WO2010022213A2

WO2010022213A2 - Compositions and methods for preserving pancreatic islet mass

Info

Publication number: WO2010022213A2
Application number: PCT/US2009/054419
Authority: WO
Inventors: Anna Louise Nolan
Original assignee: Elixir Pharmaceuticals, Inc.
Priority date: 2008-08-22
Filing date: 2009-08-20
Publication date: 2010-02-25
Also published as: WO2010022213A3

Abstract

Methods and compositions for preserving or optimizing pancreatic islet mass and other pancreas parameters are described.

Description

COMPOSITIONS AND METHODS FOR PRESERVING PANCREATIC ISLET MASS

CROSS REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Application Serial No. 61/091,003, filed on August 22, 2008. The disclosure of the prior application is considered part of (and is incorporated by reference in) the disclosure of this application.

BACKGROUND

The ghrelin receptor (GhrR), also known as the growth hormone secretagogue receptor (GHS-R), regulates a number of physiological processes, including growth hormone (GH) release, metabolism, and appetite. Ghrelin is a 28 amino acid peptide that is an endogenous ligand for ghrelin receptor. Ghrelin has been shown to stimulate feeding in humans. In rodents, ghrelin induces body weight gain and adiposity. See, e.g., Asakawa (2003) Gut 52:947. In addition to regulating feeding, ghrelin can stimulate GH secretion by activating GHS-R, particularly in somatotrophic tissue.

SUMMARY

The present invention is based, in part, on the discovery that administering a ghrelin pathway antagonist (optionally in combination with an additional treatment) can preserve pancreatic islet mass and/or islet function in a subject, e.g., a subject with insulin resistance or developing insulin resistance. The invention relates, inter alia, to compositions and methods for preserving pancreatic islet mass and/or islet function in a subject.

Administration of a ghrelin pathway antagonist (optionally in combination with an additional treatment) can lead to pancreas sparing in a subject, e.g., a subject with insulin resistance or developing insulin resistance. The invention relates, inter alia, to compositions and methods for sparing the pancreas of a subject.

Increased dietary fat intake is associated with obesity and/or insulin resistance. As insulin resistance develops, e.g., as insulin sensitivity decreases, and/or as a subject enters a pre-diabetic stage, the pancreatic islet attempts to compensate for increased blood glucose concentrations by increasing the rate of insulin secretion. The islet achieves this by increasing the number of beta (β) cells and thus increasing its mass. The pancreatic islet mass may also, or alternatively, increase due to increased lipid content, as dietary fat intake increases, e.g., and obesity develops.

Increased islet lipid content can cause an impairment of islet cell (e.g., β cell) function, e.g., reduced insulin secretion, a decreased response to glucose stimulated insulin secretion, increased metabolism of glucose and free fatty acids (FFAs), e.g., causing glucolipotoxicity, lipotoxicity, oxidative stress, and/or dysregulation of triglyceride and/or FFA cycling, β-cell exhaustion and/or endoplasmic reticulum (ER) stress can also lead to islet dysfunction. Eventually, as a subject progresses through a pre-diabetic stage and towards developing diabetes (e.g., type 2 diabetes), the cells are less able, or no longer able, to compensate for the impaired islet cell function, and islet cell mass is reduced by increased apoptosis and/or necrosis. By preserving islet mass (e.g., by preventing an increase in islet mass, e.g., in a pre-diabetic stage and/or by preventing a decrease in islet mass, e.g., as a subject develops diabetes, or due to β-cell exhaustion and/or ER stress), e.g., by preventing a change in islet lipid content, the impairment of islet cell function is delayed and/or decreased (e.g., the duration of a pre- diabetic stage is increased (e.g., progression to diabetes is delayed) and/or the severity of islet impairment is decreased).

In one aspect, the disclosure provides a method for preserving or optimizing pancreatic islet mass in a subject (e.g., mammal, e.g., human). The method includes administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves or optimizes pancreatic islet mass (e.g., mass of beta (β) cells, alpha (α) cells, delta (δ) cells, PP cells, and/or epsilon (ε) cells), e.g., relative to a standard. For example, the standard can be a cohort of subjects that do not have diabetes, a cohort of subjects that do not have prediabetes, a cohort of subjects that do not have obesity, a cohort of subjects that do not have insulin resistance, a cohort of subjects that do not have an impairment of pancreas function, a cohort of subjects that have diabetes, a cohort of subjects that have prediabetes, a cohort of subjects that have obesity, a cohort of subjects that have insulin resistance, a cohort of subjects that have an impairment of pancreas function, or a cohort of subjects that progressed from prediabetes to diabetes.

In some embodiments, the method further includes evaluating (e.g., measuring) pancreatic islet mass in the subject prior to, during (e.g., during a course of administration, e.g., a treatment regimen), and/or after administration of the ghrelin pathway antagonist.In some embodiments, the ghrelin pathway antagonist prevents or delays an increase in pancreatic islet mass in a subject with prediabetes. In some embodiments, administration of the ghrelin pathway antagonist begins prior to an increase in pancreatic islet mass. In some embodiments, administration of the ghrelin pathway antagonist extends throughout the period in which an increase in mass would be expected, e.g., and extends when an increase would otherwise be seen e.g., in the absence of administration of the ghrelin pathway antagonist.

In some embodiments, the ghrelin pathway antagonist prevents or delays a decrease in pancreatic islet mass in a subject with diabetes. In some embodiments, administration of the ghrelin pathway antagonist begins prior to a decrease in pancreatic islet mass. In some embodiments, administration of the ghrelin pathway antagonist extends throughout the period in which a decrease in mass would be expected, e.g., and extends when a decrease would otherwise be seen e.g., in the absence of administration of the ghrelin pathway antagonist. In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject has obesity or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist includes a peptide, polypeptide, protein, antibody, antibody fragment (e.g., Fc fragment), peptidomimetic, peptoid, nucleic acid, or other chemical compound or a combination thereof (e.g., a combination of two or more such antagonists).

In some embodiments, the ghrelin pathway antagonist comprises a protein. In some preferred embodiments, the protein is an antibody, a peptide (e.g., a fragment of naturally occurring ligand, a random or semi-randomly generated binding peptide), or a molecule (e.g., protein) which binds to a regulatory element for the subject component (e.g., an artificial transcription factor, e.g., a zinc finger protein).

