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WO1999034764A2 - Compositions pharmaceutiques a agoniste d'amyline, contenant de l'insuline - Google Patents

Compositions pharmaceutiques a agoniste d'amyline, contenant de l'insuline Download PDF

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Publication number
WO1999034764A2
WO1999034764A2 PCT/US1998/000662 US9800662W WO9934764A2 WO 1999034764 A2 WO1999034764 A2 WO 1999034764A2 US 9800662 W US9800662 W US 9800662W WO 9934764 A2 WO9934764 A2 WO 9934764A2
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WIPO (PCT)
Prior art keywords
insulin
pramlintide
separate
nph
pram
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PCT/US1998/000662
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English (en)
Inventor
James L'italian
Shankar Musunuri
Cale Ruby
Orville Kolterman
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Amylin Pharmaceuticals, Inc.
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Filing date
Publication date
Application filed by Amylin Pharmaceuticals, Inc. filed Critical Amylin Pharmaceuticals, Inc.
Priority to AU59162/98A priority Critical patent/AU5916298A/en
Priority to PCT/US1998/000662 priority patent/WO1999034764A2/fr
Priority to EP98902526A priority patent/EP1051141A4/fr
Priority to ZA98221A priority patent/ZA98221B/xx
Publication of WO1999034764A2 publication Critical patent/WO1999034764A2/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/28—Insulins

Definitions

  • the present invention relates to pharmaceutical compositions containing an agonist of amylin and an intermediate-acting insulin. More particularly, the invention relates to pharmaceutical compositions containing one or more amylin agonists or amylins and one or more intermediate-acting insulins, such as NPH insulin. The invention also relates to the preparation and use of such pharmaceutical compositions in the treatment of a mammal, preferable a human, who use insulin to control their blood glucose concentration, particularly people with diabetes, either alone or in combination with another insulin or glucose-lowering agent.
  • Diabetes Diabetes mellitus is a major global health problem which is inadequately treated by available drugs.
  • the International Diabetes Federation has estimated that over 100 million people worldwide are afflicted with this disease. Diabetes costs the American economy over $100 billion annually, according to a study reported in the Journal of Clinical Endocrinology and Metabolism, which went on to say that ". . . health care expenditures for people with diabetes constituted about one in seven health care dollars spent in 1992.”
  • the American Medical Association reports that the incidence of diagnosed diabetes as a percentage of the American population has tripled since 1958, and that the total number of diagnosed and undiagnosed cases has grown to about 16 million. Diabetes is the name given to the clinical description of patients with a number of symptoms arising from raised blood glucose levels.
  • diabetes The metabolism of glucose involves many organs, in each of which several important metabolic steps occur. Key regulatory points include hepatic regulation of glucose uptake and release, muscle utilization of fuels, control by pancreatic production of insulin and glucagon, and neurogenic controls. Diabetes may arise from abnormalities at one or several sites in the complex feedback loops in this system. Two main types of diabetes can be distinguished: (1) insulin-dependent diabetes mellitus (previously termed “juvenile-onset,” and now called “IDDM” or “Type 1”), and (2) non-insulin-dependent diabetes mellitus (previously termed "maturity-onset,” and now called “NIDDM” or “Type 2").
  • IDDM insulin-dependent diabetes mellitus
  • NIDDM non-insulin-dependent diabetes mellitus
  • diabetes Other forms include (3) maturity-onset or non- insulin-dependent diabetes in the young (a rare dominantly inherited, mild type of disease) ; (4) diabetes mellitus or carbohydrate intolerance associated with certain genetic syndromes; (5) secondary diabetes mellitus (e.g., drug- induced, from pancreatic disease, hormonal or receptor abnormalities, etc.), and (6) gestational diabetes mellitus.
  • diabetes occurs when the pancreas no longer produces enough insulin, a hormone that regulates the metabolism of blood glucose.
  • Type 1 diabetes which afflicts about 10% of all people with diagnosed diabetes in developed countries, the pancreatic beta cells that make insulin have been destroyed.
  • Type 2 diabetes the insulin-producing cells are unable to produce enough insulin to compensate for the patient's poor sensitivity to the hormone in glucose-using tissues such as skeletal muscle (a condition called insulin resistance) .
  • insulin resistance a condition called insulin resistance
  • the insulin deficiency results in an abnormally high blood-glucose concentration (a condition called hyperglycemia) which is an important cause of the degenerative complications associated with diabetes, including blindness, kidney failure and nerve damage.
  • hyperglycemia plays a role in the development of heart disease .
  • Type 1 diabetes One of the main features of Type 1 diabetes is the sudden appearance in non-obese children or young adults of a severe disease which only responds satisfactorily to insulin therapy.
  • Type 2 patients tend to present at an older age, are often obese and then respond to diet, without need for insulin therapy.
  • the "juvenile” diabetes and “maturity” diabetes nomenclature based on the age of onset has fallen out of fashion with the realization that auto-immunity to the islets is the characteristic pathology of Type 1 diabetes, and that this is not confined to juvenile-onset but can occur in maturity-onset diabetes and may present at any age. However, the distinction based on age is still sometimes used as a shorthand description of presentation.
  • insulin-dependent and “non-insulin-dependent , " which relate to the empirical requirement for insulin therapy. This was introduced because some maturity-onset diabetic patients are of normal weight, have severe disease which requires insulin, and resemble juvenile-onset diabetic patients.
  • IDDM' is defined by the patient's dependence on insulin for survival, in practice this definition is often extended to include patients who require insulin therapy to prevent symptoms.
  • NIDDM is often restricted to patients who can be maintained symptom-free either by diet or by tablet therapy. This usage is not strictly correct because, as noted below, patients who present with NIDDM may later develop more severe diabetes that requires insulin therapy.
  • a Type 2 patient initially treated by diet or tablets, but later transferred to insulin is often termed an insulin-treated NIDDM patient. Insulin
  • insulin replacement therapy has played a central role in treating diabetes.
  • insulin injections are essential, since these patients would otherwise die.
  • oral medications that either stimulate greater insulin production or enhance insulin sensitivity may improve metabolic control.
  • As many as 20% of people with newly diagnosed Type 2 diabetes do not respond to oral therapy.
  • patients who do respond to oral therapy become progressively resistant over time, with as many as 10% each year ceasing to derive a therapeutic benefit.
  • an estimated 40% of people diagnosed with Type 2 diabetes are using insulin injections to manage their disease. It has been estimated that in North America, Europe and Japan alone, as many as two million people with Type 1 diabetes and five million people with Type 2 diabetes use insulin to help control their blood-glucose concentrations. Because insulin given by mouth is digested as a dietary protein, it has to be administered by injection.
  • Various advances and changes have been made in the United States and
  • 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.
  • protamine zinc insulin By permitting insulin and zinc to react with the basic protein protamine, Hagedorn and associates prepared a protein complex, protamine zinc insulin .
  • this complex 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 Hagedorn' 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.
  • 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 than 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.
  • 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.
  • 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 8 I
  • Humalog ® Injection insulin lyspro, recombinant DNA origin
  • Humulin * R regular insulin, recombinant DNA origin, 100 Units
  • Fast-acting insulins available from Novo Nordisk include (1) Novolin ® R (Regular, Human Insulin Injection (recombinant DNA origin) 100 Units) ; (2) Novolin 8 R PenFill 1.5 ml Cartridges (Regular, Human Insulin Injection (recombinant DNA origin) 100 Units) ; (3) Novolin” R PrefilledTM (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" BR (Buffered Regular Human Insulin Injection, 100 Units/ml) .
  • Novolin ® R Regular, Human Insulin Injection (recombinant DNA origin) 100 Units)
  • Intermediate-acting insulins available from Eli Lilly include (1) Humulin 8 50/50 (50% human insulin isophane suspension and 50% human insulin injection (rDNA origin) , 100 Units) ; (2) Humulin 8 70/30 (70% human insulin isophane suspension and 30% human insulin injection (rDNA origin) , 100 Units) ; (3) Humulin ® L (lente; human insulin (rDNA origin) zinc suspension, 100 Units) ; ) ; (4) Humulin 8 N (NPH; human insulin (rDNA origin) isophane suspension, 100 Units) ; (5) Lente ® Iletin I, (insulin zinc suspension, beef-pork); (6) NPH Iletin I (isophane insulin suspension, beef-pork) ; (7) Lente Iletin ® II (insulin zinc suspension, purified pork) ; and (8)
  • NPH Iletin ® II isophane insulin suspension, purified pork.
  • Novo Nordisk Intermediate-acting insulins available from Novo Nordisk include (1) Novolin ® L (Lente, Human Insulin Zinc Suspension (recombinant DNA origin) , 100 Units/ml) ; (2) Novolin ® N (NPH,
  • 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 ® U (Ultralente ® human insulin (recombinant DNA origin) extended zinc suspension) .
  • each insulin injection should be adjusted to reflect the person's pre-meal blood- glucose concentration, the carbohydrate content of the meal, and the individual's planned level of physical activity.
  • the basal insulin supply can be given to diabetic patients using a long-acting, crystalline insulin which is slowly absorbed.
