U.S. patent application number 16/910945 was filed with the patent office on 2020-10-08 for propylene glycol-containing peptide formulations which are optimal for production and for use in injection devices.
The applicant listed for this patent is Novo Nordisk A/S. Invention is credited to Claude Bonde, Dorthe Kot Engelund, Tina Bjeldskov Pedersen.
Application Number | 20200316204 16/910945 |
Document ID | / |
Family ID | 1000004915680 |
Filed Date | 2020-10-08 |
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United States Patent
Application |
20200316204 |
Kind Code |
A1 |
Pedersen; Tina Bjeldskov ;
et al. |
October 8, 2020 |
PROPYLENE GLYCOL-CONTAINING PEPTIDE FORMULATIONS WHICH ARE OPTIMAL
FOR PRODUCTION AND FOR USE IN INJECTION DEVICES
Abstract
The present invention relates to pharmaceutical formulations
comprising a peptide and propylene glycol, to methods of preparing
such formulations, and to uses of such formulations in the
treatment of diseases and conditions for which use of the peptide
contained in such formulations is indicated. The present invention
further relates to methods for reducing the clogging of injection
devices by a peptide formulation and for reducing deposits on
production equipment during production of a peptide
formulation.
Inventors: |
Pedersen; Tina Bjeldskov;
(Smoerum, DK) ; Bonde; Claude; (Lyngby, DK)
; Engelund; Dorthe Kot; (Holte, DK) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Novo Nordisk A/S |
Bagsvaerd |
|
DK |
|
|
Family ID: |
1000004915680 |
Appl. No.: |
16/910945 |
Filed: |
June 24, 2020 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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16260204 |
Jan 29, 2019 |
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16910945 |
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16050827 |
Jul 31, 2018 |
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16260204 |
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13362745 |
Jan 31, 2012 |
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16050827 |
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11435977 |
May 17, 2006 |
8114833 |
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13362745 |
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PCT/DK04/00792 |
Nov 18, 2004 |
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11435977 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 47/26 20130101;
A61K 47/20 20130101; A61K 38/26 20130101; A61K 47/10 20130101; A61K
9/0019 20130101; A61K 47/183 20130101; A61K 38/28 20130101 |
International
Class: |
A61K 47/10 20060101
A61K047/10; A61K 9/00 20060101 A61K009/00; A61K 38/26 20060101
A61K038/26; A61K 38/28 20060101 A61K038/28; A61K 47/18 20060101
A61K047/18; A61K 47/20 20060101 A61K047/20; A61K 47/26 20060101
A61K047/26 |
Foreign Application Data
Date |
Code |
Application Number |
Nov 20, 2003 |
DK |
PA 2003 01719 |
Claims
1. A pharmaceutical formulation comprising: Arg34,
Lys26(N-.epsilon.-(.gamma.-Glu(N-.alpha.-hexadecanoyl)))-GLP-1(7-37)
in a concentration of about 0.1 mg/ml to about 10 mg/ml; and
propylene glycol in a concentration of from about 8 mg/ml to about
16 mg/ml, wherein said formulation has a pH in the range of from
about 7.0 to about 9.5.
2. The formulation according to claim 1, wherein the concentration
of propylene glycol is from about 13 mg/ml to about 15 mg/ml.
3. The formulation according to claim 1, wherein the concentration
of propylene glycol is from about 13.5 mg/ml to about 14.5
mg/ml.
4. The formulation according to claim 1, wherein said formulation
has a pH of from about 7.0 to about 8.3 mg/ml.
5. The formulation according to claim 3, further comprising a
buffer.
6. The formulation according to claim 5, wherein the buffer is
sodium acetate.
7. The formulation according to claim 5, wherein the buffer is
sodium carbonate.
8. The formulation according to claim 5, wherein the buffer is
citrate.
9. The formulation according to claim 5, wherein the buffer is
glycylglycine.
10. The formulation according to claim 5, wherein the buffer is
histidine.
11. The formulation according to claim 5, wherein the buffer is
glycine.
12. The formulation according to claim 5, wherein the buffer is
lysine.
13. The formulation according to claim 5, wherein the buffer is
arginine.
14. The formulation according to claim 5, wherein the buffer is
sodium dihydrogen phosphate.
15. The formulation according to claim 5, wherein the buffer is
disodium hydrogen phosphate.
16. The formulation according to claim 5, wherein the buffer is
sodium phosphate.
17. The formulation according to claim 5, wherein the buffer is
tris(hydroxymethyl)-aminomethane.
18. The formulation according to claim 5, wherein the buffer is
selected from the group consisting of sodium acetate, sodium
carbonate, citrate, glycylglycine, histidine, glycine, lysine,
arginine, sodium dihydrogen phosphate, disodium hydrogen phosphate,
sodium phosphate, tris(hydroxymethyl)-aminomethane, or mixtures
thereof.
19. A pharmaceutical formulation consisting essentially of: Arg34,
Lys26(N-.epsilon.-(.gamma.-Glu(N-.alpha.-hexadecanoyl)))-GLP-1(7-37)
in a concentration of about 0.1 mg/ml to about 10 mg/ml; propylene
glycol in a concentration of from about 13 mg/ml to about 15 mg/ml;
a buffer, a preservative; and water for injection; wherein said
formulation has a pH in the range of from about 7.0 to about
9.5.
20. The formulation according to claim 19, wherein the buffer is
selected from the group consisting of sodium acetate, sodium
carbonate, citrate, glycylglycine, histidine, glycine, lysine,
arginine, sodium dihydrogen phosphate, disodium hydrogen phosphate,
sodium phosphate, tris(hydroxymethyl)-aminomethane, or mixtures
thereof, and wherein the preservative is selected from the group
consisting of phenol, m-cresol, methyl p-hydroxybenzoate, propyl
p-hydroxybenzoate, 2-phenoxyethanol, butyl p-hydroxybenzoate,
2phenylethanol, benzyl alcohol, chlorobutanol, and thimerosal, or
mixtures thereof.
21. The formulation according to claim 20, wherein the buffer is
glycylglycine, sodium dihydrogen phosphate, disodium hydrogen
phosphate, sodium phosphate or mixtures thereof, and the
preservative is phenol or m-cresol.
22. The formulation according to claim 21, wherein said formulation
has a pH of 8.15.
23. A pharmaceutical formulation for use in an injection device
containing a needle, the formulation comprising: Arg34,
Lys26(N-.epsilon.-(.gamma.-Glu(N-.alpha.-hexadecanoyl)))-GLP-1(7-37)
in a concentration of about 0.1 mg/ml to about 10 mg/ml; and a
propylene glycol isotonic agent in a concentration of from about 13
mg/ml to about 15 mg/ml; wherein said formulation has a pH of from
about 7.0 to about 9.5; and wherein said formulation exhibits, in a
simulated in-use study, reduced clogging of the needle relative to
a formulation of the same components but containing mannitol as the
isotonic agent.
24. The formulation of claim 23, wherein said formulation exhibits
reduced deposits in production filling equipment relative to a
formulation of the same components but containing mannitol as the
isotonic agent.
25. The formulation according to claim 23, wherein the propylene
glycol is present in said formulation in a concentration of from
about 13.5 mg/ml to about 14.5 mg/ml.
26. The formulation according to claim 23, further comprising a
buffer and a preservative.
27. The formulation according to claim 23, wherein said formulation
has a pH of 8.15.
28. A parenteral administration device for administering Arg34,
Lys26(N-.epsilon.-(.gamma.-Glu(N-.alpha.-hexadecanoyl)))-GLP-1(7-37)
to a patient in need thereof, the device comprising: an injection
device containing a needle with a needle top; a pharmaceutical
formulation in the injection device, said formulation comprising:
Arg34,
Lys26(N-.epsilon.-(.gamma.-Glu(N-.alpha.-hexadecanoyl)))-GLP-1(7-37)
in a concentration of about 0.1 mg/ml to about 10 mg/ml; and a
propylene glycol isotonic agent in a concentration of from about 13
mg/ml to about 15 mg/ml; wherein said formulation has a pH of from
about 7.0 to about 9.5; and wherein the device, in a simulated
in-use study, after multiple uses exhibits no clogging of the
needle, no drops at the needle top, no dried drops at the needle
top, no gel-like drops on the needle, and no deposits on the
needle.
29. The parenteral administration device of claim 28, wherein the
propylene glycol is present in said formulation in a concentration
of from about 13.5 mg/ml to about 14.5 mg/ml.
30. The parenteral administration device of claim 28, wherein said
formulation further comprises a buffer and a preservative.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This Application is a continuation of U.S. application Ser.
No. 16/260,204 filed Jan. 29, 2019, which is a continuation of U.S.
application Ser. No. 16/050,827, filed Jul. 31, 2018 (now
abandoned), which is a continuation of U.S. application Ser. No.
13/362,745, filed Jan. 31, 2012 (now abandoned), which is a
continuation of U.S. application Ser. No. 11/435,977, filed May 17,
2006 (U.S. Pat. No. 8,114,833, issued Feb. 14, 2012), which is a
continuation of International Application serial no.
PCT/DK2004/000792 filed Nov. 18, 2004, which claims priority from
Danish Application serial no. PA 2003 01719, filed Nov. 20, 2003;
the contents of which are incorporated herein by reference.
SEQUENCE LISTING
[0002] The instant application contains a Sequence Listing which
has been submitted in ASCII format via EFS-Web and is hereby
incorporated by reference in its entirety. Said ASCII copy, created
on Jun. 11, 2020, is named 6683US04_SeqList.txt and is 3 kilobytes
in size.
FIELD OF THE INVENTION
[0003] The present invention relates to pharmaceutical formulations
comprising a peptide and propylene glycol, to methods of preparing
such formulations, and to uses of such formulations in the
treatment of diseases and conditions for which use of the peptide
contained in such formulations is indicated. The present invention
further relates to methods for reducing the clogging of injection
devices by a peptide formulation and for reducing deposits on
production equipment during production of a peptide
formulation.
BACKGROUND OF THE INVENTION
[0004] The inclusion of isotonicity agents in peptide-containing
pharmaceutical formulations is widely known and one of the more
common isotonic agents used in such formulations is mannitol.
However, the present inventors have observed that mannitol causes
problems during the production of peptide formulations as it
crystallizes resulting in deposits in the production equipment and
in the final product. Such deposits increase the need to clean the
filling equipment during production of the formulation and this
results in reduced production capability. In addition, such
deposits may also result in reduced yield of the final product
since vials/cartridges containing the peptide formulation may need
to be discarded if particles are present. Finally, the present
inventors have observed that in peptide formulations to be
administered by injection, the presence of mannitol results in
clogging of injection devices.
