may be adjusted
by the addition of a base, such as NaOH. The neutral solution was
then washed with a non-polar organic solvent, such as methylene
chloride. It should be noted that selective pH adjustment to
neutral pH ranges also allowed the compound to be extracted into
the organic solvent while retaining undesired .beta.-epimer and by
products in the aqueous phase.
[0105] In addition, antioxidants may also be added to the aqueous
solutions of compounds described herein. The antioxidants may be
provided to prevent oxidative degradation of the compounds.
Antioxidants such as ammonium sulfites or bisulfites can be
used.
3. METHODS OF PREPARING POLYMORPHIC FORMS OF COMPOUND 1
[0106] The invention also pertains to methods of preparing
polymorphic forms of crystalline Compound 1.
[0107] In one embodiment, form 1 of the tosylate salt of Compound 1
can be made by any method of making form 1 apparent to those of
skill in the art based upon the teachings herein. In certain
embodiments, form 1 can be formed from maturation of the amorphous
tosylate salt of Compound 1 in isopropanol, acetone, ethyl acetate,
methyl pentanone, toluene or acetonitrile solution. Form 1 can also
be obtained from recrystallization of the amorphous tosylate salt
slurried in isopropanol. Form 1 can also be obtained by dissolving
the freebase in an appropriate solvent or combination of solvents
such as two alcohols or an alcohol and an anti-solvent such as a
ketone, ether, ester, etc. After addition of the acid, the salt can
be crystallized slowly in the correct form.
[0108] Solvent system in which impurities and free base of Compound
1 are soluble while the stable crystalline salt of Compound 1 is
insoluble, e.g., crystalline slurry can be formed by precipitation,
can be selected.
[0109] In another embodiment, form 2 of the tosylate salt of
Compound 1 can be made by any method of making form 2 apparent to
those of skill in the art based upon the teachings herein. In
certain embodiments, form 2 can be formed from maturation of the
amorphous tosylate salt of Compound 1 in dichloromethane.
[0110] In another embodiment, form 3 of the tosylate salt of
Compound 1 can be made by any method of making form 3 apparent to
those of skill in the art based upon the teachings herein. In
certain embodiments, form 3 can be formed from maturation of the
amorphous tosylate salt of Compound 1 in methyl ethyl ketone, ethyl
acetate or methyl pentanone. Form 3 can also be obtained from
maturation of form 1 in methyl pentanone.
[0111] In a further embodiment, the polymorphic forms of tosylate
salt of Compound 1 described above may be produced by methods that
include steps of combining Compound 1 with a solvent to produce a
slurry, and adding p-toluenesulfonic acid.
[0112] Any suitable solvent may be used to create the slurry.
Solvents that may be used in embodiments include alcoholic
solvents, such as isopropanol. Any suitable combination of solvent
may be used to create the solution from which the salt
crystallizes. Solvent combinations that may be used in embodiments
include, but are not limited to, methanol and methyl-t-butyl ether
or ethanol and isopropanol.
[0113] For example, a slurry of Compound 1 in a solvent or a
combination of solvents may be produced at a temperature from about
0.degree. C. to about 60.degree. C., such as from about 15.degree.
C. to about 45.degree. C., or from about 20.degree. C. to about
25.degree. C. After it is produced, the slurry may optionally be
warmed and/or maintained at a temperature from about 15.degree. C.
to about 60.degree. C., such as from about 20.degree. C. to about
50.degree. C., or about 45.degree. C.
[0114] Once the slurry is created, the p-toluenesulfonic acid may
be added in an amount sufficient to produce a p-toluenesulfonic
acid salt of Compound 1. In one embodiment, the p-toluenesulfonic
acid is provided in an amount from about 25 to about 75 wt %, from
about 25 to about 50 wt %, from about 30 to about 40 wt %, or about
33 wt % relative to the amount of Compound 1. The p-toluenesulfonic
acid may be added in the form of a p-toluenesulfonic acid
monohydrate.
[0115] Polymorphic forms of the tosylate salt of Compound 1 can be
formed by a solution method. For example, a solution of Compound I
may be produced at a temperature from about 0.degree. C. to about
60.degree. C., such as from about 15.degree. C. to about 45.degree.
C., or from about 20.degree. C. to about 25.degree. C. After it is
produced, the solution may optionally be warmed and/or maintained
at a temperature from about 15.degree. C. to about 60.degree. C.,
such as from about 20.degree. C. to about 50.degree. C., or about
45.degree. C.
[0116] Once the solution is created, the p-toluenesulfonic acid may
be added in an amount sufficient to produce a p-toluenesulfonic
acid salt of Compound, 1. In one embodiment, the p-toluenesulfonic
acid is provided in an amount from about 25 to about 75 wt %, from
about 30 to about 50 wt %, from about 35 to about 45 wt %, or about
40 wt % relative to the amount of Compound 1. The p-toluenesulfonic
acid may be added in the form of a p-toluenesulfonic acid
monohydrate.
[0117] In one embodiment, form 1 polymorph used to seed the
solution may be added. Any suitable solvent may be used to form the
p-toluenesulfonic acid solution. Suitable solvents include
alcoholic solvents, such as isopropanol or solvent combinations
such as methanol and methyl-t-butyl ether. In a preferred
embodiment, the vol./vol. ratio of methanol to methyl-t-butyl ether
is 1:1.2. Suitable solvents include a combination of two or more
alcoholic solvents, such as a combination of ethanol and
isopropanol. In a preferred embodiment, the vol./vol. ratio of
ethanol to isopropanol is 2:1. In particular embodiments, the
p-toluenesulfonic acid solution includes the same solvent used to
create the slurry or the solution of Compound 1.
[0118] After addition of p-toluenesulfonic acid in the appropriate
solvent, a slurry of the form 1 polymorph of the tosylate salt of
Compound is formed. The water content of the supernatant of the
slurry may be adjusted to a suitable level following the addition
of the p-toluenesulfonic acid. Typically, the water content of the
slurry supernatant may be in a range from about 0.2 to about 1.0
mg/mL, such as from about 0.4 to about 0.8 mg/mL, e.g., about 0.6
mg/mL, about 0.54 mg/mL, etc.
[0119] Following the addition of the p-toluenesulfonic acid, the
slurry or the solution may be stirred to produce a crystalline
slurry. Stirring may be conducted for more than 48 hours. However,
stirring is typically conducted for a period of from about 5 to
about 36 hours, such as from about 10 to about 24 hours or about 18
hours.
[0120] Stirring may be conducted at any temperature suitable for
producing the crystalline slurry. For example, the slurry may be
stirred at a temperature from about 0.degree. C. to about
60.degree. C., such as from about 15.degree. C. to about 45.degree.
C. or from about 20.degree. C. to about 25.degree. C.
[0121] After crystal formation, the crystalline slurry may be
filtered to remove the supernatant, and the crystals may be washed
with any suitable solvent. In embodiments, the crystals may be
washed one to four times, and the solvent may be any solvent
suitable for the preparation of the crystalline slurry. In
particular, the solvent used to wash the crystals may be the same
solvent or solvents used to form the original slurry or solution,
or the p-toluenesulfonic acid solution.
[0122] The crystals produced may then be dried to remove excess
solvent by any suitable method. For example, drying may be
accomplished by one or more methods including but not limited to
elevated temperatures in a range from about 0.degree. C. to about
60.degree. C., such as from about 15.degree. C. to about 45.degree.
C.; blowing dry nitrogen over the crystals; and blowing humidified
nitrogen over the crystals.
[0123] Maturation studies were performed in which a sample of the
tosylate salt of Compound 1 was slurried in different solvents,
filtered, and the wet solid was analyzed by XRPD. Three polymorphic
forms of the tosylate salt of Compound 1 were observed. FIG. 2
depicts the XRPD spectra of the starting material (E00285), form 1
tosylate salt, form 2 tosylate salt, form 3 tosylate salt, and the
amorphous form of Compound 1.
[0124] Table 4 lists the solvents used for maturation
experiments.
TABLE-US-00004 TABLE 4 Example No. Solvent Form 1 water Form 1 2
nitromethane Amorphous 3 anisole Form 2 4 2-propanol Form 1 5
methylethyl ketone Form 3 6 acetone Form 1 7 ethyl acetate Form 1 8
dioxane Amorphous 9 acetonitrile Form 1 10 toluene Form 1 11
dichloromethane Form 2 12 chloroform Amorphous 13 TBME Amorphous 14
isopropyl acetate Form 2 15 NMP dissolved 16 4-methyl-2-pentanone
Form 3 17 THF Gum 18 10% EtOAc Amorphous 19 10% water Amorphous 20
10% water/THF Amorphous 21 10% water/CAN Amorphous 22 10%
water/2-propanol Amorphous 23 10% water/acetone Amorphous 24 10%
water/dioxane Amorphous
[0125] Recrystallization of the amorphous material of Compound 1
was performed in various solvents. Only recrystallization in
2-propanol (isopropyl alcohol, IPA) gave form 1 tosylate salt, as
shown in Table 5. FIG. 3 compares XRPD spectra of reference
Compound 1 (E00285) and the recrystallized form 1 tosylate salt
from IP A.
