U.S. patent application number 11/962034 was filed with the patent office on 2009-01-15 for formulations for amylin agonist peptides.
This patent application is currently assigned to Amylin Pharmaceuticals, Inc.. Invention is credited to James L. L'ITALIEN, Gregg STETSKO.
Application Number | 20090018053 11/962034 |
Document ID | / |
Family ID | 29709686 |
Filed Date | 2009-01-15 |
United States Patent
Application |
20090018053 |
Kind Code |
A1 |
L'ITALIEN; James L. ; et
al. |
January 15, 2009 |
FORMULATIONS FOR AMYLIN AGONIST PEPTIDES
Abstract
The present invention is concerned with a pharmaceutical
formulation comprising an amylin agonist and optionally a buffer, a
tonicifier or stabilizer, and a preservative in a container, for
example, a vial, prefilled cartridge, prefilled syringe or
disposable pen. This formulation may be in liquid, gel, solid or
powdered form for delivery, for example, via nasal, pulmonary,
oral, sublingual, buccal, transdermal, or parenteral routes.
Formulation with biocompatible polymers and release modifiers, such
as sugars, can facilitate controlled release after injection,
minimizing the number of administrations to a patient. These
formulations maintain stability upon storage under refrigerated or
room temperature conditions. Such formulations can be further
combined with insulin for administration to a patient.
Inventors: |
L'ITALIEN; James L.; (Del
Mar, CA) ; STETSKO; Gregg; (San Diego, CA) |
Correspondence
Address: |
Intellectual Property Department;Amylin Pharmaceuticals, Inc.
9360 Towne Centre Drive
San Diego
CA
92121
US
|
Assignee: |
Amylin Pharmaceuticals,
Inc.
San Diego
CA
|
Family ID: |
29709686 |
Appl. No.: |
11/962034 |
Filed: |
December 20, 2007 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10159779 |
May 31, 2002 |
7312196 |
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11962034 |
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09005262 |
Jan 9, 1998 |
6410511 |
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10159779 |
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Current U.S.
Class: |
514/1.1 |
Current CPC
Class: |
A61K 2300/00 20130101;
A61K 2300/00 20130101; A61K 38/22 20130101; A61K 38/22 20130101;
Y10S 514/866 20130101; A61K 9/1647 20130101; A61K 47/02 20130101;
A61K 38/28 20130101; A61K 9/0019 20130101; A61K 9/0014 20130101;
A61K 9/0024 20130101; A61K 47/10 20130101; A61K 47/12 20130101;
A61P 43/00 20180101; A61K 47/26 20130101; Y10S 514/864 20130101;
A61P 3/10 20180101; A61K 9/19 20130101; A61K 38/28 20130101 |
Class at
Publication: |
514/4 ;
514/2 |
International
Class: |
A61K 38/28 20060101
A61K038/28; A61K 38/00 20060101 A61K038/00; A61P 3/10 20060101
A61P003/10 |
Claims
1. A liquid pharmaceutical formulation comprising about 0.5 to 1.0%
(w/v) of an amylin agonist, a pharmaceutically acceptable
excipient, and about 0.005% to 1.0% (w/v) of a preservative.
2. The liquid pharmaceutical formulation according to claim 1,
wherein said amylin agonist is pramlintide.
3. The liquid pharmaceutical formulation according to claim 1,
further comprising insulin.
4. The liquid pharmaceutical formulation according to claim 3,
wherein said insulin is glargine.
5. The liquid pharmaceutical formulation according to claim 1
wherein said preservative is selected from the group consisting of
m-cresol, benzyl alcohol, ethyl, ethyl, propyl and butyl parabens
and phenol.
6. The liquid pharmaceutical formulation according to claim 1,
wherein said preservative is between about 0.1 to 0.3% (w/v) of
m-cresol.
7. The liquid pharmaceutical formulation according to claim 1,
wherein said preservative is at least 0.15% (w/v) of m-cresol.
8. The liquid pharmaceutical formulation according to claim 1,
wherein said amylin agonist is stable for at least 30 days at room
temperature.
9. The liquid pharmaceutical formulation according to claim 8,
wherein said amylin agonist is pramlintide.
10. A liquid pharmaceutical formulation comprising about 0.5 to
1.0% (w/v) of an amylin agonist, a pharmaceutically acceptable
excipient, and insulin.
11. The liquid pharmaceutical formulation according to claim 10,
wherein said amylin agonist is pramlintide.
12. The liquid pharmaceutical formulation according to claim 10,
wherein said insulin is glargine.
13. A formulation comprising about 1% to 98% (w/w) of an amylin
agonist and about 1.0% to 20% (w/w) of at least one
pharmaceutically acceptable excipient selected from the group
consisting of a preservative, a penetration enhancer and a
stabilizer.
14. The formulation according to claim 13 comprising about 80% to
90% (w/w) of said amylin agonist and about 10% to 20% (w/w) of said
at least one pharmaceutically acceptable excipient.
15. The formulation according to claim 13 for oral administration,
wherein said formulation is solid, wherein said pharmaceutically
acceptable excipient is powdered, wherein said pharmaceutically
acceptable excipient comprises about 1.0 to 20% (w/w) of said
formulation, wherein said formulation comprises a penetration
enhancer or stabilizer.
16. The formulation according to claim 15 wherein said formulation
is encapsulated within a polymeric coating.
17. The formulation according to claim 15, wherein said powdered
excipient is lactose.
18. The formulation according to claim 15, wherein said formulation
comprises glargine.
19. The formulation according to claim 13, wherein said amylin
agonist is pramlintide.
20. The formulation according to claim 13 which is a solid, liquid,
gel or semi-solid.
21. The formulation according to claim 13 which is lyophilized.
22. The formulation according to claim 13, further comprising
insulin.
23. The formulation according to claim 22, wherein said insulin is
glargine.
24. The formulation according to claim 13 for nasal, pulmonary or
buccal delivery.
25. The formulation according to claim 24 comprising a penetration
enhancer, or a stabilizer.
26. The formulation according to claim 24 further comprising a
polymer.
27. The formulation according to claim 24 further comprising a
preservative.
28. The formulation according to claim 13 for administration to a
subject, wherein said formulation is administered in the form
selected from the group consisting of oral delivery, a microcapsule
delivery, a microsphere, an implant delivery, and a polymer matrix
delivery.
29. The formulation according to claim 13 formulated in tablets or
capsules for oral, sublingual or buccal delivery.
30. The formulation according to claim 13 formulated in powder form
for nasal or pulmonary delivery.
31. The formulation according to claim 13 formulated for
transdermal delivery.
32. A method for treating diabetes in a patient in need of such
treatment comprising administering a therapeutically effective
amount of the formulation according to claim 14 to the patient in
need of such treatment.
33. The method according to claim 32 wherein insulin is admixed
with said formulation.
34. The method according to claim 33 wherein said admixing step is
simultaneous, or concerted and sequential.
35. A liquid suspension formulation comprising about 0.5 to 90%
(w/v) of an amylin agonist, insulin, about 1.0 to 10% (w/v) of a
carbohydrate or a polyhydric alcohol, and about 0.02 to 0.5% (w/v)
of an acetate, phosphate, citrate or glutamate buffer, and a liquid
vehicle, wherein the formulation has a pH of about 3.5 to 5.0.
36. The formulation according to claim 35, wherein said amylin
agonist is pramlintide.
37. The formulation according to claim 35, wherein said insulin is
glargine.
Description
CROSS-REFERENCE TO RELATED PATENT APPLICATIONS
[0001] This application is a continuation of U.S. Ser. No.
10/159,779, filed May 31, 2002, now issued as U.S. Pat. No.
7,312,196, which in turn is a continuation in part of U.S. Ser. No.
09/005,262, filed Jan. 9, 1998, now issued as U.S. Pat. No.
6,401,511, the contents of which are hereby incorporated by
reference in their entirety.
FIELD OF INVENTION
[0002] This invention relates to pharmaceutical formulations of
amylin agonist peptides, particularly in solid, suspension and
liquid formulations. Additionally, this invention relates to amylin
agonist pharmaceutical formulations which are compatible upon
mixing with insulin or are prepared to allow co-administration with
insulin or other therapeutic or antidiabetic compositions.
BACKGROUND OF THE INVENTION
[0003] Deposition of amyloid in pancreatic islets is a common
feature in human Type II diabetic patients. The major protein
forming these amyloid particles, called amylin, has a propensity to
form fibril amyloid structures. Proceedings of the National Academy
of Sciences USA 84(23):8628-32, 1987. Amylin is a 37 amino acid
protein which, in its fully active form, is carboxy-amidated and
has a disulfide bridge between the cysteine residues found at
positions 2 and 7. Amylin plays a role in control of systemic
concentrations of glucose, and has been proposed as a useful
therapeutic agent. See, e.g., Leighton and Cooper, 15 TIBS 295,
1990. Human amylin is described and claimed in U.S. Pat. No.
5,367,052, entitled "Amylin Peptides," and U.S. Pat. No. 5,124,314,
entitled "Pharmaceutical Compositions Containing Amylin." Amylin
has been reviewed in the literature, for example, in Gaeta, L. S.
L. and Rink, T. J., 3 Med. Chem. Res. 483-490, 1994, Pittner, R. A.
et al., 55S J. Cell. Biochem. 19-28, 1994, and Rink, T. J. et al.,
14 TIPS 113-118, 1993.
[0004] Therapeutic opportunities for insulin-using and other people
with diabetes who are deficient in amylin or for whom amylin
therapy would be of benefit, and hormone blockage for other people,
for example, the obese and Type II diabetics and those with insulin
resistance who may have elevated plasma amylin or undesired amylin
activity have been pursued. The use of amylin agonists, including
amylin itself, for the treatment of diabetes is described and
claimed in U.S. Pat. No. 5,175,145. The use of amylin antagonists
for the treatment of Type II diabetes mellitus, obesity and
essential hypertension, and insulin resistance, are described and
claimed in U.S. Pat. Nos. 5,266,561, 5,280,014, 5,281,581, and
5,364,841.
[0005] The most severe form of the disease is Type I
(juvenile-onset) diabetes. There are an estimated 1 million Type I
diabetics in the U.S. who need daily insulin injections for
survival. Their quality of life is often markedly affected by the
rigors of their daily metabolic imbalances, in particular
hypoglycemic attacks (dangerously low blood glucose) and by the
onset of serious long-term complications, including blindness,
kidney failure, impotence, ulcers, amputations and atherosclerosis
(NIH Diabetes Complications and Control Trial).