[0005] Accordingly, it is desirable to identify an alternative
isotonic agent to mannitol for inclusion in peptide-containing
formulations and in particular, for inclusion in peptide
formulations which are administered by injection.
SUMMARY OF THE INVENTION
[0006] The present inventors have discovered that peptide
formulations containing propylene glycol at certain concentrations
exhibit reduced deposits in production equipment and in the final
product and also exhibit reduced clogging of injection devices. The
present compositions may be formulated with any peptide and are
also physically and chemically stable thus rendering them
shelf-stable and suitable for invasive (eg. injection, subcutaneous
injection, intramuscular, intraveneous or infusion) as well as
non-invasive (eg nasal, oral, pulmonary, transdermal or
transmucosal e.g. buccal) means of administration.
[0007] The present invention therefore relates to a pharmaceutical
formulation comprising a peptide and propylene glycol, where the
propylene glycol is present in a concentration of 1-100 mg/ml and
the pH of the formulation is from 7-10. In a preferred embodiment,
the pharmaceutical formulations of the invention further contain a
buffer and a preservative.
[0008] The present invention also relates to methods for producing
the pharmaceutical formulations of the invention. In one
embodiment, the method for preparing a peptide formulation
comprises:
[0009] a) preparing a first solution by dissolving preservative,
propylene glycol and buffer in water;
[0010] b) preparing a second solution by dissolving the peptide in
water;
[0011] c) mixing the first and second solutions; and
[0012] d) adjusting the pH of the mixture in c) to the desired
pH.
[0013] In another embodiment, the method for preparing a peptide
formulation comprises:
[0014] a) preparing a first solution by dissolving preservative and
buffer in water;
[0015] b) adding propylene glycol to the first solution;
[0016] c) mixing the first solution with a second solution
containing peptide dissolved in water; and
[0017] d) adjusting the pH of the mixture in c) to the desired
pH.
[0018] In yet another embodiment, the method for preparing a
peptide formulation comprises:
[0019] a) preparing a solution by dissolving preservative, buffer
and propylene glycol in water;
[0020] b) adding the peptide to the solution of step a); and
[0021] c) adjusting the pH of the solution of step b) to the
desired pH.
[0022] The present invention further relates to methods of
treatment using the pharmaceutical formulations of the invention
where the compositions are administered in an amount effective to
combat the disease, condition, or disorder for which administration
of the peptide contained in the formulation is indicated. In
addition the present invention also relates to a method for
reducing deposits on production equipment during production of a
peptide formulation, where the method comprises replacing the
isotonicity agent previously utilized in said formulation with
propylene glycol at a concentration of between 1-100 mg/ml.
[0023] In one embodiment, the reduction in deposits on the
production equipment during production by the propylene
glycol-containing formulation relative to that observed for the
formulation containing the previously utilized isotonicity agent is
measured by a simulated filling experiment.
[0024] The present invention also relates to a method for reducing
deposits in the final product during production of a peptide
formulation, where the method comprises replacing the isotonicity
agent previously utilized in said formulation with propylene glycol
at a concentration of between 1-100 mg/ml.
[0025] In one embodiment, the reduction in deposits in the final
product is measured by a reduction in the number of vials and/or
cartridges of the propylene glycol-containing formulation that must
be discarded due to deposits relative to number of vials and/or
cartridges of the formulation containing the previously utilized
isotonicity agent that must be discarded due to deposits.
[0026] The present invention further relates to a method for
reducing the clogging of injection devices by a peptide
formulation, where the method comprises replacing the isotonicity
agent previously utilized in said formulation with propylene glycol
at a concentration of between 1-100 mg/ml.
[0027] In one embodiment, the reduction in clogging of the
injection device by the propylene glycol-containing formulation
relative to that observed for the formulation containing the
previously utilized isotonicity agent is measured in a simulated in
use study.
BRIEF DESCRIPTION OF THE FIGURES
[0028] FIG. 1 shows a photograph of dried droplets on microscope
slides of from left to right, placebo (no peptide) formulations
containing no isotonic agent (e only water, preservative and
buffer), mannitol, sorbitol, xylitol, sucrose or glycerol as the
isotonic agent with the far right slide containing mannitol with
peptide Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37).
[0029] FIG. 2 shows light microscopy pictures of from left to
right, some of the dried droplets of placebo formulations
containing mannitol, arginin, inositol or glycerol as the isotonic
agent.
[0030] FIG. 3 shows light microscopy pictures of clogged needles
dosed with placebo formulations containing myoinositol, maltose or
glycerol as the isotonic agent.
[0031] FIG. 4 shows light microscopy pictures of deposits on
needles dosed with placebo formulations containing glycine, lactose
or mannitol as the isotonic agent.
[0032] FIG. 5 shows filling equipment after 24 hours simulated
filling with Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.hexadecanoyl)))-GLP--
1(7-37) medium containing myo-inositol.
[0033] FIG. 6 shows deposits on filling equipment after 24 hours
simulated filling with a mannitol-containing placebo
formulation.
[0034] FIG. 7 shows deposits on needles dosed with mannitol (top
panel) and propylene glycol (bottom panel)-containing Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.hexadecanoyl)))-GLP--
1(7-37) formulations.
DESCRIPTION OF THE INVENTION
[0035] The present invention relates to a pharmaceutical
formulation comprising a peptide or a mixture of peptides and
propylene glycol where the final concentration of propylene glycol
in the formulation is 1-100 mg/ml and the pH of the formulation is
in the range of from 7-10.
[0036] The pharmaceutical formulations of the invention are found
to be optimal for production because they exhibit reduced deposits
in production equipment relative to formulations containing other
isotonicity agents as measured by the simulated filling studies
described in the Examples. In addition, the pharmaceutical
formulations of the invention are found to be optimal for use in
injection devices because they exhibit reduced clogging of the
injection devices relative to formulations containing other
isotonicity agents as measured by the simulated in use studies
described in the Examples.
[0037] The formulations of the present invention may be formulated
with any peptide where examples of such peptides include, but are
not limited to, glucagon, human growth hormone (hGH), insulin,
aprotinin, FactorVll, tissue plasminogen activator (TPA),
FactorVIIa, FFRFactorVIIa, heparinase, ACTH, Heparin Binding
Protein, corticotropin-releasing factor, angiotensin, calcitonin,
glucagon-like peptide-1, glucagon-like peptide-2, insulin-like
growth factor-1, insulin-like growth factor-2, fibroblast growth
factors, gastric inhibitory peptide, growth hormone-releasing
factor, pituitary adenylate cyclase activating peptide, secretin,
enterogastrin, somatostatin, somatomedin, parathyroid hormone,
thrombopoietin, erythropoietin, hypothalamic releasing factors,
prolactin, thyroid stimulating hormones, endorphins, enkephalins,
vasopressin, oxytocin, opiods, DPP IV, interleukins,
immunoglobulins, complement inhibitors, serine protease inhibitors,
cytokines, cytokine receptors, PDGF, tumor necrosis factors, tumor
necrosis factors receptors, growth factors and analogues as well as
derivatives thereof where each of these peptides constitutes an
alternative embodiment of the present invention.
[0038] In the present application, the designation "an analogue" is
used to designate a peptide wherein one or more amino acid residues
of the parent peptide have been substituted by another amino acid
residue and/or wherein one or more amino acid residues of the
parent peptide have been deleted and/or wherein one or more amino
acid residues have been added to the parent peptide. Such addition
can take place either at the N-terminal end or at the C-terminal
end of the parent peptide or both. Typically "an analogue" is a
peptide wherein 6 or less amino acids have been substituted and/or
added and/or deleted from the parent peptide, more preferably a
peptide wherein 3 or less amino acids have been substituted and/or
added and/or deleted from the parent peptide, and most preferably,
a peptide wherein one amino acid has been substituted and/or added
and/or deleted from the parent peptide.
[0039] In the present application, "a derivative" is used to
designate a peptide or analogue thereof which is chemically
modified by introducing an organic substituent e.g. ester, alkyl or
lipophilic functionalities, on one or more amino acid residues of
the peptide or analogue thereof.
[0040] In one embodiment, the peptide to be included in the
formulation of the invention is a GLP-1 agonist where "a GLP-1
agonist" is understood to refer to any peptide which fully or
partially activates the human GLP-1 receptor. In a preferred
embodiment, the "GLP-1 agonist" is any peptide that binds to a
GLP-1 receptor, preferably with an affinity constant (K.sub.D) or a
potency (EC.sub.50) of below 1 .mu.M, e.g. below 100 nM as measured
by methods known in the art (see e.g. WO 98/08871) and exhibits
insulinotropic activity, where insulinotropic activity may be
measured in vivo or in vitro assays known to those of ordinary
skill in the art. For example, the GLP-1 agonist may be
administered to an animal and the insulin concentration measured
over time.
[0041] Methods for identifying GLP-1 agonists are described in WO
93/19175 (Novo Nordisk A/S) and examples of suitable GLP-1
analogues and derivatives which can be used according to the
present invention includes those referred to in WO 99/43705 (Novo
Nordisk A/S), WO 99/43706 (Novo Nordisk A/S), WO 99/43707 (Novo
Nordisk A/S), WO 98/08871 (analogues with lipophilic substituent)
and in WO 02/46227 (analogues fused to serum albumin or to Fc
portion of an Ig).(Novo Nordisk A/S), WO 99/43708 (Novo Nordisk
A/S), WO 99/43341 (Novo Nordisk A/S), WO 87/06941 (The General
Hospital Corporation), WO 90/11296 (The General Hospital
Corporation), WO 91/11457 (Buckley et al.), WO 98/43658 (Eli Lilly
& Co.), EP 0708179-A2 (Eli Lilly & Co.), EP 0699686-A2 (Eli
Lilly & Co.), WO 01/98331 (Eli Lilly & Co).
[0042] In one embodiment, the GLP-1 agonist is selected from the
group consisting of GLP-1(7-36)-amide, GLP-1(7-37), a
GLP-1(7-36)-amide analogue, a GLP-1(7-37) analogue, or a derivative
of any of these.
[0043] In one embodiment, the GLP-1 agonist is a derivative of
GLP-1(7-36)-amide, GLP-1(7-37), a GLP-1(7-36)-amide analogue or a
GLP-1(7-37) analogue, which comprises a lipophilic substituent.