TABLE-US-00005 TABLE 5 XRPD of Experiment precipitate No. Solvent
Volume At 50.degree. C. of solid 1 nitromethane 24 Soluble 2
anisole 200 Insoluble Amorphous 3 2-propanol 60 Soluble Form 1 4
methylethyl ketone 100 Soluble Amorphous 5 acetone 80 Soluble
Amorphous 6 ethyl acetate 200 Insoluble Amorphous 7 dioxane 120
Soluble Amorphous 8 acetonitrile 80 Soluble 9 toluene 200 Insoluble
Amorphous 10 dichloromethane 5 Soluble Amorphous 11 chloroform 5
Soluble Amorphous 12 TBME 200 Insoluble Amorphous 13 isopropyl
acetate 200 Insoluble Amorphous 14 4-methyl-2-pentanone 200
Insoluble Amorphous 15 THF 100 Soluble Amorphous 16 10% EtOAc/ 200
Insoluble Amorphous cyclohexane
[0126] A summary of the polymorphism of the tosylate salt of
Compound 1 is presented in FIG. 11.
[0127] After recrystallization, samples of form 1, form 2, and form
3 tosylate salt were dried overnight in a vacuum and analyzed by
XRPD, as shown in FIG. 4. There was no change in form after
drying.
[0128] Variable temperature XRPD was performed on form 2 and form 3
tosylate salt of Compound 1. See FIG. 5 and FIG. 6,
respectively.
[0129] The relative stabilities of the polymorphic forms of
Compound 1 were analyzed. For example, form 1 tosylate salt was
subjected to a maturation experiment for 24 hours in either IPA or
methyl pentanone, seeded with either form 2 or form 3 tosylate
salt. During this experiment, there was no change of form 1 into
form 2 or 3. A slurry of a 50:50 mixture of form 1 and form 3 were
analyzed in IPA for 18 hours at 0.degree. C., 25.degree. C.,
40.degree. C., and 60.degree. C., as shown in FIG. 7. There was no
change of form 1 into form 3.
[0130] Recrystallization experiments indicated that form 1 tosylate
salt can be obtained reproducibly from the amorphous tosylate salt
by slurrying in IPA. Form 1 of Compound 1 can also be obtained
reproducibly by the addition of tosic acid. A high resolution XRPD
scan of form 1 is depicted in FIG. 8 and the characteristic of the
diffraction pattern are shown Table 1.
4. PHARMACEUTICAL COMPOSITIONS COMPRISING COMPOUNDS, SALTS,
CRYSTALLINE FORMS OR POLYMORPHS THEREOF OF THE INVENTION
[0131] In a further embodiment, the invention pertains to
pharmaceutical compositions comprising a tetracycline compound of
the invention (e.g., synthesized, or purified by the methods of the
invention) or a pharmaceutically acceptable salt, prodrug or ester
thereof. The pharmaceutical compositions may comprise a
pharmaceutically acceptable carrier.
[0132] The term "composition" as used herein is intended to
encompass a product comprising the specified ingredients (and in
the specified amounts, if indicated), as well as any product which
results, directly or indirectly, from combination of the specified
ingredients in the specified amounts. By "pharmaceutically
acceptable" it is meant the diluent, excipient or carrier must be
compatible with the other ingredients of the formulation and not
deleterious to the recipient thereof.
[0133] As set out above, certain embodiments of the present
compounds can contain a basic functional group, such as amino or
alkylamino, and are, thus, capable of forming pharmaceutically
acceptable salts with pharmaceutically acceptable acids. The term
"pharmaceutically acceptable salts" is recognized in the art and
includes relatively non-toxic, inorganic and organic acid addition
salts of compounds of the present invention. These salts can be
prepared in situ during the final isolation and purification of the
compounds of the invention, or by separately reacting a purified
compound of the invention in its free base form with a suitable
organic or inorganic acid, and isolating the salt thus formed.
Representative salts include the hydro bromide, hydrochloride,
sulfate, bisulfate, phosphate, nitrate, acetate, valerate, oleate,
palmitate, stearate, laurate, benzoate, lactate, phosphate,
tosylate, citrate, maleate, fumarate, succinate, tartrate,
napthylate, mesylate, glucoheptonate, lactobionate, and
laurylsulphonate salts, and the like. (See, e.g., Berge et at.
(1977) "Pharmaceutical Salts", J. Farm. SCI. 66:1-19).
[0134] In other cases, the compounds of the present invention may
contain one or more acidic functional groups and, thus, are capable
of forming pharmaceutically acceptable salts with pharmaceutically
acceptable bases. The term "pharmaceutically acceptable salts" in
these instances includes relatively non-toxic, inorganic and
organic base addition salts of compounds of the present invention.
These salts can likewise be prepared in situ during the final
isolation and purification of the compounds, or by separately
reacting the purified compound in its free acid form with a
suitable base, such as the hydroxide, carbonate or bicarbonate of a
pharmaceutically acceptable metal cation, with ammonia, or with a
pharmaceutically acceptable organic primary, secondary or tertiary
amine. Representative alkali or alkaline earth salts include the
lithium, sodium, potassium, calcium, magnesium, and aluminum salts
and the like. Representative organic amines useful for the
formation of base addition salts include ethylamine, diethylamine,
ethylenediamine, ethanolamine, diethanolamine, piperazine, and the
like.
[0135] The term "pharmaceutically acceptable esters" refers to the
relatively nontoxic, esterified products of the compounds of the
present invention. These esters can be prepared in situ during the
final isolation and purification of the compounds, or by separately
reacting the purified compound in its free acid form or hydroxyl
with a suitable esterifying agent. Carboxylic acids can be
converted into esters via treatment with an alcohol in the presence
of a catalyst. Hydroxyls can be converted into esters via treatment
with an esterifying agent such as alkanoyl halides. The term also
includes lower hydrocarbon groups capable of being solvated under
physiological conditions, e.g., alkyl esters, methyl, ethyl and
propyl esters. (See, for example, Berge et at., supra.).
[0136] The invention also pertains to tetracycline compounds, which
are synthesized and/or purified by the methods of the invention,
and pharmaceutically acceptable salts thereof.
[0137] The phrase "pharmaceutically acceptable carrier" is
recognized in the art and includes a pharmaceutically acceptable
material, composition or vehicle, suitable for administering
compounds of the present invention to mammals. The carriers include
liquid or solid filler, diluent, excipient, solvent or
encapsulating material, involved in carrying or transporting the
subject agent from one organ, or portion of the body, to another
organ, or portion of the body. Each carrier must be "acceptable" in
the sense of being compatible with the other ingredients of the
formulation and not injurious to the patient. Some examples of
materials which can serve as pharmaceutically acceptable carriers
include: sugars, such as lactose, glucose and sucrose; starches,
such as corn starch and potato starch; cellulose and its
derivatives, such as sodium carboxymethyl cellulose, ethyl
cellulose and cellulose acetate; powdered tragacanth; malt;
gelatin; talc; excipients, such as cocoa butter and suppository
waxes; oils, such as peanut oil, cottonseed oil, safflower oil,
sesame oil, olive oil, corn oil and soybean oil; glycols, such as
propylene glycol; polyols, such as glycerin, sorbitol, mannitol and
polyethylene glycol; esters, such as ethyl oleate and ethyllaurate;
agar; buffering agents, such as magnesium hydroxide and aluminum
hydroxide; alginic acid; pyrogen-free water; isotonic saline;
Ringer's solution; ethyl alcohol; phosphate buffer solutions; and
other nontoxic compatible substances employed in pharmaceutical
formulations.
[0138] Wetting agents, emulsifiers and lubricants, such as sodium
lauryl sulfate and magnesium stearate, as well as coloring agents,
releasing agents, coating agents, sweetening agents, flavoring and
perfuming agents, preservatives and antioxidants can also be
present in the compositions.
[0139] Examples of pharmaceutically acceptable antioxidants
include: water soluble antioxidants, such as ascorbic acid,
cysteine hydrochloride, sodium bisulfate, sodium metabisulfite,
sodium sulfite and the like; oil-soluble antioxidants, such as
ascorbyl palmitate, butylated hydroxyanisole (BRA), butylated
hydroxytoluene (BHT), lecithin, propyl gallate, .alpha.-tocopherol,
and the like; and metal chelating agents, such as citric acid,
ethylenediamine tetraacetic acid (EDTA), sorbitol, tartaric acid,
phosphoric acid, and the like.