[0006] Type II (adult-onset) diabetes afflicts over 10 million
Americans, who are also subject to the same complications. Impaired
glucose tolerance, a risk factor for Type II diabetes and
cardiovascular disease, is thought to affect another 20 million
people in the U.S. and is not treatable by any known regimen. There
is also an alarming increase in the incidence of Type II diabetics
in groups of populations around the world, whose standard of living
is increasing through economic development or migration.
Sulfonylureas are the primary oral antihyperglycemic diabetic
medications sold in the U.S. Discovered in the 1940's, these
compounds do not address the underlying causes of Type II diabetes
and, in many cases, are not effective or lose their efficacy after
a few years of treatment. Type II diabetics do not lack insulin,
rather they are insulin resistant, so that insulin does not work
properly and the insulin secretory responses are disordered.
[0007] After a meal, the pancreas secretes insulin in response to a
rise in glucose. Insulin stimulates the uptake of glucose into
muscle and fat, and signals the liver to reduce glucose production;
this results in a return of blood glucose to normal levels. In
muscle, large amounts of glucose are stored as glycogen. Some of
the glycogen is broken down into lactate, which circulates to the
liver and can be converted back into glucose and stored as
glycogen. Between meals the liver breaks down these glycogen stores
to provide glucose to the brain and other tissues. This cycle in
which glycogen is effectively transferred from muscle to liver is
known as the Cori cycle. The stimulus for this flux from muscle to
liver under resting conditions remain unidentified; recent results
indicate that amylin provides a major stimulus to this pathway.
[0008] Amylin has been demonstrated to have direct metabolic
effects in both skeletal muscle and the pancreas. In skeletal
muscle, amylin acts as a non-competitive antagonist of insulin,
reducing insulin-stimulated incorporation of glucose into glycogen.
In vitro studies indicate that amylin reduces glycogen synthase
activity and favors the formation of an active form of glycogen
phosphorylase, the enzyme that converts glycogen into glucose
6-phosphate. The actions of amylin on skeletal muscle promote
glycogen breakdown, thus stimulating lactate formation and
increasing turnover of the Cori cycle. Amylin is co-secreted with
insulin from pancreatic beta cells and has been demonstrated to
suppress insulin secretion. It appears to provide feedback
regulation of the beta-cell, in order to modulate insulin secretory
activity.
[0009] It is believed that amylin plays a role in the regulation of
glucose uptake from ingested food into blood, and that amylin or
amylin agonist therapy in diabetics, particularly insulin-using
diabetics, such as Type I diabetics and late-stage Type II
diabetics, will smooth the excessive glucose rises that these
patients typically experience after meals. Deficiency of an
important hormone such as amylin which has been reported to have
effects on carbohydrate, fat and bone metabolism, may also disrupt
other important physiological mechanisms. The co-administration of
amylin, or an amylin agonist which exerts the physiological effects
of amylin, will significantly improve existing insulin therapy by
restoring the appropriate metabolic balance.
[0010] Many factors affect the stability of a pharmaceutical
product, including the chemical reactivity of the active
ingredient(s), the potential interaction between active and
inactive ingredients, the manufacturing process, the dosage form,
the container-closure system, and the environmental conditions
encountered during shipment, storage, handling and length of time
between manufacture and usage. Pharmaceutical product stability is
determined by the chemical stability as well as the physical
stability of the formulation. Physical factors including heat and
light may initiate or accelerate chemical reactions.
[0011] Optimal physical stability of a formulation is very
important for at least three primary reasons. First, a
pharmaceutical product must appear fresh, elegant and professional,
when it is administered to a patient. Any changes in physical
appearance such as color changes or haziness can cause a patient or
consumer to lose confidence in the product. Second, because some
products are dispensed in multiple-dose containers, uniformity of
dose content of the active ingredient over time must be assured. A
cloudy solution or a broken emulsion can lead to a non-uniform
dosage pattern. Third, the active ingredient must be available to
the patient throughout the expected shelf life of the preparation.
A breakdown of the product to inactive or otherwise undesired forms
can lead to non-availability of the medicament to the patient.
[0012] Stability of a pharmaceutical product, then, may be defined
as the capability of a particular formulation to remain within its
physical, chemical, microbiological, therapeutic and toxicological
specifications. A stable solution retains its original clarity,
color and odor throughout its shelf life. Retention of clarity of a
solution is a main concern in maintaining physical stability.
Solutions should remain clear over a relatively wide temperature
range such as about 4.degree. C. to about 37.degree. C. At the
lower range an ingredient may precipitate due to a lower solubility
at that temperature, while at higher temperatures homogeneity may
be destroyed by extractables from the glass containers or rubber
closures. Thus, solutions of active pharmaceutical ingredients must
be able to handle cycling temperature conditions. Similarly, a
formulation should retain its color throughout this temperature
range, and its odor should be stably maintained.
[0013] Small peptides are typically unstable and are susceptible to
degradation in aqueous solution. In this regard, once a human
amylin agonist or amylin has less than approximately 90% of its
labeled potency, it is no longer considered to be suitable for
administration to a patient. Various types of molecules such as
sugars, surfactant, amino acids and fatty acids, used singly or in
combination, have been used in efforts to stabilize protein and
peptide products against degradation. See Wang and Hanson, J.
Parenteral Science and Technology Supplement, 1988, Technical
Report No. 10 (describing parenteral formulations of proteins and
peptides); Manning et al., 6 Pharmaceutical Research, 1989.
Examples of excipients such as buffers, preservatives, isotonic
agents, and surfactants are also known in the art. See 21 C.F.R.
.sctn. 180.22 et seq. (defining recognized food additives); Wang
and Kowal, 34 J. Parenteral Drug Association 452, 1980 (describing
various excipients); A. R. Gennaro et al., 17th Remington's
"Pharmaceutical Sciences," 1985; Avis et al., Pharmaceutical Dosage
Forms: Parenteral Medications, Vol. 1, 1992, all of which,
including the definitions of various useful excipients, are hereby
incorporated by reference herein.
SUMMARY OF THE INVENTION
[0014] The inventors have discovered novel pharmaceutical
formulations and dosage forms useful for the administration of
amylin agonist peptides to a patient or subject in need of
treatment with such compounds. These dosage forms include
approximately 0.01 to 1.0% (w/v), preferably 0.5 to 1.0%,
respectively, of an amylin agonist, or amylin, as the active
ingredient in an aqueous system. This concentration may vary based
upon mode of administration and type of formulations, however,
formulations of the present description, whether liquid or solid,
may contain amylin or amylin agonist in a concentration range from
about 0.01 (w/w) to about 98% (w/w).
[0015] The amylin or amylin agonist is frequently included along
with approximately 0.02 to 0.5% (w/v) of an acetate, phosphate,
citrate or glutamate buffer to obtain a pH of the final composition
of about 3.0 to about 7.0, as well as approximately 1.0 to 10%
(w/v) of a carbohydrate or polyhydric alcohol tonicifier in an
aqueous continuous phase, provided, however, that if the amylin
agonist or amylin has similar physiochemical characteristics
similar to those of human amylin, it should be formulated and
lyophilized for storage (and used immediately upon reconstitution).
Approximately 0.005 to 1.0% (w/v) of an antimicrobial preservative
selected from the group consisting of m-cresol, benzyl alcohol,
methyl, ethyl, propyl and butyl parabens and phenol may also be
present in the formulation of product designed to allow the patient
to withdraw or administer multiple doses. Importantly, a stabilizer
is not required in this amylin agonist product formulation. A
sufficient amount of a liquid vehicle, such as water for injection
may be used to obtain the desired concentration of solution. Sodium
chloride, as well as other excipients, may also be present, if
desired. Such excipients, however, must maintain the overall
stability of the amylin agonist or amylin peptide.
[0016] The liquid formulations of the invention should be
substantially isotonic. An isotonic solution may be defined as a
solution that has a concentration of electrolytes,
non-electrolytes, or a combination of the two that will exert
equivalent osmotic pressure as that into which it is being
introduced, here, for example in the case of parenteral injection
of the formulation, a mammalian tissue. By "substantially isotonic"
is meant within .+-.20% of isotonicity, preferably within .+-.10%.
The formulated product is included within a container, typically,
for example, a vial, cartridge, prefilled syringe, or a two
chambered syringe, cartridge or disposable pen injector.
[0017] It has been found that this novel parenteral dosage form
surprisingly maintains the stability of the peptide for up to four
years at refrigerated temperatures, e.g., approximately 5.degree.
C., and over 30 days at room temperature, e.g., approximately
30.degree. C.
[0018] The inventors have further discovered that the above
described pharmaceutical formulation can be further mixed with
insulin product and retain short term stability. This mixture may
occur, e.g., in a syringe. This short term mixing compatibility is
extremely advantageous. This allows administration of a single
injection of an amylin agonist, or amylin, along with insulin to a
patient.
[0019] This invention describes formulations of amylin agonists, or
amylins, which are stabilized with respect to aggregation,
adsorption and degradation, and thus enhance preservation of their
biological activities, although formulations of either an amylin
agonist or an amylin which has physicochemical properties similar
to those of human amylin, which is poorly soluble and highly
unstable should also include a stabilizer, comprising 1.0 to 10%
(w/v) of a carbohydrate or a polyhydric alcohol, and a surfactant,
preferably about 0.05 to about 1.0% (w/v) of polysorbate 80 or
other non-ionic detergent, should be lyophilized promptly after
formulation, and should be used promptly upon reconstitution and
are therefore less preferred. Generally, the presently described
formulations may be lyophilized according to methods known in the
art.
[0020] A further aspect of the invention features a formulation
including an amylin agonist, mixed with a stabilizing compound
which reduces loss of biological potency of the peptide in, for
example, an amylin specific receptor binding assay, reduces loss of
biological activity as measured in, for example, the in vitro
soleus muscle bioassay, and general loss of material by, for
example, an HPLC assay, as compared to a formulation consisting of
the amylin alone.
[0021] In a related aspect, the invention features a method for
formulating amylin agonists that retain short term (e.g., up to 24
hours) mixing compatibility with insulin. Certain insulins, such as
insulin glargine (Lantus.TM.), may allow for longer term stability
(the term "insulin," as used in the presently described
formulations, refers to various types of insulins including, for
example, insulin glargine).
[0022] In another aspect, the present disclosure provides
therapeutic methods for treating patients, for example, a method
for treating a diabetic patient in need of such treatment
comprising administering a therapeutically effective amount of an
amylin or amylin agonist formulation described herein.