  • normoglycemia a condition called normoglycemia
  • hypoglycemia a condition called hypoglycemia
  • patients need to be attentive to their life-style and assess the response to their insulin therapy by measuring their blood glucose. This is done by pricking a finger and placing the blood onto a strip containing the enzyme glucose oxidase; the glucose concentration is determined either by an electronic sensor or by a color change monitored visually.
  • Diet therapy inducing weight reduction, may be sufficient to reduce the blood glucose to below the renal threshold and to make people with Type 2 diabetes symptom- free, although it is rarely sufficient to induce normal fasting glucose levels. If symptoms persist despite dietary therapies, then most physicians treat with tablets containing sulphonylureas to stimulate insulin secretion. This approximately doubles the ⁇ -cell efficiency, but, nevertheless, continued symptom-free hyperglycemia with a fasting glucose level of 9-10 mmol/1 is common. "Second generation” drugs such as glibenclamide or glipizide are no more effective than "first generation” drugs such as tolbutamide or chlorpropamide . Biguanide therapy with metformin, to improve glucose uptake is an alternative, but like a sulphonylurea it only induces a modest decrease of blood glucose. If symptoms recur on diet and tablet therapy, patients are transferred to insulin therapy.
  • Amylin is a 37 amino acid protein hormone that is co-secreted with insulin from the beta cells of the pancreas in response to a meal. This hormone in healthy individuals is believed to work in concert with insulin in controlling glucose metabolism.
  • the structure and biology of amylin have previously been reviewed. See, for example, Rink _t al .. Trends in Pharmaceutical Sciences, 14:113-118 (1993); Gaeta and Rink, Med. Chem . Res . , 3:483-490 (1994); and, Pittner et al . , J". Cell . Biochem .
  • Amylin is the subject of United States Patent No. 5,367,052, issued November 22, 1995. Excess amylin action has been said to mimic key features of Type 2 diabetes and amylin blockade has been proposed as a novel therapeutic strategy. It has been disclosed in United States Patent No. 5,266,561, issued November 30, 1993, that amylin causes reduction in both basal and insulin-stimulated incorporation of labeled glucose into glycogen in skeletal muscle. The latter effect was also disclosed to be shared by calcitonin gene related peptide (CGRP) (see also Leighton and Cooper, Nature, 335:632-635 (1988)).
  • CGRP calcitonin gene related peptide
  • Amylin and CGRP were approximately equipotent, showing marked activity at 1 to 10 nM. Amylin is also reported to reduce insulin-stimulated uptake of glucose into skeletal muscle and reduce glycogen content (Young et al . , Amer. J. Physiol . , 259:45746-1 (1990) ) . The treatment of Type 2 diabetes and insulin resistance with amylin antagonists is disclosed.
  • amylin has been shown to be missing or deficient and combined replacement with insulin has been proposed as a preferred treatment over insulin alone in all forms of diabetes. It has been proposed that the lack of amylin contributes to poor glucose control, especially after eating. Indeed, amylin has been shown to have at least two effects believed to be important for normal glucose metabolism: it slows glucose inflow into the bloodstream from the gastrointestinal tract, and it suppresses glucagon secretion and thereby helps to lower glucose production by the liver.
  • glucose inflow rate is insulin, . which is secreted by pancreatic beta-cells in response to rising blood glucose concentrations.
  • the endocrine regulator of glucose inflow rate has, until recently, been unknown.
  • the simultaneous secretion of both insulin and amylin by the pancreatic beta-cells acts to regulate both inflow and outflow, thereby keeping post- meal blood glucose concentrations within a narrow and healthy range .
  • the liver produces glucose which is carried by the bloodstream to the brain and other tissues that do not store glucose.
  • the endocrine regulator of liver glucose production is glucagon, a peptide hormone secreted by pancreatic alpha-cells in response to falling blood glucose concentrations.
  • glucagon secretion must be suppressed to avoid hyperglycemia induced by excess liver glucose production, and a known regulator of glucagon suppression is insulin.
  • amylin too is an endocrine regulator of glucagon secretion.
  • increasing amylin blood concentrations slows pancreatic alpha-cell secretion of glucagon, an effect which amplifies the same regulatory effect of insulin.
  • Gedulin _____. Metabolism 46:67-70 (1997) .
  • the simultaneous secretion of both insulin and amylin by the pancreatic beta-cells can act to suppress glucagon and curtail liver glucose production, thereby helping to keep post-meal blood glucose concentrations within a narrow and healthy range.
  • the use of amylin and other amylin agonists for the treatment of diabetes mellitus is the subject of United States Patent No. 5,175,145, issued December 29, 1992. Pharmaceutical compositions containing amylin and amylin plus insulin are described in United States Patent No. 5,124,314, issued June 23, 1992.
  • amylin agonist pramlintide ( 25,28,29 Pro-human amylin, also previously referred to as "AC137")
  • AC137 a synthetic analog of human amylin in which select modifications have been made
  • amylin agonists for example, the amylin agonist analog 25,28, 9 Pro-h-amylin (also known as “pramlintide” and previously referred to as “AC- 0137”)
  • an intermediate-acting insulin zinc suspension or protamine zinc insulin for example, lente insulin or NPH insulin
  • the present invention is directed to novel pharmaceutical mixtures of amylin agonists or amylins and intermediate-acting insulins. These pharmaceuticals have unique attributes when used to treat people with diabetes. As described more fully herein, this amylin agonist/intermediate-acting insulin mixture leads to a glucose lowering effect of greater magnitude and longer duration in comparison to the glucose lowering effects of equivalent amounts of these drugs administered separately.
  • the invention also includes methods for treating people who use insulin to control their blood glucose which comprises the administration of an amylin agonist, for example, the amylin agonist analog 25,28,29 Pro-h-amylin, or an amylin, that has been mixed together with an intermediate- acting insulin (or any insulin preparation which contains an intermediate-acting insulin) in a single injection.
  • an amylin agonist for example, the amylin agonist analog 25,28,29 Pro-h-amylin, or an amylin
  • intermediate- acting insulin or any insulin preparation which contains an intermediate-acting insulin
  • intermediate-acting insulin is meant an insulin formulated as an insulin zinc suspension or as a modified protamine zinc insulin suspension that is crystalline, preferably having a pH of from about 7.0 to about 7.4, more preferably a pH of about 7.2.
  • Examples of preferred intermediate acting insulins include Eli Lilly's & Company's Humulin N ® (human insulin (recombinant DNA origin) isophane suspension) , NPH Iletin ® I (isophane insulin suspension, USP, beef-pork) , NPH Iletin ® II (isophane insulin suspension, USP, purified pork) , and Novo Nordisk' s Novolin ® N (NPH, human insulin isophane suspension (recombinant DNA origin) ) and NPH Purified Pork Isophane Insulin Suspension USP (100 units/ml) .
  • Other intermediate acting insulins include Eli Lilly's & Company's Humulin ® L (lente; human insulin (rDNA origin) zinc suspension, 100 Units), Lente ® Iletin ® I, (insulin zinc suspension, beef-pork) , Lente Iletin ® II (insulin zinc suspension, purified pork), and Novo Nordisk' s Novolin ® L (Lente, Human Insulin Zinc Suspension (recombinant DNA origin) , 100 Units/ml) and Lente Purified Pork Insulin (Zinc Suspension, USP 100 Units/ml) .
  • Still other intermediate- acting insulins available from Eli Lilly include Humulin ® 50/50 (50% human insulin isophane suspension and 50% human insulin injection (rDNA origin) , 100 Units) ; Humulin ® 70/30 (70% human insulin isophane suspension and 30% human insulin injection (rDNA origin) , 100 Units) ; Intermediate-acting insulins available from Novo Nordisk include Novolin ® 70/30 (70% NPH, Human Insulin Isophane Suspension and 30% Regular, Human Insulin Injection (recombinant DNA origin) , 100 Units/ml) .]
  • amylin is understood to include compounds such as those defined in U.S. Patent No. 5,234,906, issued August 10, 1993, for “Hyperglycemic Compositions," and U.S. Patent No. 5,367,052, issued November 22,1994 for "Amylin Peptides,” the contents of which are hereby incorporated by reference.
  • it includes the human peptide hormone referred to as amylin and secreted from the beta cells of the pancreas, and species variations of it.
  • Amylin agonist is also a term known in the art, and refers to a compound which mimics effects of amylin.
  • amylin agonist may be a peptide or a non-peptide compound, and includes amylin agonist analogs.
  • amylin agonist analog is understood to refer to derivatives of an amylin which act as amylin agonists, normally, it is presently believed, by virtue of binding to or otherwise directly or indirectly interacting with an amylin receptor or other receptor or receptors with which amylin itself may interact to elicit a biological response.
  • Amylin agonist analogs include those described and claimed in U.S. Patent No. 5,686,411, entitled “Amylin Agonist Peptides And Uses Therefor, " the contents of which is also hereby incorporated by reference.
  • the amylin agonist is an amylin agonist analog, preferably,
  • the invention is directed to a method of treating an insulin-using mammalian subject comprising administering together to said subject effective glucose- lowering amounts of an amylin, preferably an amylin agonist having superior physicochemical and other properties compared to those of human amylin, such as pramlintide, and an intermediate-acting insulin.
  • effective glucose-lowering amount is meant an amount effective to reduce or normalize glucose.