[0044] In this embodiment of the invention, the GLP-1 derivative
preferably has three lipophilic substituents, more preferably two
lipophilic substituents, and most preferably one lipophilic
substituent attached to the parent peptide (ie GLP-1(7-36)-amide,
GLP-1(7-37), a GLP-1(7-36)-amide analogue or a GLP-1(7-37)
analogue), where each lipophilic substituent(s) preferably has 4-40
carbon atoms, more preferably 8-30 carbon atoms, even more
preferably 8-25 carbon atoms, even more preferably 12-25 carbon
atoms, and most preferably 14-18 carbon atoms.
[0045] In one embodiment, the lipophilic substituent comprises a
partially or completely hydrogenated cyclopentanophenathrene
skeleton.
[0046] In another embodiment, the lipophilic substituent is a
straight-chain or branched alkyl group.
[0047] In yet another embodiment, the lipophilic substituent is an
acyl group of a straight-chain or branched fatty acid. Preferably,
the lipophilic substituent is an acyl group having the formula
CH.sub.3(CH.sub.2).sub.nCO--, wherein n is an integer from 4 to 38,
preferably an integer from 12 to 38, and most preferably is
CH.sub.3(CH.sub.2).sub.12CO--, CH.sub.3(CH.sub.2).sub.14CO--,
CH.sub.3(CH.sub.2).sub.16CO--, CH.sub.3(CH.sub.2).sub.18CO--,
CH.sub.3(CH.sub.2).sub.20CO-- and CH.sub.3(CH.sub.2).sub.22CO--. In
a more preferred embodiment, the lipophilic substituent is
tetradecanoyl. In a most preferred embodiment, the lipophilic
substituent is hexadecanoyl.
[0048] In a further embodiment of the present invention, the
lipophilic substituent has a group which is negatively charged such
as a carboxylic acid group. For example, the lipophilic substituent
may be an acyl group of a straight-chain or branched alkane
.alpha.,.omega.-dicarboxylic acid of the formula
HOOC(CH.sub.2).sub.mCO--, wherein m is an integer from 4 to 38,
preferably an integer from 12 to 38, and most preferably is
HOOC(CH.sub.2).sub.14CO--, HOOC(CH.sub.2).sub.16CO--,
HOOC(CH.sub.2).sub.18CO--, HOOC(CH.sub.2).sub.20CO-- or
HOOC(CH.sub.2).sub.22CO--.
[0049] In the GLP-1 derivatives of the invention, the lipophilic
substituent(s) contain a functional group which can be attached to
one of the following functional groups of an amino acid of the
parent GLP-1 peptide:
[0050] (a) the amino group attached to the alpha-carbon of the
N-terminal amino acid,
[0051] (b) the carboxy group attached to the alpha-carbon of the
C-terminal amino acid,
[0052] (c) the epsilon-amino group of any Lys residue,
[0053] (d) the carboxy group of the R group of any Asp and Glu
residue,
[0054] (e) the hydroxy group of the R group of any Tyr, Ser and Thr
residue,
[0055] (f) the amino group of the R group of any Trp, Asn, Gln,
Arg, and His residue, or
[0056] (g) the thiol group of the R group of any Cys residue.
[0057] In one embodiment, a lipophilic substituent is attached to
the carboxy group of the R group of any Asp and Glu residue.
[0058] In another embodiment, a lipophilic substituent is attached
to the carboxy group attached to the alpha-carbon of the C-terminal
amino acid.
[0059] In a most preferred embodiment, a lipophilic substituent is
attached to the epsilon-amino group of any Lys residue.
[0060] In a preferred embodiment of the invention, the lipophilic
substituent is attached to the parent GLP-1 peptide by means of a
spacer. A spacer must contain at least two functional groups, one
to attach to a functional group of the lipophilic substituent and
the other to a functional group of the parent GLP-1 peptide.
[0061] In one embodiment, the spacer is an amino acid residue
except Cys or Met, or a dipeptide such as Gly-Lys. For purposes of
the present invention, the phrase "a dipeptide such as Gly-Lys"
means any combination of two amino acids except Cys or Met,
preferably a dipeptide wherein the C-terminal amino acid residue is
Lys, His or Trp, preferably Lys, and the
[0062] N-terminal amino acid residue is Ala, Arg, Asp, Asn, Gly,
Glu, Gln, Ile, Leu, Val, Phe, Pro, Ser, Tyr, Thr, Lys, His and Trp.
Preferably, an amino group of the parent peptide forms an amide
bond with a carboxylic group of the amino acid residue or dipeptide
spacer, and an amino group of the amino acid residue or dipeptide
spacer forms an amide bond with a carboxyl group of the lipophilic
substituent.
[0063] Preferred spacers are lysyl, glutamyl, asparagyl, glycyl,
beta-alanyl and gamma-aminobutanoyl, each of which constitutes an
individual embodiment. Most preferred spacers are glutamyl and
beta-alanyl. When the spacer is Lys, Glu or Asp, the carboxyl group
thereof may form an amide bond with an amino group of the amino
acid residue, and the amino group thereof may form an amide bond
with a carboxyl group of the lipophilic substituent. When Lys is
used as the spacer, a further spacer may in some instances be
inserted between the .epsilon.-amino group of Lys and the
lipophilic substituent. In one embodiment, such a further spacer is
succinic acid which forms an amide bond with the .epsilon.-amino
group of Lys and with an amino group present in the lipophilic
substituent. In another embodiment such a further spacer is Glu or
Asp which forms an amide bond with the a-amino group of Lys and
another amide bond with a carboxyl group present in the lipophilic
substituent, that is, the lipophilic substituent is a
N.sup..epsilon.-acylated lysine residue.
[0064] In another embodiment, the spacer is an unbranched alkane
.alpha.,.omega.-dicarboxylic acid group having from 1 to 7
methylene groups, which spacer forms a bridge between an amino
group of the parent peptide and an amino group of the lipophilic
substituent. Preferably, the spacer is succinic acid.
[0065] In a further embodiment, the lipophilic substituent with the
attached spacer is a group of the formula
CH.sub.3(CH.sub.2).sub.pNH--CO(CH.sub.2).sub.qCO--, wherein p is an
integer from 8 to 33, preferably from 12 to 28 and q is an integer
from 1 to 6, preferably 2.
[0066] In a further embodiment, the lipophilic substituent with the
attached spacer is a group of the formula
CH.sub.3(CH.sub.2).sub.rCO--NHCH(COOH)(CH.sub.2).sub.2CO--, wherein
r is an integer from 4 to 24, preferably from 10 to 24.
[0067] In a further embodiment, the lipophilic substituent with the
attached spacer is a group of the formula
CH.sub.3(CH.sub.2).sub.sCO--NHCH((CH.sub.2).sub.2COOH)CO--, wherein
s is an integer from 4 to 24, preferably from 10 to 24.
[0068] In a further embodiment, the lipophilic substituent is a
group of the formula COOH(CH.sub.2).sub.tCO-- wherein t is an
integer from 6 to 24.
[0069] In a further embodiment, the lipophilic substituent with the
attached spacer is a group of the formula
--NHCH(COOH)(CH.sub.2).sub.4NH--CO(CH.sub.2).sub.uCH.sub.3, wherein
u is an integer from 8 to 18.
[0070] In a further embodiment, the lipophilic substituent with the
attached spacer is a group of the formula
CH.sub.3(CH.sub.2).sub.vCO--NH--(CH.sub.2).sub.z--CO, wherein v is
an integer from 4 to 24 and z is an integer from 1 to 6.
[0071] In a further embodiment, the lipophilic substituent with the
attached spacer is a group of the formula
--NHCH(COOH)(CH.sub.2).sub.4NH--COCH((CH.sub.2).sub.2COOH)NH--CO(CH.sub.2-
).sub.wCH.sub.3, wherein w is an integer from 10 to 16.
[0072] In a further embodiment, the lipophilic substituent with the
attached spacer is a group of the formula
--NHCH(COOH)(CH.sub.2).sub.4NH--CO(CH.sub.2).sub.2CH(COOH)NHCO(CH.sub.2).-
sub.xCH.sub.3, wherein x is zero or an integer from 1 to 22,
preferably 10 to 16.
[0073] In yet another embodiment the GLP-1 agonist is Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37).
[0074] In yet another embodiment the GLP-1 agonist is selected from
the group consisting of Gly.sup.8-GLP-1(7-36)-amide,
Gly.sup.8-GLP-1(7-37), Val.sup.8-GLP-1(7-36)-amide,
Val.sup.8-GLP-1(7-37), Val.sup.8Asp.sup.22-GLP-1(7-36)-amide,
Val.sup.8Asp.sup.22-GLP-1(7-37) ,
Val.sup.8Glu.sup.22-GLP-1(7-36)-amide ,
Val.sup.8Glu.sup.22-GLP-1(7-37),
Val.sup.8Lys.sup.22-GLP-1(7-36)-amide,
Val.sup.8Lys.sup.22-GLP-1(7-37),
Val.sup.8Arg.sup.22-GLP-1(7-36)-amide,
Val.sup.8Arg.sup.22-GLP-1(7-37),
Val.sup.8His.sup.22-GLP-1(7-36)-amide,
Val.sup.8His.sup.22-GLP-1(7-37), analogues thereof and derivatives
of any of these.