[0023] Other features and advantages of the invention will be
apparent from the following description of the preferred
embodiments thereof, and from the claims.
BRIEF DESCRIPTION OF THE DRAWINGS
[0024] FIG. 1 shows the degradation rates of pramlintide at various
pH formulations.
[0025] FIG. 2 shows the stability of the pramlintide formulation
over time at 30.degree. C.
[0026] FIG. 3 shows the stability of the pramlintide formulation
over time at 40.degree. C.
[0027] FIG. 4 shows the stability of the pramlintide formulation
over time at 50.degree. C.
[0028] FIG. 5 shows an Arrenhius plot of the K.sub.obs values from
FIGS. 2-4.
[0029] FIG. 6 shows plasma concentrations versus time for insulin
following mixing the amylin agonist peptide pramlintide formulated
according to Table A.
DETAILED DESCRIPTION
[0030] It is understood that the development of a suitable
pharmaceutical formulation for administration to a subject is
complex. A need exists in the art for pharmaceutical formulations
of amylin agonist peptides, or amylins (both are also referred to
herein as "peptides"), designed to provide single or multiple doses
having substantial stability when refrigerated and at room
temperature. Further, a need exists in the art for a liquid
pharmaceutical formulation packaged with a container/closure system
that also minimizes the physical and chemical degradation of such
peptides. Liquid formulations are suitable for, e.g., injection,
nasal or pulmonary delivery. A need also exists for a
pharmaceutical formulation for amylin agonist peptides, or amylins,
which can be mixed with insulin or other therapeutics, such as
antidiabetic agents prior to administration. The mixture of the
amylin or amylin agonist with insulin may be simultaneous, in
either order or concerted and sequential. In addition, this mixture
may occur prior to preparing the presently described formulations,
concurrently therewith or at a later time. Other formulations, such
as solid dosage forms (including powders), allow for administration
to a subject via oral, pulmonary, sublingual or buccal routes.
Formulations that entrap or coat the desired peptide in a polymer,
wax or fatty material are suitable for controlled release. See
e.g., U.S. Pat. No. 6,368,630 and related patents, U.S. Pat. No.
6,379,704 and related patents. Formulations which are produced in a
fine, solid particulate form (fine powders) may be suitable for
pulmonary or nasal delivery. Formulations that incorporate the
desired peptide in a patch or device are suitable for transdermal
delivery. The invention described and claimed herein meets these
needs.
[0031] Peptide drugs undergo physical and chemical degradation in
solution, and lose their biological activity. A preferred dosage
form reported in this invention minimizes the chemical degradation
of amylin, for example, pramlintide, through different pathways,
for example, deamidation and peptide bond hydrolysis, and keeps the
peptide biologically active for up to four years when stored at
approximately 5.degree. C. This dosage form is well-tolerated by
patients. The container/closure system used to store this
formulation also minimizes the physical loss of drug product
through adsorption onto the container surface or adsorption into
the rubber closure.
[0032] People with Type I diabetes must be treated with exogenous
insulin. Generally, persons who initially require insulin tend to
be younger than 30 years of age at the time of diagnosis, lean,
prone to developing ketoacidosis, and markedly hyperglycemic even
in the fasting state. Insulin is also indicated for Type II
diabetics who do not respond to diet and exercise therapy either
alone or in combination with oral hypoglycemic drugs. Insulin
therapy is also necessary in some Type II diabetic patients who are
subject to stresses such as infections, pregnancy, or surgery. In
Type II diabetics, doses of 10-20 units of intermediate-acting
insulins are occasionally needed to bring hyperglycemia under
control.
[0033] Any functional macromolecular component of a patient may
serve operationally as a drug receptor. A particularly important
group of drug receptors are proteins that normally serve as
receptors for endogenous regulatory ligands (e.g., hormones,
neurotransmitters). Many drugs act on such physiological receptors.
Those that mimic the effects of the endogenous regulatory compound
are termed agonists.
[0034] By "amylin agonist" is meant peptide analogues of human
amylin useful as agonists of amylin, including but not limited to
those amylin agonists which are represented by the formula
TABLE-US-00001
.sup.1A.sub.1-X-Asn-Thr-.sup.5Ala-Thr-Y-Ala-Thr-.sup.10Gln-Arg-Leu-B.sub.1-
-
Asn-.sup.15Phe-Leu-C.sub.1-D.sub.1-E.sub.1-.sup.20F.sub.1-G.sub.1-Asn-H.su-
b.1-Gly-.sup.25I.sub.1-J.sub.1-
Leu-K.sub.1-L.sub.1-.sup.30Thr-M.sub.1-Val-Gly-Ser-.sup.35Asn-Thr-Tyr-Z
wherein A.sub.1 is hydrogen Lys, Ser, Ala, des-.alpha.-amino Lys,
or acetylated Lys; B.sub.1 is Ala, Ser or Thr; C.sub.1 is Val, Leu
or Ile; D.sub.1 is His or Arg; E.sub.1 is Ser or Thr; F.sub.1 is
Ser, Thr, Gln or Asn; G.sub.1 is Asn, Gln or His; H.sub.1 is Phe,
Leu or Tyr; T.sub.1 is Ala or Pro; J, is Ile, Val, Ala or Leu;
K.sub.1 is Ser, Pro, Leu, Ile or Thr; L.sub.1 is Ser, Pro or Thr;
M.sub.1 is Asn, Asp or Gln; X and Y are independently selected
residues having side chains which are chemically bonded to each
other to form an intramolecular linkage; and Z is hydroxy, amino,
alkylamino, dialkylamino, cycloalkylamino, arylamino, aralkylamino,
alkyloxy, aryloxy or aralkyloxy; provided that (a) when A.sub.1 is
Lys, B.sub.1 is Ala, C.sub.1 is Val, D.sub.1 is His, E.sub.1 is
Ser, F.sub.1 is Ser, G.sub.1 is Asn, H.sub.1 is Phe, T.sub.1 is
Ala, J.sub.1 is Ile, K.sub.1 is Ser, L.sub.1 is Ser, and M.sub.1 is
Asn; (b) when A.sub.1 is Lys, B.sub.1 is Ala, C.sub.1 is Ile,
D.sub.1 is Arg, E.sub.1 is Ser, F.sub.1 is Ser, G.sub.1 is Asn,
H.sub.1 is Leu, T.sub.1 is Ala, J.sub.1 is Ile, K.sub.1 is Ser,
L.sub.1 is Pro, and M.sub.1 is Asn; (c) when A.sub.1 is Lys,
B.sub.1 is Ala, C.sub.1 is Val, D.sub.1 is Arg, E.sub.1 is Thr,
F.sub.1 is Ser, G.sub.1 is Asn, H.sub.1 is Leu, T.sub.1 is Ala,
J.sub.1 is Ile, K.sub.1 is Ser, L.sub.1 is Pro, and M.sub.1 is Asn;
(d) when A.sub.1 is Lys, B.sub.1 is Ala, C.sub.1 is Val, D.sub.1 is
Arg, E.sub.1 is Ser, F.sub.1 is Ser, G.sub.1 is Asn, H.sub.1 is
Leu, T.sub.1 is Pro, J.sub.1 is Val, K.sub.1 is Pro, L.sub.1 is
Pro, and M.sub.1 is Asn; (e) when A.sub.1 is Lys, B.sub.1 is Ala,
C.sub.1 is Val, D.sub.1 is His, E.sub.1 is Ser, F.sub.1 is Asn,
G.sub.1 is Asn, H.sub.1 is Leu, T.sub.1 is Pro, J, is Val, K.sub.1
is Ser, L.sub.1 is Pro and M.sub.1 is Asn; or (f) when A.sub.1 is
Lys, B.sub.1 is Thr, C.sub.1 is Val, D.sub.1 is Arg, E.sub.1 is
Ser, F.sub.1 is Ser, G.sub.1 is His, H.sub.1 is Leu, T.sub.1 is
Ala, J, is Ala, K.sub.1 is Leu, L.sub.1 is Pro and M.sub.1 is Asp;
then one or more of any of A.sub.1 to M.sub.1 is not an L-amino
acid and Z is not amino, it being further noted that single-proline
substituted peptides at T.sub.1, K.sub.1, and L.sub.1 are not
preferred.
[0035] Suitable side chains for X and Y include groups derived from
alkyl sulfhydryls which may form disulfide bonds; alkyl acids and
alkyl amines which may form cyclic lactams; alkyl aldehydes or
alkyl halides and alkylamines which may condense and be reduced to
form an alkyl amine bridge; or side chains which may be connected
to form an alkyl, alkenyl, alkynyl, ether or thioether bond.
Preferred alkyl chains include lower alkyl groups having from about
1 to about 6 carbon atoms.
[0036] As used herein, the following terms have the following
meanings unless expressly stated to the contrary: The term "alkyl"
refers to both straight- and branched-chain alkyl groups. The term
"lower alkyl" refers to both straight- and branched-chain alkyl
groups having a total of from 1 to 6 carbon atoms and includes
primary, secondary and tertiary alkyl groups. Typical lower alkyls
include, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl,
isobutyl, t-butyl, n-pentyl, n-hexyl, and the like. The term "aryl"
refers to carbocyclic aromatic groups of 6 to 14 carbon atoms such
as phenyl and naphthyl, as well as heterocyclic aromatic groups
containing 1 to 3 heteroatoms (nitrogen, oxygen, sulfur, etc.) such
as pyridyl, triazolopyrazine, pyrimidine and the like. The term
"aralkyl" refers to an "aryl" group of 6 to 10 carbon atoms
directly attached to an "alkyl" group of 1 to 4 carbon atoms and
includes for example benzyl, p-chlorobenzyl, p-methylbenzyl, and
2-phenylethyl. The term "cycloalkyl" refers to cyclic alkyl groups
of 5 to 8 carbon atoms.
[0037] Biologically active derivatives of the above formula agonist
analogues are also included in which the stereochemistry of
individual amino acids may be inverted from (L)/S to (D)/R at one
or more specific sites. Also included are the agonist analogues
modified by glycosylation of Asn, Ser and/or Thr residues.
[0038] Biologically active agonist analogues of amylin are included
which contain less peptide character. Such peptide mimetics may
include, for example, one or more of the following substitutions
for --CO--NH-- amide bonds: depsipeptides (--CO--O--),
iminomethylenes (--CH.sub.2--NH--), trans-alkenes (--CH.dbd.CH--),
-enaminonitriles (--C(.dbd.CH--CN)--NH--), thioamides (--CS--NH--),
thiomethylenes (--S--CH.sub.2-- or --CH.sub.2--S--), methylenes
(CH.sub.2--CH.sub.2--) and retro-amides (NH--CO--).