  • the invention is directed to a method of enhancing the glucose-lowering activity of an intermediate-acting insulin comprising administering said intermediate-acting insulin along with an amylin, preferably an amylin agonist having superior physicochemical and other properties compared to those of human amylin, such as pramlintide, as described herein.
  • an amylin preferably an amylin agonist having superior physicochemical and other properties compared to those of human amylin, such as pramlintide, as described herein.
  • This co-administration enables the use of lower and less frequent doses of either or both drugs with a concomitant reduction in the risk of possible side effects.
  • Such co-administration is performed by drawing such intermediate-acting insulin and such amylin or amylin agonist into the same container for administration 5 together.
  • the amylin agonist is an amylin agonist analog, preferably, pramlintide
  • the container is a syringe.
  • compositions comprising a therapeutically
  • compositions may include pharmaceutically acceptable salts of an amylin
  • compositions may further comprise a pharmaceutically acceptable carrier.
  • FIGURE 2 shows plasma pramlintide concentrations after
  • FIGURE 5 shows serum free insulin concentrations after subcutaneous administration of pramlintide, NPH insulin, and regular insulin in separate injections in patients with Type
  • FIGURE 10 shows plasma glucose concentrations after subcutaneous administration of pramlintide, NPH insulin, and regular insulin in separate injections in patients with Type
  • FIGURE 24 shows plasma glucose concentrations after subcutaneous administration of pramlintide, isophane insulin, and soluble insulin in separate injections in patients with
  • amylin agonist peptides can be formulated as described herein and in an application filed concurrently herewith entitled "Formulations for Amylin Agonist Peptides," the contents of which are hereby incorporated in their entirety by reference, to yield a compatible insulin/amylin agonist or amylin peptide pharmaceutical having superior glucose lowering properties when compared to insulin alone and when compared to insulin and an amylin agonist or amylin peptide administered separately.
  • amylin agonist peptide pramlintide when mixed with an intermediate-acting insulin, such as NPH insulin or isophane insulin, amylin agonist peptides are rapidly absorbed and eliminated following administration of agonist peptide, regular insulin, and NPH insulin administered as separate and in various combined subcutaneous injections. Additionally, when mixed with an intermediate-acting insulin, such as NPH or isophane insulin, amylin agonist peptides have increased bioavailability compared to other treatments as evidenced by increased values for AUC (0 _ 300) .
  • the clinical results described herein demonstrate that there is an increase in median insulin C max associated with mixing an amylin agonist peptide according to the present invention with regular and/or an intermediate-acting insulin, such as NPH insulin or isophane insulin, compared to administration in separate injections.
  • the increase in median C ma ⁇ is greatest when the amylin agonist peptide is mixed with the intermediate-acting insulin.
  • There is also a delay in median insulin T max which is greatest when an amylin agonist peptide, regular insulin, and an intermediate-acting insulin are all mixed in the same syringe prior to injection.
  • insulin AUC (0 _ 600) has been demonstrated for an amylin agonist peptide mixed with regular insulin and/or an intermediate-acting insulin compared to administration in separate injections, and when the amylin agonist peptide is mixed with an intermediate-acting insulin in one syringe and regular insulin is administered by separate injection, insulin has increased bioavailability compared to other treatments between 0 and 300 minutes.
  • the results of the clinical trials described in Examples 1 and 2 also demonstrate that there is a decrease in median glucose C ⁇ values associated with mixing an amylin agonist peptide according to the present invention with NPH insulin compared to administration in separate injections. For example, when an intermediate-acting insulin, NPH insulin, was administered in a separate injection from an amylin agonist peptide, pramlintide, a higher glucose peak was observed after lunch than when the intermediate-acting insulin was administered in the same syringe with pramlintide. Additionally, when an intermediate-acting insulin was administered in the same syringe with an amylin agonist peptide, median glucose C ma ⁇ X values were lower. The results also demonstrated that over the entire time period (0 to 600 minutes) the glucose profile was clinically optimal with the combination of an amylin agonist peptide and an intermediate-acting insulin in one syringe with or without regular insulin compared to the other treatments.
  • AUC, C, ⁇ , and T ⁇ values determined from plasma glucose concentrations during the breakfast period (0 to 300 minutes after dosing) and during the lunch period (300 to 600 minutes after dosing) support the same conclusions as those drawn from the 0 to 600 minute data.
  • Data regarding plasma glucose during the breakfast period showed that median glucose AUC (0 _ 300) , C max , and T raax values are comparable for an amylin agonist mixed with regular and/or and intermediate -acting insulin (e.g., NPH insulin and isophane insulin) compared to administration in separate injections.
  • regular and/or and intermediate -acting insulin e.g., NPH insulin and isophane insulin
  • glucose AUC (300 _ 600) values The data for plasma glucose during the lunch period demonstrated equivalence in glucose AUC (300 _ 600) values following administration of an amylin agonist peptide mixed with regular insulin without an intermediate-acting insulin compared to administration in separate injections. Additionally, median glucose AUC (300 _ 600) values following administration of an amylin agonist peptide mixed with an intermediate-acting insulin were lower compared to administration in separate injections. Importantly, after lunch, a lower glucose peak was observed for both treatments when an amylin agonist and an intermediate-acting insulin were mixed than when an intermediate-acting insulin was administered in a separate injection.
  • an amylin agonist can be mixed according to the present invention with a regular insulin and/or an intermediate-acting insulin (e.g., NPH insulin and/or iosphane insulin) prior to injection; and, importantly that there is an advantage, with respect to lowered and extended glucose control , to mixing an amylin agonist (e.g., pramlintide) and an intermediate-acting insulin prior to injection.
  • an amylin agonist e.g., pramlintide
  • This invention will allow patients to administer insulin and an amylin agonist or amylin less frequently and with fewer injections.
  • Amylin agonists useful in this invention include amylin agonist analogs disclosed and claimed in the above-noted U.S. Patent No. 5,686,411, entitled “New Amylin Agonist Peptides And Uses Therefor.”
  • Preferred amylin agonist analogs include 25 ' 28 ' 29 Pro-h-amylin, 18 Arg 25 ' 28 ' 29 Pro-h-amylin, and 18 Arg 25 ' 28 Pro-h- amylin.
  • Activity as amylin agonists can be confirmed and quantified by performing various screening assays, including the nucleus accumbens receptor binding assay described below in Example 6, followed by the soleus muscle assay described below in Example 7, a gastric emptying assay described below in Example 8 or 9, or by the ability to induce hypocalcemia or reduce postprandial hyperglycemia in mammals, as described herein.
  • the receptor binding assay a competition assay which measures the ability of compounds to bind specifically to membrane-bound amylin receptors, is described in United States Patent No. 5,264,372, issued November 23, 1993, the disclosure of which is incorporated herein by reference.
  • the receptor binding assay is also described in Example 6 below.
  • a preferred source of the membrane preparations used in the assay is the basal forebrain which comprises membranes from the nucleus accumbens and surrounding regions . Compounds being assayed compete for binding to these receptor preparations with 125 I Bolton Hunter rat amylin.
  • Competition curves wherein the amount bound (B) is plotted as a function of the log of the concentration of ligand are analyzed by computer, using analyses by nonlinear regression to a 4- parameter logistic equation (Inplot program; GraphPAD Software, San Diego, California) or the ALLFIT program of DeLean et al . (ALLFIT, Version 2.7 (NIH, Bethesda, MD 20892)). Munson and Rodbard, Anal. Biochem. 107:220-239 (1980) .
  • Assays of biological activity of amylin agonists in the soleus muscle may be performed using previously described methods (Leighton, B. and Cooper, Nature, 335:632-635 (1988); Cooper, et al . , Proc . Natl . Acad. Sci . USA 85:7763-7766 (1988) ) , in which amylin agonist activity may be assessed by measuring the inhibition of insulin-stimulated glycogen synthesis.
  • the soleus muscle assay is also described below. Methods of measuring the rate of gastric emptying are disclosed in, for example, Young et al .. Diabetologia. 38 (6) :642-648 (1995).
  • a phenol red method which is described below, conscious rats receive by gavage an aggregateric gel containing methyl cellulose and a phenol red indicator. Twenty minutes after gavage, animals are anesthetized using halothane, the stomach exposed and clamped at the pyloric and lower esophageal sphincters, removed and opened into an alkaline solution. Stomach content may be derived from the intensity of the phenol red in the alkaline solution, measured by absorbance at a wavelength of 560 n .
  • a tritiated glucose method which is described in Example 9 below, conscious rats are gavaged with tritiated glucose in water. The rats are gently restrained by the tail, the tip of which is anesthetized using lidocaine. Tritium in the plasma separated from tail blood is collected at various timepoints and detected in a beta counter. Test compounds are normally administered about one minute before gavage.
  • amylin agonists or amylins can be identified, evaluated, or screened for using the methods described below, or other art-known or equivalent methods for determining glucose lowering effect.
  • Preferred amylin agonist compounds exhibit activity in the receptor binding assay on the order of less than about 1 to 5 nM, preferably less than about 1 nM and more preferably less than about 50 pM.
  • preferred amylin agonist compounds show EC 50 values on the order of less than about 1 to 10 micromolar.
  • preferred agonist compounds show ED 50 values on the order of less than 100 ⁇ g/rat.