[0140] Formulations of the present invention include those suitable
for oral, nasal, topical, transdermal, buccal, sublingual, rectal,
vaginal and/or parenteral administration. The formulations may
conveniently be presented in unit dosage form and may be prepared
by any methods well known in the art of pharmacy. The amount of
active ingredient which can be combined with a carrier material to
produce a single dosage form will generally be that amount of the
compound which produces a therapeutic effect. Generally, out of one
hundred percent, this amount will range from about 1 percent to
about ninety-nine percent of active ingredient, preferably from
about 5 percent to about 70 percent, most preferably from about 10
percent to about 30 percent.
[0141] Methods of preparing these formulations or compositions
include the step of bringing into association a compound of the
present invention with the carrier and, optionally, one or more
accessory ingredients. In general, the formulations are prepared by
uniformly and intimately bringing into association a compound of
the present invention with liquid carriers, or finely divided solid
carriers, or both, and then, if necessary, shaping the product.
[0142] Formulations of the invention suitable for oral
administration may be in the form of capsules, cachets, pills,
tablets, lozenges (using a flavored basis, usually sucrose and
acacia or tragacanth), powders, granules, or as a solution or a
suspension in an aqueous or non-aqueous liquid, or as an
oil-in-water or water-in-oil liquid emulsion, or as an elixir or
syrup, or as pastilles (using an inert base, such as gelatin and
glycerin, or sucrose and acacia) and/or as mouth washes and the
like, each containing a predetermined amount of a compound of the
present invention as an active ingredient. A compound of the
present invention may also be administered as a bolus, electuary or
paste.
[0143] In solid dosage forms of the invention for oral
administration (capsules, tablets, pills, dragees, powders,
granules and the like), the active ingredient is mixed with one or
more pharmaceutically acceptable carriers, such as sodium citrate
or dicalcium phosphate and/or any of the following: fillers or
extenders, such as starches, lactose, sucrose, glucose, mannitol
and/or silicic acid; binders, such as carboxymethylcellulose,
alginates, gelatin, polyvinyl pyrrolidone, sucrose and/or acacia;
humectants, such as glycerol; disintegrating agents, such as
agar-agar, calcium carbonate, potato or tapioca starch, alginic
acid, certain silicates, and sodium carbonate; solution retarding
agents, such as paraffin; absorption accelerators, such as
quaternary ammonium compounds; wetting agents, such as cetyl
alcohol and glycerol monostearate; absorbents, such as kaolin and
bentonite clay; lubricants, such as talc, calcium stearate,
magnesium stearate, solid polyethylene glycols, sodium lauryl
sulfate, and mixtures thereof; and coloring agents. In the case of
capsules, tablets and pills, the pharmaceutical compositions may
also comprise buffering agents. Solid compositions of a similar
type may also be employed as fillers in soft and hard-filled
gelatin capsules using such excipients as lactose or milk sugars,
as well as high molecular weight polyethylene glycols and the
like.
[0144] A tablet may be made by compression or molding, optionally
with one or more accessory ingredients. Compressed tablets may be
prepared using binder (for example, gelatin or hydroxypropylmethyl
cellulose), lubricant, inert diluent, preservative, disintegrant
(for example, sodium starch glycolate or cross-linked sodium
carboxymethyl cellulose), surface-active or dispersing agent.
Molded tablets may be made by molding in a suitable machine a
mixture of the powdered compound moistened with an inert liquid
diluent.
[0145] The tablets, and other solid dosage forms of the
pharmaceutical compositions of the present invention, such as
dragees, capsules, pills and granules, may optionally be scored or
prepared with coatings and shells, such as enteric coatings and
other coatings well known in the pharmaceutical-formulating art.
They may also be formulated so as to provide slow or controlled
release of the active ingredient therein using, for example,
hydroxypropylmethyl cellulose in varying proportions to provide the
desired release profile, other polymer matrices, liposomes and/or
microspheres. They may be sterilized by, for example, filtration
through a bacteria-retaining filter, or by incorporating
sterilizing agents in the form of sterile solid compositions which
can be dissolved in sterile water, or some other sterile injectable
medium immediately before use. These compositions may also
optionally contain opacifying agents and may be of a composition
that they release the active ingredient(s) only, or preferentially,
in a certain portion of the gastrointestinal tract, optionally, in
a delayed manner. Examples of embedding compositions which can be
used include polymeric substances and waxes. The active ingredient
can also be in microencapsulated form, if appropriate, with one or
more of the above-described excipients.
[0146] Liquid dosage forms for oral administration of the compounds
of the invention include pharmaceutically acceptable emulsions,
microemulsions, solutions, suspensions, syrups and elixirs. In
addition to the active ingredient, the liquid dosage forms may
contain inert diluent commonly used in the art, such as water or
other solvents, solubilizing agents and emulsifiers, such as ethyl
alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl
alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol,
oils (in particular, cottonseed, groundnut, corn, germ, olive,
castor and sesame oils), glycerol, tetrahydrofuryl alcohol,
polyethylene glycols and fatty acid esters of sorbitan, and
mixtures thereof.
[0147] Besides inert diluents, the oral compositions can also
include adjuvants such as wetting agents, emulsifying and
suspending agents, sweetening agents, flavoring agents, coloring
agents, perfuming and preservative agents.
[0148] Suspensions, in addition to the active compounds, may
contain suspending agents as, for example, ethoxylated isostearyl
alcohols, polyoxyethylene sorbitol and sorbitan esters,
microcrystalline cellulose, aluminum metahydroxide, bentonite,
agar-agar and tragacanth, and mixtures thereof.
[0149] Formulations of the pharmaceutical compositions of the
invention for rectal or vaginal administration may be presented as
a suppository, which may be prepared by mixing one or more
compounds of the invention with one or more suitable nonirritating
excipients or carriers comprising, for example, cocoa butter,
polyethylene glycol, a suppository wax or a salicylate, and which
is solid at room temperature, but liquid at body temperature and,
therefore, will melt in the rectum or vaginal cavity and release
the active compound.
[0150] Formulations of the present invention which are suitable for
vaginal administration also include pessaries, tampons, creams,
gels, pastes, foams or spray formulations containing such carriers
as are known in the art to be appropriate.
[0151] Dosage forms for the topical or transdermal administration
of a compound of this invention include powders, sprays, ointments,
pastes, creams, lotions, gels, solutions, patches and inhalants.
The active compound may be mixed under sterile conditions with a
pharmaceutically acceptable carrier, and with any preservatives,
buffers or propellants which may be required.
[0152] The ointments, pastes, creams and gels may contain, in
addition to an active compound of this invention, excipients, such
as animal and vegetable fats, oils, waxes, paraffins, starch,
tragacanth, cellulose derivatives, polyethylene glycols, silicones,
bentonites, silicic acid, talc and zinc oxide, or mixtures
thereof.
[0153] Powders and sprays can contain, in addition to a compound of
this invention, excipients such as lactose, talc, silicic acid,
aluminum hydroxide, calcium silicates and polyamide powder, or
mixtures of these substances. Sprays can additionally contain
customary propellants, such as chlorofluorohydrocarbons and
volatile unsubstituted hydrocarbons, such as butane and
propane.
[0154] Transdermal patches have the added advantage of providing
controlled delivery of a compound of the present invention to the
body. Such dosage forms can be made by dissolving or dispersing the
compound in the proper medium. Absorption enhancers can also be
used to increase the flux of the compound across the skin. The rate
of such flux can be controlled by either providing a rate
controlling membrane or dispersing the active compound in a polymer
matrix or gel.
[0155] Ophthalmic formulations, eye ointments, powders, solutions
and the like, are also contemplated as being within the scope of
this invention.
[0156] Pharmaceutical compositions of this invention suitable for
parenteral administration comprise one or more compounds of the
invention in combination with one or more pharmaceutically
acceptable sterile isotonic aqueous or nonaqueous solutions,
dispersions, suspensions or emulsions, or sterile powders which may
be reconstituted into sterile injectable solutions or dispersions
just prior to use, which may contain antioxidants, buffers,
bacteriostats, solutes which render the formulation isotonic with
the blood of the intended recipient or suspending or thickening
agents.
[0157] Examples of suitable aqueous and nonaqueous carriers which
may be employed in the pharmaceutical compositions of the invention
include water, ethanol, polyols (such as glycerol, propylene
glycol, polyethylene glycol, and the like), and suitable mixtures
thereof, vegetable oils, such as olive oil, and injectable organic
esters, such as ethyl oleate. Proper fluidity can be maintained,
for example, by the use of coating materials, such as lecithin, by
the maintenance of the required particle size in the case of
dispersions, and by the use of surfactants.