[0039] Amylin agonist compounds form salts with various inorganic
and organic acids and bases. Such salts include salts prepared with
organic and inorganic acids, for example, HCl, HBr,
H.sub.2SO.sub.4, H.sub.3PO.sub.4, trifluoroacetic acid, acetic
acid, formic acid, methanesulfonic acid, toluenesulfonic acid,
maleic acid, fumaric acid and camphorsulfonic acid. Salts prepared
with bases include, for example, ammonium salts, alkali metal salts
(such as sodium and potassium salts) and alkali earth salts (such
as calcium and magnesium salts). Acetate, hydrochloride, and
trifluoroacetate salts are preferred.
[0040] The salts may be formed by conventional means, as by
reacting the free acid or base forms of the product with one or
more equivalents of the appropriate base or acid in a solvent or
medium in which the salt is insoluble, or in a solvent such as
water which is then removed in vacuo or by freeze-drying or by
exchanging the ions of an existing salt for another ion on a
suitable ion exchange resin.
[0041] Amylin agonist compounds include various stereoisomers. In
the preferred compounds of this invention, the chiral centers on
the peptide backbone are all S.
[0042] Amylin agonists may be prepared by those of ordinary skill
in the art, as described in "Amylin Agonist Peptides and Uses
Therefor," U.S. Pat. No. 5,686,411, the contents of which is hereby
incorporated by reference in its entirety.
[0043] By "human amylin" is meant the 37 amino acid amylin set
forth in U.S. Pat. No. 5,357,052.
[0044] The nomenclature of the compounds of the present invention
can be used to indicate both the peptide that the sequence is based
on and the modifications made to any basic peptide amylin sequence,
such as human amylin. An amino acid preceded by a superscript
number indicates that the named amino acid replaces the amino acid
normally present at the amino acid position of the superscript in
the basic amino acid sequence. For example,
".sup.18Arg.sup.25,28Pro-h-amylin" refers to a peptide based on the
sequence of "h-amylin" or "human-amylin" having the following
substitutions: Arg replacing His at residue 18, Pro replacing Ala
at residue 25 and Pro replacing Ser at residue 28. The term
"des-1Lys-h-amylin" refers to a peptide based on the sequence of
human amylin, with the first, or N-terminal, amino acid
deleted.
[0045] In addition to the description of compounds pursuant to the
above formula, certain preferred compounds may be identified,
including .sup.25,28,29Pro-h-amylin,
.sup.18Arg.sup.25,28Pro-h-amylin,
des-.sup.1Lys.sup.18Arg.sup.25,28Pro-h-amylin,
des-.sup.1Lys-h-amylin and
.sup.25Pro.sup.26Val.sup.28,29Pro-h-amylin. These peptides
preferably have a reduced tendency to form aggregates or to
precipitate under pressure compared to human amylin.
[0046] Compounds described herein which are especially preferred
include .sup.18Arg.sup.25,28Pro-h-amylin,
des-.sup.1Lys.sup.18Arg.sup.25,28Pro-h-amylin,
.sup.18Arg.sup.25,28,29Pro-h-amylin,
des-.sup.1Lys.sup.18Arg.sup.25,28,29Pro-h-amylin,
.sup.25,28,29Pro-h-amylin, des-.sup.1Lys.sup.25,28,29Pro-h-amylin,
and .sup.25Pro.sup.26Val.sup.25,28Pro-h-amylin. Still further
amylin agonist peptide compounds include: [0047]
.sup.23Leu.sup.25Pro.sup.26Val.sup.21,29Pro-h-amylin; [0048]
.sup.23Leu.sup.25Pro.sup.26Val.sup.28Pro-h-amylin; [0049]
des-.sup.1Lys.sup.23Leu.sup.25Pro.sup.26Val.sup.28Pro-h-amylin;
[0050] .sup.18Arg.sup.23Leu.sup.25Pro.sup.26Val.sup.28Pro-h-amylin;
[0051] .sup.18Arg.sup.25,28,29Pro-h-amylin; [0052]
.sup.18Arg.sup.23Leu.sup.25,28Pro-h-amylin; [0053]
.sup.17Ile.sup.23Leu.sup.25,28,29Pro-h-amylin; [0054]
.sup.17Ile.sup.25,28,29Pro-h-amylin; [0055]
des-.sup.1Lys.sup.17Ile.sup.23Leu.sup.25,28,29Pro-h-amylin; [0056]
.sup.17Ile.sup.18Arg.sup.23Leu-h-amylin; [0057]
.sup.17Ile.sup.18Arg.sup.23Leu.sup.26Val.sup.29Pro-h-amylin; [0058]
.sup.17Ile.sup.18Arg.sup.23Leu.sup.25Pro.sup.26Val.sup.28,29Pro-h-amylin;
[0059]
.sup.13Thr.sup.21His.sup.23Leu.sup.26Ala.sup.28Leu.sup.29Pro.sup.3-
1Asp-h-amylin; [0060]
.sup.13Thr.sup.21His.sup.23Leu.sup.26Ala.sup.29Pro.sup.31Asp-h-amylin;
[0061]
des-.sup.1Lys.sup.13Thr.sup.21His.sup.23Leu.sup.26Ala.sup.28Pro.su-
p.31Asp-h-amylin; [0062]
.sup.13Thr.sup.18Arg.sup.21His.sup.23Leu.sup.26Ala.sup.29Pro.sup.31Asp-h--
amylin; [0063]
.sup.13Thr.sup.18Arg.sup.21His.sup.23Leu.sup.28,29Pro.sup.31Asp-h-amylin;
and, [0064]
.sup.13Thr.sup.18Arg.sup.21His.sup.23Leu.sup.25Pro.sup.26Ala.sup.28,29Pro-
.sup.31Asp-h-amylin.
[0065] .sup.25,28,29Pro-h-amylin which is also referred to as
"pramlintide," is a particularly preferred agonist of human amylin.
.sup.25,28,29Pro-h-amylin will be referred to as "pramlintide"
hereafter. Pramlintide is substantially different from and better
than human amylin, retaining the desired biological properties of
human amylin with superior attributes, including superior
pharmaceutical properties (L. S. L. Gaeta and T. J. Rink, Medicinal
Chemistry Research, 1994). Pramlintide and other amylin agonists
set forth herein are described and claimed in U.S. Pat. No.
5,686,411, issued Nov. 11, 1997.
[0066] Amylin agonists may be formulated into a stable, safe
pharmaceutical composition for administration to a patient. The
novel pharmaceutical formulations of the present invention may
comprise approximately 0.01 to 1.0% (w/v), preferably 0.05 to 1.0%,
of an amylin agonist, or amylin, approximately 0.02 to 0.5% (w/v)
of an acetate, phosphate, citrate or glutamate buffer allowing a pH
of the final composition of from about 3.0 to about 7.0;
approximately 1.0 to 10% (w/v) of a carbohydrate or polyhydric
alcohol tonicifier and, optionally, approximately 0.005 to 1.0%
(w/v) of a preservative selected from the group consisting of
m-cresol, benzyl alcohol, methyl, ethyl, propyl and butyl parabens
and phenol. Such a preservative is generally included if the
formulated peptide is to be included in a multiple use product.
[0067] In a particular embodiment of the present invention, a
pharmaceutical formulation of the present invention may contain a
range of concentrations of amylin agonist, e.g., between about
0.01% to about 98% w/w, or between about 1 to about 98% w/w, or
preferably between 80% and 90% w/w, or preferably between about
0.01% to about 50% w/w, or more preferably between about 10% to
about 25% w/w in this embodiment. A sufficient amount of water for
injection may be used to obtain the desired concentration of
solution.
[0068] Additional tonicifying agents such as sodium chloride, as
well as other known excipients, may also be present, if desired. It
is preferred, however, if such excipients maintain the overall
tonicity of the amylin or amylin agonist. An excipient may be
included in the presently described formulations at various
concentrations. For example, an excipient may be included in the
concentration range from about 0.02% to about 20% w/w, preferably
between about 0.02% and 0.5% w/w, about 0.02% to about 10% w/w, or
about 1% to about 20% w/w. In addition, similar to the present
formulations themselves, an excipient may be included in solid
(including powdered), liquid, semi-solid or gel form.
[0069] In another aspect, the present formulations may be composed
in various forms, e.g., solid, liquid, semisolid or liquid. The
term "solid", as used herein, is meant to encompass all normal uses
of this term including, for example, powders and lyophilized
formulations. In a related aspect, the presently described
formulations may be lyophilized.
[0070] The terms buffer, buffer solution and buffered solution,
when used with reference to hydrogen-ion concentration or pH, refer
to the ability of a system, particularly an aqueous solution, to
resist a change of pH on adding acid or alkali, or on dilution with
a solvent. Characteristic of buffered solutions, which undergo
small changes of pH on addition of acid or base, is the presence
either of a weak acid and a salt of the weak acid, or a weak base
and a salt of the weak base. An example of the former system is
acetic acid and sodium acetate. The change of pH is slight as long
as the amount of hydronium or hydroxyl ion added does not exceed
the capacity of the buffer system to neutralize it.
[0071] As described herein, a variety of liquid vehicles are
suitable for use in the present peptide formulations, for example,
water or an aqueous/organic solvent mixture or suspension.
[0072] The stability of a peptide formulation of the present
invention is enhanced by maintaining the pH of the formulation in
the range of about 3.0 to about 7.0 when in liquid form.
Preferably, the pH of the formulation is maintained in the range of
about 3.5 to 5.0, or about 3.5 to 6.5, most preferably from about
3.7 to 4.3, or about 3.8 to 4.2. A frequently preferred pH may be
about 4.0. While not seeking be bound by theory, it is presently
understood that where the pH of the pharmaceutical formulation
exceeds 5.5, chemical degradation of the peptide may be accelerated
such that the shelf life is less than about two years.
[0073] The buffer used in the practice of the present invention is
an acetate buffer (preferably at a final formulation concentration
of from about 1-5 to about 60 mM), phosphate buffer (preferably at
a final formulation concentration of from about 1-5 to about to
about 30 mM) or glutamate buffer (preferably at a final formulation
concentration of from about 1-5 to about to about 60 mM). The most
preferred buffer is acetate (preferably at a final formulation
concentration of from about 5 to about 30 mM).