[0075] In yet another embodiment the GLP-1 agonist is selected from
the group consisting of Arg.sup.26-GLP-1(7-37); Are-GLP-1(7-37);
Lys.sup.36-GLP-1(7-37); Arg.sup.26,34Lys.sup.36-GLP-1(7-37);
Arg.sup.26,34-GLP-1(7-37); Arg.sup.26,34Lys.sup.40-GLP-1(7-37);
Arg.sup.26Lys.sup.36-GLP-1(7-37); Arg.sup.34Lys.sup.36-GLP-1(7-37);
Val.sup.8Arg.sup.22-GLP-1(7-37); Met.sup.8Arg.sup.22-GLP-1(7-37);
Gly.sup.8His.sup.22-GLP-1(7-37); Val.sup.8His.sup.22-GLP-1(7-37);
Met.sup.8His.sup.22-GLP-1(7-37); His.sup.37-GLP-1(7-37);
Gly.sup.8-GLP-1(7-37); Val.sup.8-GLP-1(7-37);
Met.sup.8-GLP-1(7-37);Gly.sup.8Asp.sup.22-GLP-1(7-37);
Val.sup.8Asp.sup.22-GLP-1(7-37);
Met.sup.8Asp.sup.22-GLP-1(7-37);Gly.sup.8Glu.sup.22-GLP-1(7-37);
Val.sup.8Glu.sup.22-GLP-1(7-37); Met.sup.8Glu.sup.22-GLP-1(7-37);
Gly.sup.8Lys.sup.22-GLP-1(7-37); Val.sup.8Lys.sup.22-GLP-1(7-37);
Met.sup.8Lys.sup.22-GLP-1(7-37); Gly.sup.8Arg.sup.22-GLP-1(7-37);
Val.sup.8Lys.sup.22His.sup.37-GLP-1(7-37);
Gly.sup.8Glu.sup.22His.sup.37-GLP-1(7-37);
Val.sup.8Glu.sup.22His.sup.37-GLP-1(7-37);
Met.sup.8Glu.sup.22His.sup.37-GLP-1(7-37);Gly.sup.8Lys.sup.22
His.sup.37-GLP-1(7-37);
Met.sup.8Lys.sup.22His.sup.37-GLP-1(7-37);Gly.sup.8Arg.sup.22His.sup.37-G-
LP-1(7-37); Val.sup.8Arg.sup.22His.sup.37-GLP-1(7-37);
Met.sup.8Arg.sup.22His.sup.37-GLP-1(7-37);
Gly.sup.8His.sup.22His.sup.37-GLP-1(7-37);
Val.sup.8His.sup.22His.sup.37-GLP-1(7-37);
Met.sup.8His.sup.22His.sup.37-GLP-1(7-37);
Gly.sup.8His.sup.37-GLP-1(7-37); Val.sup.8His.sup.37-GLP-1(7-37);
Met.sup.8His.sup.37-GLP-1(7-37);Gly.sup.8Asp.sup.22
His.sup.37-GLP-1(7-37); Val.sup.8Asp.sup.22His.sup.37-GLP-1(7-37);
Met.sup.8Asp.sup.22His.sup.37-GLP-1(7-37);
Arg.sup.26-GLP-1(7-36)-amide; Arg.sup.34-GLP-1(7-36)-amide;
Lys.sup.36-GLP-1(7-36)-amide;
Arg.sup.26,34Lys.sup.36-GLP-1(7-36)-amide;
Arg.sup.26,34-GLP-1(7-36)-amide;
Arg.sup.26,34Lys.sup.40-GLP-1(7-36)-amide;
Arg.sup.26Lys.sup.36-GLP-1(7-36)-amide;
Arg.sup.34Lys.sup.36-GLP-1(7-36)-amide;
Gly.sup.8-GLP-1(7-36)-amide; Val.sup.8-GLP-1(7-36)-amide;
Met.sup.8-GLP-1(7-36)-amide; Gly.sup.8Asp.sup.22-GLP-1(7-36)-amide;
Gly.sup.8Glu.sup.22His.sup.37-GLP-1(7-36)-amide;
Val.sup.8Asp.sup.22-GLP-1(7-36)-amide;
Met.sup.8Asp.sup.22-GLP-1(7-36)-amide;
Gly.sup.8Glu.sup.22-GLP-1(7-36)-amide;
Val.sup.8Glu.sup.22-GLP-1(7-36)-amide;
Met.sup.8Glu.sup.22-GLP-1(7-36)-amide;
Gly.sup.8Lys.sup.22-GLP-1(7-36)-amide;
Val.sup.8Lys.sup.22-GLP-1(7-36)-amide;
Met.sup.8Lys.sup.22-GLP-1(7-36)-amide;
Gly.sup.8His.sup.22His.sup.37-GLP-1(7-36)-amide;
Gly.sup.8Arg.sup.22-GLP-1(7-36)-amide;
Val.sup.8Arg.sup.22-GLP-1(7-36)-amide;
Met.sup.8Arg.sup.22-GLP-1(7-36)-amide;
Gly.sup.8His.sup.22-GLP-1(7-36)-amide;
Val.sup.8His.sup.22-GLP-1(7-36)-amide;
Met.sup.8His.sup.22-GLP-1(7-36)-amide;
His.sup.37-GLP-1(7-36)-amide;
Val.sup.8Arg.sup.22His.sup.37-GLP-1(7-36)-amide;
Met.sup.8Arg.sup.22His.sup.37-GLP-1(7-36)-amide;
Gly.sup.8His.sup.37-GLP-1(7-36)-amide;
Val.sup.8His.sup.37-GLP-1(7-36)-amide; Met8His37-GLP-1(7-36)-amide;
Gly.sup.8Asp.sup.22 His.sup.37-GLP-1(7-36)-amide;
Val.sup.8Asp.sup.22His.sup.37-GLP-1(7-36)-amide;
Met.sup.8Asp.sup.22His.sup.37-GLP-1(7-36)-amide;
Val.sup.8Glu.sup.22His.sup.37-GLP-1(7-36)-amide;
Met.sup.8Glu.sup.22His.sup.37-GLP-1(7-36)-amide;
Gly.sup.8Lys.sup.22 His.sup.37-GLP-1(7-36)-amide;
Val.sup.8Lys.sup.22His.sup.37-GLP-1(7-36)-amide;
Met.sup.8Lys.sup.22His.sup.37-GLP-1(7-36)-amide;
Gly.sup.8Arg.sup.22His.sup.37-GLP-1(7-36)-amide;
Val.sup.8His.sup.22His.sup.37-GLP-1(7-36)-amide;
Met.sup.8His.sup.22His.sup.37-GLP-1(7-36)-amide; and derivatives
thereof.
[0076] In yet another embodiment the GLP-1 agonist is selected from
the group consisting of Val.sup.8Trp.sup.19Glu.sup.22-GLP-1(7-37),
Val.sup.8Glu.sup.22Val.sup.25-GLP-1(7-37),
Val.sup.8Tyr.sup.16Glu.sup.22-GLP-1(7-37),
Val.sup.8Trp.sup.16Glu.sup.22-GLP-1(7-37),
Val.sup.8Leu.sup.16Glu.sup.22-GLP-1(7-37),
Val.sup.8Tyr.sup.18Glu.sup.22-GLP-1(7-37),
Val.sup.8Glu.sup.22His.sup.37-GLP-1(7-37),
Val.sup.8Glu.sup.22Ile.sup.33-GLP-1(7-37),
Val.sup.8Trp.sup.16Glu.sup.22Val.sup.25Ile.sup.33-GLP-1(7-37),
Val.sup.8Trp.sup.16Glu.sup.22Ile.sup.33-GLP-1(7-37),
Val.sup.8Glu.sup.22Val.sup.25Ile.sup.33-GLP-1(7-37),
Val.sup.8Trp.sup.16Glu.sup.22Val.sup.25-GLP-1(7-37), analogues
thereof and derivatives of any of these.
[0077] In yet another embodiment the GLP-1 agonist is exendin-4 or
exendin-3, an exen-din-4 or exendin-3 analogue or a derivative of
any of these.
[0078] Examples of exendins as well as analogues, derivatives, and
fragments thereof to be included within the present invention are
those disclosed in WO 97/46584, U.S. Pat. No. 5,424,286 and WO
01/04156. U.S. Pat. No. 5,424,286 describes a method for
stimulating insulin release with an exendin polypeptide. The
exendin polypeptides disclosed include
HGEGTFTSDLSKQMEEEAVRLFIEWLKNGGX (SEQ ID NO.: 1); wherein X=P or Y;
HSDGTFITSDLSKQMEEEAVRLFIEWLKNGGPSSGAPPPS (SEQ ID NO.: 2)
(exendin-3); and HGEGT-FITSDLSKQMEEEAVRLFIEWLKNGGPSSGAPPPS (SEQ ID
NO.: 4) (exendin-4). WO 97/46584 describes truncated versions of
exendin peptide(s). The disclosed peptides increase secretion and
biosynthesis of insulin, but reduce those of glucagon. WO 01/04156
describes exendin-4 analogues and derivatives as well as the
preparation of these molecules. Exendin-4 analogues stabilized by
fusion to serum albumin or Fc portion of an Ig are disclosed in WO
02/46227.
[0079] In one embodiment, the exendin-4 analogue is
HGEGTFTSDLSKQMEEEAVRL-FIEWLKNGGPSSGAPPSKKKKKK-amide (SEQ ID NO.:
3).
[0080] Where the peptide to be included in the formulation of the
invention is a GLP-1 agonist, the GLP-1 agonist is present in a
concentration from about 0.1 mg/ml to about 100 mg/ml, more
preferably in a concentration from about 0.1 mg/ml to about 50
mg/ml, and most preferably in a concentration of from about 0.1
mg/ml to about 10 mg/ml.
[0081] In another embodiment, the peptide to be included in the
formulation of the invention is insulin , where "insulin" is
understood to mean human insulin, [where "human insulin" means
insulin having the amino acid sequence shown in D S H W Nicol and L
F Smith: Nature, (1960) 4736: 483-485, which is hereby incorporated
by reference], human insulin analogs, human insulin derivatives or
mixtures thereof, where examples of insulin analogs and derivatives
are those disclosed in EP 0 792 290 (Novo Nordisk A/S), EP 0 214
826 and EP 0 705 275 (Novo Nordisk A/S), US 5,504,188 (Eli Lilly),
EP 0 368 187 (Aventis), U.S. Pat. Nos. 5,750,497 and 6,011,007, EP
375437 and EP 383472 and where such insulins may include, but are
not limited to, NPH insulin, Lys .beta.29
(N.epsilon.-tetradecanoyl) des(B30) human insulin,
LysB.sup.29-(N.sup..epsilon.-(.gamma.-glutamyl-N.sup..alpha.-lithocholyl)
des(B30) human insulin, N.sup..quadrature.B29-octanoyl insulin,
30/70 mixtures of prompt insulin zinc (SemiLente.RTM.) with
extended insulin zinc (Ultralente.RTM.), sold commercially as
Lente.RTM., insulin glargine (Lantus.RTM.) or extended insulin zinc
(Ultralente.RTM.), Lys.sup.B28 Pro.sup.B29 human insulin
(Hu-malog.RTM.) , Asp.sup.B28 human insulin, insulin aspart
(Novolog.RTM.), or a 30/70 mixture of insulin aspart and insulin
aspart protamine (NovoMix.RTM.) .
[0082] In one embodiment, the insulin is a derivative of human
insulin or a human insulin analogue where the derivative contains
at least one lysine residue and a lipophilic substituent is
attached to the epsilon amino group of the lysine residue.