In some embodiments, the ghrelin pathway antagonist comprises a non- proteinaceous molecule, e.g., a molecule less than 3000, 2000, 1000, 700, 500, 400, 300, or 200 in molecular weight.

In some embodiments, the ghrelin pathway antagonist comprises a nucleic acid. In some preferred embodiments, the nucleic acid comprises a nucleic acid aptamer, RNAi, an antisense RNA, or a ribozyme molecule.

In some embodiments, the ghrelin pathway antagonist binds to ghrelin. In some embodiments, the ghrelin pathway antagonist binds to the ghrelin receptor (GHS-R).

In some embodiments, the ghrelin pathway antagonist is ghrelin receptor antagonist [D-Lys³]-GHRP-6.

In some embodiments, the ghrelin pathway antagonist is used in combination with another therapeutic agent. In some preferred embodiments, the ghrelin pathway antagonist is used in combination with another ghrelin pathway antagonist, e.g., another ghrelin pathway antagonist described herein.

In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for obesity. In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for metabolic syndrome.

In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for prediabetes.

In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for diabetes.

In some embodiments, the ghrelin pathway antagonist is a compound described herein, e.g., a compound of a formula described herein, e.g., a compound of formula (I), (II), (III), (IV), (V), or (VI).

In another aspect, the disclosure provides a method for preserving or optimizing pancreatic islet function (e.g., insulin secretion, response to glucose stimulated insulin secretion, metabolism of glucose and/or free fatty acids (FFAs)) in a subject (e.g., mammal, e.g., human). The method includes administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves or optimizes pancreatic islet function, e.g., relative to a standard. For example, the standard can be a cohort of subjects that do not have diabetes, a cohort of subjects that do not have prediabetes, a cohort of subjects that do not have obesity, a cohort of subjects that do not have insulin resistance, a cohort of subjects that do not have an impairment of pancreas function, a cohort of subjects that have diabetes, a cohort of subjects that have prediabetes, a cohort of subjects that have obesity, a cohort of subjects that have insulin resistance, a cohort of subjects that have an impairment of pancreas function, or a cohort of subjects that progressed from prediabetes to diabetes.

In some embodiments, the method further includes evaluating (e.g., measuring) pancreatic islet function (e.g., insulin secretion, response to glucose stimulated insulin secretion, metabolism of glucose and/or free fatty acids (FFAs)) in the subject prior to, during (e.g., during a course of administration, e.g., a treatment regimen), and/or after administration of the ghrelin pathway antagonist.

In some embodiments, administration of the ghrelin pathway antagonist begins prior to a decrease in pancreatic islet function. In some embodiments, administration of the ghrelin pathway antagonist extends throughout the period in which a decrease in function would be expected, e.g., and extends when a decrease would otherwise be seen e.g., in the absence of administration of the ghrelin pathway antagonist.

In some embodiments, the method decreases and/or delays glucolipotoxicity, lipotoxicity, oxidative stress, and/or dysregulation of triglyceride and/or FFA cycling.

In some embodiments, the subject has insulin resistance. In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject has obesity or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist includes a peptide, polypeptide, protein, antibody, antibody fragment (e.g., Fc fragment), peptidomimetic, peptoid, nucleic acid, or other chemical compound or a combination thereof. In some embodiments, the ghrelin pathway antagonist comprises a protein. In some preferred embodiments, the protein is an antibody, a peptide (e.g., a fragment of naturally occurring ligand, a random or semi-randomly generated binding peptide), or a molecule (e.g., protein) which binds to a regulatory element for the subject component (e.g., an artificial transcription factor, e.g., a zinc finger protein).

In some embodiments, the ghrelin pathway antagonist binds to ghrelin.

In some embodiments, the ghrelin pathway antagonist binds to the ghrelin receptor (GHS-R). In some embodiments, the ghrelin pathway antagonist is ghrelin receptor antagonist [D-Lys³]-GHRP-6.

In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for obesity.

In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for metabolic syndrome. In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for prediabetes.

In some embodiments, the ghrelin pathway antagonist is a compound described herein, e.g., a compound of a formula described herein, e.g., a compound of formula (I), (II), (III), (IV), (V), or (VI). In another aspect, the disclosure provides a method for preserving or optimizing pancreas weight in a subject (e.g., mammal, e.g., human). The method includes administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves or optimizes pancreas weight, e.g., relative to a standard. For example, the standard can be a cohort of subjects that do not have diabetes, a cohort of subjects that do not have prediabetes, a cohort of subjects that do not have obesity, a cohort of subjects that do not have insulin resistance, a cohort of subjects that do not have an impairment of pancreas function, a cohort of subjects that have diabetes, a cohort of subjects that have prediabetes, a cohort of subjects that have obesity, a cohort of subjects that have insulin resistance, a cohort of subjects that have an impairment of pancreas function, or a cohort of subjects that progressed from prediabetes to diabetes.

In some embodiments, the method further includes evaluating (e.g., measuring) pancreas weight in the subject prior to, during (e.g., during a course of administration, e.g., a treatment regimen), and/or after administration of the ghrelin pathway antagonist.

In some embodiments, administration of the ghrelin pathway antagonist begins prior to an increase in pancreas weight. In some embodiments, administration of the ghrelin pathway antagonist extends throughout the period in which an increase in pancreas weight would be expected, e.g., and extends when an increase would otherwise be seen e.g., in the absence of administration of the ghrelin pathway antagonist. In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has prediabetes or a risk factor thereof. In some embodiments, the subject has diabetes or a risk factor thereof. In some embodiments, the subject has obesity or a risk factor thereof. In some embodiments, the ghrelin pathway antagonist includes a peptide, polypeptide, protein, antibody, antibody fragment (e.g., Fc fragment), peptidomimetic, peptoid, nucleic acid, or other chemical compound or a combination thereof.