[0074] A stabilizer may be included in the present formulation but,
and importantly, is not necessarily needed. If included, however, a
stabilizer useful in the practice of the present invention is a
carbohydrate or a polyhydric alcohol. The present inventors have
discovered that a suitable stabilizer useful in the practice of the
present invention is approximately 1.0 to 10% (w/v) of a
carbohydrate or polyhydric alcohol. The polyhydric alcohols and
carbohydrates share the same feature in their backbones, i.e.,
--CHOH--CHOH--, which is responsible for stabilizing the proteins.
The polyhydric alcohols include such compounds as sorbitol,
mannitol, glycerol, and polyethylene glycols (PEGs). These
compounds are straight-chain molecules. The carbohydrates, such as
mannose, ribose, sucrose, fructose, trehalose, maltose, inositol,
and lactose, on the other hand, are cyclic molecules that may
contain a keto or aldehyde group. These two classes of compounds
have been demonstrated to be effective in stabilizing protein
against denaturation caused by elevated temperature and by
freeze-thaw or freeze-drying processes. Suitable carbohydrates
include: galactose, arabinose, lactose or any other carbohydrate
which does not have an adverse affect on a diabetic patient, i.e.,
the carbohydrate is not metabolized to form unacceptably large
concentrations of glucose in the blood. Such carbohydrates are well
known in the art as suitable for diabetics. Sucrose and fructose
are suitable for use with amylins in non-diabetic applications
(e.g. treating obesity).
[0075] Preferably, if a stabilizer is included, the peptide of the
present invention is stabilized with a polyhydric alcohol such as
sorbitol, mannitol, inositol, glycerol, xylitol, and
polypropylene/ethylene glycol copolymer, as well as various
polyethylene glycols (PEG) of molecular weight 200, 400, 1450,
3350, 4000, 6000, and 8000). Mannitol is the preferred polyhydric
alcohol. Another useful feature of the lyophilized formulations of
the present invention is the maintenance of the tonicity of the
lyophilized formulations described herein with the same formulation
component that serves to maintain their stability. Mannitol is the
preferred polyhydric alcohol used for this purpose.
[0076] The United States Pharmacopeia (USP) states that
anti-microbial agents in bacteriostatic or fungistatic
concentrations must be added to preparations contained in multiple
dose containers. They must be present in adequate concentration at
the time of use to prevent the multiplication of microorganisms
inadvertently introduced into the preparation while withdrawing a
portion of the contents with a hypodermic needle and syringe, or
using other invasive means for delivery, such as pen injectors.
Antimicrobial agents should be evaluated to ensure compatibility
with all other components of the formula, and their activity should
be evaluated in the total formula to ensure that a particular agent
that is effective in one formulation is not ineffective in another.
It is not uncommon to find that a particular antimicrobial agent
will be effective in one formulation but not effective in another
formulation.
[0077] A preservative is, in the common pharmaceutical sense, a
substance that prevents or inhibits microbial growth and may be
added to pharmaceutical formulations for this purpose to avoid
consequent spoilage of the formulation by microorganisms. While the
amount of the preservative is not great, it may nevertheless affect
the overall stability of the peptide. Thus, even selection of a
preservative can be difficult.
[0078] While the preservative for use in the practice of the
present invention can range from 0.005 to 1.0% (w/v), the preferred
range for each preservative, alone or in combination with others,
is: benzyl alcohol (0.1-1.0%), or m-cresol (0.1-0.6%), or phenol
(0.1-0.8%) or combination of methyl (0.05-0.25%) and ethyl or
propyl or butyl (0.005%-0.03%) parabens. The parabens are lower
alkyl esters of para-hydroxybenzoic acid.
[0079] A detailed description of each preservative is set forth in
"Remington's Pharmaceutical Sciences" as well as Pharmaceutical
Dosage Forms: Parenteral Medications, Vol. 1, 1992, Avis et al.
[0080] Pramlintide (previously referred to as "AC-137") does not
have a tendency to adsorb onto the glass in a glass container when
in a liquid form, therefore, a surfactant is not required to
further stabilize the pharmaceutical formulation. However, with
regard to amylin agonists or amylins which do have such a tendency
when in liquid form, a surfactant should be used in their
formulation. These formulations may then be lyophilized.
Surfactants frequently cause denaturation of protein, both of
hydrophobic disruption and by salt bridge separation. Relatively
low concentrations of surfactant may exert a potent denaturing
activity, because of the strong interactions between surfactant
moieties and the reactive sites on proteins. However, judicious use
of this interaction can stabilize proteins against interfacial or
surface denaturation. Surfactants which could further stabilize the
peptide may optionally be present in the range of about 0.001 to
0.3% (w/v) of the total formulation and include polysorbate 80
(i.e., polyoxyethylene(20) sorbitan monooleate), CHAPS.RTM. (i.e.,
3-[(3-cholamidopropyl)dimethylammonio]1-propanesulfonate), Brij
(e.g., Brij 35, which is (polyoxyethylene (23) lauryl ether),
poloxamer, or another non-ionic surfactant.
[0081] It may also be desirable to add sodium chloride or other
salt to adjust the tonicity of the pharmaceutical formulation,
depending on the tonicifier selected. However, this is optional and
depends on the particular formulation selected. Parenteral
formulations must be isotonic or substantially isotonic otherwise
significant irritation and pain would occur at the site of
administration.
[0082] The vehicle of greatest importance for parenteral products
is water. Water of suitable quality for parenteral administration
must be prepared either by distillation or by reverse osmosis. Only
by these means is it possible to separate adequately various
liquid, gas and solid contaminating substances from water. Water
for injection is the preferred aqueous vehicle for use in the
pharmaceutical formulation of the present invention.
[0083] It is possible that other ingredients may be present in the
peptide pharmaceutical formulation of the present invention. Such
additional ingredients may include, e.g., wetting agents,
emulsifiers, oils, antioxidants, bulking agents, tonicity
modifiers, chelating agents, metal ions, oleaginous vehicles,
proteins (e.g., human serum albumin, gelatin or proteins) and a
zwitterion (e.g., an amino acid such as betaine, taurine, arginine,
glycine, lysine and histidine). Additionally, polymer solutions, or
mixtures with polymers provide the opportunity for controlled
release of the peptide. Such additional ingredients, of course,
should not adversely affect the overall stability of the
pharmaceutical formulation of the present invention.
[0084] Containers are also an integral part of the formulation of
an injection and may be considered a component, for there is no
container that is totally inert, or does not in some way affect the
liquid it contains, particularly if the liquid is aqueous.
Therefore, the selection of a container for a particular injection
must be based on a consideration of the composition of the
container, as well as of the solution, and the treatment to which
it will be subjected. Adsorption of the peptide to the glass
surface of the vial can also be minimized, if necessary, by use of
borosilicate glass, for example, Wheaton Type I borosilicate glass
#33 (Wheaton Type 1-33) or its equivalent (Wheaton Glass Co.).
Other vendors of similar borosilicate glass vials and cartridges
acceptable for manufacture include Kimbel Glass Co., West Co.,
Bunder Glas GMBH and Form a Vitrum. The biological and chemical
properties of amylin may be stabilized by formulation and
lyophilization in a Wheaton Type 1-33 borosilicate serum vial to a
final concentration of 0.1 mg/ml and 10 mg/ml of amylin in the
presence of 5% mannitol, and 0.02% Tween 80.
[0085] In order to permit introduction of a needle from a
hypodermic syringe into a multiple-dose vial and provide for
resealing as soon as the needle is withdrawn, the open end of each
vial is preferably sealed with a rubber stopper closure held in
place by an aluminum band.
[0086] Stoppers for glass vials, such as, West 4416/50, 4416/50
(Teflon faced) and 4406/40, Abbott 5139 or any equivalent stopper
can be used as the closure for pharmaceutical for injection. These
stoppers are compatible with the peptide as well as the other
components of the formulation. The inventors have also discovered
that these stoppers pass the stopper integrity test when tested
using patient use patterns, e.g., the stopper can withstand at
least about 100 injections. Alternatively, the peptide can be
lyophilized in to vials, syringes or cartridges for subsequent
reconstitution. Liquid formulations of the present invention can be
filled into one or two chambered cartridges, or one or two chamber
syringes.
[0087] Each of the components of the pharmaceutical formulation
described above is known in the art and is described in
Pharmaceutical Dosage Forms: Parenteral Medications, Vol. 1, 2nd
ed., Avis et al. Ed., Mercel Dekker, New York, N.Y. 1992, which is
incorporated by reference in its entirety herein.
[0088] The manufacturing process for the above liquid formulations
generally involves compounding, sterile filtration and filling
steps. The compounding procedure involves dissolution of
ingredients in a specific order (preservative followed by
stabilizer/tonicity agents, buffers and pramlintide) or dissolving
at the same time.
[0089] Alternative formulations, e.g., non-parenteral, may not
require sterilization. However, if sterilization is desired or
necessary, any suitable sterilization process can be used in
developing the peptide pharmaceutical formulation of the present
invention. Typical sterilization processes include filtration,
steam (moist heat), dry heat, gases (e.g., ethylene oxide,
formaldehyde, chlorine dioxide, propylene oxide,
beta-propiolacctone, ozone, chloropicrin, peracetic acid methyl
bromide and the like), exposure to a radiation source, and aseptic
handling. Filtration is the preferred method of sterilization for
liquid formulations of the present invention. The sterile
filtration involves filtration through 0.45 .mu.m and 0.22 .mu.m (1
or 2) which may be connected in series. After filtration, the
solution is filled into appropriate vials or containers.
[0090] The liquid pharmaceutical formulations of the present
invention are intended for parenteral administration. Suitable
routes of administration include intramuscular, intravenous,
subcutaneous, intradermal, intraarticular, intrathecal and the
like. The subcutaneous route of administration is preferred.
Mucosal delivery is also preferred. These routes include, but are
not limited to, oral, nasal, sublingual, pulmonary and buccal
routes which may include administration of the peptide in liquid,
semi-solid or solid form. Administration via these routes requires
substantially more peptide to obtain the desired biological effects
due to decreased bioavailability compared to parenteral delivery.
In addition, parenteral controlled release delivery can be achieved
by forming polymeric microcapsules, matrices, solutions, implants
and devices and administering them parenterally or by surgical
means. These dosage forms would typically have a lower
bioavailability due to entrapment of some of the peptide in the
polymer matrix or device. See e.g., U.S. Pat. Nos. 6,379,704,
6,379,703, and 6,296,842.