[0083] In one embodiment, the lysine residue to which the
lipophilic substituent is attached is present at position B28 of
the insulin peptide.
[0084] In an alternative embodiment, the lysine residue to which
the lipophilic substituent is attached is present at position B29
of the insulin peptide.
[0085] In yet another embodiment, lipophilic substituent is an acyl
group corresponding to a carboxylic acid having at least 6 carbon
atoms.
[0086] In another preferred embodiment, the lipophilic substituent
is an acyl group, branched or unbranched, which corresponds to a
carboxylic acid having a chain of carbon atoms 8 to 24 atoms
long.
[0087] In another preferred embodiment, the lipophilic substituent
is an acyl group corresponding to a fatty acid having at least 6
carbon atoms.
[0088] In another preferred embodiment, the lipophilic substituent
is an acyl group corresponding to a linear, saturated carboxylic
acid having from 6 to 24 carbon atoms.
[0089] In another preferred embodiment, the lipophilic substituent
is an acyl group corresponding to a linear, saturated carboxylic
acid having from 8 to 12 carbon atoms.
[0090] In another preferred embodiment, the lipophilic substituent
is an acyl group corresponding to a linear, saturated carboxylic
acid having from 10 to 16 carbon atoms. In another preferred
embodiment, the lipophilic substituent is an oligo oxyethylene
group comprising up to 10, preferably up to 5, oxyethylene
units.
[0091] In another preferred embodiment, the lipophilic substituent
is an oligo oxypropylene group comprising up to 10, preferably up
to 5, oxypropylene units.
[0092] In one preferred embodiment, the invention relates to a
human insulin derivative in which the B30 amino acid residue is
deleted or is any amino acid residue which can be coded for by the
genetic code except Lys, Arg and Cys; the A21 and the B3 amino acid
residues are, independently, any amino acid residues which can be
coded for by the genetic code except Lys, Arg and Cys; Phe.sup.B1
may be deleted; the .quadrature.-amino group of Lys.sup.B29 has a
lipophilic substituent which comprises at least 6 carbon atoms; and
2-4 Zn.sup.2+ ions may be bound to each insulin hexamer with the
proviso that when B30 is Thr or Ala and A21 and B3 are both Asn,
and Phe.sup.B1 is not deleted, then 2-4 Zn.sup.2+ ions are bound to
each hexamer of the insulin derivative.
[0093] In another preferred embodiment, the invention relates to a
human insulin derivative in which the B30 amino acid residue is
deleted or is any amino acid residue which can be coded for by the
genetic code except Lys, Arg and Cys; the A21 and the B3 amino acid
residues are, independently, any amino acid residues which can be
coded for by the genetic code except Lys, Arg and Cys, with the
proviso that if the B30 amino acid residue is Ala or Thr, then at
least one of the residues A21 and B3 is different from Asn;
Phe.sup.B1 may be deleted; and the .quadrature.-amino group of
Lys.sup.B29 has a lipophilic substituent which comprises at least 6
carbon atoms.
[0094] In another preferred embodiment, the invention relates to a
human insulin derivative in which the B30 amino acid residue is
deleted or is any amino acid residue which can be coded for by the
genetic code except Lys, Arg and Cys; the A21 and the B3 amino acid
residues are, independently, any amino acid residues which can be
coded for by the genetic code except Lys, Arg and Cys; Phe.sup.B1
may be deleted; the .quadrature.-amino group of Lys.sup.B29 has a
lipophilic substituent which comprises at least 6 carbon atoms; and
2-4 Zn.sup.2+ ions are bound to each insulin hexamer.
[0095] Where the peptide to be included in the formulation of the
invention is an insulin, the insulin is present in a concentration
from about 0.5 mg/ml to about 20 mg/ml, more preferably in a
concentration from about 1 mg/ml to about 15 mg/ml.
[0096] In another embodiment, the peptide to be included in the
formulations of the invention is hGH or Met-hGH.
[0097] Where the peptide to be included in the formulation of the
invention is hGH or MethGH, the hGH or Met-hGH is present in a
concentration from about 0.5 mg/ml to about 50 mg/ml, more
preferably in a concentration from about 1 mg/ml to about 10
mg/ml.
[0098] In yet another embodiment, the peptide to be included in the
formulations of the invention is GLP-2 or an analogue or derivative
thereof.
[0099] Where the peptide to be included in the formulation of the
invention is GLP-2 or an analogue or derivative thereof, the GLP-2
or an analogue or derivative thereof is present in a concentration
from about 1 mg/ml to about 100 mg/ml, more preferably in a
concentration from about 1 mg/ml to about 10 mg/ml.
[0100] In yet a further embodiment, the peptide to be included in
the formulations of the invention is Factor VII or Factor VIIa or
an analogue or derivative thereof.
[0101] Where the peptide to be included in the formulation of the
invention is Factor VII or Factor VIIa or an analogue or derivative
thereof, the Factor VII or Factor VIIa or an analogue or derivative
thereof is present in a concentration from about 0.1 mg/ml to about
10 mg/ml, more preferably in a concentration from about 0.5 mg/ml
to about 5 mg/ml.
[0102] In one embodiment, the final concentration of propylene
glycol in the formulations of the invention is from about 1 to
about 50 mg/ml.
[0103] In another embodiment, the final concentration of propylene
glycol in the formulations of the invention is from about 5 to
about 25 mg/ml.
[0104] In yet another embodiment, the final concentration of
propylene glycol in the formulations of the invention is from about
8 to about 16 mg/ml.
[0105] In yet a further embodiment, the final concentration of
propylene glycol in the formulations of the invention is from about
13 to about 15 mg/ml.
[0106] In still another embodiment, the final concentration of
propylene glycol in the formulations of the invention is from about
13.5 to about 14.5 mg/ml.
[0107] In another embodiment of the invention, the formulation has
a pH in the range from about 7.0 to about 9.5 where the term
"about" as used in connection with pH means + or -0.1 pH units from
the stated number.
[0108] In a further embodiment of the invention, the formulation
has a pH in the range from about 7.0 to about 8.0.
[0109] In yet a further embodiment of the invention, the
formulation has a pH in the range from about 7.2 to about 8.0.
[0110] In a further embodiment of the invention, the formulation
has a pH in the range from about 7.0 to about 8.3.
[0111] In yet a further embodiment of the invention, the
formulation has a pH in the range from about 7.3 to about 8.3.
[0112] In a preferred embodiment of the invention, the formulations
contain, in addition to a peptide and propylene glycol, a buffer
and/or a preservative.
[0113] Where a buffer is to be included in the formulations of the
invention, the buffer is selected from the group consisting of
sodium acetate, sodium carbonate, citrate, glycylglycine,
histidine, glycine, lysine, arginin, sodium dihydrogen phosphate,
disodium hydrogen phosphate, sodium phosphate, and
tris(hydroxymethyl)-aminomethan, or mixtures thereof. Each one of
these specific buffers constitutes an alternative embodiment of the
invention. In a preferred embodiment of the invention the buffer is
glycylglycine, sodium dihydrogen phosphate, disodium hydrogen
phosphate, sodium phosphate or mixtures thereof.
[0114] Where a pharmaceutically acceptable preservative is to be
included in the formulations of the invention, the preservative is
selected from the group consisting of phenol, m-cresol, methyl
p-hydroxybenzoate, propyl p-hydroxybenzoate, 2-phenoxyethanol,
butyl p-hydroxybenzoate, 2-phenylethanol, benzyl alcohol,
chlorobutanol, and thiomerosal, or mixtures thereof. Each one of
these specific preservatives constitutes an alternative embodiment
of the invention. In a preferred embodiment of the invention the
preservative is phenol or m-cresol.
[0115] In a further embodiment of the invention the preservative is
present in a concentration from about 0.1 mg/ml to about 50 mg/ml,
more preferably in a concentration from about 0.1 mg/ml to about 25
mg/ml, and most preferably in a concentration from about 0.1 mg/ml
to about 10 mg/ml
[0116] The use of a preservative in pharmaceutical compositions is
well-known to the skilled person. For convenience reference is made
to Remington: The Science and Practice of Pharmacy, 19.sup.th
edition, 1995.
[0117] In a further embodiment of the invention the formulation may
further comprise a chelating agent where the chelating agent may be
selected from salts of ethlenediaminetetraacetic acid (EDTA),
citric acid, and aspartic acid, and mixtures thereof. Each one of
these specific chelating agents constitutes an alternative
embodiment of the invention.
[0118] In a further embodiment of the invention the chelating agent
is present in a concentration from 0.1 mg/ml to 5mg/ml. In a
further embodiment of the invention the chelating agent is present
in a concentration from 0.1 mg/ml to 2mg/ml. In a further
embodiment of the invention the chelating agent is present in a
concentration from 2 mg/ml to 5 mg/ml.
[0119] The use of a chelating agent in pharmaceutical compositions
is well-known to the skilled person. For convenience reference is
made to Remington: The Science and Practice of Pharmacy, 19.sup.th
edition, 1995.
[0120] In a further embodiment of the invention the formulation may
further comprise a stabiliser selected from the group of high
molecular weight polymers or low molecular compounds where such
stabilizers include, but are not limited to, polyethylene glycol
(e.g. PEG 3350), polyvinylalcohol (PVA), polyvinylpyrrolidone,
carboxymethylcellulose, different salts (e.g. sodium chloride),
L-glycine, L-histidine, imidazole, arginine, lysine, isoleucine,
aspartic acid, tryptophan, threonine and mixtures thereof. Each one
of these specific stabilizers constitutes an alternative embodiment
of the invention. In a preferred embodiment of the invention the
stabiliser is selected from the group consisting of L-histidine,
imidazole and arginine.
[0121] In a further embodiment of the invention the high molecular
weight polymer is present in a concentration from 0.1 mg/ml to 50
mg/ml. In a further embodiment of the invention the high molecular
weight polymer is present in a concentration from 0.1 mg/ml to 5
mg/ml. In a further embodiment of the invention the high molecular
weight polymer is present in a concentration from 5 mg/ml to 10
mg/ml. In a further embodiment of the invention the high molecular
weight polymer is present in a concentration from 0 mg/ml to 20
mg/ml. In a further embodiment of the invention the high molecular
weight polymer is present in a concentration from 20 mg/ml to 30
mg/ml. In a further embodiment of the invention the high molecular
weight polymer is present in a concentration from 30 mg/ml to 50
mg/ml.