  • Amylin and peptide amylin agonists may be prepared using standard solid-phase peptide synthesis techniques and preferably an automated or semiautomated peptide synthesizer, taking care to use appropriate reactants and conditions to ensure that any amylin or amylin agonist analogue is prepared to include a C-terminal amide (typically in the form of a tyrosinamide residue) and to include a bridge between the residues normally found at positions 2 and 7 (typically a disulfide bridge between the cysteine amino acids found at these positions) , both of which are required for full biological activity.
  • a C-terminal amide typically in the form of a tyrosinamide residue
  • bridge between the residues normally found at positions 2 and 7 typically a disulfide bridge between the cysteine amino acids found at these positions
  • an ⁇ - N-carbamoyl protected amino acid and an amino acid attached to the growing peptide chain on a resin are coupled at room temperature in an inert solvent such as dimethylformamide, N- methylpyrrolidinone or methylene chloride in the presence of coupling agents such as dicyclohexylcarbodiimide and 1- hydroxybenzotriazole in the presence of a base such as diisopropylethylamine.
  • an inert solvent such as dimethylformamide, N- methylpyrrolidinone or methylene chloride
  • coupling agents such as dicyclohexylcarbodiimide and 1- hydroxybenzotriazole in the presence of a base such as diisopropylethylamine.
  • the ⁇ -N-carbamoyl protecting group is removed from the resulting peptide-resin using a reagent such as trifluoroacetic acid or piperidine, and the coupling reaction repeated with the next desired N-protected amino acid to be added to the peptide chain.
  • a reagent such as trifluoroacetic acid or piperidine
  • Suitable N-protecting groups are well known in the art, with t-butyloxycarbonyl (tBoc) and fluorenylmethoxycarbonyl (Fmoc) being preferred herein.
  • the solvents, amino acid derivatives and 4- methylbenzhydryl-amine (Rink) resin used in the peptide synthesizer may be purchased from Applied Biosystems Inc.
  • Boc-His may be purchased from Applied Biosystems, Inc. or Bachem Inc. (Torrance, CA) .
  • Anisole, dimethylsulfide, methylsulfide, phenol, ethanedithiol, and thioanisole may be obtained from Aldrich Chemical Company (Milwaukee, WI) . Air Products and Chemicals (Allentown, PA) supplies HF.
  • Ethyl ether, acetic acid and methanol may be purchased from Fisher Scientific (Pittsburgh, PA) .
  • Solid phase peptide synthesis may be carried out with an automated peptide synthesizer (Model 43 OA, Applied Biosystems Inc., Foster City, CA) using the NMP/HOBt (Option 1) system and Tboc or Fmoc chemistry (see, Applied Biosystems User's Manual for the ABI 430A Peptide Synthesizer, Version 1.3B July 1, 1988, section 6, pp. 49-70, Applied Biosystems, Inc., Foster City, CA) with capping. Boc-peptide-resins may be cleaved with HF (-5°C to 0°C, 1 hour) . The peptide may be extracted from the resin with alternating water and acetic acid, and the filtrates lyophilized.
  • an automated peptide synthesizer Model 43 OA, Applied Biosystems Inc., Foster City, CA
  • NMP/HOBt Option 1
  • Tboc or Fmoc chemistry see, Applied Biosystems User's Manual for the ABI
  • the Fmoc-peptide resins may be cleaved according to standard methods (Introduction to Cleavage Techniques , Applied Biosystems, Inc., 1990, pp. 6- 12) .
  • Peptides may be also be assembled using an Advanced Chem Tech Synthesizer (Model MPS 350, Louisville, Kentucky) .
  • Peptides may be purified by RP-HPLC (preparative and analytical) using a Waters Delta Prep 3000 system.
  • a C4 , C8 or C18 preparative column (10 F, 2.2 x 25 cm; Vydac, Hesperia, CA) may be used to isolate peptides, and purity may be determined using a C4 , C8 or C18 analytical column (5 F, 0.46 x 25 cm; Vydac) .
  • Amino acid analyses may be performed on the Waters Pico Tag system and processed using the Maxima program.
  • Peptides may be hydrolyzed by vapor-phase acid hydrolysis (115°C, 20-24 h) .
  • Hydrolysates may be derivatized and analyzed by standard methods (Cohen, et al . , The Pico Tag
  • Fast atom bombardment analysis may be carried out by M-Scan, Incorporated (West Chester, PA) .
  • Mass calibration may be performed using cesium iodide or cesium iodide/glycerol .
  • Plasma desorption ionization analysis using time of flight detection may be carried out on an Applied Biosystems Bio- Ion 20 mass spectrometer.
  • Peptide compounds useful in the invention may also be prepared using recombinant DNA techniques, using methods now known in the art . See, e.g. , Sambrook e al . , Molecular
  • Non-peptide compounds useful in the present invention may be prepared by art-known methods.
  • the compounds referenced above may form salts with various inorganic and organic acids and bases.
  • Such salts include salts prepared with organic and inorganic acids, for example, HCI, HBr, H 2 S0 4 , H 3 P0 4 , trifluoroacetic acid, acetic acid, formic acid, methanesulfonic acid, toluenesulfonic acid, maleic acid, fumaric acid and camphorsulfonic acid.
  • Salts prepared with bases include ammonium salts, alkali metal salts, e.g. , sodium and potassium salts, and alkali earth salts, e.g., calcium and magnesium salts.
  • Acetate, hydrochloride, and trifluoroacetate salts are preferred.
  • 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.
  • compositions useful in the invention may conveniently be provided in the form of formulations suitable for parenteral (including intravenous, intramuscular and subcutaneous) or nasal or oral administration.
  • a suitable administration format may best be determined by a medical practitioner for each patient individually.
  • Suitable pharmaceutically acceptable carriers and their formulation are described in standard formulation treatises, e.g. , Remington ' s Pharmaceutical Sciences by E.W. Martin. See also Wang, Y.J. and Hanson, M.A. "Parenteral Formulations of Proteins and Peptides: Stability and Stabilizers," Journal of Parenteral Science and Technology, Technical Report No. 10, Supp. 42:2S (1988) .
  • compositions for injection or infusion can, for example, be suspended in an inert oil, suitably a vegetable oil such as sesame, peanut, olive oil, or other acceptable carrier. These compositions may be sterilized by conventional sterilization techniques, or may be sterile filtered.
  • the compositions may contain pharmaceutically acceptable auxiliary substances as required to approximate physiological conditions, such as pH buffering agents.
  • Useful buffers include for example, sodium acetate/acetic acid buffers.
  • a form of repository or "depot" slow release preparation may be used so that therapeutically effective amounts of the preparation are delivered into the bloodstream over many hours or days following transdermal injection or delivery.
  • these parenteral dosage forms are prepared according to the concurrently filed application, "Formulations for Amylin Agonist Peptides,” and include approximately 0.01 to 0.5% (w/v) , respectively, of an amylin agonist, or amylin, as the active ingredient, in an aqueous system 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 approximately 3.0 to 6.0 (more preferably 3.5 to 5.5, and most preferably 4.0), provided, however, that if the amylin or amylin agonist has physicochemical characteristics similar to those of human amylin, it should be formulated and lyophilized for storage
  • the desired isotonicity may be accomplished using polyols (for example, mannitol and sorbitol) sodium chloride or other pharmaceutically acceptable agents such as dextrose, boric acid, sodium tartrate, propylene glycol, or other inorganic or organic solutes.
  • Mannitol is preferred, and at 1.0 to 10.0% (w/v).
  • an antimicrobial preservative selected from the group consisting of m-cresol, benzyl alcohol, methyl, ethyl, propyl and butyl parabens and phenol is also present in the preferred formulation of product designed to allow the patient to withdraw multiple doses, but is not required for single-use containers.
  • the tonicity agent is mannitol
  • the buffer is an acetate buffer
  • the preservative is approximately 0.1 to 0.3 w/v of m-cresol
  • the pH is approximately 3.7 to 4.3, most preferably 4.0.
  • 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.
  • 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 disposable pen.
  • solutions of the above compositions may be thickened with a thickening agent such as methyl cellulose .
  • a thickening agent such as methyl cellulose .
  • They may be prepared in emulsified form, either water in oil or oil in water.
  • Any of a wide variety of pharmaceutically acceptable emulsifying agents may be employed including, for example, acacia powder, a non- ionic surfactant (such as a Tween) , or an ionic surfactant (such as alkali polyether alcohol sulfates or sulfonates, e.g. , a Triton) .
  • compositions useful in the invention are prepared by mixing the ingredients following generally accepted procedures.
  • the selected components may be simply mixed in a blender or other standard device to produce a concentrated mixture which may then be adjusted to the final concentration and viscosity by the addition of water or thickening agent and possibly a buffer to control pH or an additional solute to control tonicity.
  • compositions will be provided in dosage unit form containing an amount of an amylin or amylin agonist, for example, an amylin agonist analog compound which will be effective in one or multiple doses to control glucose at the selected level .
  • an amylin or amylin agonist such as an amylin agonist analog
  • an effective amount of therapeutic agent will vary with many factors including the age and weight of the patient, the patient's physical condition, the action to be obtained and other factors.