[0091] Some formulations of the present invention are especially
advantageous because they retain short term mixing compatibility
with insulin, or in the case of insulins formulated at similar pH,
for longer term storage. Currently, there are over thirty insulin
products available in the United States. All regular insulin
preparations in the United States are now supplied at neutral pH.
This has resulted in improved stability of the hormone, and
patients need no longer refrigerate the vial of insulin in use.
Furthermore, neutral regular insulin can be mixed in any desired
proportion with other, modified insulin preparations since all
marketed insulin preparations will be at the same pH. Preparations
of insulin have been divided into three general categories
according to promptness, duration, and intensity of action
following subcutaneous administration. They are classified as
fast-, intermediate-, and long-acting insulins. There are also
various types of insulins within these categories. They include
regular insulins, protamine zinc insulins, NPH insulins, semilente
insulins (prompt insulin zinc suspensions), lente insulins (insulin
zinc suspensions), and ultralente insulins (extended insulin zinc
suspensions). Recently, a new type of insulin, insulin glargine
(Lantus.RTM.), formulated at a pH of approximately 4.0 has become
available, and may allow for longer term storage of peptide-insulin
mixtures.
[0092] Crystalline insulin is prepared by the precipitation of the
hormone in the presence of zinc (as zinc chloride) in a suitable
buffer medium. Crystalline insulin when dissolved in water is also
known as regular insulin. Following subcutaneous injection it is
rapidly absorbed (15-60 minutes). Its action is prompt in onset and
relatively short in duration, i.e., it reaches its peak effect in
about 1.5 to 4 hours, and lasts for about 5-9 hours.
[0093] By permitting insulin and zinc to react with the basic
protein protamine, Hagedorn and associates prepared a protein
complex, protamine zinc insulin. When this complex is injected
subcutaneously in an aqueous suspension, it dissolves only slowly
at the site of deposition, and the insulin is absorbed at a
retarded but steady rate. Protamine zinc suspension insulin has
largely been replaced by isophane insulin suspension, also known as
NPH insulin; the N denotes a neutral solution (pH 7.2), the P
refers to the protamine zinc insulin content, and the H signifies
the origin in Hagedom's laboratory. It is a modified protamine zinc
insulin suspension that is crystalline. The concentrations of
insulin, protamine, and zinc are so arranged that the preparation
has an onset and a duration of action intermediate between those of
regular insulin and protamine zinc insulin suspension. Its effects
on blood sugar are indistinguishable from those of an
extemporaneous mixture of 2 to 3 units of regular insulin and 1
unit of protamine zinc insulin suspension.
[0094] Chemical studies have revealed that the solubility of
insulin is determined in important measure by its physical state
(amorphous, crystalline, size of the crystals) and by the zinc
content and the nature of the buffer in which it is suspended.
Insulin can thus be prepared in a slowly absorbed, slow-acting form
without the use of other proteins, such as protamine, to bind it.
Large crystals of insulin with high zinc content, when collected
and resuspended in a solution of sodium acetate-sodium chloride (pH
7.2 to 7.5), are slowly absorbed after subcutaneous injection and
exert an action of long duration. This crystal preparation is named
extended insulin zinc suspension (ultralente insulin). Amorphous
insulin precipitated at high pH is almost as rapid in onset as
regular insulin, but has a somewhat longer duration of action. This
amorphous preparation is named prompt insulin zinc suspension
(semilente insulin). These two forms of insulin may be mixed to
yield a stable mixture of crystalline (7 parts) and amorphous (3
parts) insulin--called insulin zinc suspension (lente
insulin)--that is intermediate in onset and duration of action
between semilente and ultralente preparations and is similar to NPH
insulin.
[0095] In summary, the fast-acting insulins include the regular
insulins and the prompt insulin zinc suspensions (semilente
insulins). The intermediate-acting insulins include the isophane
insulin suspensions (NPH insulins, isophane insulin) and the
insulin zinc suspensions (lente insulins). The long-acting insulins
include protamine zinc insulin suspensions, and extended insulin
zinc suspensions (ultralente insulins). Most of these preparations
are available as either porcine or bovine insulins. Human insulins
of recombinant DNA origin are available as regular and isophane
insulins and as insulin zinc suspensions. Recently, a modified
insulin (Lys(B28), Pro(B29) human insulin analog, created by
reversing the amino acids at positions 28 and 29 on the insulin
B-chain) has been introduced. It is a fast-acting insulin, with a
more rapid onset of glucose lowering action, an earlier peak
action, and a shorter duration of action than regular human
insulin.
[0096] Many insulins are available from a number of companies.
These include Eli Lilly & Company and Novo Nordisk, two of the
largest suppliers of insulin in the world. Fast-acting insulins
available from Eli Lilly include (1) Iletin.RTM. I (Regular); (2)
Regular Iletin.RTM. II (Pork, 100 Units); (3) Regular Iletin.RTM.
II (Concentrated, Pork, 500 Units); (4) Humalog.RTM. Injection
(insulin lyspro, recombinant DNA origin); and (5) Humulin.RTM. R
(regular insulin, recombinant DNA origin, 100 Units). Fast-acting
insulins available from Novo Nordisk include (1) Novolin.RTM. R
(Regular, Human Insulin Injection (recombinant DNA origin) 100
Units); (2) Novolin.RTM. R PenFill 1.5 ml Cartridges (Regular,
Human Insulin Injection (recombinant DNA origin) 100 Units); (3)
Novolin.RTM. R Prefilled.TM. (Regular, Human Insulin Injection
(recombinant DNA origin) in a 1.5 ml Prefilled Syringe, 100
units/ml); (4) Regular Purified Pork Insulin (100 Units/ml); and
(5) Velosulin.RTM. BR (Buffered Regular Human Insulin Injection,
100 Units/ml). Intermediate-acting insulins available from Eli
Lilly include (1) Humulin.RTM. 50/50 (50% human insulin isophane
suspension and 50% human insulin injection (rDNA origin), 100
Units); (2) Humulin.RTM. 70/30 (70% human insulin isophane
suspension and 30% human insulin injection (rDNA origin), 100
Units); (3) Humulin.RTM. L (lente; human insulin (rDNA origin) zinc
suspension, 100 Units); (4) Humulin.RTM. N(NPH; human insulin (rDNA
origin) isophane suspension, 100 Units); (5) Lente.RTM. Iletin.RTM.
I, (insulin zinc suspension, beef-pork); (6) NPH Iletin.RTM. I
(isophane insulin suspension, beef-pork); (7) Lente Iletin.RTM. II
NPH (insulin zinc suspension, purified pork); and (8) NPH
Iletin.RTM. II, (isophane insulin suspension, purified pork).
Intermediate-acting insulins available from Novo Nordisk include
(1) Novolin.RTM. L (Lente, Human Insulin Zinc Suspension
(recombinant DNA origin), 100 Units/ml); (2) Novolin.RTM. N(NPH,
Human Insulin Isophane Suspension (recombinant DNA origin), 100
Units/ml); (3) Novolin.RTM. N PenFill.RTM. 1.5 ml Cartridges; (4)
Novolin.RTM. N Prefilled.TM. (NPH, Human Insulin Isophane
Suspension (recombinant DNA origin) in a 1.5 ml Prefilled Syringe,
100 Units/ml); (5) Novolin.RTM. 70/30 (70% NPH, Human Insulin
Isophane Suspension and 30% Regular, Human Insulin Injection
(recombinant DNA origin), 100 Units/ml); (6) Novolin.RTM. 70/30
PenFill.RTM. 1.5 ml Cartridges; (7) Novolin.RTM. 70/30
Prefilled.TM. (70% NPH, Human Insulin Isophane Suspension and 30%
Regular, Human Insulin Injection (recombinant DNA origin) in a 1.5
ml Prefilled Syringe, 100 Units/ml); (8) Lente Purified Pork
Insulin (Zinc Suspension, USP 100 Units/ml); and (9) NPH Purified
Pork Isophane Insulin Suspension (100 Units/ml). Long acting
insulins include Eli Lilly's Humulin.RTM. U (Ultralente.RTM. human
insulin (recombinant DNA origin) extended zinc suspension); and
insulin glargine (Lantus.TM.)(Aventis).
[0097] This invention provides unique peptide formulations,
preferably pramlintide or other amylin agonist peptide
formulations, which facilitate the short-term mixing compatibility
of all types of insulin products including regular insulin products
(e.g., Humulin.RTM. R and Novolin.RTM. R), intermediate-acting
insulin products (e.g., Humulin.RTM. 70/30 and Novolin.RTM. 70/30)
and long-acting insulin products (e.g., Humulin.RTM. U) with the
peptide formulation before administration. The desired
bioavailability of insulin and amylin agonist peptide or amylin is
maintained. This results in a significant reduction in the number
of injections per day in patients who are undergoing treatment with
insulin and an amylin agonist (e.g., pramlintide) or amylin.
[0098] Insulin as well as insulin analogs are useful in the
practice of the present invention. Monomeric insulin analogs have
been developed, e.g., Lys.sup.B28Pro.sup.B29-human insulin, which
is also referred to as "LysPro insulin" or "insulin lispro." These
analogs are said to be advantageous because they can be stabilized
by ligands which induce the otherwise monomeric analog to associate
under pharmaceutically useful conditions. This stabilized analog
remains fast acting in an associated state.
[0099] Thus, any type of insulin may be drawn into a syringe along
with the peptide formulation of the present invention. Any order of
mixing or introduction of peptide and insulin into a syringe is
also possible, such as simultaneous or concerted and sequential
admixing, but the preferred order is to place the insulin in the
syringe first, followed by peptide in order to reduce the potential
for cross-contamination and precipitation.
[0100] The amount of insulin and peptide formulation in the syringe
depends on the individual needs of a particular patient.
Accordingly, the amount present in a syringe is an amount
sufficient to maintain a proper insulin level for a patient. The
pharmaceutical formulations of the present invention can be
administered to any human or mammal in need of such treatment.
[0101] As has been demonstrated with pramlintide, and depending
upon the type of insulin and the volume ratio of the mixture, the
insulin and peptide formulation can remain in the same syringe for
at least about 24 hours and the insulin and the peptide will retain
their activity and stability.