In another aspect, the disclosure provides a method for preserving or optimizing body weight in a subject (e.g., mammal, e.g., human). The method includes administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves or optimizes body weight, e.g., relative to a standard. For example, the standard can be a cohort of subjects that do not have diabetes, a cohort of subjects that do not have prediabetes, a cohort of subjects that do not have obesity, a cohort of subjects that do not have insulin resistance, a cohort of subjects that do not have an impairment of pancreas function, a cohort of subjects that have diabetes, a cohort of subjects that have prediabetes, a cohort of subjects that have obesity, a cohort of subjects that have insulin resistance, a cohort of subjects that have an impairment of pancreas function, or a cohort of subjects that progressed from prediabetes to diabetes. In some embodiments, the method further includes evaluating (e.g., measuring) body weight in the subject prior to, during (e.g., during a course of administration, e.g., a treatment regimen), and/or after administration of the ghrelin pathway antagonist.

In some embodiments, administration of the ghrelin pathway antagonist begins prior to an increase in body weight. In some embodiments, administration of the ghrelin pathway antagonist extends throughout the period in which an increase in body weight would be expected, e.g., and extends when an increase would otherwise be seen e.g., in the absence of administration of the ghrelin pathway antagonist.

In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has prediabetes or a risk factor thereof. In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject has obesity or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist includes a peptide, polypeptide, protein, antibody, antibody fragment (e.g., Fc fragment), peptidomimetic, peptoid, nucleic acid, or other chemical compound or a combination thereof. In some embodiments, the ghrelin pathway antagonist comprises a protein. In some preferred embodiments, the protein is an antibody, a peptide (e.g., a fragment of naturally occurring ligand, a random or semi-randomly generated binding peptide), or a molecule (e.g., protein) which binds to a regulatory element for the subject component (e.g., an artificial transcription factor, e.g., a zinc finger protein). In some embodiments, the ghrelin pathway antagonist comprises a non- proteinaceous molecule, e.g., a molecule less than 3000, 2000, 1000, 700, 500, 400, 300, or 200 in molecular weight.

In some embodiments, the ghrelin pathway antagonist binds to ghrelin.

In another aspect, the disclosure provides a method for preserving or optimizing pancreatic islet triglyceride (TG) content in a subject (e.g., mammal, e.g., human). The method includes administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves or optimizes pancreatic islet TG content, e.g., relative to a standard. For example, the standard can be a cohort of subjects that do not have diabetes, a cohort of subjects that do not have prediabetes, a cohort of subjects that do not have obesity, a cohort of subjects that do not have insulin resistance, a cohort of subjects that do not have an impairment of pancreas function, a cohort of subjects that have diabetes, a cohort of subjects that have prediabetes, a cohort of subjects that have obesity, a cohort of subjects that have insulin resistance, a cohort of subjects that have an impairment of pancreas function, or a cohort of subjects that progressed from prediabetes to diabetes.

In some embodiments, the method further includes evaluating (e.g., measuring) pancreatic islet TG content in the subject prior to, during (e.g., during a course of administration, e.g., a treatment regimen), and/or after administration of the ghrelin pathway antagonist.

In some embodiments, administration of the ghrelin pathway antagonist begins prior to an increase in pancreatic islet triglyceride content. In some embodiments, administration of the ghrelin pathway antagonist extends throughout the period in which an increase in pancreatic islet triglyceride content would be expected, e.g., and extends when an increase would otherwise be seen e.g., in the absence of administration of the ghrelin pathway antagonist.

In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject has obesity or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist includes a peptide, polypeptide, protein, antibody, antibody fragment (e.g., Fc fragment), peptidomimetic, peptoid, nucleic acid, or other chemical compound or a combination thereof.

In some embodiments, the ghrelin pathway antagonist comprises a protein. In some preferred embodiments, the protein is an antibody, a peptide (e.g., a fragment of naturally occurring ligand, a random or semi-randomly generated binding peptide), or a molecule (e.g., protein) which binds to a regulatory element for the subject component (e.g., an artificial transcription factor, e.g., a zinc finger protein). In some embodiments, the ghrelin pathway antagonist comprises a non- proteinaceous molecule, e.g., a molecule less than 3000, 2000, 1000, 700, 500, 400, 300, or 200 in molecular weight.

In some embodiments, the ghrelin pathway antagonist binds to ghrelin.

In one aspect, the disclosure provides a method for preserving or optimizing pancreatic islet lipid content in a subject (e.g., mammal, e.g., human). The method includes administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves or optimizes pancreatic islet lipid content, e.g., relative to a standard. For example, the standard can be a cohort of subjects that do not have diabetes, a cohort of subjects that do not have prediabetes, a cohort of subjects that do not have obesity, a cohort of subjects that do not have insulin resistance, a cohort of subjects that do not have an impairment of pancreas function, a cohort of subjects that have diabetes, a cohort of subjects that have prediabetes, a cohort of subjects that have obesity, a cohort of subjects that have insulin resistance, a cohort of subjects that have an impairment of pancreas function, or a cohort of subjects that progressed from prediabetes to diabetes.

In some embodiments, the method further includes evaluating (e.g., measuring) pancreatic islet lipid content in the subject prior to, during (e.g., during a course of administration, e.g., a treatment regimen), and/or after administration of the ghrelin pathway antagonist.

In some embodiments, administration of the ghrelin pathway antagonist begins prior to an increase in pancreatic islet lipid content. In some embodiments, administration of the ghrelin pathway antagonist extends throughout the period in which an increase in pancreatic lipid triglyceride content would be expected, e.g., and extends when an increase would otherwise be seen e.g., in the absence of administration of the ghrelin pathway antagonist.

In some embodiments, the ghrelin pathway antagonist prevents or delays an increase in pancreatic islet lipid content in a subject with prediabetes. In some embodiments, the ghrelin pathway antagonist prevents or delays an increase in pancreatic islet lipid content in a subject with diabetes.

In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject has diabetes or a risk factor thereof. In some embodiments, the subject has obesity or a risk factor thereof.