[0158] These compositions may also contain adjuvants such as
preservatives, wetting agents, emulsifying agents and dispersing
agents. Prevention of the action of microorganisms may be ensured
by the inclusion of various antibacterial and antifungal agents,
for example, paraben, chlorobutanol, phenol sorbic acid, and the
like. It may also be desirable to include isotonic agents, such as
sugars, sodium chloride, and the like into the compositions. In
addition, prolonged absorption of the injectable pharmaceutical
form may be brought about by the inclusion of agents which delay
absorption such as aluminum monostearate and gelatin.
[0159] In some cases, in order to prolong the effect of a drug, it
is desirable to slow the absorption of the drug from subcutaneous
or intramuscular injection. This may be accomplished by the use of
a liquid suspension of crystalline or amorphous material having
poor water solubility. The rate of absorption of the drug then
depends upon its rate of dissolution which, in turn, may depend
upon crystal size and crystalline form. Alternatively, delayed
absorption of a parenterally-administered drug form may be
accomplished by dissolving or suspending the drug in an oil
vehicle.
[0160] Injectable depot forms are made by forming microencapsule
matrices of the subject compounds in biodegradable polymers such as
polylactide-polyglycoJide. Depending on the ratio of drug to
polymer, and the nature of the particular polymer employed, the
rate of drug release can be controlled. Examples of other
biodegradable polymers include poly(orthoesters) and
poly(anhydrides). Depot injectable formulations are also prepared
by entrapping the drug in liposomes or microemulsions which are
compatible with body tissue.
[0161] The preparations of the present invention may be given
orally, parenterally, topically or rectally. They are given by
forms suitable for each administration route. For example, they are
administered in tablets or capsule form. For example, they are
administered by injection, infusion, inhalation, lotion, ointment,
suppository, etc. Oral administration is preferred.
[0162] The phrases "parenteral administration" and "administered
parenterally" as used herein means modes of administration other
than enteral and topical administration, usually by injection, and
includes, without limitation, intravenous, intramuscular,
intraarterial, intrathecal, intracapsular, intraorbital,
intracardiac, intradermal, intraperitoneal, transtracheal,
subcutaneous, subcuticular, intraarticular, subcapsular,
subarachnoid, intraspinal and intrasternal injection and
infusion.
[0163] The phrases "systemic administration", "administered
systemically", "peripheral administration" and "administered
peripherally" as used herein mean the administration of a compound,
drug or other material other than directly into the central nervous
system, such that it enters the patient's system and, thus, is
subject to metabolism and other like processes, for example,
subcutaneous administration.
[0164] These compounds may be administered to humans and other
animals for therapy by any suitable route of administration,
including orally, nasally (as by, for example, a spray), rectally,
intravaginally, parenterally, intracisternally and topically (as by
powders, ointments or drops, including buccally and
sublingually).
[0165] Regardless of the route of administration selected, the
compounds of the present invention, which may be used in a suitable
hydrated form, and/or the pharmaceutical compositions of the
present invention, are formulated into pharmaceutically acceptable
dosage forms by conventional methods known to those of skill in the
art.
[0166] The term "therapeutically effective amount" refers to the
amount of the subject salt or polymorph that will elicit the
biological or medical response of a tissue, system, animal or human
that is being sought by the researcher, veterinarian, medical
doctor or other clinician or that is sufficient to prevent
development of or alleviate to some extent one or more of the
symptoms of the disease being treated.
[0167] Actual dosage levels of the active ingredients in the
pharmaceutical compositions of this invention may be varied so as
to obtain an amount of the active ingredient which is effective to
achieve the desired therapeutic response for a particular patient,
composition, and mode of administration, without being toxic to the
patient.
[0168] The selected dosage level will depend upon a variety of
factors including the activity of the particular compound of the
present invention employed, or the ester, salt or amide thereof,
the route of administration, the time of administration, the rate
of excretion of the particular compound being employed, the
duration of the treatment, other drugs, compounds and/or materials
used in combination with the particular compound employed, the age,
sex, weight, condition, general health and prior medical history of
the patient being treated, and like factors well known in the
medical arts.
[0169] A physician or veterinarian having ordinary skill in the art
can readily determine and prescribe the effective amount of the
pharmaceutical composition required. For example, the physician or
veterinarian could start doses of the compounds of the invention
employed in the pharmaceutical composition at levels lower than
that required in order to achieve the desired therapeutic effect
and gradually increase the dosage until the desired effect is
achieved.
[0170] In general, a suitable daily dose of a compound of the
invention will be that amount of the compound which is the lowest
dose effective to produce a therapeutic effect. Such an effective
dose will generally depend upon the factors described above.
Generally, intravenous and subcutaneous doses of the compounds of
this invention for a patient, when used for the indicated analgesic
effects, will range from about 0.0001 to about 100 mg per kilogram
of body weight per day, more preferably from about 0.01 to about 50
mg per kg per day, and still more preferably from about 0.1 to
about 10 mg per kg per day. For example, in some embodiments the
doses are between 0.5 and 4.0 mg per kg day. If desired, the
effective daily dose of the active compound may be administered as
one, two, three, four, five, six or more sub-doses administered
separately at appropriate intervals throughout the day or week or
other suitable time period, optionally, in unit dosage forms.
[0171] While it is possible for a compound of the present invention
to be administered alone, it is preferable to administer the
compound as a pharmaceutical composition.
5. METHODS OF USING THE TETRACYCLINE COMPOUNDS OF THE INVENTION
[0172] The invention also pertains to a method for treating a
tetracycline responsive state in a subject, by administering to the
subject an effective amount of a composition comprising a Compound
1 according to the invention or a pharmaceutically acceptable salt
thereof, such that the state is treated.
[0173] The terms "treat", "treating" or "treatment", as used
herein, refer to a method of alleviating or abrogating a disease or
disorder (e.g., the tetracycline compound responsive state) and/or
its attendant symptoms. The terms "prevent", "preventing" or
"prevention", as used herein, refer to a method of barring a
subject from acquiring a disease or disorder. The "subject", as
used herein, includes a mammal. The mammal can be e.g., any mammal,
e.g., a human, primate, mouse, rat, dog, cat, cow, horse, goat,
camel, sheep or a pig. Preferably, the mammal is a human.
[0174] The language "tetracycline compound responsive state" or
"tetracycline responsive state" includes states which can be
treated, prevented, or otherwise ameliorated by the administration
of a tetracycline compound of the invention. Tetracycline compound
responsive states include bacterial, viral, and fungal infections
(including those which are resistant to other tetracycline
compounds), cancers (e.g., prostate cancer, breast cancer, colon
cancer, lung cancer, melanoma, lymph cancers and other disorders
characterized by unwanted cellular proliferation, including, but
not limited to, those described in U.S. Pat. No. 6,100,248),
arthritis, osteoporosis, diabetes, and other states for which
tetracycline compounds have been found to be active (see, for
example, U.S. Pat. Nos. 5,789,395; 5,834,450; 6,277,061 and
5,532,227, each of which is expressly incorporated herein by
reference). Compounds of the invention can be used to prevent or
control important mammalian and veterinary diseases such as
diarrhea, urinary tract infections, infections of skin and skin
structure, ear, nose and throat infections, wound infection,
mastitis and the like. In addition, methods for treating neoplasms
using tetracycline compounds of the invention are also included
(van der Bozert et ai., Cancer Res., 1998,48:6686-6690). In one
embodiment, the tetracycline responsive state is not a bacterial
infection. In another embodiment, the tetracycline compounds of the
invention are essentially non-antibacterial. For example,
non-antibacterial tetracycline compounds of the invention may have
MIC values greater than about 4 .mu.g/ml as measured by assays
known in the art.
[0175] Tetracycline compound responsive states also include
inflammatory process associated states (IPAS). The term
"inflammatory process associated state" includes states in which
inflammation or inflammatory factors (e.g., matrix
metalloproteinases (MMPs), nitric oxide (NO), TNF, interleukins,
plasma proteins, cellular defense systems, cytokines, lipid
metabolites, proteases, toxic radicals, adhesion molecules, etc.)
are involved or present in an area in aberrant amounts. The
inflammatory process is the response of living tissue to damage.
The cause of inflammation may be due to physical damage, chemical
substances, micro-organisms, tissue necrosis, cancer or other
agents. Acute inflammation is short-lasting for only a few days. If
it is longer lasting, however, then it may be referred to as
chronic inflammation.
[0176] IPAS's include inflammatory disorders Inflammatory disorders
are generally characterized by heat, redness, swelling, pain or
loss of function. Examples of causes of inflammatory disorders
include, but are not limited to, microbial infections (e.g.,
bacterial and fungal infections), physical agents (e.g., burns,
radiation and trauma), chemical agents (e.g., toxins and caustic
substances), tissue necrosis and various types of immunologic
reactions. In a further embodiment, the IPAS includes disorders
described in U.S. Pat. Nos. 5,929,055 and 5,532,227, incorporated
herein by reference in their entirety.