[0122] In a further embodiment of the invention the low molecular
weight compound is present in a concentration from 0.1 mg/ml to 50
mg/ml. In a further embodiment of the invention the low molecular
weight compound is present in a concentration from 0.1 mg/ml to 5
mg/ml.
[0123] In a further embodiment of the invention the low molecular
weight compound is present in a concentration from 5 mg/ml to 10
mg/ml. In a further embodiment of the invention the low molecular
weight compound is present in a concentration from 10 mg/ml to 20
mg/ml. In a further embodiment of the invention the low molecular
weight compound is present in a concentration from 20 mg/ml to 30
mg/ml. In a further embodiment of the invention the low molecular
weight compound is present in a concentration from 30 mg/ml to 50
mg/ml.
[0124] The use of a stabilizer in pharmaceutical compositions is
well-known to the skilled person. For convenience reference is made
to Remington: The Science and Practice of Pharmacy, 19.sup.th
edition, 1995.
[0125] In a further embodiment of the invention the formulation of
the invention may further comprise a surfactant where a surfactant
may be selected from a detergent, ethoxylated castor oil,
polyglycolyzed glycerides, acetylated monoglycerides, sorbitan
fatty acid esters, poloxamers, such as 188 and 407, polyoxyethylene
sorbitan fatty acid esters, polyoxyethylene derivatives such as
alkylated and alkoxylated derivatives (tweens, e.g. Tween-20, or
Tween-80), monoglycerides or ethoxylated derivatives thereof,
diglycerides or polyoxyethylene derivatives thereof, glycerol,
cholic acid or derivatives thereof, lecithins, alcohols and
phospholipids, glycerophospholipids (lecithins, kephalins,
phosphatidyl serine), glyceroglycolipids (galactopyransoide),
sphingophospholipids (sphingomyelin), and sphingoglycolipids
(ceramides, gangliosides), DSS (docusate sodium, docusate calcium,
docusate potassium, SDS (sodium dodecyl sulfate or sodium lauryl
sulfate), dipalmitoyl phosphatidic acid, sodium caprylate, bile
acids and salts thereof and glycine or taurine conjugates,
ursodeoxycholic acid, sodium cholate, sodium deoxycholate, sodium
taurocholate, sodium glycocholate,
N-Hexadecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate, anionic
(alkyl-aryl-sulphonates) monovalent surfactants, palmitoyl
lysophosphatidyl-L-serine, lysophospholipids (e.g.
1-acyl-sn-glycero-3-phosphate esters of ethanolamine, choline,
serine or threonine), alkyl, alkoxyl (alkyl ester), alkoxy (alkyl
ether)-derivatives of lysophosphatidyl and phosphatidylcholines,
e.g. lauroyl and myristoyl derivatives of lysophosphatidylcholine,
dipalmitoylphosphatidylcholine, and modifications of the polar head
group, that is cholines, ethanolamines, phosphatidic acid, serines,
threonines, glycerol, inositol, and the postively charged DODAC,
DOTMA, DCP, BISHOP, lysophosphatidylserine and
lysophosphatidylthreonine, zwitterionic surfactants (e.g.
N-alkyl-N,N-dimethylammonio-1-propanesulfonates,
3-cholamido-1-propyldimethylammonio-1-propanesulfonate,
dodecylphosphocholine, myristoyl lysophosphatidylcholine, hen egg
lysolecithin), cationic surfactants (quarternary ammonium bases)
(e.g. cetyl-trimethylammonium bromide, cetylpyridinium chloride),
non-ionic surfactants, polyethyleneoxide/polypropyleneoxide block
copolymers (Pluronics/Tetronics, Triton X-100, Dodecyl
.beta.-D-glucopyranoside) or polymeric surfactants (Tween-40,
Tween-80, Brij-35), fusidic acid derivatives- (e.g. sodium
tauro-dihydrofusidate etc.), long-chain fatty acids and salts
thereof C6-C12 (eg. oleic acid and caprylic acid), acylcarnitines
and derivatives, N.sup..alpha.-acylated derivatives of lysine,
arginine or histidine, or side-chain acylated derivatives of lysine
or arginine, N.sup..alpha.-acylated derivatives of dipeptides
comprising any combination of lysine, arginine or histidine and a
neutral or acidic amino acid, N.sup..alpha.-acylated derivative of
a tripeptide comprising any combination of a neutral amino acid and
two charged amino acids, or the surfactant may be selected from the
group of imidazoline derivatives, or mixtures thereof. Each one of
these specific surfactants constitutes an alternative embodiment of
the invention.
[0126] The use of a surfactant in pharmaceutical compositions is
well-known to the skilled person. For convenience reference is made
to Remington: The Science and Practice of Pharmacy, 19.sup.th
edition, 1995.
[0127] The formulations of the invention may be prepared by
conventional techniques, e.g. as described in Remington's
Pharmaceutical Sciences, 1985 or in Remington: The Science and
Practice of Pharmacy, 19.sup.th edition, 1995, where such
conventional techniques of the pharmaceutical industry involve
dissolving and mixing the ingredients as appropriate to give the
desired end product.
[0128] As mentioned above, in a preferred embodiment, the
formulations of the invention-contain, in addition to a peptide and
propylene glycol, a buffer and/or a preservative.
[0129] In one embodiment, the method for preparing such a peptide
formulation comprises:
[0130] a) preparing a first solution by dissolving preservative,
propylene glycol and buffer in water;
[0131] b) preparing a second solution by dissolving the peptide in
water;
[0132] c) mixing the first and second solutions; and
[0133] d) adjusting the pH of the mixture in c) to the desired
pH.
[0134] In another embodiment, the method for preparing such a
peptide formulation comprises:
[0135] a) preparing a first solution by dissolving preservative and
buffer in water;
[0136] b) adding propylene glycol to the first solution;
[0137] c) mixing the first solution with a second solution
containing peptide dissolved in water; and
[0138] d) adjusting the pH of the mixture in c) to the desired
pH.
[0139] In yet another embodiment, the method for preparing a
peptide formulation comprises:
[0140] a) preparing a solution by dissolving preservative, buffer
and propylene glycol in water;
[0141] b) adding the peptide to the solution of step a); and
[0142] c) adjusting the pH of the solution of step b) to the
desired pH.
[0143] As the formulations of the invention are optimal for
production and for use in injection devices since they exhibit
reduced deposits of production equipment and reduced clogging of
injection devices, the above methods of production can be used to
produce peptide formulations suitable for use in production and/or
for use in injection devices.
[0144] The formulations of the invention are suitable for
administration to a mammal, preferably a human. The route of
administration of the formulations of the invention may be any
route which effectively transports the peptide contained in the
formulation to the appropriate or desired site of action, such as
oral, nasal, buccal, pulmonal, transdermal or parenteral.
[0145] Due to the ability of propylene glycol to reduce clogging of
injection devices when compared to other isotonic agents and to
mannitol in particular, in a preferred embodiment, the formulations
of the invention are to be administered parenterally to a patient
in need thereof. Parenteral administration may be performed by
subcutaneous, intramuscular or intravenous injection by means of a
syringe, optionally a pen-like syringe. Alternatively, parenteral
administration can be performed by means of an infusion pump.
[0146] A further option is a composition which may be a powder or a
liquid for the administration of the formulation in the form of a
nasal or pulmonal spray. As a still further option, the formulation
can also be administered transdermally, e.g. from a patch,
optionally a iontophoretic patch, or transmucosally, e.g. bucally.
The above-mentioned possible ways to administer the formulations of
the invention are not to be considered as limiting the scope of the
invention.
[0147] Of course, it is understood that depending on the peptide or
peptides included in the formulations of the invention, the
formulations may be used in methods of treatment of diseases or
conditions for which use of the peptide is indicated. One skilled
in the art would understand that when used in such methods of
treatment, the formulations would have to be administered in amount
effective to treat the condition or disease for which the peptide
was being administered where an "effective amount" or an "amount .
. . effective" is understood to mean a dosage which is sufficient
in order for the treatment of the patient with the disease or
condition to be treated to be effective compared to treatment
without the administered dosage. It is to be understood that "an
effective amount" is the effective dose to be determined by a
qualified practitioner, who may titrate dosages to achieve the
desired response. Factors for consideration of dose will include
potency, bioavailability, desired pharmacokinetic/pharmacodynamic
profiles, the condition or disease to be treated (e.g. diabetes,
obesity, weight loss, gastric ulcers), patient-related factors
(e.g. weight, health, age, etc.), presence of co-administered
medications (e.g. insulin), time of administration, or other
factors known to a medical practitioner.
[0148] The present invention also relates to a method for reducing
deposits on production equipment during production of a peptide
formulation, where the method comprises replacing the isotonicity
agent previously utilized in said formulation with propylene glycol
at a concentration of between 1-100 mg/ml.
[0149] In one embodiment, the reduction in deposits on the
production equipment during production by the propylene
glycol-containing formulation relative to that observed for the
formulation containing the previously utilized isotonicity agent is
measured by a simulated filling experiment as described in the
Examples.
[0150] In another embodiment, the isotonicity agent to be replaced
by propylene glycol is selected from the group consisting of
sorbitol, sucrose, glycine, mannitol, lactose monohydrate, arginin,
myo-inositol and dimethylsulfon.
[0151] In a further embodiment, the isotonicity agent previously
utilized in said formulation is replaced with propylene glycol in a
concentration of from about 1 to about 50 mg/ml.
[0152] In another embodiment, the isotonicity agent previously
utilized in said formulation is replaced with propylene glycol in a
concentration of from about 5 to about 25 mg/ml.
[0153] In yet another embodiment, the isotonicity agent previously
utilized in said formulation is replaced with propylene glycol in a
concentration of from about 8 to about 16 mg/ml.
[0154] In another embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from about 7.0
to about 9.5.
[0155] In a further embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from about 7.0
to about 8.0.
[0156] In yet a further embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from 7.2 to
about 8.0.
[0157] In a further embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from about 7.0
to about 8.3.
[0158] In a further embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from 7.3 to
about 8.3.
[0159] The present invention also relates to a method for reducing
deposits in the final product during production of a peptide
formulation, where the method comprises replacing the isotonicity
agent previously utilized in said formulation with propylene glycol
at a concentration of between 1-100 mg/ml.
[0160] In one embodiment, the reduction in deposits in the final
product is measured by a reduction in the number of vials and/or
cartridges of the propylene glycol-containing formulation that must
be discarded due to deposits relative to number of vials and/or
cartridges of the formulation containing the previously utilized
isotonicity agent that must be discarded due to deposits.