In another aspect, the disclosure provides a method for pancreas sparing in a subject (e.g., mammal, e.g., human). The method includes administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist causes pancreas sparing, e.g., relative to a standard. The term "pancreas sparing" refers to the ability to delay the need for treatment with insulin. For example, subjects that no longer respond to oral anti-diabetic therapy need insulin treatment, e.g., are insulin dependent. Once insulin therapy is needed, the pancreas is exhausted and not able to produce sufficient amounts of insulin, e.g., amounts needed to control blood glucose levels. For example, the ghrelin pathway antagonist can delay the need for insulin therapy, e.g., by about 1 week, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 24 weeks, about 28 weeks, about 32 weeks, about 36 weeks, about 40 weeks, about 44 weeks, about 48 weeks, about 52 weeks, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 3.5 years, about 4 years, about 4.5 years, about 5 years, about 5.5 years, about 6 years, about 6.5 years, about 7 years, about 7.5 years, about 8 years, about 9.5 years, about 10 years, about 12 years, about 14 years, about 16 years, about 18 years, about 20 years or longer, e.g., as compared to a standard, e.g., the average time for initiation of insulin treatment in a reference, e.g., a cohort of subjects (e.g., that were treated with a diabetes therapy other than a ghrelin pathway antagonist) that progressed from prediabetes or diabetes to insulin resistance.

In some embodiments, the method preserves or optimizes pancreas function (e.g., increases insulin secretion and/or increase glucose-responsive insulin secretion, e.g., as compared to the amount of secretion in the absence of the ghrelin pathway antagonist). In some embodiments, the method further includes evaluating (e.g., measuring) pancreas function (e.g., insulin production or insulin sensitivity to glucose levels) in the subject prior to, during (e.g., during a course of administration, e.g., a treatment regimen), and/or after administration of the ghrelin pathway antagonist. In some embodiments, administration of the ghrelin pathway antagonist begins prior to a decrease in pancreas functon. In some embodiments, administration of the ghrelin pathway antagonist extends throughout the period in which a decrease in pancreas function would be expected, e.g., and extends when a decrease would otherwise be seen e.g., in the absence of administration of the ghrelin pathway antagonist. In some embodiments, the ghrelin pathway antagonist prevents or delays a decrease in pancreas function in a subject with diabetes. In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject has obesity or a risk factor thereof. In some embodiments, the ghrelin pathway antagonist includes a peptide, polypeptide, protein, antibody, antibody fragment (e.g., Fc fragment), peptidomimetic, peptoid, nucleic acid, or other chemical compound or a combination thereof.

In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for obesity. In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for metabolic syndrome. In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for prediabetes.

In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for diabetes. In some embodiments, the ghrelin pathway antagonist is a compound described herein, e.g., a compound of a formula described herein, e.g., a compound of formula (I), (II), (III), (IV), (V), or (VI).

In another aspect, the disclosure provides a method for selecting a subject for treatment with a ghrelin pathway antagonist. Optionally, the method includes selecting a subject for such treatment on the basis that the subject is in need of preservation of pancreatic islet mass. The method includes evaluating (e.g., measuring) pancreatic islet mass in the subject, and comparing the pancreatic islet mass to a standard (e.g., a standard described herein), wherein the subject is selected for treatment with a ghrelin pathway antagonist if the difference between the pancreatic islet mass and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. Optionally, the ghrelin pathway antagonist is selected on the basis that it acts to preserve pancreatic islet mass. In some embodiments, the subject is evaluated to determine if the subject is in need of preservation of pancreatic islet mass or if the subject has altered (e.g., increased or decreased) pancreatic islet mass, e.g., as compared to a standard (e.g., a standard described herein). For example, in some embodiments, a subject is in need of preservation of pancreatic islet mass if the subject has prediabetes or a risk factor thereof, the subject's pancreatic islet mass is increased relative to the standard, the subject has diabetes or a risk factor thereof, the subject's pancreatic islet mass is decreased relative to the standard, the subject has insulin resistance, and/or the subject has obesity or a risk factor thereof.

In some embodiments, the subject has prediabetes or a risk factor thereof. In some embodiments, the subject's pancreatic islet mass is increased relative to the standard. In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject's pancreatic islet mass is decreased relative to the standard.

In some embodiments, the subject has insulin resistance. In some embodiments, the subject has obesity or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist comprises a nucleic acid. In some preferred embodiments, the nucleic acid comprises a nucleic acid aptamer, RNAi, an antisense RNA, or a ribozyme molecule. In some embodiments, the ghrelin pathway antagonist binds to ghrelin.

In some embodiments, the ghrelin pathway antagonist binds to the ghrelin receptor (GHS-R).

In some embodiments, the ghrelin pathway antagonist is ghrelin receptor antagonist [D-Lys³]-GHRP-6. In some embodiments, the ghrelin pathway antagonist is used in combination with another therapeutic agent. In some preferred embodiments, the ghrelin pathway antagonist is used in combination with another ghrelin pathway antagonist, e.g., another ghrelin pathway antagonist described herein.

In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for prediabetes. In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for diabetes.

In one aspect, the disclosure provides a method for selecting a subject for treatment with a ghrelin pathway antagonist. Optionally, the method includes selecting a subject for such treatment on the basis that the subject is in need of preservation of pancreas weight. The method includes evaluating (e.g., measuring) pancreas weight in the subject, and comparing the pancreas weight to a standard (e.g., a standard described herein), wherein the subject is selected for treatment with a ghrelin pathway antagonist if the difference between the pancreas weight and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. Optionally, the ghrelin pathway antagonist is selected on the basis that it acts to preserve pancreas weight.

In some embodiments, the subject is evaluated to determine if the subject is in need of preservation of pancreas weight or if the subject has altered (e.g., increased or decreased) pancreas weight, e.g., as compared to a standard (e.g., a standard described herein). For example, in some embodiments, a subject is in need of preservation of pancreas weight if the subject has prediabetes or a risk factor thereof, the subject's pancreas weight is increased relative to the standard, the subject has diabetes or a risk factor thereof, the subject's pancreas weight is decreased relative to the standard, the subject has insulin resistance, and/or the subject has obesity or a risk factor thereof. In some embodiments, the subject has prediabetes or a risk factor thereof. In some embodiments, the subject's pancreas weight is increased relative to the standard.

In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject's pancreas weight is decreased relative to the standard.

In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has obesity or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist binds to ghrelin.

In some embodiments, the ghrelin pathway antagonist is used in combination with another therapeutic agent. In some preferred embodiments, the ghrelin pathway antagonist is used in combination with another ghrelin pathway antagonist, e.g., another ghrelin pathway antagonist described herein. In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for obesity.