[0177] Examples of inflammatory disorders include, but are not
limited to, osteoarthritis, rheumatoid arthritis, acute and chronic
infections (bacterial and fungal, including diphtheria and
pertussis), acute and chronic bronchitis, sinusitis, upper
respiratory infections (the common cold, etc.), acute and chronic
gastroenteritis and colitis, acute and chronic cystitis and
urethritis, acute and chronic dermatitis, acute and chronic
conjunctivitis, acute and chronic serositis (pericarditis,
peritonitis, synovitis, pleuritis and tendinitis), uremic
pericarditis, acute and chronic cholecystis, acute and chronic
vaginitis, acute and chronic uveitis, drug reactions, insect bites,
burns (thermal, chemical and electrical) and sunburn.
[0178] The term "inflammatory process associated state" includes,
in one embodiment, NO associated states. The term "NO associated
state" includes states which involve or are associated with nitric
oxide (NO) or inducible nitric oxide synthase (iNOS). NO associated
state includes states which are characterized by aberrant amounts
of NO and/or iNOS. Preferably, the NO associated state can be
treated by administering tetracycline compounds of the invention.
The disorders, diseases and states described in U.S. Pat. Nos.
6,231,894; 6,015,804; 5,919,774; and 5,789,395 are also included as
NO associated states. The entire contents of each of these patents
are hereby incorporated herein by reference.
[0179] Other examples of NO associated states include, but are not
limited to, malaria, senescence, diabetes, vascular stroke,
neurodegenerative disorders (Alzheimer's disease, Huntington's
disease and Parkinson's disease), cardiac disease
(reperfusion-associated injury following infarction), juvenile
diabetes, inflammatory disorders, osteoarthritis, rheumatoid
arthritis, acute, recurrent and chronic infections (bacterial,
viral and fungal), acute and chronic bronchitis, sinusitis, and
respiratory infections (the common cold, etc.), acute and chronic
gastroenteritis and colitis, acute and chronic cystitis and
urethritis, acute and chronic dermatitis, acute and chronic
conjunctivitis, acute and chronic serositis (pericarditis,
peritonitis, synovitis, pleuritis and tendonitis), uremic
pericarditis, acute and chronic cholecystis, cystic fibrosis, acute
and chronic vaginitis, acute and chronic uveitis, drug reactions,
insect bites, burns (thermal, chemical and electrical), and
sunburn.
[0180] The term "inflammatory process associated state" also
includes, in one embodiment, matrix metalloproteinase associated
states (MMPAS). MMPAS include states characterized by aberrant
amounts of MMPs or MMP activity. These are also included as
tetracycline compound responsive states which may be treated using
compounds of the invention.
[0181] Examples of matrix metalloproteinase associated states
(MMPAS's) include, but are not limited to, arteriosclerosis,
corneal ulceration, emphysema, osteoarthritis, multiple sclerosis
(Liedtke et al., Ann. Neurol. 1998, 44:35-46; Chandler et al., J.
Neuroimmunol. 1997, 72:155-71), osteosarcoma, osteomyelitis,
bronchiectasis, chronic pulmonary obstructive disease, skin and eye
diseases, periodontitis, osteoporosis, rheumatoid arthritis,
ulcerative colitis, inflammatory disorders, tumor growth and
invasion (Stetler-Stevenson et al., Annu. Rev. Cell Biol.
1993,9:541-73; Tryggvason et al., Biochim. Biophys. Acta 1987,
907:191-217; Li et al., Mol. Carcinog. 1998,22:84-89), metastasis,
acute lung injury, stroke, ischemia, diabetes, aortic or vascular
aneurysms, skin tissue wounds, dry eye, bone and cartilage
degradation (Greenwald et al., Bone 1998,22:33-38; Ryan et al.,
Curr. Op. Rheumatol. 1996,8:238-247). Other MMPAS include those
described in U.S. Pat. Nos. 5,459,135; 5,321,017; 5,308,839;
5,258,371; 4,935,412; 4,704,383; 4,666,897 and RE34,656,
incorporated herein by reference in their entirety.
[0182] In another embodiment, the tetracycline compound responsive
state is cancer. Examples of cancers which the tetracycline
compounds of the invention may be useful to treat include all solid
tumors, i.e., carcinomas e.g., adenocarcinomas and sarcomas.
Adenocarcinomas are carcinomas derived from glandular tissue or in
which the tumor cells form recognizable glandular structures.
Sarcomas broadly include tumors whose cells are embedded in a
fibrillar or homogeneous substance like embryonic connective
tissue. Examples of carcinomas which may be treated using the
compounds of the invention include, but are not limited to,
carcinomas of the prostate, breast, ovary, testis, lung, colon and
breast. The methods of the invention are not limited to the
treatment of these tumor types, but extend to any solid tumor
derived from any organ system. Examples of treatable cancers
include, but are not limited to, colon cancer, bladder cancer,
breast cancer, melanoma, ovarian carcinoma, prostatic carcinoma,
lung cancer, and a variety of other cancers as well. The methods of
the invention also cause the inhibition of cancer growth in
adenocarcinomas, such as, for example, those of the prostate,
breast, kidney, ovary, testes and colon.
[0183] In an embodiment, the invention pertains to a method for
treating a subject suffering or at risk of suffering from cancer,
by administering an effective amount of a substituted tetracycline
compound such that inhibition of cancer cell growth occurs., i.e.,
cellular proliferation, invasiveness, metastasis, or tumor
incidence is decreased, slowed, or stopped. The inhibition may
result from inhibition of an inflammatory process, downregulation
of an inflammatory process, some other mechanism or a combination
of mechanisms. Alternatively, the tetracycline compounds may be
useful for preventing cancer recurrence, for example, to treat
residual cancer following surgical resection or radiation therapy.
The tetracycline compounds useful according to the invention are
especially advantageous as they are substantially non-toxic
compared to other cancer treatments. In a further embodiment, the
compounds of the invention are administered in combination with
standard cancer therapy, such as, but not limited to,
chemotherapy.
[0184] Examples of tetracycline responsive states also include
neurological disorders which include both neuropsychiatric and
neurodegenerative disorders, but are not limited to, Alzheimer's
disease, dementias related to Alzheimer's disease such as Pick's
disease, Parkinson's diseases and other Lewy diffuse body diseases,
senile dementia, Huntington's disease, Gilles de la Tourette's
syndrome, multiple sclerosis, amylotrophic lateral sclerosis (ALS),
progressive supranuclear palsy, epilepsy, and Creutzfeldt-Jakob
disease, autonomic function disorders such as hypertension and
sleep disorders, neuropsychiatric disorders (depression,
schizophrenia, schizoaffective disorder, Korsakoffs psychosis,
mania, anxiety disorders, phobic disorders, etc.), learning or
memory disorders (amnesia or age-related memory loss, attention
deficit disorder, etc.), dysthymic disorder, major depressive
disorder, obsessive-compulsive disorder, psychoactive substance use
disorders, anxiety, phobias, panic disorder, as well as bipolar
affective disorder (severe bipolar affective (mood) disorder (BPI),
bipolar affective neurological disorders, e.g., migraine and
obesity, etc.). Further neurological disorders include, for
example, those listed in the American Psychiatric Association's
Diagnostic and Statistical manual of Mental Disorders (DSM), the
most current version of which is incorporated herein by reference
in its entirety.
[0185] In another embodiment, the tetracycline compound responsive
state is diabetes, e.g., juvenile diabetes, diabetes mellitus,
diabetes type I, or diabetes type II. In a further embodiment,
protein glycosylation is not affected by the administration of the
tetracycline compounds of the invention. In another embodiment, the
tetracycline compound of the invention is administered in
combination with standard diabetic therapies, such as, but not
limited to insulin therapy.
[0186] In another embodiment, the tetracycline compound responsive
state is a bone mass disorder. Bone mass disorders include
disorders where a subjects bones are disorders and/or states where
the formation, repair or remodeling of bone is advantageous. For
example, bone mass disorders include osteoporosis (e.g., a decrease
in bone strength and density), bone fractures, bone formation
associated with surgical procedures (e.g., facial reconstruction),
osteogenesis imperfecta (brittle bone disease), hypophosphatasia,
Paget's disease, fibrous dysplasia, osteopetrosis, myeloma bone
disease, and the depletion of calcium in bone, such as that which
is related to primary hyperparathyroidism. Bone mass disorders
include all states in which the formation, repair or remodeling of
bone is advantageous to the subject as well as all other disorders
associated with the bones or skeletal system of a subject which can
be treated with the tetracycline compounds of the invention. In a
further embodiment, the bone mass disorders include those described
in U.S. Pat. Nos. 5,459,135; 5,231,017; 5,998,390; 5,770,588; RE
34,656; 5,308,839; 4,925,833; 3,304,227 and 4,666,897, each of
which is hereby incorporated herein by reference in its
entirety.