[0161] In another embodiment, the isotonicity agent to be replaced
by propylene glycol is selected from the group consisting of
sorbitol, sucrose, glycine, mannitol, lactose monohydrate, arginin,
myo-inositol and dimethylsulfon.
[0162] In a further embodiment, the isotonicity agent previously
utilized in said formulation is replaced with propylene glycol in a
concentration of from about 1 to about 50 mg/ml.
[0163] In another embodiment, the isotonicity agent previously
utilized in said formulation is replaced with propylene glycol in a
concentration of from about 5 to about 25 mg/ml.
[0164] In yet another embodiment, the isotonicity agent previously
utilized in said formulation is replaced with propylene glycol in a
concentration of from about 8 to about 16 mg/ml.
[0165] In another embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from about 7.0
to about 9.5.
[0166] In a further embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from about 7.0
to about 8.0.
[0167] In yet a further embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from 7.2 to
about 8.0.
[0168] In a further embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from about 7.0
to about 8.3.
[0169] In a further embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from 7.3 to
about 8.3.
[0170] The present invention further relates to a method for
reducing the clogging of injection devices by a peptide
formulation, where the method comprises replacing the isotonicity
agent previously utilized in said formulation with propylene glycol
at a concentration of between 1-100 mg/ml.
[0171] In one embodiment, the reduction in clogging of the
injection device by the propylene glycol-containing formulation
relative to that observed for the formulation containing the
previously utilized isotonicity agent is measured in a simulated in
use study as described in the Examples.
[0172] In another embodiment, the isotonicity agent to be replaced
by propylene glycol is selected from the group consisting of
inositol, maltose, glycine, lactose and mannitol.
[0173] In a further embodiment, the isotonicity agent previously
utilized in said formulation is replaced with propylene glycol in a
concentration of from about 1 to about 50 mg/ml.
[0174] In another embodiment, the isotonicity agent previously
utilized in said formulation is replaced with propylene glycol in a
concentration of from about 5 to about 25 mg/ml.
[0175] In yet another embodiment, the isotonicity agent previously
utilized in said formulation is replaced with propylene glycol in a
concentration of from about 8 to about 16 mg/ml.
[0176] In another embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from about 7.0
to about 9.5.
[0177] In a further embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from about 7.0
to about 8.0.
[0178] In yet a further embodiment of the invention, the propylene
glycol-containing formulation has a pH in the range from 7.2 to
about 8.0.
[0179] All scientific publications and patents cited herein are
specifically incorporated by reference. The following examples
illustrate various aspects of the invention but are in no way
intended to limit the scope thereof.
EXAMPLES
Example 1
Simulated Filling Experiments, Drop and Clogging Tests of
Replacement Candidates for Mannitol
[0180] As laboratory experiments have shown that with regards to
clogging of needles and deposits on needles, formulations without
peptide ("placebo") give the same conclusions as formulations with
peptide at 0.3-5.0 mg/ml, the screening studies in Example 1 have
been done using placebo except where indicated otherwise.
Preparation of Formulations With Different Isotonic Agents
[0181] Preservative (5.5 mg/ml phenol) and buffer 1.24 mg/ml
disodium hydrogen phosphate, dihydrate) were dissolved in water and
the isotonic agent was added while stirring. pH was adjusted to pH
7.9 using Sodium Hydroxide and/or Hydrochloric acid. Finally, the
formulation was filtered through a 0.22 .mu.m filter. The isotonic
agents tested in each formulation and their concentrations are
shown in Table 1.
TABLE-US-00001 TABLE 1 Composition of the tested formulations
Formulation no. Tonicity modifier 1 Glucose monohydrate (38.0
mg/ml) 2 Laktose monohydrate (65.0 mg/ml) 3 Maltose (67.2 mg/ml) 4
Glycine (15.1 mg/ml) 5 Polyethylenglycol 400 (77.5 mg/ml) 6
L-arginin (24.6 mg/ml) 7 Myo-Inositol (35.2 mg/ml) 8 Propylene
glycol (13.7 mg/ml) 9 Dimethylsulfon (18 mg/ml) 10 Mannitol (35.9
mg/ml) 11 Sorbitol (39.5 mg/ml) 12 Xylitol (39.5 mg/ml) 13 Sucrose
(79.1 mg/ml 14 Glycerol (16 mg/ml)
Osmolarity
[0182] The osmolarity of the different placebo formulations was
determined and the results are shown in Table 2. An isotonic
solution has an osmolarity of around 0.286 osmol/L. As can be seen
from Table 2 three of the formulations (PEG 400, sucrose and
xylitol) are more than 20% from being isotonic (0.229-0.343
osmol/l), however for these kind of experiments the osmolarity is
not expected to influence the results, though, the tonicity of the
formulations should be adjusted in future experiments.
TABLE-US-00002 TABLE 2 The measured osmolarity of the formulations
Formulation no. Isotonic agent Osmolarity 1 Glucose monohydrate
(38.0 mg/ml) 0.315 2 Laktose monohydrate (65.0 mg/ml) 0.283 3
Maltose (67.2 mg/ml) 0.306 4 Glycine (15.1 mg/ml) 0.286 5
Polyethylenglykol 400 (77.5 mg/ml) 0.370 6 L-arginin(24.6 mg/ml)
0.318 7 Myo-Inositol (35.2 mg/ml) 0.285 8 Propylene glycol (13.7
mg/ml) 0.268 9 Dimethylsulfon (18 mg/ml) 0.274 10 Mannitol (35.9
mg/ml) 0.284 11 Sorbitol (39.5 mg/ml) 0.310 12 Xylitol (39.5 mg/ml)
0.351 13 Sucrose (79.1 mg/ml 0.346 14 Glycerol (16 mg/ml) 0.262
Drop Test
[0183] A droplet of each formulation is placed on a microscope
slide and let to dry. The deposit is visually examined by eye and
light microscope. A photograph of the dried droplets of some of the
formulations is shown in FIG. 1. In this figure it is clearly
observed that mannitol cause deposits on the microscope slide when
let to dry. No deposits were observed for sorbitol, xylitol,
sucrose and glycerol. The droplet on the far right (Form 1)
contains mannitol and Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37). In FIG. 2, the candidates causing the most deposits on
the microscope slide are shown. For comparison glycerol, which does
not cause deposits, is shown (mannitol, arginine, inositol).
Clogging Test
[0184] In this test 10 NovoPens.RTM. 1.5 ml mounted with NovoFine
30.RTM. G (G 30 needle) were tested for each formulation, 5 of them
placed in upright and 5 in horizontal position. The Pensystems were
stored at room temperature in between testing. Each day the needle
was examined for deposits and an air shot was performed prior to
injection into a tissue. Degree of resistance and clogging, if any,
was noted. Injections were made on a daily basis with the same
needle, and this was done for 9 working days for all the
formulations. The results from the clogging test are shown in Table
3.
TABLE-US-00003 TABLE 3 Clogging test in NovoPen 1.5 using 30G
NovoFine Isotonic Drop at Dried Gel-like agent (no. of Some Much
top of drop at drop on Deposits observations) resistance Resistance
resistance Clogged needle needle top needle on needle Mannitol (90)
10 0 0 0 0 2 0 43 Glycerol (90) 13 0 0 0 1 0 3 0 Sucrose (90) 23 0
0 0 0 0 21 0 Propylene 20 0 0 0 0 0 0 0 glycol (90) PEG 400 (90) 25
1 0 0 12 (5 at 0 0 0 needle) arginin (90) 26 2 0 0 3 (2 at 1 0 0
needle) Xylitol (90) 14 0 0 0 5 0 0 0 Dimethylsulfon 21 0 0 0 4 0 0
0 (90) sorbitol (90) 12 0 0 0 9 1 0 1 Myoinositol 20 1 2 6 6 0 0 47
(90) Glucose (90) 32 11 5 0 16 (7 at 1 0 (1 at needle) needle)
glycine (90) 41 9 2 0 1 (2 at 0 0 31 (2 at needle) needle) maltose
(90) 35 8 7 4 16 (6 at 0 0 1 (5 at needle) needle) laktose (90) 44
10 8 0 5 0 0 31 (2 at needle)
In Table 3 and in FIG. 3 it was observed that inositol and maltose
clogged the needle. For comparison glycerol which does not clog the
needle is shown in FIG. 3. In FIG. 4, and in Table 3, it was
observed that formulations containing glycine, lactose and mannitol
gave rise to a lot of deposits on the needle. For glycine, the
deposits were a droplet deposited down the needle, whereas for
lactose and mannitol the deposits occurred at the top of the
needle.
Simulated Filling
[0185] 1 L of each formulation was subjected to a simulated filling
experiment which lasted for 24 hours. After 24 hours the filling
equipment was inspected for the presence of deposits. Based on the
results from the simulated filling studies (data not shown), the
placebo formulations can be divided into three categories. 1. Those
isotonic agents that do not cause deposits on the filling
equipment: Xylitol, glycerol, glucose monohydrate, maltose, PEG 400
and propylene glycol. 2. Those isotonic agent that cause few
deposits and have superior filling properties compared to mannitol:
Sorbitol, sucrose and glycine. 3. Those isotonic agent that are
comparable or worse than mannitol: Mannitol, lactose monohydrate,
arginin, myo-inositol and dimethylsulfon.