In another aspect, the disclosure provides a method for selecting a subject for treatment with a ghrelin pathway antagonist. Optionally, the method includes selecting a subject for such treatment on the basis that the subject is in need of preservation of pancreatic islet function. The method includes evaluating (e.g., measuring) pancreatic islet function in the subject, and comparing the pancreatic islet function to a standard (e.g., a standard described herein), wherein the subject is selected for treatment with a ghrelin pathway antagonist if the difference between the pancreatic islet function and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. Optionally, the ghrelin pathway antagonist is selected on the basis that it acts to preserve pancreatic islet function. In some embodiments, the subject is evaluated to determine if the subject is in need of preservation of pancreatic islet function or if the subject has altered (e.g., increased or decreased) pancreatic islet function, e.g., as compared to a standard (e.g., a standard described herein). For example, in some embodiments, a subject is in need of preservation of pancreatic islet function if the subject has prediabetes or a risk factor thereof, the subject's pancreatic islet function is increased relative to the standard, the subject has diabetes or a risk factor thereof, the subject's pancreatic islet function is decreased relative to the standard, the subject has insulin resistance, and/or the subject has obesity or a risk factor thereof.

In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject's pancreatic islet function is increased relative to the standard.

In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject's pancreatic islet function is decreased relative to the standard.

In another aspect, the disclosure provides a method for selecting a subject for treatment with a ghrelin pathway antagonist. Optionally, the method includes selecting a subject for such treatment on the basis that the subject is in need of preservation of body weight. The method includes evaluating (e.g., measuring) body weight in the subject, and comparing the body weight to a standard (e.g., a standard described herein), wherein the subject is selected for treatment with a ghrelin pathway antagonist if the difference between the body weight and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. Optionally, the ghrelin pathway antagonist is selected on the basis that it acts to preserve body weight.

In some embodiments, the subject is evaluated to determine if the subject is in need of preservation of body weight or if the subject has altered (e.g., increased or decreased) body weight, e.g., as compared to a standard (e.g., a standard described herein). For example, in some embodiments, a subject is in need of preservation of body weight if the subject has prediabetes or a risk factor thereof, the subject' s body weight is increased relative to the standard, the subject has diabetes or a risk factor thereof, the subject's body weight is decreased relative to the standard, the subject has insulin resistance, and/or the subject has obesity or a risk factor thereof.

In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject's body weight is increased relative to the standard.

In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject's body weight is decreased relative to the standard.

In another aspect, the disclosure provides a method for selecting a subject for treatment with a ghrelin pathway antagonist. Optionally, the method includes selecting a subject for such treatment on the basis that the subject is in need of preservation of pancreatic islet triglyceride content. The method includes evaluating (e.g., measuring) pancreatic islet triglyceride content in the subject, and comparing the pancreatic islet triglyceride content to a standard (e.g., a standard described herein), wherein the subject is selected for treatment with a ghrelin pathway antagonist if the difference between the pancreatic islet triglyceride content and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. Optionally, the ghrelin pathway antagonist is selected on the basis that it acts to preserve pancreatic islet triglyceride content.

In some embodiments, the subject is evaluated to determine if the subject is in need of preservation of pancreatic islet triglyceride content or if the subject has altered (e.g., increased or decreased) pancreatic islet triglyceride content, e.g., as compared to a standard (e.g., a standard described herein). For example, in some embodiments, a subject is in need of preservation of pancreatic islet triglyceride content if the subject has prediabetes or a risk factor thereof, the subject's pancreatic islet triglyceride content is increased relative to the standard, the subject has diabetes or a risk factor thereof, the subject's pancreatic islet triglyceride content is decreased relative to the standard, the subject has insulin resistance, and/or the subject has obesity or a risk factor thereof. In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject's pancreatic islet triglyceride content is increased relative to the standard.

In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject's pancreatic islet triglyceride content is decreased relative to the standard.

In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has obesity or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist binds to ghrelin.

In some embodiments, the ghrelin pathway antagonist binds to the ghrelin receptor (GHS-R). In some embodiments, the ghrelin pathway antagonist is ghrelin receptor antagonist [D-Lys³]-GHRP-6. In some embodiments, the ghrelin pathway antagonist is used in combination with another therapeutic agent. In some preferred embodiments, the ghrelin pathway antagonist is used in combination with another ghrelin pathway antagonist, e.g., another ghrelin pathway antagonist described herein. In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for obesity.

In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for metabolic syndrome.

In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for prediabetes .

In another aspect, the disclosure provides a method for selecting a subject for treatment with a ghrelin pathway antagonist. Optionally, the method includes selecting a subject for such treatment on the basis that the subject is in need of preservation of pancreatic islet lipid content. The method includes evaluating (e.g., measuring) pancreatic islet lipid content in the subject, and comparing the pancreatic islet lipid content to a standard (e.g., a standard described herein), wherein the subject is selected for treatment with a ghrelin pathway antagonist if the difference between the pancreatic islet lipid content and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. Optionally, the ghrelin pathway antagonist is selected on the basis that it acts to preserve pancreatic islet lipid content.

In some embodiments, the subject is evaluated to determine if the subject is in need of preservation of pancreatic islet lipid content or if the subject has altered (e.g., increased or decreased) pancreatic islet lipid content, e.g., as compared to a standard (e.g., a standard described herein). For example, in some embodiments, a subject is in need of preservation of pancreatic islet lipid content if the subject has prediabetes or a risk factor thereof, the subject's pancreatic islet lipid content is increased relative to the standard, the subject has diabetes or a risk factor thereof, the subject's pancreatic islet lipid content is decreased relative to the standard, the subject has insulin resistance, and/or the subject has obesity or a risk factor thereof.

In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject's pancreatic islet lipid content is increased relative to the standard. In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject's pancreatic islet lipid content is decreased relative to the standard.

In some embodiments, the subject has insulin resistance.

In some embodiments, the ghrelin pathway antagonist binds to ghrelin. In some embodiments, the ghrelin pathway antagonist binds to the ghrelin receptor (GHS-R). In some embodiments, the ghrelin pathway antagonist is ghrelin receptor antagonist [D-Lys³]-GHRP-6.