[0187] In another embodiment, the tetracycline compound responsive
state is acute lung injury. Acute lung injuries include adult
respiratory distress syndrome (ARDS), postpump syndrome (PPS) and
trauma. Trauma includes any injury to living tissue caused by an
extrinsic agent or event. Examples of trauma include, but are not
limited to, crush injuries, contact with a hard surface, or cutting
or other damage to the lungs.
[0188] The invention also pertains to a method for treating acute
lung injury by administering a substituted tetracycline compound of
the invention.
[0189] The tetracycline responsive states of the invention also
include chronic lung disorders. The invention pertains to methods
for treating chronic lung disorders by administering a tetracycline
compound, such as those described herein. The method includes
administering to a subject an effective amount of a substituted
tetracycline compound such that the chronic lung disorder is
treated. Examples of chronic lung disorders include, but are not
limited, to asthma, cystic fibrosis and emphysema. In a further
embodiment, the tetracycline compounds of the invention used to
treat acute and/or chronic lung disorders such as those described
in U.S. Pat. Nos. 5,977,091; 6,043,231; 5,523,297; and 5,773,430,
each of which is hereby incorporated herein by reference in its
entirety.
[0190] In yet another embodiment, the tetracycline compound
responsive state is ischemia, stroke or ischemic stroke. The
invention also pertains to a method for treating ischemia, stroke
or ischemic stroke by administering an effective amount of a
substituted tetracycline compound of the invention. In a further
embodiment, the tetracycline compounds of the invention are used to
treat such disorders as described in U.S. Pat. Nos. 6,231,894;
5,773,430; 5,919,775 and 5,789,395, incorporated herein by
reference.
[0191] In another embodiment, the tetracycline compound responsive
state is a skin wound. The invention also pertains, at least in
part, to a method for improving the healing response of the
epithelialized tissue (e.g., skin, mucusae, etc.) to acute
traumatic injury (e.g., cut, burn, scrape, etc.). The method may
include using a tetracycline compound of the invention (which mayor
may not have antibacterial activity) to improve the capacity of the
epithelialized tissue to heal acute wounds. The method may increase
the rate of collagen accumulation of the healing tissue. The method
may also decrease the proteolytic activity in the epthithelialized
tissue by decreasing the collagenolytic and/or gellatinolytic
activity of MMPs. In a further embodiment, the tetracycline
compound of the invention is administered to the surface of the
skin (e.g., topically). In a further embodiment, the tetracycline
compound of the invention used to treat a skin wound, and other
such disorders as described in, for example, U.S. Pat. Nos.
5,827,840; 4,704,383; 4,935,412; 5,258,371; 5,308,8391 5,459,135;
5,532,227 and 6,015,804, each of which is incorporated herein by
reference in its entirety.
[0192] In yet another embodiment, the tetracycline compound
responsive state is an aortic or vascular aneurysm in vascular
tissue of a subject (e.g., a subject having or at risk of having an
aortic or vascular aneurysm, etc.). The tetracycline compound may
be effective to reduce the size of the vascular aneurysm or it may
be administered to the subject prior to the onset of the vascular
aneurysm such that the aneurysm is prevented. In one embodiment,
the vascular tissue is an artery, e.g., the aorta, e.g., the
abdominal aorta. In a further embodiment, the tetracycline
compounds of the invention are used to treat disorders described in
U.S. Pat. Nos. 6,043,225 and 5,834,449, incorporated herein by
reference in their entirety.
[0193] Bacterial infections may be caused by a wide variety of gram
positive and gram negative bacteria. The compounds of the invention
are useful as antibiotics against organisms which are resistant to
other tetracycline compounds. The antibiotic activity of the
tetracycline compounds of the invention may be determined using the
in vitro standard broth dilution method described in Waitz, J. A.,
National Commission/or Clinical Laboratory Standards, Document
M7-A2, vol. 10, no. 8, pp. 13-20, 2nd edition, Villanova, Pa.
(1990).
[0194] The tetracycline compounds may also be used to treat
infections traditionally treated with tetracycline compounds such
as, for example, rickettsiae infection, a number of gram-positive
and gram-negative bacterial infection, lymphogranuloma venereum,
inclusion conjunctivitis and psittacosis. The tetracycline
compounds may be used to treat infections of, e.g., K. pneumoniae,
Salmonella, E. hirae, A. baumanii, B. catarrhalis, H influenzae, P.
aeruginosa, E. faecium, E. coli, S. aureus or E. faecalis. In one
embodiment, the tetracycline compound is used to treat a bacterial
infection that is resistant to other tetracycline antibiotic
compounds. The tetracycline compound of the invention may be
administered with a pharmaceutically acceptable carrier.
[0195] The language "in combination with" another therapeutic agent
or treatment includes co-administration of the tetracycline
compound, (e.g., inhibitor) and with the other therapeutic agent or
treatment, administration of the tetracycline compound first,
followed by the other therapeutic agent or treatment and
administration of the other therapeutic agent or treatment first,
followed by the tetracycline compound. The other therapeutic agent
may be any agent which is known in the art to treat, prevent, or
reduce the symptoms of a disease or disorder, such as IPAS.
Furthermore, the other therapeutic agent may be any agent of
benefit to the patient when administered in combination with the
administration of a tetracycline compound. In one embodiment, the
diseases, such as cancer, treated by methods of the invention
include those described in U.S. Pat. Nos. 6,100,248; 5,843,925;
5,837,696 and 5,668,122, incorporated herein by reference in their
entirety.
[0196] The language "effective amount" of the compound is that
amount necessary or sufficient to treat or prevent a tetracycline
compound responsive state. The effective amount can vary depending
on such factors as the size and weight of the subject, the type of
illness, or the particular tetracycline compound. For example, the
choice of the tetracycline compound can affect what constitutes an
"effective amount". One of ordinary skill in the art would be able
to study the aforementioned factors and make the determination
regarding the effective amount of the tetracycline compound without
undue experimentation.
[0197] In the therapeutic methods of the invention, one or more
tetracycline compounds of the invention may be administered alone
to a subject, or more typically a compound of the invention will be
administered as part of a pharmaceutical composition in mixture
with conventional excipient, i.e., pharmaceutically acceptable
organic or inorganic carrier substances suitable for parenteral,
oral or other desired administration and which do not deleteriously
react with the active compounds and are not deleterious to the
recipient thereof.
[0198] The invention is further illustrated by the following
examples, which should not be construed as further limiting.
6. EXEMPLIFICATION OF THE INVENTION
Example 1
Synthesis of 9-Alkyl Aminomethyl Minocycline
##STR00005##
[0200] Minocycline hydrochloride (compound 2) was dissolved in
methylsulfonic acid or hydrofluoric acid with methylsulfonic
anhydride or similar water scavenger acid such as triflic acid.
N-hydroxymethyl phthalimide was added to the reaction mixture. The
mixture was stirred at 20-35.degree. C. until the reaction was
complete. The acid solution was added to an ice/water mixture, and
triflic salt can be readily precipitated, filtered and collected.
The solid was re-dissolved in acetone and brought to a neutral pH
with base. The product was precipitated by the addition of water.
If the triflic acid was present as scavenger the product can be
precipitated without neutralization. The product was isolated as a
mixture of the bis and tris alkylated product. The isolated
material of this reaction was enriched in the desired bis ratio
(90%).
[0201] The solid was suspended in the EtOH or MeOH. Aminolysis was
carried out by using methylamine. A phthalamide by-product
precipitated as the reaction progressed and was removed by
filtration. The light yellow solid product was precipitated out by
the addition of about 1.5 volumes of t-butylmethylether to the
reaction mixture, and collected through a simple filtration that
left many small impurities and methylamine reagent in the solution.
Further purification of the compound was performed through
re-slurrying with a lower aliphatic alcohol such as methanol.
[0202] Compound 4 as freebase was transferred to a hydrogenation
vessel which was charged with methanol and aldehyde. An inactivated
Pd/C catalyst was charged and the vessel was pressurized with
hydrogen gas. The reaction mixture was hydrogenated under hydrogen
pressure around 30 Psi for about 24 hours. When conversion of
compound 4 to 1 was complete, the solution was filtered and washed
through a Celite pad. At this point the reaction mixture contained
very low .about.C-4 epimer, around 3-7%.