[0102] For instance, regular insulin products may be mixed with a
peptide, e.g., pramlintide, at a pH of approximately 4.0 with 20 or
30 mM acetate buffer to maintain the solubility of the insulin. The
pH of the mixture would then be less than 4.5. A preferred peptide
formulation of pramlintide with a higher buffer capacity (30 mM
acetate) and at low label strength, e.g., 0.1 mg/ml, forms a clear
solution instantaneously (under a minute) when mixed with regular
insulin products in the range of five to 20 units. This low label
strength of pramlintide results in a high dose volume, 300 .mu.L.
This increased volume may be advantageous for bringing the pH down
to less than 4.5 almost immediately by increasing insulin dilution
factor and facilitating the transition of insulin from hexamer to
monomer before injection. This modulation of insulin is believed to
be advantageous in permitting increased rate of absorption and
causing rapid-time action without affecting bioavailability of
insulin. This effect may approach the rapid-time action observed in
LyS.sup.B28Pro.sup.B29 human insulin.
[0103] Peptide formulations, e.g., pramlintide, at a pH from 4.0 to
5.5, 2 to 30 mM buffer concentration and high potency, can be mixed
with regular insulin products before injection to yield solutions
with a pH greater than 6.8 so that the properties of insulin are
not affected. These mixtures would not affect the rate of
absorption or bioavailability of insulin nor the bioavailability of
the peptide.
[0104] An alternative to mixing of the peptide with insulin prior
to administration is to utilize a two chambered cartridge or
syringe. In such a system, the peptide (e.g. pramlintide) is filled
during manufacturing into one cartridge or syringe chamber in
contact with the plunger. In this cartridge or syringe, a divider
comprised of rubber or another suitable material known in the art,
preferably isolates the solution of peptide contained in the first
chamber from the second chamber. Prior to administration, the
needed amount of insulin is measured into the second chamber.
Preferably, the insulin is measured into the two chambered
cartridge or syringe immediately prior to administration to a
subject. When both chambers are filled with the appropriate amount
of peptide and insulin, the two chambers may be administered to a
subject, together or in series.
[0105] Another alternative is the formulation of peptide for use
with insulin pumps. Patients using insulin pumps frequently must
administer an extra dose of insulin in close proximity to meals.
Amylin peptides are typically also administered at this time.
Formulations of amylin peptide can be filled into cartridges or
syringes or other devices that allow the user of an insulin pump to
co-administer the peptide as needed.
[0106] Formulations of the invention are generally described above.
Below are provided examples of various formulations useful in the
invention. These examples are not limiting to the invention and
those of ordinary skill in the art can readily construct other
formulations within the ambit of the claims.
[0107] The invention will now be described in greater detail by
reference to the following non-limiting examples.
EXAMPLES
Example 1
Liquid Pramlintide Formulation
[0108] This example describes two preferred liquid formulations for
pramlintide. Major degradation pathways for the peptide are
deamidation and peptide bond hydrolysis. Therefore, the stability
of the peptide was investigated in the pH region of 4.0-5.5 at
45.degree. C. The pH-rate profile for the peptide in 60 mM acetate
buffer, 4.1% mannitol, 0.3% m-cresol is shown in FIG. 1. It can be
observed from this figure that Pro.sup.25,28,29 h-amylin over the
pH range studied is most stable at pH 4.0. The following
formulation was developed:
TABLE-US-00002 TABLE A INGREDIENT Weight (%)/Range Pramlintide
0.01-0.2 Acetate (30 mM, pH 4.0 .+-. 0.1): sodium acetate
trihydrate 0.061 glacial acetic acid 0.153 mannitol 4.3 m-cresol
0.225 Water For Injection (qs) 100 mL
[0109] The above formulation with 0.01% drug showed an acceptably
low irritancy in a rabbit subcutaneous irritancy study. The placebo
of this formulation when tested in humans also showed an acceptably
high level of tolerability and low irritancy. The preservative used
in the formulation meets BP criteria for preservative efficacy at
m-cresol levels as low as 0.15%.
[0110] The stability of the above formulation with 0.01% peptide
was evaluated at 4.degree. C. and accelerated conditions,
30.degree. C., 40.degree. C. and 50.degree. C. Based on strong
cation exchange HPLC analysis for purity and potency,
semi-logarithmic plots of % Initial purity or potency versus time
for the peptide were constructed as shown in FIGS. 2, 3 and 4. It
can be observed that the degradation of the peptide follows
pseudo-first order or zero order kinetics and that the degradation
rate (represented by slopes of these plots), increases
significantly with an increase in temperature. FIG. 5 shows plot of
ln(k.sub.obs) versus 1/Temperature, constructed based on the
Arrhenius relationship. It can be observed from this plot that the
degradation of pramlintide in the formulation follows Arrhenius
kinetics. By extrapolation of the plot to 5.degree. C. rate
constant was calculated and shelf-life of up to four years for the
peptide was predicted using this rate constant. Approximately 10%
loss of potency and/or 5% to 7% degradation was considered
acceptable. The shelf-life of the pharmaceutical formulation at
30.degree. C. based on direct measurement is at least 60 days.
Example 2
Liquid Pramlintide Formulation
[0111] Table B describes second the peptide formulation with a
shelf-life of greater than 4 years at 4.degree. C. and greater than
60 days at 30.degree. C. This formulation differs from the one in
Table A in acetate buffer concentration. This formulation also
showed no irritancy in a rabbit subcutaneous irritancy study.
Additionally, the placebo did not show significant irritancy in
humans. The shelf-lives of these formulations are at least as great
as the formulation given in Table A, both formulations being novel
parenteral peptide dosage forms with substantial shelf-lives.
TABLE-US-00003 TABLE B INGREDIENT Weight (%)/Range Pramlintide
0.01-0.2 Acetate (20 mM, pH 4.0 .+-. 0.1): sodium acetate
trihydrate 0.049 glacial acetic acid 0.0985 mannitol 4.3 m-cresol
0.225 Water for injection (qs) 100 mL
Example 3
Liquid Pramlintide Formulation and Insulin
[0112] The formulations reported in Tables A and B are also
compatible when mixed in a syringe with commercially available
insulin products. Table C provides results of insulin compatibility
study with Humulin.RTM. R. The results indicate the time to form a
clear solution and criteria for compatibility, when the peptide
formulation is mixed with Humulin R in a specific ratio in a
syringe. As outlined by Brange et al, "Insulin Structure and
Stability," in Stability and Characterization of Protein and
Peptide Drugs: Case Histories, 1993, Wang et al. (Ed), Plenum
Press, NY, insulin has an isoelectric precipitation zone pH range
of 4.5-6.5, within 1 pH unit from isoelectric point. Therefore,
while not seeking to be bound by theory, if the pH of the insulin
product drops from 7.2.+-.0.2 to pH in the isoelectric
precipitation range it may cause insulin to precipitate. Thus,
clarity of the solution mixture was used as a criteria for
preliminary compatibility studies. The insulin volumes used in this
study (Table C) cover the maximum and minimum ranges typical for
Type I diabetic patients. The formulation with higher buffer
capacity (Table C) forms a clear solution faster than the lower
buffer capacity formulation. However, at 0.01% of the peptide label
strength both formulations form a clear solution with insulin
within approximately one minute. In a syringe, one minute can be
considered to be instantaneous mixing. The higher label strength of
the peptide (0.015% or higher) results in lower dose volume and may
require longer than a minute to form a clear solution. The final pH
of all solution mixtures fall outside of insulin isoelectric
precipitation range (pH 4.5-6.5).
TABLE-US-00004 TABLE C Pramlintide Formulation Insulin: Variants
Pramlintide Time to Form Label Acetate Mixing Volume pH of the
Clear Solution Strength (%) (mM) Ratio (.mu.L:.mu.L) Mixture (sec)
0.01 30 50 (5 units):300 4.03 0 150:300 4.10 15 200:300 4.13 35 20
50:300 4.09 0 150:300 4.15 22 200:300 4.20 60 0.015 30 50:200 4.05
0 150:200 4.12 99 200:200 4.24 192 20 50:200 4.13 10 150:200 4.27
197 200:200 4.30 300
[0113] The formulation with 0.03% label strength (Table A) was
subjected to in vivo compatibility study in rats. FIG. 6 shows the
plasma concentration versus time for insulin. The study indicated
no significant differences in the bioavailability of either insulin
or peptide when injected together after mixing or separately as
individual injections. Therefore, these novel formulations are not
only stable but also compatible with regular insulin products such
as, Humulin.RTM. R and Novolin.RTM. R.
[0114] The preferred formulations (Tables A and B) were also tested
for compatibility with intermediate acting (Humulin.RTM. 70/30) and
long acting (Humulin.RTM. N) insulin products. These two insulin
products are suspensions and they both contain soluble and
insoluble insulin portions in specific ratios, and these ratios are
required to achieve targeted bioavailability. Therefore, when mixed
with the peptide formulation the ratio of soluble to insoluble
insulin is preferably maintained. The sample preparation to
separate soluble to insoluble portions from peptide/insulin
mixtures was adopted from Arakawa et al., Diabetes Research and
Clinical Practice, 1989. Samples were analyzed using a reverse
phase HPLC assay. Table D summarizes the results of the study. Both
formulations maintain the soluble to insoluble ratios as in control
formulations when Humulin.RTM. 70/30 and Pro.sup.25".sup.28".sup.29
h-amylin formulations are mixed in a 3:1 ratio. Therefore, in a
preferred formulation the dose volume should be maintained by
choosing an appropriate peptide label strength. For example, using
0.6 mg/mL or 0.06% label strength formulation to mix with 15 units
of Humulin 70/30.
TABLE-US-00005 TABLE D Pramlintide Formulation Variants (Label
Strength: 0.3 Insulin: mg/mL or 0.03%) Pramlintide Percent Acetate
Ratio pH of the Soluble % Soluble pH (mM) (.mu.L:.mu.L) Mixture
Insulin Pramlintide 4.0 20 300(30 6.77 12 101 Units):100 150:100
5.74 4 100 4.0 30 300:100 6.41 10 97 150:100 5.15 4 98 Humulin
N:Humulin R N/A 7.12 11 N/A (70:30) (control) Premixed Humulin
100:0 7.46 16 N/A 70/30 (control)
[0115] Table E shows compatibility data for formulations with long
acting insulin product, Humulin.RTM. N. The Humulin.RTM. N covers a
normal dosage range for Type I diabetic patients. The soluble to
insoluble ratios do not change significantly compared to the
control when preferred formulations, in Tables A and B, are mixed
with Humulin.RTM. N. The preferred peptide dose volume is 100
.mu.L.
[0116] The formulations described in Tables A and B of Example 1
are preferred formulations with necessary stability and insulin
compatibility requirements.