In another aspect, the disclosure provides a method for identifying a subject as suitable for (e.g., being a candidate for) treatment with a ghrelin pathway antagonist. The method includes evaluating (e.g., measuring) pancreatic islet mass in the subject (e.g., as described herein), and comparing the pancreatic islet mass to a standard (e.g., a standard described herein), wherein the subject is suitable for treatment with a ghrelin pathway antagonist if the difference between the pancreatic islet mass and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%.

In some embodiments, the subject has prediabetes or a risk factor thereof. In some embodiments, the subject's pancreatic islet mass is increased relative to the standard.

In some embodiments, the subject has diabetes or a risk factor thereof. In some embodiments, the subject's pancreatic islet mass is decreased relative to the standard.

In some embodiments, the subject has insulin resistance.

In another aspect, the disclosure provides a method for identifying a subject as suitable for (e.g., being a candidate for) treatment with a ghrelin pathway antagonist. The method includes evaluating (e.g., measuring) pancreatic islet function in the subject (e.g., as described herein), and comparing the pancreatic islet function to a standard (e.g., a standard described herein), wherein the subject is suitable for treatment with a ghrelin pathway antagonist if the difference between the pancreatic islet function and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%.

In some embodiments, the subject has prediabetes or a risk factor thereof. In some embodiments, the subject's pancreatic islet function is increased relative to the standard.

In some embodiments, the subject has diabetes or a risk factor thereof. In some embodiments, the subject's pancreatic islet function is decreased relative to the standard.

In another aspect, the disclosure provides a method for identifying a subject as suitable for (e.g., being a candidate for) treatment with a ghrelin pathway antagonist. The method includes evaluating (e.g., measuring) pancreas weight in the subject (e.g., as described herein), and comparing the pancreas weight to a standard (e.g., a standard described herein), wherein the subject is suitable for treatment with a ghrelin pathway antagonist if the difference between the pancreas weight and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%.

In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject's pancreas weight is increased relative to the standard.

In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist comprises a nucleic acid. In some preferred embodiments, the nucleic acid comprises a nucleic acid aptamer, RNAi, an antisense RNA, or a ribozyme molecule. In some embodiments, the ghrelin pathway antagonist binds to ghrelin. In some embodiments, the ghrelin pathway antagonist binds to the ghrelin receptor (GHS-R).

In one aspect, the disclosure provides a method for identifying a subject as suitable for (e.g., being a candidate for) treatment with a ghrelin pathway antagonist. The method includes evaluating (e.g., measuring) body weight in the subject, and comparing the body weight to a standard (e.g., a standard described herein), wherein the subject is suitable for treatment with a ghrelin pathway antagonist if the difference between the body weight and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. In some embodiments, the subject has prediabetes or a risk factor thereof. In some embodiments, the subject's body weight is increased relative to the standard. In some embodiments, the subject has diabetes or a risk factor thereof.

In another aspect, the disclosure provides a method for identifying a subject as suitable for (e.g., being a candidate for) treatment with a ghrelin pathway antagonist. The method includes evaluating (e.g., measuring) pancreatic islet triglyceride content in the subject (e.g., as described herein), and comparing the pancreatic islet triglyceride content to a standard (e.g., a standard described herein), wherein the subject is suitable for treatment with a ghrelin pathway antagonist if the difference between the pancreatic islet triglyceride content and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%.

In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject has diabetes or a risk factor thereof. In some embodiments, the subject's pancreatic islet triglyceride content is decreased relative to the standard.

In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has obesity or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist binds to ghrelin.

In some embodiments, the ghrelin pathway antagonist is a compound described herein, e.g., a compound of a formula described herein, e.g., a compound of formula (I), (II), (III), (IV), (V), or (VI). In one aspect, the disclosure provides a method for identifying a subject as suitable for (e.g., being a candidate for) treatment with a ghrelin pathway antagonist. The method includes evaluating (e.g., measuring) pancreatic islet lipid content in the subject (e.g., as described herein), and comparing the pancreatic islet lipid content to a standard (e.g., a standard described herein), wherein the subject is suitable for treatment with a ghrelin pathway antagonist if the difference between the pancreatic islet lipid content and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%.

In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject's pancreatic islet lipid content is increased relative to the standard.

In some embodiments, the subject has diabetes or a risk factor thereof. In some embodiments, the subject's pancreatic islet lipid content is decreased relative to the standard.

In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has obesity or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist comprises a non- proteinaceous molecule, e.g., a molecule less than 3000, 2000, 1000, 700, 500, 400, 300, or 200 in molecular weight. In some embodiments, the ghrelin pathway antagonist comprises a nucleic acid. In some preferred embodiments, the nucleic acid comprises a nucleic acid aptamer, RNAi, an antisense RNA, or a ribozyme molecule.

In one aspect, the disclosure provides a method for identifying a subject as suitable for (e.g., being a candidate for) treatment with a ghrelin pathway antagonist. The method includes identifying the subject as need pancreas sparing (e.g., by measuring pancreas function, e,g., insulin production), and comparing the pancreas function of the subject to a standard (e.g., a standard described herein), wherein the subject is suitable for treatment with a ghrelin pathway antagonist if the difference between the pancreas function and the standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. For example, the standard can be a cohort of subjects that do not have diabetes, prediabetes or an impairment of pancreas function. In some embodiments, the subject has prediabetes or a risk factor thereof.

In some embodiments, the subject has diabetes or a risk factor thereof.

In some embodiments, the subject has insulin resistance.

In some embodiments, the subject has obesity or a risk factor thereof.

In some embodiments, the ghrelin pathway antagonist comprises a non- proteinaceous molecule, e.g., a molecule less than 3000, 2000, 1000, 700, 500, 400, 300, or 200 in molecular weight. In some embodiments, the ghrelin pathway antagonist comprises a nucleic acid.

In some preferred embodiments, the nucleic acid comprises a nucleic acid aptamer, RNAi, an antisense RNA, or a ribozyme molecule.

In some embodiments, the ghrelin pathway antagonist binds to ghrelin.