[0203] The product (1) was worked up as follows to isolate the
product selectively from its impurities. The pH of the solution was
adjusted to about 4.5 with concentrated HCl and the solution was
extracted with dichloromethane. The aqueous layer was extracted
with dichloromethane to selectively recover the preferred epimer
product (e.g., a). The dichloromethane layers were combined and
concentrated, and 2 L of n-heptane was added to precipitate the
product. Further purification was obtained by repeating the work-up
procedure with or without t-butylmethylether to dissolve the crude
product.
Example 2
Purification of Compound 1
[0204] Crude 9-(2',2'-dimethylpropyl aminomethyl) minocycline
freebase (40 g) was dissolved in 150 mL of buffer A (0.1% aqueous
solution of methane sulfonic acid--MSA) and the pH was adjusted to
2-3 with MSA.
[0205] The solution was filtered and injected into an HPLC and the
product was eluted with an isocratic gradient of 94% buffer A and
6% acetonitrile. The product fraction collection was initiated when
the product peak was detected. Each fraction was analyzed and an
acceptance criterion of greater than 80% AVC of the main peak was
used for the early product fractions. When combining fractions, the
level of impurities and relative concentration of the pooled
fractions was factored into the selection criteria that meet the
final product specifications. To the product fractions was added a
10% aqueous solution of sodium sulfite equal to 10% of the original
volume of the collected fractions.
[0206] The following example represents the output of a single
injection. The output from multiple injections may also be combined
and worked up. A product fraction volume of 3.5 L (including sodium
sulfite) was collected and the pH was adjusted to 4.0-4.5 using a
solution of sodium hydroxide. The aqueous solution was washed with
2 L of dichloromethane and the organic layer was separated and
discarded.
[0207] The pH of the aqueous layer is adjusted to 7.5-8.5 using
sodium hydroxide and the product was extracted four times with 2.4
L of dichloromethane. The pH was readjusted to 7.5-8.5 with sodium
hydroxide or MSA, prior to each extraction.
[0208] The four dichloromethane layers were combined and
concentrated to about 200 mL, which was then added slowly (over a
period of about 10 minutes) to a vigorously stirred n-heptane (2.5
L). The suspension was stirred for about 10 minutes at room
temperature and diluted slowly (over a period of 5 minutes) with
n-heptane 1.5 L. The slurry is cooled to 0-5.degree. C. and stirred
for 1-2 hours. The suspended solid was filtered and washed with
3.times.150 mL portions of n-heptane. The product was dried under
vacuum at 40.degree. C. for at least 24 hours until a constant
weight was achieved and the levels of all residual solvents were
within specification. Approximately 13.6 g of
9-(2',2'-dimethylpropyl aminomethyl) minocycline free base was
isolated as a yellow solid.
Example 3
Preparation of Crystalline HCl Salt of Compound 1
[0209] Compound 1 (13 g) was dissolved in acetone (300 mL),
filtered and the filter was additionally washed with acetone. The
combined filtrate and washes was cooled to 5.degree. C. To the
combined filtrate and washes, a solution of concentrated HCl (3.9
mL) in acetone (79 mL) was slowly added with vigorous stirring. The
resultant slurry was stirred in an ice bath for 15 minutes and
filtered off.
[0210] The first crop of solid was washed with cold acetone and
pentane and dried in vacuo for 48 hours, yielding 13.7 g of yellow
amorphous solid. The flask with saturated filtrate was covered with
aluminum foil and left for 2 weeks during which the growth of
single crystals was observed. Crystals were collected by filtration
and washed with hexane.
Example 4
Preparation of Crystalline Methanesulfonic Acid Salt of Compound
1
[0211] To a 25-mL 3-neck flask under an inert nitrogen atmosphere,
3 mL of isopropanol (IPA) was charged. A slurry was prepared by
adding 225 mg amorphous free base of Compound 1 to the flask. The
slurry was warmed to a temperature of 45.degree. C. Methanesulfonic
acid hydrate (98.0 mg) was then added to the slurry. The slurry was
stirred at 45.degree. C. for one hour, then cooled to 22.degree. C.
to produce a thick crystalline slurry. The slurry was filtered and
washed with IPA (2 xl mL). Excess IPA was removed from the
crystalline cake by drying at 55.degree. C. for more than two hours
to achieve a constant weight. Crystalline mesylate (methanesulfonic
acid) salt of Compound 1 (180 mg) was isolated. It was determined
that the crystalline mesylate salt was unstable at 5.degree. C.
Example 5
Preparation of Crystalline Tosylate Salt of Compound 1 (Using a
Slurry Method)
[0212] To a 5-L 3-neck flask under an inert nitrogen atmosphere,
2.0 L of isopropanol (IPA) was charged. A slurry was prepared by
adding 289 g of the amorphous free base of Compound 1 to the flask.
A solution of p-toluenesulfonic acid hydrate (97.0 g) in IPA (400
mL) was then added to the slurry. The water content of the slurry
supernatant was adjusted to 0.6 g/L with the addition of water (9
mL), and the slurry was stirred at 20-25.degree. C. for 18 hours to
produce a thick crystalline slurry. The slurry was filtered and
washed with IPA (2.times.500 mL). Excess IPA was removed from the
crystalline cake by blowing dry nitrogen through the cake for 24
hours. With the solids containing 3 weight-percent (wt %) of IP A,
the cake was further dried by blowing humidified nitrogen through
the cake at a relative humidity of 70-75% for 24 hours. The cake
retained 0.9 wt % of IPA that was not further reduced by this
method. Excess water was then removed from the cake by blowing dry
nitrogen through the cake for 24 hours. The tosylate salt of
Compound 1 was isolated as an orange powder (337 g). The isolated
tosylate salt of Compound 1 was crystalline with only one observed
form: a non-stoichiometric hemihydrate, as determined by x-ray
powder diffraction (XPRD) and thermogravimetric (TG) analysis.
Example 6
Preparation of Crystalline Tosylate Salt of Compound 1 (Using a
Solution Method)
[0213] To a 5-L 3-neck flask under an inert nitrogen atmosphere,
1.7 L of methanol and 1.7 L ofmethyl-t-butyl ether were charged.
P-toluene sulfonic acid monohydrate (209 g) and the amorphous free
base of Compound 1 (556 g) were added with stirring to the flask to
obtain a clear solution. A seed quantity (3 g) of the monotosylate
salt of compound 1 was added to initiate crystallization and a
further quantity of methanol (0.1 L) and methyl-t-butyl ether (0.5
L) was added. The resulting slurry was stirred at about 20.degree.
C. for 22 hours to produce a thick crystalline slurry. The slurry
was filtered and washed with a mixture of 1.1 L of methanol and 1.3
L ofmethyl-t-butyl ether followed by methyl-t-butyl ether
(2.times.2.4 L). The tosylate salt of Compound 1 was isolated as an
orange powder. Excess solvent was removed from the crystalline cake
by blowing dry nitrogen through the cake for 24 hours. The cake was
then dried under vacuum at about 30.degree. C. until the solids
contained about 6 weight-percent (wt %) of solvent. The cake was
further dried under vacuum at about 45.degree. C. until the solids
contained less than 3 weight-percent (wt %) of solvent. The
isolated tosylate salt of Compound 1 was crystalline with the
observed form 1 as determined by x-ray powder diffraction
(XPRD).
Example 7
Characterization of Tosylate Salt of Compound 1
[0214] The XRPD pattern of the isolated tosylate salt of Compound 1
comprised 28 values in degrees of 5.6, 8.0, 8.6, 11.4, 13.0, 15.5,
18.8, 20.4 and 24.5.
[0215] The crystalline tosylate salt was subjected to
thermogravimetric (TO) analysis under nitrogen flow at a heating
rate of 10.degree. C./minute. A weight loss of 3.9% was observed up
to 81.3.degree. C., which was due to water loss.
[0216] Upon drying, water content in the crystalline tosylate salt
of Compound 1 was calculated to be 0.5%. After standing at room
temperature for 24 hours, this value increased to 5%. In contrast
to the amorphous Compound 1, the crystalline tosylate salt of
Compound 1 was stable for weeks and months at room temperature,
maintaining a water content of approximately 5%.
[0217] The hygroscopicity of the subject crystalline tosylate salt
was determined using a symmetric vapor sorption analyzer, and
reported as wt % gained as a function of percent relative humidity
(%RH) from 5% to 95% to 1%, at 5% intervals at 25.0.degree. C. A
maximum weight gain of 12 wt % at 95% RH, with slight hysteresis
upon desorption, was observed, and a 2 wt % loss due to IPA was
observed.
[0218] The crystalline tosylate salt was determined to have the
solubilities summarized in Table 6 below. The solubilities were
determined by mixing excess solids with solvent, at ambient
temperature for two hours, followed by filtration of the
supernatant. The supernatant concentration was determined by
high-performance liquid chromatography (HPLC). Equilibration of
56.8 mg of the subject crystalline tosylate salt in 0.5 mL water
and 122.1 mg of the subject crystalline tosylate salt in 1.0 mL
water resulted in clear solutions. The pH of 122.1 mg of the
subject crystalline tosylate salt in 1.0 mL of water was determined
to be 5.70.