TABLE-US-00006 TABLE E Pramlintide Formulation Variants (Label
Strength: 0.3 Insulin: mg/mL or 0.03%) Pramlintide Percent Acetate
Ratio pH of the Soluble % Soluble pH (mM) (.mu.L:.mu.L) Mixture
Insulin Pramlintide 4.0 20 160(16 5.51 2 100 Units):100 120:100
5.10 4 105 4.0 30 160:100 5.06 3 99 120:100 4.82 4 97 Humulin N
(control) 100:0 7.20 2 N/A
[0117] As provided above, a co-formulation with insulin glargine,
formulated at approximately pH 4.0, provides an alternative mixture
which exhibits long term stability and allows for storage of the
formulation for greater than 24 hours.
Example 4
Liquid Peptide Formulation and Insulin
[0118] The formulation in this example is the preferred formulation
for mixing with intermediate acting (Humulin.RTM. 70/30 &
Novolin.RTM. 70/30) and long acting (Humulin.RTM. N and
Novolin.RTM. N) insulin products.
TABLE-US-00007 TABLE F INGREDIENT Weight (%)/Range Pramlintide 0.03
Acetate (12 mM, pH 4.4) sodium acetate trihydrate 0.0555 glacial
acetic acid 0.0476 mannitol 4.4 m-cresol 0.225 Water for injection
(qs) 100 mL
[0119] The stability of this formulation is 3.3 years at 5.degree.
C., calculated by extrapolation using accelerated stability data,
and found to be 33 days at 30.degree. C. using the real time data.
The insulin compatibility studies with Humulin.RTM. 70/30 and
Humulin.RTM. N were conducted following the procedure outlined in
Example 3. The formulation maintains insulin soluble to insoluble
ratios in the range of controls when mixed with Humulin.RTM. 70/30
(Table G) or Humulin.RTM. N (Table H). The pH of the mixture does
not deviate from the control samples.
TABLE-US-00008 TABLE G Pramlintide Formulation Variants (Label
Strength: 0.3 Insulin: mg/mL or 0.03%) Pramlintide Percent Acetate
Ratio pH of the Soluble % Soluble pH (mM) (.mu.L:.mu.L) Mixture
Insulin Pramlintide 4.4 12 300:100 7.02 13 88 150:100 6.82 11 92
Humulin .RTM. N: N/A 7.12 11 N/A Humulin .RTM. R (70:30) (Control)
Premixed Humulin .RTM. 100:0 7.46 16 N/A 70/30 (Control)
TABLE-US-00009 TABLE H Pramlintide Formulation Variants (Label
Strength: 0.3 Insulin: mg/mL or 0.03%) Pramlintide Percent Acetate
Ratio pH of the Soluble % Soluble pH (mM) (.mu.L:.mu.L) Mixture
Insulin Pramlintide 4.4 12 160:100 6.54 3 99 120:100 6.26 1 99
Humulin .RTM. N 100:0 7.20 2 N/A (Control)
Example 5
Liquid Pramlintide Formulation
[0120] This is a formulation with low buffer concentration, 10 mM
of acetate buffer and pH closer to physiological pH compared to the
formulations in examples 1-4. While not seeking to be bound by
theory, the stability of this formulation is calculated by
extrapolation to be 2.76 years 5.degree. C. based on accelerated
stability data and found to be at least 32 days at 30.degree. C.
using real time data.
TABLE-US-00010 TABLE I INGREDIENT Weight (%)/Range Pramlintide
0.01-0.4 Acetate (10 mM, pH 4.7) sodium acetate trihydrate 0.066
glacial acetic acid 0.0309 mannitol 4.5 m-cresol 0.225-0.3 Water
for injection (qs) 100 mL
Example 6
Lyophilized Peptide Formulation
[0121] Amylin agonists may be stabilized by lyophilization. In
addition, amylin agonist formulations described herein may be
stabilized by lyophilization. For example, see the formulation
listed in Table J. This formulation is intended for amylin agonists
or amylins having physicochemical characteristics that include
lower solubility and/or higher instability.
[0122] Pramlintide, however, a superior amylin agonist peptide, may
also be lyophilized in a formulation, e.g., listed in Table J.
TABLE-US-00011 TABLE J INGREDIENT WEIGHT (%)/RANGE Amylin Agonist
0.03-1.0 Mannitol 5.0 Polysorbate 80 0.02 Water for Injection 100
ml (qs)
Example 7
Liquid Peptide Formulation
[0123] Amylin agonists such as pramlintide may be minimally
formulated with 30 mM acetate buffer as described in Table K.
TABLE-US-00012 TABLE K INGREDIENT WEIGHT (%)/RANGE Pramlintide
0.01-0.5 Glacial Acetic Acid 0.153 Sodium Acetate 0.061
Trihydrate
Example 8
Liquid Peptide Formulation
[0124] Amylin agonists such as pramlintide may be minimally
formulated with a buffer. In addition, a penetration enhancer, a
preservative or a polymer may be added individually or in some
combination to facilitate nasal or buccal delivery in a liquid drop
or spray, as described in Table L. Alternate buffer systems, known
in the art, can also be used.
TABLE-US-00013 TABLE L INGREDIENT WEIGHT (%)/RANGE Pramlintide
0.01-1.0 Glacial Acetic Acid 0.153 Sodium Acetate 0.061 Trihydrate
Penetration enhancer As needed. Preservative As needed. Polymer As
needed.
[0125] A composition suitable for buccal, pulmonary and/or
sublingual delivery can be produced by formulating a solid dosage
form with solid peptide (i.e., a tablet), through lyophilizing the
peptide formulation of Table L to produce a solid form or adsorbing
this formulation on to a solid carrier (e.g. a powdered excipient,
such as lactose) or a polymer, a polymer mixture or a polymeric
substrate, if needed. See A. R. Gennaro et al., 17th Remington's
"Pharmaceutical Sciences."
[0126] Any of the above described processes of formulating a solid
dosage form will be suitable to produce sublingual tablets which
dissolve rapidly and deliver the solid peptide to the mucosa under
the tongue.
[0127] Pulmonary delivery is typically facilitated by inhalation of
a fine powder formulation or fine liquid droplets. In some cases, a
surfactant or enzyme inhibitor is included in the fine powder
formulation to prevent or slow the chemical degradation of the
peptide when in contact with biological tissues.
[0128] Formulations for transdermal delivery can be solid (e.g.,
powder), semi-solid, or liquid, and can be delivered via any
suitable technology, for example as described in U.S. Pat. Nos.
6,183,434, 6,142,939, 6,173,202, and related patents.
TABLE-US-00014 TABLE M INGREDIENT WEIGHT (%)/RANGE Pramlintide
0.01-90% Lactose 1.0-20% Stabilizer or enzyme As needed. inhibitor
Penetration enhancer As needed. Preservative As needed. Polymer As
needed.
[0129] Buccal formulations can be prepared in a similar manner as
the solid formulations described above, but with the addition of
bioadhesive polymers or related formulations that allow the dosage
formulation to adhere to the buccal mucosa. See e.g., U.S. Pat. No.
6,153,211.
[0130] Producing solid dosage forms for oral or sublingual
administration involves the utilization of a lyophilized or
crystalline form of the peptide which is preferably blended with
additional powders which are known in the art, to formulate a
capsule or tablet. Preferably, the solid dosage form additionally
includes an enzyme inhibitor, a complexing agent and/or stabilizer.
These ingredients are useful for protection of the peptide from
degradation in the gastrointestinal tract. Alternatively, the
capsule or tablet may be enteric coated or coated with a polymer
that releases the formulation of the peptide in a desired location
in the gastrointestinal tract. See e.g., A. R. Gennaro et al., 17th
Remington's "Pharmaceutical Sciences." A suitable formulation could
be created by modification of the formulation in Table M in the
above described manner.
Example 9
Injectable Controlled Release Formulation
[0131] It is advantageous to patients to utilize a formulation of
peptide that requires administration less frequently than multiple
times per day. Specifically, the peptide can be formulated in a
mixture with biocompatible polymers or oils that slowly release the
peptide in a timeframe from days to months. An example of these
formulations are shown on Table N and O. Table N represents an
injectable microsphere product and Table 0 represents an oil based
formulation.
TABLE-US-00015 TABLE N INGREDIENT WEIGHT (%)/RANGE Pramlintide
0.01-40.0 Glacial Acetic Acid 0.153 Sodium Acetate 0.061 Trihydrate
Sugar or polyhydric 0.1-10% alcohol Release modifier 0.005-2% (e.g.
Ammonium sulfate) Polymer 1-98% (e.g. PLGA)
TABLE-US-00016 TABLE O INGREDIENT WEIGHT (%)/RANGE Pramlintide
0.01-40.0 Oil vehicle 1-98% Non-aqueous solvent 0.01-90% Sugar or
polyhydric 0.1-10% alcohol
[0132] Any pharmaceutically acceptable emulsifiers, such as oils,
may be utilized in the formulation of Table 0 and other similar
formulations described herein. See e.g., Remington's
"Pharmaceutical Sciences"; and Idson, Pharmaceutical Emulsions, Ch.
6, Pharmaceutical Dosage Forms, Disperse Systems. Vol. 1., New
York, 1988.
Example 10
Transdermal Delivery
[0133] In certain instances it is beneficial to eliminate the need
for injections through the use of transdermal delivery of the
peptide. This may be facilitated through the formulation of the
peptide into a patch or device that is attached to the skin and
delivers the peptide into the skin at a desired rate.
[0134] All references cited herein are hereby incorporated by
reference in their entireties, whether previously specifically
incorporated or not. As used herein, the terms "a", "an", and "any"
are each intended to include both the singular and plural forms and
the term "or" is intended to refer to alternatives and
combinations.
[0135] Having now fully described this invention, it will be
appreciated by those skilled in the art that the same can be
performed within a wide range of equivalent parameters,
concentrations, and conditions without departing from the spirit
and scope of the invention and without undue experimentation.
[0136] While this invention has been described in connection with
specific embodiments thereof, it will be understood that it is
capable of further modifications. This application is intended to
cover any variations, uses, or adaptations of the invention
following, in general, the principles of the invention and
including such departures from the present disclosure as come
within known or customary practice within the art to which the
invention pertains and as may be applied to the essential features
hereinbefore set forth.
[0137] Citation of the above publications or documents is not
intended as an admission that any of the foregoing is pertinent
prior art, nor does it constitute any admission as to the contents
or date of these publications or documents.
* * * * *