Increased dietary fat intake is associated with an increase in islet cell mass by increasing the cell number and/or the size of the islet cells. Changes in pancreatic islet mass can occur due to a change in the number and/or size of cells that make up the islet.

By the term "preserved," it is meant that change does not occur, or occurs to a lesser extent or more slowly (e.g., with a delay) than it would have occurred in the absence of preserving. For example, preserving can prevent a change (e.g., an increase or a decrease) in a parameter (e.g., a parameter described herein) of greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or about 95% as compared to a standard (e.g., a reference value). E.g., the parameter changes by less than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or about 95% as compared to a standard (e.g., a reference value). As another example, preserving can delay a change in a parameter (e.g., a parameter described herein). For example, preserving can the delay the change in the parameter by about 1 week, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 24 weeks, about 28 weeks, about 32 weeks, about 36 weeks, about 40 weeks, about 44 weeks, about 48 weeks, about 52 weeks, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 3.5 years, about 4 years, about 4.5 years, about 5 years, about 5.5 years, about 6 years, about 6.5 years, about 7 years, about 7.5 years, about 8 years, about 9.5 years, about 10 years, about 12 years, about 14 years, about 16 years, about 18 years, about 20 years or longer, e.g., as compared to a standard, e.g., the average time for the parameter change in a reference, e.g., a cohort of subjects that progressed from prediabetes to diabetes (e.g., type 2 diabetes).

For example, the pancreatic islet mass is preserved as opposed to the mass changing. Preserved can indicate, e.g., that the mass is unchanged, or similar to, or altered (e.g., increased or decreased) less than it otherwise would have been altered, e.g., relative to a standard. The standard can be, for example, pancreatic islet mass present in the subject before the first administration of the ghrelin pathway antagonist (alone or in combination with another agent), or the pancreatic islet mass present in the subject before the onset of obesity or the onset of prediabetes (or a symptom thereof) or the onset of diabetes (or a symptom thereof), e.g., type 2 diabetes. For example, the ghrelin pathway antagonist (alone or in combination with another agent) can preserve the pancreatic islet mass, e.g., by preventing a change (e.g., an increase or a decrease) in islet mass of greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90% as compared to a standard, e.g., the subject's pancreatic islet mass before the onset of obesity or the onset of prediabetes (or a symptom thereof) or the onset of diabetes (or a symptom thereof), e.g., type 2 diabetes. As another example, the administration of the ghrelin pathway antagonist (alone or in combination with another agent) can preserve the pancreatic islet mass, e.g., prevent a change in pancreatic islet mass in a subject (e.g., a pre-diabetic subject) over time. The preservation can, e.g., extend or ameliorate a subject's prediabetes and/or prevent or delay the development of diabetes or insulin dependence.

For example, the ghrelin pathway antagonist (alone or in combination with another agent) can prevent a change (e.g., an increase) in islet mass of greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90% as compared to a standard, e.g., the subject's pancreatic islet mass before the onset of prediabetes.

As another example, the the ghrelin pathway antagonist (alone or in combination with another agent) can prevent a change (e.g., a decrease) in islet mass of greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90% as compared to a standard, e.g., the subject's pancreatic islet mass before the onset of diabetes (e.g., type 2 diabetes).

As a further example, the administration of the ghrelin pathway antagonist (alone or in combination with another agent) can preserve the pancreatic islet mass, e.g., delay a change in pancreatic islet mass in a subject (e.g., a pre-diabetic subject). For example, the ghrelin pathway antagonist can delay the increase in pancreatic islet mass by about 1 week, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 24 weeks, about 28 weeks, about 32 weeks, about 36 weeks, about 40 weeks, about 44 weeks, about 48 weeks, about 52 weeks, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 3.5 years, about 4 years, about 4.5 years, about 5 years, about 5.5 years, about 6 years, about 6.5 years, about 7 years, about 7.5 years, about 8 years, about 9.5 years, about 10 years, about 12 years, about 14 years, about 16 years, about 18 years, about 20 years or longer, e.g., as compared to a standard, e.g., the average time to the onset of diabetes (e.g., type 2 diabetes) in a reference, e.g., a cohort of subjects that progressed from prediabetes to diabetes. As another example, the ghrelin pathway antagonist can delay the decrease (e.g., in a diabetic subject) in pancreatic islet mass by about 1 week, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 24 weeks, about 28 weeks, about 32 weeks, about 36 weeks, about 40 weeks, about 44 weeks, about 48 weeks, about 52 weeks, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 3.5 years, about 4 years, about 4.5 years, about 5 years, about 5.5 years, about 6 years, about 6.5 years, about 7 years, about 7.5 years, about 8 years, about 9.5 years, about 10 years, about 12 years, about 14 years, about 16 years, about 18 years, about 20 years or longer, e.g., as compared to a standard, e.g., a standard described herein. As a further example, the administration of the ghrelin pathway antagonist (alone or in combination with another agent) can preserve the pancreas weight, e.g., prevent a change in pancreas weight in a subject over time. For example, the ghrelin pathway antagonist (alone or in combination with another agent) can prevent a change (e.g., an 5 increase) in pancreas weight of greater than about 5%, about 10%, about 15%, about

20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90% as compared to a standard, e.g., the subject's pancreas weight before the onset of prediabetes. As another example, the administration of the ghrelin pathway antagonist o (alone or in combination with another agent) can preserve the pancreas weight, e.g., delay a change in pancreas weight in a subject (e.g., a pre-diabetic subject). For example, the ghrelin pathway antagonist can delay a change in pancreas weight by about 1 week, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 24 weeks, about 28 weeks, about 32 weeks, about 36 weeks, about 40 weeks, about 445 weeks, about 48 weeks, about 52 weeks, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 3.5 years, about 4 years, about 4.5 years, about 5 years, about 5.5 years, about 6 years, about 6.5 years, about 7 years, about 7.5 years, about 8 years, about 9.5 years, about 10 years, about 12 years, about 14 years, about 16 years, about 18 years, about 20 years or longer, e.g., as compared to a standard, e.g., the average time to the0 onset of diabetes (e.g., type 2 diabetes) in a reference, e.g., a cohort of subjects that progressed from prediabetes to diabetes.