  • the effective single, divided or continuous glucose- lowering doses of the compounds for example, including pramlintide, 18 Arg 25,28 ' 29 Pro-h-amylin and 18 Arg 25 ' 28 Pro-h-amylin will typically be in the range of 0.01 or 0.03 to about 5 mg/day, preferably about 0.01 or 0.5 to 2 mg/day and more preferably about 0.01 or 0.1 to 1 mg/day, for a 70 kg patient, administered in a single, divided or continuous doses.
  • the exact dose to be administered is determined by the attending clinician and is dependent upon a number of factors, including, those noted above. Administration may be by injection or infusion, preferably by subcutaneous or intramuscular injection.
  • Orally active compounds may be taken orally, however dosages should be increased 5-10 fold.
  • the compounds of this invention may be administered to patients in need of such treatment in a dosage ranges similar to those given above, however, the compounds may be administered more frequently, for example, one, two, or three times a day or continuously.
  • This clinical trial was designed to evaluate syringe mixing of short-acting and/or intermediate-acting insulins and the amylin agonist pramlintide.
  • Pramlintide was synthesized by standard solid phase peptide synthesis methods. Insulins were obtained from their manufacturers, as noted. The trial was conducted as an open-label, single- center, five-period, randomized, crossover study with 1-week washout periods between treatments to compare pharmacokinetic profiles for plasma pramlintide, serum free insulin, and plasma glucose after pramlintide, NPH insulin (Humulin ® N) , and regular insulin (Humulin ® R) were mixed and administered together and as separate subcutaneous injections.
  • Placebo matching pramlintide
  • NPH insulin and regular insulin in a single injection [PBO+NPH+R mixed] .
  • Treatments were administered in randomized sequence at approximately 7:00 AM (Time 0), 15 minutes prior to breakfast on study days separated by a one week washout period. (Time 0 minutes on all figures represents the time of study drug administration.)
  • patients received each of five treatments containing pramlintide or placebo and NPH or regular insulin administered combined or as separate injections in randomized sequence.
  • the pramlintide dose was 30 ⁇ g in all treatments containing pramlintide.
  • patients maintained a stable diet and exercise program and continued to receive their usual regular and NPH insulin doses.
  • All treatments were administered by subcutaneous injection into the anterior abdominal wall. All syringes, including mixed injections, were given to the patient for self-administration within 5 minutes of preparation. Blood samples for plasma pramlintide, serum free insulin, and plasma glucose concentrations were obtained at the following time points: 0 (pre-dose) , 15, 30, 45, 60, 90, 120, 180 minutes (3 hours) , 4, and 5 hours following administration of study drug on study days 2, 4, 6, 8, and 10. In addition, blood samples were collected at 6, 7, 8.5, and 10 hours after dosing on the same days for serum free insulin and plasma glucose concentrations. On each dosing day, identical meals (breakfast, lunch, and dinner) were eaten.
  • C max the peak concentration determined as the highest observed concentration during the blood sampling interval
  • T max the blood sampling time at which C ⁇ ,. occurred
  • Glucose parameters also were calculated for the meal periods. The effect of meals on plasma glucose profiles was evaluated by calculating AUC, C max , and T, ⁇ values for the period that included breakfast (0 to 300 minutes) and the period that included lunch (300 to 600 minutes) . Mean plasma pramlintide concentration-time profiles and pharmacokinetics are described and compared between treatments .
  • Mean plasma pramlintide concentration-time profiles for all evaluable patients after all pramlintide treatments are displayed in Figures 1-4.
  • Mean plasma pramlintide concentrations following administration of pramlintide, NPH insulin, and regular insulin (PRAM, NPH, R separate) in separate syringes are displayed in Figure 1.
  • the plasma concentration profiles indicate that mean plasma pramlintide concentrations increased rapidly, reached peak concentration at 15 minutes after dosing, and declined rapidly thereafter to approach baseline at approximately 240 minutes after dosing .
  • Mean plasma pramlintide concentrations following administration of PRAM+R mixed, NPH separate compared to administration of PRAM, NPH, R separate are displayed in Figure 2.
  • the mean plasma pramlintide profile following administration of PRAM+R mixed, NPH separate was similar to that for PRAM, NPH, R separate.
  • the mean plasma pramlintide profile for PRAM+R mixed, NPH separate exhibited a slightly lower peak concentration at 15 minutes, and then declined rapidly to approach baseline at 300 minutes after dosing.
  • Mean plasma pramlintide concentrations following administration of PRAM+NPH mixed, R separate compared to PRAM, NPH, R separate are displayed in Figure 3. Following subcutaneous administration of PRAM+NPH mixed, R separate, mean plasma pramlintide concentrations increased with time, reached peak concentrations at 30 minutes after the dose, and declined rapidly thereafter to baseline at 300 minutes. Compared to PRAM, NPH, R separate, the mean peak concentration was reached later and was not as high, and the decline was not as rapid.
  • Mean plasma pramlintide concentrations following administration of PRAM+NPH+R mixed compared to PRAM, NPH, R separate are displayed in Figure 4.
  • the mean plasma pramlintide concentration profile after the administration of PRAM+NPH+R mixed increased with time, reached peak concentrations at 30 minutes after the dose, and declined rapidly thereafter to baseline at 300 minutes.
  • the mean peak concentration was reached later and was not as high, and the decline was not as rapid.
  • PRAM, NPH, R separate 3 N 27 29 29
  • pramlintide had increased bioavailability compared to other treatments as evidenced by increased values for AUC (0 . 300) .
  • Mean serum free insulin concentration-time profiles for all evaluable patients after all treatments are displayed in Figures 5-9.
  • Mean serum free insulin concentrations following administration of PRAM, NPH, R separate are displayed in Figure 5.
  • Mean serum free insulin concentrations increased rapidly during the initial 45 minutes following administration PRAM, NPH, R separate. Thereafter, mean serum free insulin concentrations continued to increase but at a slower rate until peak concentration was reached between 120 and 180 minutes. After the peak, mean serum free insulin concentrations declined slowly over the remainder of the 600-minute sampling period. At 600 minutes, the mean free insulin concentration remained above the mean baseline serum free insulin concentration.
  • Mean serum free insulin concentrations following PRAM+R mixed, NPH separate compared to PRAM, NPH, R separate are displayed in Figure 6.
  • the mean serum free insulin concentration-time profile following administration of PRAM+R mixed, NPH separate was similar to that for administration of PRAM, NPH, R separate.
  • Mean serum free insulin concentrations following PRAM+NPH mixed, R separate compared to PRAM, NPH, R separate are displayed in Figure 7.
  • Mean serum free insulin concentrations following administration of PRAM+NPH mixed, R separate increased rapidly during the initial 45 minutes similar to PRAM, NPH, R separate.
  • the mean serum free insulin concentrations continued to increase after 45 minutes more rapidly to reach a higher mean peak concentration at approximately 180 minutes compared to that reached at 120 minutes for PRAM, NPH, R separate.
  • mean serum free insulin concentrations remained higher than those for PRAM, NPH, R separate between 180 and 300 minutes. Thereafter, concentrations declined at a similar rate.
  • the mean serum free insulin concentration remained above the mean baseline serum free insulin concentration but was similar to that observed with PRAM, NPH, R separate.
  • Mean serum free insulin concentrations following PRAM+NPH+R mixed compared to PRAM, NPH, R separate are displayed in Figure 8.
  • Mean serum free insulin concentrations following administration of PRAM+NPH+R mixed increased rapidly during the initial 45 minutes similar to PRAM, NPH, R separate. However, the mean serum free insulin concentrations from PRAM+NPH+R mixed continued to increase at a slower rate, but still more rapidly than for PRAM, NPH, R separate, to reach a higher mean peak concentration at approximately 180 minutes. After an initial rapid decrease between 180 and 240 minutes for PRAM+NPH+R mixed, mean serum free insulin concentrations declined at a similar rate for the two treatments throughout the remainder of the 600-minute sampling period.
  • a Pramlintide, NPH insulin, and regular insulin in separate syringes b Pramlintide + regular insulin in one syringe, with NPH insulin separate. c Pramlintide + NPH insulin in one syringe, with regular insulin separate. d Pramlintide + NPH insulin + regular insulin in one syringe. e Placebo (matching pramlintide) + NPH insulin + regular insulin in one syringe.
  • Mean plasma glucose concentrations following administration of PRAM, NPH, R separate are displayed in Figure 10.
  • Mean plasma glucose concentrations after PRAM, NPH, R separate fluctuated between approximately 210 and 285 mg/dL.
  • Mean plasma glucose concentrations following PRAM+R mixed, NPH separate compared to PRAM, NPH, R separate are displayed in Figure 11.
  • the mean plasma glucose concentration profile following administration of PRAM+R mixed, NPH separate was similar in shape to that for PRAM, NPH, R separate and mean concentrations fluctuated between approximately 200 and 275 mg/dL. The two profiles were almost superimposable for the first 120 minutes following administration. From 180 to 600 minutes after dosing, mean plasma glucose concentrations for PRAM+R mixed, NPH separate were slightly elevated but followed the same time course as the profile for PRAM, NPH, R separate and declined to approximately 250 mg/dL at 600 minutes.
  • Mean plasma glucose concentrations following administration of PRAM+NPH mixed, R separate compared to PRAM, NPH, R separate are displayed in Figure 12.