TABLE-US-00006 TABLE 6 Solvent Solubility (mg/mL) Water >100
Acetonitrile 5.9 Methanol 86.1 IPA 5.8
Example 8
XRPD of Compound 1
[0219] X-Ray Powder Diffraction patterns were collected on a
Siemens D5000 diffractometer using Cu Ka radiation (40 kV, 40 mA),
8-8 goniometer, divergence of V20 and receiving slits, a graphite
secondary monochromator and a scintillation counter. The instrument
is performance checked using a certified Corundum standard (NIST
1976). The software used for data collection was Diffrac Plus XRD
Commander v2.3.1 and the data were analyzed and presented using
Diffrac Plus EVA v 11,0.0.2 or v 13.0.0.2.
[0220] Powder samples were prepared as flat plate specimens using.
Approximately 35 mg of the sample was gently packed into a cavity
cut into polished, zero-background (510) silicon wafer. The sample
was rotated in its own plane during analysis. The details of the
data collection are: [0221] Angular range: 2 to 42 .degree. 28
[0222] Step size: 0.05 .degree. 28 [0223] Collection time: 4
s/step. High resolution X-Ray Powder Diffraction patterns were
collected on a Bruker AXS C2 GADDS diffractometer using Cu Ka
radiation (40 kV, 40 rnA), automated XYZ stage, laser video
microscope for auto-sample positioning and a HiStar 2-dimensional
area detector. X-ray optics consists of a single Gobel multilayer
mirror coupled with a pinhole collimator of 0.3 mm.
[0224] The beam divergence, i.e. the effective size of the X-ray
beam on the sample, was approximately 4 mm. A 8-8 continuous scan
mode was employed with a sample--detector distance of 20 cm which
gives an effective 28 range of 3.2.degree.-29.7.degree.. Typically,
the sample would be exposed to the X-ray beam for 120 seconds. The
software used for data collection was GADDS for WNT 4.1.16 and the
data were analyzed and presented using Diffrac Plus EVA v 9.0.0.2
or v 13.0.0.2.
Example 9
Temperature and Moisture Stability Study
[0225] A stability study was conducted on tosylate salt and
mesylate salt of Compound 1, by monitoring changes in the high
performance liquid chromatography (HPLC) impurity profiles for each
salt when exposed to various temperatures and/or humidity
conditions. Each sample was placed in a closed container and
exposed to three controlled environments: refrigerated (5.degree.
C.), 20.degree. C. and 60% relative humidity, and 40.degree. C. and
75% relative humidity. After two weeks, the crystalline tosylate
salt was determined to be the most stable crystalline form of
Compound 1. The stability study of the crystalline tosylate salt
was continued.
[0226] Samples were analyzed by the following reversed-phase HPLC
impurity profile method at time points 0, 1, 2, 4 weeks (Table 7)
and 3 months (Table 8) for the tosylate salt; and 0, 1 and 2 weeks
for the mesylate salt (Table 9).
[0227] Reversed-phase HPLC analysis was conducted using a SYMMETRY
SHIELD RP18 column (4.6.times.250 mm, 5 pm particle size), although
analysis using a similar or equivalent column would be expected to
yield similar results. The mobile phase components were 0.01 M
ammonium acetate in water at pH 3.3 (A) and acetonitrile (B). The
mobile phase composition was a gradient that increased from 6% to
15% B in five minutes, held at 15% B for 15 minutes, increased from
15% to 60% B in ten minutes, then from 60% to 90% B in two minutes,
and then the system was re-equilibrated at 6% B for four minutes.
The flow rate was 1.0 mLiminute, and the injection volume was 1 0.0
.about.L. The column temperature was maintained at 30.degree. C.
Detection was by ultraviolet light (UV) at 280 nm. The retention
time for Compound 1 was approximately 15.7 minutes. The sample
solution was prepared in mobile phase A to a final concentration of
2.0 mg/mL.
TABLE-US-00007 TABLE 7 RRT RRT RRT RRT RRT*** RRT Total 0.52 0.77
0.93 1.19 1.26 1.77 Compound Time Point Impurities** (%) (%) (%)
(%) (%) (%) (%) Initial 1.60 0.06 ND ND 0.24 1.2 0.10 98.4 1 week
5.degree. C. 1.51 0.09 ND ND 0.21 1.1 0.11 98.5 20.degree. C. &
60% RH* 1.63 0.09 ND ND 0.21 1.2 0.12 98.3 40.degree. C. & 75%
RH 1.80 0.09 0.05 ND 0.21 1.3 0.14 98.2 2 weeks 5.degree. C. 1.62
0.08 ND 0.08 0.24 1.1 0.12 98.4 20.degree. C. & 60% RH 1.66
0.08 ND 0.08 0.25 1.1 0.15 98.4 40.degree. C. & 75% RH 1.68
0.08 ND 0.09 0.25 1.1 0.16 98.3 4 weeks 5.degree. C. 1.61 0.09 0.08
0.07 0.23 1.0 0.14 98.3 20.degree. C. & 60% RH 1.84 0.10 0.10
0.12 0.24 1.1 0.18 98.2 40.degree. C. & 75% RH 1.96 0.10 0.17
0.13 0.26 1.1 0.20 98.1 *RH denotes relative humidity **Total
impurities included all impurities in a specific batch ***RRT 1.26
denotes beta-epimer.
TABLE-US-00008 TABLE 8 20.degree. C. & 60% RH* 40.degree. C.
& 75% RH 4-keto M-2 Beta- 4-keto M-2 Beta-epimer compound
impurity Total epimer compound impurity Total Time Point RRT = 1.26
RRT = 1.57 RRT = 0.84 impurity** RRT = 1.26 RRT = 1.57 RRT = 0.84
impurity 0 month 1.76% 0.05% -- 2.12% 1.76% 0.05% -- 2.12% 3 month
1.74% 0.10% 0.18% 2.86% 1.66% 0.14% 0.31% 3.12% *RH denotes
relative humidity **Total impurities included all impurities in a
specific batch
TABLE-US-00009 TABLE 9 Total RRT RRT RRT RRT RRT RRT Impurities
0.52 0.77 0.93 1.19 1.26 1.77 Compound 1 Time Point (%) (%) (%) (%)
(%) (%) (%) (%) Initial 1.70 ND ND ND 0.20 1.5 ND 98.3 1 week
5.degree. C. 1.85 ND ND ND 0.17 1.6 0.08 98.2 20.degree. C. &
60% RH* 2.06 ND ND ND 0.17 1.8 0.09 98.0 40.degree. C. & 75% RH
2.97 0.07 0.05 ND 0.16 2.6 0.09 97.1 2 weeks 5.degree. C. 1.65 ND
ND 0.06 0.20 1.3 0.09 98.3 20.degree. C. & 60% RH 1.87 ND ND
0.06 0.20 0.11 0.11 98.1 40.degree. C. & 75% RH 3.38 0.05 0.07
0.06 0.19 0.11 0.11 96.6
[0228] The above data demonstrates that the tosylate salt of
Compound 1 is stable when stored refrigerated and/or 20.degree. C.
& 60% RH conditions for at least four weeks. The tosylate salt
of Compound 1 is stable at temperature in a range from 0.degree.
C.-70.degree. C., or in a range from 5.degree. C.-50.degree. C., or
in a range from 20.degree. C.-30.degree. C.
Example 10
Photostability Study
[0229] A photostability study was conducted on the tosylate salt of
Compound 1. Two samples were prepared in clear glass Petri dishes.
One sample was wrapped in aluminum foil to serve as a control
sample. Both samples were placed in the ES 2000 Environmental Light
10 Chamber and exposed to 12 kilo lux of cool white fluorescent
light for a total of 47 hours. Samples were then analyzed by HPLC
impurity profile method, and the results are summarized in Table
10.
TABLE-US-00010 TABLE 10 RRT RRT RRT RRT RRT RRT RRT RRT 0.52 0.0.64
0.77 0.83 0.93 1.19 1.26 1.77 Compound 1 Sample (%) (%) (%) (%) (%)
(%) (%) (%) (%) Control 0.06 ND 0.08 ND 0.12 0.26 1.2 0.16 98.2
Exposed 0.06 ND 0.09 ND 0.13 0.26 1.1 0.16 98.2
Equivalents
[0230] Those skilled in the art will recognize, or be able to
ascertain using no more than routine experimentation, many
equivalents to the specific embodiments and methods described
herein. Such equivalents are intended to be encompassed by the
scope of the following claims.
[0231] All patents, patent applications, and literature references
cited herein are hereby expressly incorporated by reference.
* * * * *