CONCLUSION
[0186] In the simulated filling experiment xylitol, glycerol,
glucose, maltose, PEG 400, propylene glycol, sorbitol, sucrose and
glycine were found to be suitable replacements candidates for
mannitol. However, as glucose is a reducing saccharide, and
therefore is able to initiate unwanted degradation in the
formulation, this tonicity modifier is ruled out. Furthermore,
maltose is ruled out due to clogging of needles. This leads to the
following candidates: glycerol, xylitol, sorbitol, sucrose,
glycine, propylene glycol and PEG 400, which are found to have
suitable properties as replacements candidates for mannitol in
peptide formulations with regards to drop test, clogging of needles
and simulated filling. However, on the basis of the following
considerations, propylene glycol was chosen as the isotonic agent
over the other candidates to be further investigated in head to
head comparison studies with mannitol: [0187] a. propylene glycol
was observed to have no influence on the physical and chemical
stability of Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37)-containing formulations; [0188] b. propylene glycol was
observed to have no influence on antimicrobial preservative
testing; and [0189] c. use of propylene glycol would no require
that further toxicity studies be tested
Example 2
Comparison Of Mannitol and Propylene Glycol-Containing Placebo
Formulations In Simulated Filling Studies and Simulated Use
Studies
Preparation of Formulations
[0190] Preservative and buffer were dissolved in water and the
isotonic agent was added while stirring. pH was adjusted to the
aimed pH using Sodium Hydroxide and/or Hydrochloric acid. Finally,
the formulation was filtered through a 0.22 .mu.m filter. The
compositions of the formulations were as follows: [0191] Disodium
hydrogen phosphate, dihydrate: 1.42 mg/ml [0192] Phenol: 5.5 mg/ml
[0193] Propylene glycol or mannitol: 13.7 or 35.9 mg/ml [0194]
Water for Injection: up to 1.0 ml. [0195] pH: 7.90
Simulated Filling Study
[0196] A simulated filling study lasting 24 hours was performed as
described in Example 1 and after 24 hours, the filling equipment
was inspected for the presence of deposits. No deposits were
observed on the filling equipment for the propylene glycol
formulation. By comparison, after 24 hours, a lot of deposits were
observed on the filling equipment for the mannitol formulation (see
FIG. 6).
Simulated In Use Study
[0197] For the simulated in use study, a clogging test was
conducted as described in Example 1. The same needle was used
during the study period of ten working days and each day, the
needle was inspected for the presence of deposits. FIG. 7 shows
photographs of needles dosed with the propylene glycol (top panel)
or mannitol (bottom panel) containing formulations. Deposits on the
needle were observed in 48% of the cases when mannitol was used as
an isotonic agent whereas no deposits were observed when propylene
glycol was used as the isotonic agent.
Example 3
Comparison of Propylene Glycol to Mannitol In Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37) Containing Formulations
Preparation of Formulations
[0198] Preservative, isotonic agent (mannitol or propylene glycol)
and buffer were dissolved in water and pH was adjusted to the
desired pH. Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.hexadecanoyl)))-GLP--
1(7-37) was dissolved in water while stirring slowly. The two
solutions were then mixed and pH adjusted to the desired pH using
sodium hydroxide and/or hydrochloric acid. Finally, the formulation
was filtered through a 0.22 .mu.m filter. The compositions of the
formulations were as follows:
[0199] Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37) (6.25 mg/ml),
[0200] Disodium hydrogen phosphate, dihydrate (1.42 mg/ml),
[0201] Phenol (5.5 mg/ml),
[0202] mannitol or propylene glycol (35.9 or 14.0 mg/ml),
[0203] Water for Injection (up to 1.0 ml),
[0204] pH: 8.15
Simulated In Use Study
[0205] For the simulated in use study, a clogging test was
conducted as described in Example 1 except that a G31 needle was
used. The same G31 needle was used during the study period of ten
working days and each day, the needle was inspected for the
presence of deposits. FIG. 7 shows photographs of needles with no
deposits when dosed with the propylene glycol (bottom panel) or
showing deposits when dosed with the mannitol (top panel)
containing formulations.
[0206] For the mannitol containing formulation, clogging of the
needle was observed in 1 out of 10 cases on day 4, 2 out of 10
cases on day 5, 3 out of 10 cases on day 8 and 4 out of 10 cases on
day 9. By comparison, no clogging of needles was observed for the
propylene glycol containing formulation.
[0207] It is believed that similar results to those obtained with
the above-described propylene glycol-containing formulation would
also be obtained if the pH was adjusted to 7.40, 7.70 or 7.90. In
addition, additional formulations which could be tested include
those having the following compositions:
[0208] Buffering agents: glycylglycine (1.32 mg/ml), L-Histidine
(1.55 mg/ml), Hepes (2.38 mg/ml), or bicine (1.63 mg/ml)
[0209] Preservatives: phenol (5.0 or 5.5 mg/ml), benzylalcohol (18
mg/ml) or a mixture of m-cresol and phenol (2.5/2.0 mg/ml)
[0210] Propylene glycol: 14.0 or 14.3 mg.ml
[0211] Water for injection: up to 1.0 ml
[0212] pH: 7.40, 7.70, 7.90 or 8.15
Example 4
Influence of Peptide Concentration On Clogging of Needles
[0213] Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37) formulations were prepared as described in Example 3 using
peptide concentrations ranging from 0-5 mg/ml of Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37). The compositions of the formulations were as follows:
Liraglutide: 0, 0.3, 3 and 5 mg/ml Disodium hydrogen phosphate,
dihydrate: 0.71 mg/ml Sodium dihydrogenphosphate, dihydrate: 0.62
mg/ml Mannitol: 36.9 mg/ml Phenol: 5.0 mg/ml Water for injection:
up to 1.0 ml pH 7.40
[0214] A simulated in use study was conducted as in Example 3
except that a G30 needle was used and the results (data not shown)
indicated that the clogging effect of the mannitol-containing
formulations relative to the absence of clogging with the propylene
glycol formulations was observed independent of the peptide
concentration.
Example 5
Clogging of Needles in Lys .beta.29 (N.epsilon.-tetradecanoyl)
des(B30) Human Insulin and NovoMix 30 Formulations Containing
Mannitol
Preparation Of Formulations
[0215] The Lys .beta.29 (N.epsilon.-tetradecanoyl) des(B30) human
insulin-containing formulation was prepared as follows: a) Prepared
a first solution by dissolving buffer, sodium chloride,
preservatives (phenol and m-cresol) and mannitol in water b)
Prepared a second solution of Lys .beta.29
(N.epsilon.-tetradecanoyl) des(B30) human insulin and zinc acetate
dissolved in water c) added the peptide-containing solution of step
b) to the solution of step a); and d) adjusted the pH of the
solution to the desired pH The composition of Lys .beta.29
(N.epsilon.-tetradecanoyl) des(B30) human insulin-containing
formulation prepared in the above manner was as follows: Lys
.beta.29 (N.epsilon.-tetradecanoyl) des(B30) human insulin (2400
nmol), Phenol (1.80 mg/ml), m-cresol (2.06 mg/ml), Mannitol (30.0
mg/ml), disodiumphosphate, dihydrate (0.890 mg/ml), Sodium chloride
(1.17 mg/ml), Zinc acetate (65.4 ug/ml), water for injection (up to
1.0 ml), pH: 7.4 The NovoMix 30-containing formulation was prepared
as follows: a) Prepared a solution by dissolving buffer, sodium
chloride, phenol, mannitol and sodium hydroxide in water b)
Prepared a solution of sodium chloride, phenol and mannitol in
water c) Prepared a solution of protamine sulphate in water d)
Prepared a solution of insulin, hydrochloric acid and zinc in water
e) Solutions b), c) and d) were mixed f) Solution e) was added to
the solution of step a) g) Adjusted the pH of the solution to the
desired pH and crystallized at room temperature h) Prepared a
solution by dissolving m-cresol, phenol and mannitol in water i)
Solution h) is added to the crystalline fraction of step g); and j)
Adjusted the pH to the desired pH The composition of the NovoMix
30-containing formulation prepared in the above manner was as
follows: Insulin aspart (100 units/ml), protamine sulphate (approx.
0.33 mg/ml), phenol (1.50 mg/ml), m-cresol (1.72 mg/ml), mannitol
(30.0 mg/ml), disodiumphosphate dihydrate (1.25 mg/ml), sodium
chloride (0.58 mg/ml), zinc (19.6 ug/ml), water for injection (up
to 1.0 ml), pH: 7.3.
Results
[0216] A simulated in use study was conducted as described in
Example 3 using G31 needles where 20 needles were investigated for
10 days. The results were as follows: Clogging of needles was
observed for Lys .beta.29 (N.epsilon.-tetradecanoyl) des(B30) human
insulin on day 2 (5%), day 3 (70%) and on day 4 (100%). Clogging of
needles for NovoMix 30 was observed on day 3 (5%), day 4 (10%), day
5 (35%), day 6 (40%), day 8 (50%), day 9 (55%) and day 10 (80%).
Thus, the effect of mannitol on the clogging of needles is
independent of the type of peptide included in the formulations
since a comparable clogging effect was observed with Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37), Lys .beta.29 (N.epsilon.-tetradecanoyl) des(B30) human
insulin and NovoMix 30.
Example 6
Testing of Lys .beta.29 (N.epsilon.-tetradecanoyl) des(B30) Human
Insulin and NovoMix 30 Formulations Containing Propylene Glycol
[0217] The preparation and composition of the Lys .beta.29
(N.epsilon.-tetradecanoyl) des(B30) human insulin and NovoMix 30
formulations will be as described in Example 5 except that mannitol
will be replaced with a concentration of propylene glycol that
assures tonicity. A simulated in use test will then be conducted as
described in Example 5. Based on the fact that the clogging effect
of Lys .beta.29 (N.epsilon.-tetradecanoyl) des(B30) human insulin
and NovoMix 30 mannitol-containing formulations was similar to that
observed with Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37) mannitol-containing formulations, it is believed that the
effect of propylene glycol on the clogging effect of Lys .beta.29
(N.epsilon.-tetradecanoyl) des(B30) human insulin and NovoMix
30-containing formulations will be similar to that observed with
Arg.sup.34,
Lys.sup.26(N.sup..epsilon.-(.gamma.-Glu(N.sup..alpha.-hexadecanoyl)))-GLP-
-1(7-37)-containing formulations.
Sequence CWU 1
1
4131PRTArtificial SequenceSyntheticMISC_FEATURE(31)..(31)Xaa = P or
Y 1His Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu1
5 10 15Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Xaa
20 25 30240PRTArtificial SequenceSynthetic 2His Ser Asp Gly Thr Phe
Ile Thr Ser Asp Leu Ser Lys Gln Met Glu1 5 10 15Glu Glu Ala Val Arg
Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro 20 25 30Ser Ser Gly Ala
Pro Pro Pro Ser 35 40344PRTArtificial
SequenceSyntheticMISC_FEATURELysine at position 44 is amidated 3His
Gly Glu Gly Thr Phe Thr Ser Asp Leu Ser Lys Gln Met Glu Glu1 5 10
15Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser
20 25 30Ser Gly Ala Pro Pro Ser Lys Lys Lys Lys Lys Lys 35
40440PRTArtificial SequenceSynthetic 4His Gly Glu Gly Thr Phe Ile
Thr Ser Asp Leu Ser Lys Gln Met Glu1 5 10 15Glu Glu Ala Val Arg Leu
Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro 20 25 30Ser Ser Gly Ala Pro
Pro Pro Ser 35 40
* * * * *