  • the mean plasma glucose concentration profile following administration of PRAM+NPH mixed, R separate was similar in shape to that for PRAM, NPH, R separate and fluctuated between approximately 170 and 265 mg/dL. The two profiles were almost superimposable for the first 60 minutes following administration. From 60 to 600 minutes after dosing, mean plasma glucose concentrations for PRAM+NPH mixed, R separate were lower but followed the same time course as those after administration of PRAM, NPH, R separate and had reached approximately 225 mg/dL at 600 minutes.
  • Mean plasma glucose concentrations following administration of PRAM+NPH+R mixed compared to PRAM, NPH, R separate are displayed in Figure 13.
  • the mean plasma glucose concentration profile following administration of PRAM+NPH+R mixed was similar in shape to that after PRAM, NPH, R separate and mean concentrations fluctuated between approximately 185 and 270 mg/dL.
  • the two profiles were almost superimposable for the first 120 minutes following administration. From 120 to 600 minutes after dosing, mean plasma glucose concentrations for PRAM+NPH+R mixed were lower but followed the same time course as those after administration of PRAM, NPH, R separate and declined to approximately 205 mg/dL at 600 minutes.
  • Mean plasma glucose concentrations following administration of PBO+NPH+R mixed compared to PRAM+NPH+R mixed are displayed in Figure 14.
  • Mean plasma glucose concentrations were higher after breakfast for up to 240 minutes after administration of PBO+NPH+R mixed compared to PRAM+NPH+R mixed. This is consistent with the effect of pramlintide to lower plasma glucose concentrations following meal ingestion within the first 180 to 240 minutes after dosing.
  • the mean plasma glucose concentration profile from PBO+NPH+R mixed was similar to that for PRAM+NPH+R mixed.
  • Mean plasma glucose concentrations were approximately 220 mg/dL at 600 minutes after PBO+NPH+R mixed compared to approximately 205 mg/dL after PRAM+NPH+R mixed.
  • a Pramlintide, NPH insulin, and regular insulin in separate syringes b Pramlintide + regular insulin in one syringe, with NPH insulin separate. c Pramlintide + NPH insulin in one syringe, with regular insulin separate. d Pramlintide + NPH insulin + regular insulin in one syringe. e Placebo (matching pramlintide) + NPH insulin + regular insulin in one syringe.
  • PRAM, NPH, R separate Although mean ratios indicated a delay in plasma glucose T mx for PRAM+R mixed, NPH separate; PRAM+NPH mixed, R separate; and PRAM+NPH+R mixed compared to PRAM, NPH, R separate, median ratios indicated T ⁇ was shorter when pramlintide, regular insulin, and NPH insulin were mixed.
  • AUC (0-600) for plasma glucose There was no period effect observed during statistical analysis of the data, The conclusions for plasma glucose are as follows. (1) There was a decrease in median glucose C max values associated with mixing pramlintide with NPH insulin compared to administration in separate injections. (2) Although mean ratios indicate a delay in T raax when pramlintide was mixed with regular and/or NPH insulin, median T max values were either comparable (PRAM+R mixed, NPH separate and PRAM+NPH mixed, R separate) or shorter (PRAM+NPH+R mixed) compared to separate injections. (3) After lunch, a lower glucose peak was observed for both treatments when pramlintide and NPH insulin were mixed prior to injection than when NPH insulin was administered in a separate injection.
  • placebo matching pramlintide
  • isophane insulin isophane insulin
  • soluble insulin in a single injection [PBO+ISO+S mixed] .
  • This trial was designed as an open-label, single-center, five-period, randomised, crossover study with 1-week washout periods between treatments to compare profiles for plasma pramlintide, serum free insulin, and plasma glucose after pramlintide, isophane (Novo Nordisk) human insulin, and soluble (Novo Nordisk) human insulin were mixed and administered together and as separate subcutaneous injections .
  • Treatments were administered in randomized sequence at approximately 7:00 AM (Time 0), 15 minutes prior to breakfast on study days separated by a one week washout period. (Time 0 minutes on all figures represents the time of study drug administration.)
  • patients received one of five treatments containing pramlintide or placebo and isophane or soluble insulin administered combined or as separate injections in randomized sequence.
  • the pramlintide dose was 30 ⁇ g in all treatments containing pramlintide.
  • patients maintained a stable diet and continued to receive their usual soluble and isophane insulin doses. All treatments were administered by subcutaneous injection into the anterior abdominal wall. All syringes were given to the patient for self-administration within 5 minutes of preparation.
  • Plasma samples for plasma pramlintide, serum free insulin, and plasma glucose concentrations were obtained at the following time points: 0 (predose) , 15, 30, 45, 60, 90, 120, 180 minutes (3 hours), 4, and 5 hours following administration of study drug on study days 2, 4, 6, 8, and 10.
  • blood samples were collected at 6, 7, 8.5, and 10 hours after dosing on the same days for serum free insulin and plasma glucose concentrations.
  • C- ⁇ the peak concentration determined as the highest observed concentration during the blood sampling interval
  • T- ⁇ the blood sampling time at which C max occurred
  • Glucose parameters were also calculated for the meal periods.
  • the mean concentration-time profiles for plasma pramlintide, serum free insulin, and plasma glucose following each treatment were calculated. Since T max varies between patients, the peak concentrations contained in the mean profiles do not correspond to median or mean C ⁇ values .
  • the effect of meals on plasma glucose profiles was evaluated by calculating AUC, C max and T max values for the periods that included breakfast (0 to 300 minutes) and the period that included lunch (300 to 600 minutes) .
  • Statistical analysis of the variables AUC (0 _ 300) , C max , and T max for glucose during the breakfast period and AUC (300 _ 600) , C raax , and T max for glucose during the lunch period was as described for glucose.
  • Plasma Pramlintide Concentrations - - Mean plasma pramlintide concentration-time profiles for all evaluable patients after all pramlintide treatments, are displayed in Figures 15-18.
  • Mean plasma pramlintide concentrations following administration of pramlintide, isophane insulin, and soluble insulin (PRAM, ISO, S separate) in separate syringes are displayed in Figure 15.
  • the plasma concentration profile indicates that mean plasma pramlintide concentrations increased rapidly, reached peak concentration at 15 minutes after dosing, and declined rapidly thereafter to approach baseline at approximately 300 minutes after dosing.
  • Mean plasma pramlintide concentrations following administration of PRAM+ISO+S mixed compared to PRAM, ISO, S separate are displayed in Figure 18.
  • the mean plasma pramlintide concentrations after the administration of PRAM+ISO+S mixed increased with time, reached peak concentrations at 30 minutes after the dose, and declined rapidly thereafter to baseline at 300 minutes.
  • the mean peak concentration was reached later and was not as high, and the decline was not as rapid.
  • Pramlintide isophane insulin, and soluble insulin in separate syringes .
  • b Pramlintide + soluble insulin in one syringe, with isophane insulin separate.
  • c Pramlintide + isophane insulin in one syringe, with soluble insulin separate.
  • d Pramlintide + isophane insulin + soluble insulin in one syringe .
  • Serum Free Insulin Concentrations Mean serum free insulin concentration-time profiles for all evaluable patients after all treatments are displayed in Figures 19-23.
  • Figure 19 Mean serum free insulin concentrations increased rapidly during the initial 45 minutes following administration PRAM, ISO, S separate. Thereafter, mean serum free insulin concentrations continued to increase but at a slower rate until peak concentration was reached at approximately 120 minutes. After the peak, mean serum free insulin concentrations declined slowly over the remainder of the 600-minute sampling period. At 600 minutes, the mean free insulin concentration remained above the mean baseline serum free insulin concentration.
  • Mean serum free insulin concentrations following PRAM+S mixed, ISO separate compared to PRAM, ISO, S separate are displayed in Figure 20.
  • the mean serum free insulin concentration-time profile following administration of PRAM+S mixed, ISO separate was similar to that for administration of PRAM, ISO, S separate.
  • Mean serum free insulin concentrations following PRAM+ISO mixed, S separate compared to PRAM, ISO, S separate are displayed in Figure 21.
  • Mean serum free insulin concentrations were higher throughout most of the profile following administration of PRAM+ISO mixed, S separate compared to the mean concentrations for PRAM, ISO, S separate.
  • Mean serum free insulin concentrations increased more rapidly for PRAM+ISO mixed, S separate to reach a higher mean peak concentration at approximately 120 minutes compared to that reached at the same time for PRAM, ISO, S separate. Following the peak, mean serum free insulin concentrations remained higher than those for PRAM, ISO, S separate up to 420 minutes after dosing. Thereafter, concentrations declined at a similar rate.
  • Mean serum free insulin concentrations following PRAM+ISO+S mixed compared to PBO+ISO+S mixed are displayed in Figure 23.
  • Mean serum free insulin concentrations following administration of PRAM+ISO+S mixed increased slightly more rapidly during the initial 45 minutes following injection than those after PBO+ISO+S mixed. Thereafter, mean serum free insulin concentrations for both treatments increased at a similar rate until reaching peak concentrations at 180 minutes. Following the peak, mean serum free insulin concentrations declined gradually throughout the remainder of the 600-minute sampling period for both treatments. At 600 minutes, mean serum free insulin concentrations remained above the mean baseline serum free insulin concentrations.
  • Serum Free Insulin Pharmacokinetic Parameters Mean ⁇ SEM serum free insulin pharmacokinetic parameter values for all evaluable patients for all treatments are displayed in Table 5. Table 5.
  • Serum Free Insulin Pharmacokinetic Parameter Values in Patients with Type I Diabetes Mellitus Following Single Doses of Pramlintide, Soluble Insulin, and Isophane Insulin Administered as Separate and Combined Subcutaneous Injections [Mean ⁇ SEM, Median, and Range; N 27]
  • a Pramlintide, isophane insulin and soluble insulin in separate syringes a Pramlintide, isophane insulin and soluble insulin in separate syringes .
  • b Pramlintide + soluble insulin in one syringe, with isophane insulin separate.
  • Median serum free insulin C raax and AUC (0 _ 600) values were comparable after pramlintide was administered in combined injections with soluble insulin with and without isophane insulin (PRAM+ISO+S mixed and PRAM+S mixed, ISO separate, respectively) compared to PRAM, ISO, S separate.
  • Median C maLX and AUC (0 _ 600) values were slightly larger (27% and 19%, respectively) when pramlintide and isophane insulin were mixed (PRAM+ISO mixed, S separate) compared to PRAM, ISO, S separate.
  • Plasma Glucose Concentrations Mean plasma glucose concentration-time profiles and parameters are described and compared between treatments .
  • Mean plasma glucose concentration-time profiles for all evaluable patients after all treatments are displayed in Figures 24-28.
  • Mean plasma glucose concentrations following administration of PRAM, ISO, S separate are displayed in Figure 24.
  • Mean plasma glucose concentrations after PRAM, ISO, S separate fluctuated between approximately 180 and 285 mg/dL.
  • Mean plasma glucose concentrations following PRAM+S mixed, ISO separate compared to PRAM, ISO, S separate are displayed in Figure 25. Although the shapes of the profiles were similar, mean plasma glucose concentrations following administration of PRAM+S mixed, ISO separate were lower compared to those for PRAM, ISO, S separate from 0 to 300 minutes after dosing. Mean concentrations for PRAM+S mixed, ISO separate fluctuated between approximately 160 and 295 mg/dL. From 360 to 600 minutes after dosing, mean plasma glucose concentrations for PRAM+S mixed, ISO separate were slightly elevated but followed the same time course as the profile for PRAM, ISO, S separate and declined to approximately 250 mg/dL at 600 minutes.
  • Mean plasma glucose concentrations following administration of PRAM+ISO mixed, S separate compared to PRAM, ISO, S separate are displayed in Figure 26. Although the shapes of the profiles were similar, mean plasma glucose concentrations following administration of PRAM+ISO mixed, S separate were lower than those for PRAM, ISO, S separate after breakfast and after lunch (from 0 to 510 minutes) . Mean concentrations for PRAM+ISO mixed, S separate fluctuated between approximately 135 and 260 mg/dL. At 600 minutes after dosing, the mean plasma glucose concentration had reached approximately 235 mg/dL for PRAM+ISO mixed, S separate, which was similar to that for PRAM, ISO, S separate (225 mg/dL) .
  • Mean plasma glucose concentrations following administration of PRAM+ISO+S mixed compared to PRAM, ISO, S separate are displayed in Figure 27.
  • the mean plasma glucose concentration profile following administration of PRAM+ISO+S mixed was similar in shape to that after PRAM, ISO, S separate.
  • Mean concentrations for PRAM+ISO+S mixed fluctuated between approximately 150 and 260 mg/dL. The two profiles were almost superimposable between 60 and 120 minutes following administration. From 120 to 600 minutes after dosing, mean plasma glucose concentrations for PRAM+ISO+S mixed were lower but followed the same time course as those after administration of PRAM, ISO, S separate and declined to approximately 225 mg/dL at 600 minutes.
  • Mean plasma glucose concentrations following administration of PBO+ISO+S mixed compared to PRAM+ISO+S mixed are displayed in Figure 28.
  • Mean plasma glucose concentrations were higher after breakfast for up to 180 minutes after administration of PBO+ISO+S mixed compared to PRAM+ISO+S mixed. This is consistent with the effect of pramlintide to lower plasma glucose concentrations following meal ingestion within the first 180 to 240 minutes after dosing.
  • the mean plasma glucose concentration profile for PBO+ISO+S mixed was similar to that for PRAM+ISO+S mixed.
  • Mean plasma glucose concentrations were approximately 230 mg/dL at 600 minutes after PBO+ISO+S mixed compared to approximately 225 mg/dL after PRAM+ISO+S mixed.
  • Plasma glucose AUC (0 . 600) , C max , and T raax values for all evaluable patients after all treatments are displayed in Table 6.
  • a Pramlintide, soluble insulin, and isophane insulin in separate syringes a Pramlintide, soluble insulin, and isophane insulin in separate syringes .
  • b Pramlintide + soluble insulin in one syringe, with isophane insulin separate.
  • Pramlintide + isophane insulin in one syringe, with soluble insulin separate.
  • d Pramlintide + isophane insulin + soluble insulin in one syringe .
  • Mean baseline plasma glucose concentrations were approximately 175 to 200 mg/dL for all treatments.
  • Median AUC (0 _ 600) and C max values were comparable when pramlintide was mixed with soluble insulin (PRAM+S mixed, ISO separate) compared to administration in separate injections.
  • the median AUC (0 . 600) and C max values for PRAM+ISO mixed, S separate and median AUC (0 . 600) value for PRAM+ISO+S mixed were slightly lower than that for PRAM, ISO, S separate.
  • median T max values were comparable when pramlintide was mixed with insulin compared to administration in separate injections, mean T max values were delayed (141% to 181%) for the combined injections compared to separate injections.
  • Significant deviations from normality were not observed for the log e - transformed values for the variables ⁇ and AUC (0.600) for plasma glucose. There was no period effect observed during statistical analysis of the data.
  • Membranes were washed three times in fresh buffer by centrifugation for 15 minutes at 48,000 x g. The final membrane pellet was resuspended in 20 mM HEPES buffer containing 0.2 mM phenylmethylsulfonyl fluoride (PMSF) .
  • PMSF phenylmethylsulfonyl fluoride
  • membranes from 4 mg original wet weight of tissue were incubated with 125 I -amylin at 12-16 pM in 20 mM HEPES buffer containing 0.5 mg/ml bacitracin, 0.5 mg/ml bovine serum albumin, and 0.2 mM PMSF.
  • Soleus Muscle Assay Determination of amylin agonist activity of compounds was carried out using the soleus muscle assay as follows. Male Harlan Sprague-Dawley rats of approximately 200g mass were used in order to maintain mass of the split soleus muscle less than 40mg. The animals were fasted for 4 hours prior to sacrifice by decapitation. The skin was stripped from the lower limb which was then pinned out on corkboard. The tendo achilles was cut just above os calcis and m. gastrocnemius reflected out from the posterior aspect of the tibia. M. soleus, a small 15-20mm long, 0.5mm thick flat muscle on the bone surface of m.
  • gastrocnemius was then stripped clear and the perimysium cleaned off using fine scissors and forceps.
  • M. soleus was then split into equal parts using a blade passed antero-posteriorly through the belly of the muscle to obtain a total of 4 muscle strips from each animal. After dissecting the muscle from the animal, it was kept for a short period in physiological saline. It was not necessary that the muscle be held under tension as this had no demonstrable effects on radioglucose incorporation into glycogen.
  • Muscles were added to 50mL Erlenmeyer flasks containing lOmL of a pregassed Krebs-Ringer bicarbonate buffer containing (each liter) NaCl 118.5mmol (6.93g), KCl 5.94mmol (443mg) , CaCl 2 2.54mmol (282mg) , MgS0 4 1.19mmol (143mg) , KH 2 P0 4 1.19mmol (162mg) , NaHC0 3 25mmol (2.1g), 5.5mmol glucose (lg) and recombinant human insulin (Humulin-R, Eli Lilly, IN) and the test compound, as detailed below. pH at 37EC was verified as being between 7.1 and 7.4.
  • Muscles were assigned to different flasks so that the 4 muscle pieces from each animal were evenly distributed among the different assay conditions.
  • the incubation media were gassed by gently blowing carbogen (95% 0 2 , 5% C0 2 ) over the surface while being continuously agitated at 37EC in an oscillating water bath. After a half-hour "preincubation" period, 0.5:Ci of U- 14 C- glucose was added to each flask which was incubated for a further 60 minutes. Each muscle piece was then rapidly removed, blotted and frozen in liquid N 2 , weighed and stored for subsequent determination of 14 C-glycogen.
  • 14 C-glycogen determination was performed in a 7mL scintillation vial. Each frozen muscle specimen was placed in a vial and digested in lmL 60% potassium hydroxide at 70EC for 45 minutes under continuous agitation. Dissolved glycogen was precipitated out onto the vial by the addition of 3mL absolute ethanol and overnight cooling at -20EC. The supernatant was gently aspirated, the glycogen washed again with ethanol, aspirated and the precipitate dried under vacuum. All ethanol is evaporated to avoid quenching during scintillation counting. The remaining glycogen was redissolved in lmL water and 4mL scintillation fluid and counted for 14 C.
  • the rate of glucose incorporation into glycogen was obtained from the specific activity of 14 C-glucose in the 5.5mM glucose of the incubation medium, and the total 14 C counts remaining in the glycogen extracted from each muscle.
  • Dose/response curves were fitted to a 4 -parameter logistic model using a least-squares iterative routine (ALLFIT, v2.7, NIH, MD) to derive EC 50 ' s . Since EC 50 is log-normally distributed, it is expressed ⁇ standard error of the logarithm. Pairwise comparisons were performed using t-test based routines of SYSTAT (Wilkinson, "SYSTAT: the system for statistics," SYSTAT Inc., Evanston IL (1989) ) .
  • Dose response curves were generated with muscles added to media containing 7. InM (1000:U/mL) insulin and each test compound added at final (nominal) concentrations of 0, 1, 3, 10, 30, 100, 300 and lOOOnM.
  • Each assay also contained internal positive controls consisting of a single batch of archived rat amylin, lyophilized and stored at -70EC.
  • Human amylin is a known hyperglycemic peptide, and EC 50 measurements of amylin preparations in the soleus muscle assay range typically from about 1-10 nM, although some commercial preparations which are less than 90% pure have higher EC 50 ' s due to the presence of contaminants that result in a lower measured activity. Results for test compounds are set forth in Table A.
  • percent of stomach contents remaining after 20 minutes were expressed as a fraction of the gastric contents recovered from control rats sacrificed immediately after gavage in the same experiment .
  • Percent gastric emptying contents remaining (absorbance at 20 min) / (absorbance at 0 min) .
  • Dose response curves for gastric emptying were fitted to a 4 -parameter logistic model using a least-squares iterative routine (ALLFIT, v2.7, NIH, Bethesda, MD) to derive ED 50 s . Since ED 50 is log-normally distributed, it is expressed + standard error of the logarithm. Pairwise comparisons were performed using one-way analysis of variance and the Student -Newman-Keuls multiple comparisons test (Instat v2.0, GraphPad Software, San Diego, CA) using P ⁇ 0.05 as the level of significance.
  • rat amylin (Bachem, Torrance, CA) dissolved in 0.15M saline, was administered as a 0.1 mL subcutaneous bolus in doses of 0 , 0.01, 0.1, 1, 10 or 100 Fg 5 minutes before gavage in Harlan Sprague Dawley (non- diabetic) rats fasted 20 hours and diabetic BB rats fasted 6 hours.
  • the ED 50 for inhibition of gastric emptying in normal rats was 0.43 Fg (0.60nmol/kg) ⁇ 0.19 log units, and was 2.2Fg (2.3nmol/kg) ⁇ 0.18 log units in diabetic rats.
  • EXAMPLE 9 TRITIATED GLUCOSE GASTRIC EMPTYING ASSAY Conscious, non-fasted, Harlan Sprague Dawley rats were restrained by the tail, the tip of which was anesthetized using 2% lidocaine. Tritium in plasma separated from tail blood collected 0, 15, 30, 60, 90 and 120 minutes after gavage was detected in a beta counter. Rats were injected subcutaneously with 0.1 mL saline containing 0, 0.1, 0.3, 1, 10 or 100 Fg of rat amylin 1 minute before gavage

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Abstract

La présente invention concerne des compositions pharmaceutiques contenant un agoniste d'amyline, par exemple 25, 28, 29 Pro-h-amyline (connu aussi sous le nom de pramlintide), et une insuline. L'invention concerne aussi les procédés de préparation et d'utilisation desdites compositions pharmaceutiques dans le traitement de mammifères, de préférence des humains, qui utilisent l'insuline pour réguler les concentrations de glucose dans le sang, notamment les personnes souffrant de diabètes.
PCT/US1998/000662 1998-01-09 1998-01-09 Compositions pharmaceutiques a agoniste d'amyline, contenant de l'insuline WO1999034764A2 (fr)

Priority Applications (4)

Application Number Priority Date Filing Date Title
AU59162/98A AU5916298A (en) 1998-01-09 1998-01-09 Amylin agonist pharmaceutical compositions containing insulin
PCT/US1998/000662 WO1999034764A2 (fr) 1998-01-09 1998-01-09 Compositions pharmaceutiques a agoniste d'amyline, contenant de l'insuline
EP98902526A EP1051141A4 (fr) 1998-01-09 1998-01-09 Compositions pharmaceutiques a agoniste d'amyline, contenant de l'insuline
ZA98221A ZA98221B (en) 1998-01-09 1998-01-12 Amylin agonist pharmaceutical compositions containing insulin

Applications Claiming Priority (2)

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PCT/US1998/000662 WO1999034764A2 (fr) 1998-01-09 1998-01-09 Compositions pharmaceutiques a agoniste d'amyline, contenant de l'insuline
ZA98221A ZA98221B (en) 1998-01-09 1998-01-12 Amylin agonist pharmaceutical compositions containing insulin

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Cited By (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2007104786A1 (fr) * 2006-03-15 2007-09-20 Novo Nordisk A/S Melanges d'amyline et d'insuline
EP2036539A1 (fr) * 2007-09-11 2009-03-18 Novo Nordisk A/S Formulations stables d'amyline et ses analogues
WO2009034117A1 (fr) * 2007-09-11 2009-03-19 Novo Nordisk A/S Mélange comprenant de l'amyline, qui est un peptide, et une insuline retard
WO2018046719A1 (fr) 2016-09-09 2018-03-15 Zealand Pharma A/S Analogues d'amyline
US10766939B2 (en) 2015-03-18 2020-09-08 Zealand Pharma A/S Amylin analogues
US11318191B2 (en) 2020-02-18 2022-05-03 Novo Nordisk A/S GLP-1 compositions and uses thereof
US11752198B2 (en) 2017-08-24 2023-09-12 Novo Nordisk A/S GLP-1 compositions and uses thereof

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of EP1051141A4 *

Cited By (25)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2007104786A1 (fr) * 2006-03-15 2007-09-20 Novo Nordisk A/S Melanges d'amyline et d'insuline
JP2009530249A (ja) * 2006-03-15 2009-08-27 ノボ・ノルデイスク・エー／エス アミリンとインスリンの混合物
EP2241327A1 (fr) * 2006-03-15 2010-10-20 Novo Nordisk A/S Melange d'amyline et d'insuline
EP2036539A1 (fr) * 2007-09-11 2009-03-18 Novo Nordisk A/S Formulations stables d'amyline et ses analogues
WO2009034118A1 (fr) * 2007-09-11 2009-03-19 Novo Nordisk A/S Formulations stables d'amyline et analogues associés
WO2009034117A1 (fr) * 2007-09-11 2009-03-19 Novo Nordisk A/S Mélange comprenant de l'amyline, qui est un peptide, et une insuline retard
JP2010539131A (ja) * 2007-09-11 2010-12-16 ノボ・ノルデイスク・エー／エス アミリンとそのアナログの安定的製剤
US8404645B2 (en) 2007-09-11 2013-03-26 Novo Nordisk As Stable formulations of amylin and its analogues
CN101854914B (zh) * 2007-09-11 2013-03-27 诺沃-诺迪斯克有限公司 胰岛淀粉样多肽及其类似物的稳定制剂
US10766939B2 (en) 2015-03-18 2020-09-08 Zealand Pharma A/S Amylin analogues
CN109863168A (zh) * 2016-09-09 2019-06-07 西兰制药公司 胰淀素类似物
US11382956B2 (en) 2016-09-09 2022-07-12 Zealand Pharma A/S Amylin analogues
US10071140B2 (en) 2016-09-09 2018-09-11 Zealand Pharma A/S Amylin analogues
JP2019534248A (ja) * 2016-09-09 2019-11-28 ジーランド ファーマ アクティーゼルスカブ アミリン類似体
WO2018046719A1 (fr) 2016-09-09 2018-03-15 Zealand Pharma A/S Analogues d'amyline
AU2017322277B2 (en) * 2016-09-09 2021-11-11 Zealand Pharma A/S Amylin analogues
US12083164B2 (en) 2016-09-09 2024-09-10 Zealand Pharma A/S Amylin analogues
KR20190045333A (ko) * 2016-09-09 2019-05-02 질랜드 파마 에이/에스 아밀린 유사체
EP4074729A1 (fr) 2016-09-09 2022-10-19 Zealand Pharma A/S Analogues de l'amyline
TWI784968B (zh) * 2016-09-09 2022-12-01 丹麥商西蘭製藥公司 澱粉素類似物
KR102498393B1 (ko) 2016-09-09 2023-02-13 질랜드 파마 에이/에스 아밀린 유사체
CN109863168B (zh) * 2016-09-09 2023-04-18 西兰制药公司 胰淀素类似物
US11752198B2 (en) 2017-08-24 2023-09-12 Novo Nordisk A/S GLP-1 compositions and uses thereof
US12214017B2 (en) 2017-08-24 2025-02-04 Novo Nordisk A/S GLP-1 compositions and uses thereof
US11318191B2 (en) 2020-02-18 2022-05-03 Novo Nordisk A/S GLP-1 compositions and uses thereof

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