As a further example, the administration of the ghrelin pathway antagonist (alone or in combination with another agent) can preserve the body weight, e.g., prevent a change in body weight in a subject over time, e.g., in a subject with pre-diabetes,5 diabetes, insulin resistance, and/or obesity. For example, the ghrelin pathway antagonist (alone or in combination with another agent) can prevent a change (e.g., an increase) in body weight of greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90% as compared to a0 standard, e.g., the subject's body weight before the onset of prediabetes. As another example, the administration of the ghrelin pathway antagonist (alone or in combination with another agent) can preserve the body weight, e.g., delay a change in pancreas weight in a subject (e.g., a pre-diabetic subject). For example, the ghrelin pathway antagonist can delay a change in body weight by about 1 week, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 24 weeks, about 28 weeks, about 32 5 weeks, about 36 weeks, about 40 weeks, about 44 weeks, about 48 weeks, about 52 weeks, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 3.5 years, about 4 years, about 4.5 years, about 5 years, about 5.5 years, about 6 years, about 6.5 years, about 7 years, about 7.5 years, about 8 years, about 9.5 years, about 10 years, about 12 years, about 14 years, about 16 years, about 18 years, about 20 years or longer, e.g., as o compared to a standard, e.g., the average time to the onset of diabetes (e.g., type 2 diabetes) in a reference, e.g., a cohort of subjects that progressed from prediabetes to diabetes.

As a further example, the administration of the ghrelin pathway antagonist (alone or in combination with another agent) can preserve islet triglyceride content, e.g., prevent5 a change in triglyceride content in a subject over time. For example, the ghrelin pathway antagonist (alone or in combination with another agent) can prevent a change (e.g., an increase) in islet triglyceride content of greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about0 90% as compared to a standard, e.g., the subject's islet triglyceride content before the onset of prediabetes or diabetes. As another example, the administration of the ghrelin pathway antagonist (alone or in combination with another agent) can preserve the islet triglyceride content, e.g., delay a change in triglyceride content in a subject (e.g., a pre- diabetic subject). For example, the ghrelin pathway antagonist can delay a change in islet5 triglyceride content by about 1 week, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 24 weeks, about 28 weeks, about 32 weeks, about 36 weeks, about 40 weeks, about 44 weeks, about 48 weeks, about 52 weeks, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 3.5 years, about 4 years, about 4.5 years, about 5 years, about 5.5 years, about 6 years, about 6.5 years, about 7 years,0 about 7.5 years, about 8 years, about 9.5 years, about 10 years, about 12 years, about 14 years, about 16 years, about 18 years, about 20 years or longer, e.g., as compared to a standard, e.g., the average time to the onset of diabetes (e.g., type 2 diabetes) in a reference, e.g., a cohort of subjects that progressed from prediabetes to diabetes.

As a further example, the administration of the ghrelin pathway antagonist (alone or in combination with another agent) can preserve islet lipid content, e.g., prevent a change in lipid content in a subject over time. For example, the ghrelin pathway antagonist (alone or in combination with another agent) can prevent a change (e.g., an increase) in islet lipid content of greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90% as compared to a standard, e.g., the subject's islet lipid content before the onset of prediabetes or diabetes. As another example, the administration of the ghrelin pathway antagonist (alone or in combination with another agent) can preserve the islet lipid content, e.g., delay a change in lipid content in a subject (e.g., a pre-diabetic subject). For example, the ghrelin pathway antagonist can delay a change in islet lipid content by about 1 week, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 24 weeks, about 28 weeks, about 32 weeks, about 36 weeks, about 40 weeks, about 44 weeks, about 48 weeks, about 52 weeks, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 3.5 years, about 4 years, about 4.5 years, about 5 years, about 5.5 years, about 6 years, about 6.5 years, about 7 years, about 7.5 years, about 8 years, about 9.5 years, about 10 years, about 12 years, about 14 years, about 16 years, about 18 years, about 20 years or longer, e.g., as compared to a standard, e.g., the average time to the onset of diabetes (e.g., type 2 diabetes) in a reference, e.g., a cohort of subjects that progressed from prediabetes to diabetes.

As another example, the standard can be a cohort of subjects, e.g., subjects with obesity, prediabetes, diabetes, or insulin resistance, e.g., such that it is below the average parameter for the selected cohort.

In some embodiments, the islet mass can be measured indirectly, e.g., by measuring pancreas parenchyma, e.g., by computer tomography (CT) density measurements (see, e.g., Saisho et al., Clin. Anat. 20:933-942 (2007)). In some embodiment, positron emission tomography (PET) can be used to estimate islet mass (see, e.g. Souza et al. (J. Clin. Invest. 116:1506-1513 (2006)). "Insulin resistance" (IR) refers to a condition in which the cells of the body become resistant to the effects of insulin, that is, the normal response to a given amount of insulin is reduced (e.g., the cells are less sensitive to insulin/have decreased insulin sensitivity). As a result, higher levels of insulin are needed in order for insulin to have its 5 effects.

In some aspects, the disclosure provides a method for evaluating a subject, e.g., a subject to whom a ghrelin pathway antagonist (e.g., a ghrelin pathway antagonist described herein) has been administered. The method includes evaluating one or more of o the following parameters of the subject: pancreatic islet mass; pancreatic islet function; pancreas weight; body weight; pancreatic islet triglyceride content; pancreatic islet lipid content; and the pancreas function (e.g., insulin production).

The evaluating can be performed, e.g., about 1 week, about 2 weeks, about 35 weeks, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 24 weeks, about 28 weeks, about 32 weeks, about 36 weeks, about 40 weeks, about 44 weeks, about 48 weeks, about 52 weeks, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 3.5 years, about 4 years, about 4.5 years, about 5 years, about 5.5 years, about 6 years, about 6.5 years, about 7 years, about 7.5 years, about 8 years,0 about 9.5 years, about 10 years, about 12 years, about 14 years, about 16 years, about 18 years, or about 20 years after the ghrelin pathway antagonist was administered to the subject. The evaluating can also, or alternatively, be performed at the time administration of the ghrelin pathway antagonist commenced (e.g., within 1, 2, 3, 4, or 5 days of commencing) and/or ceased (e.g., at the time of the last administration of the antagonist,5 (e.g., within 1, 2, 3, 4, or 5 days of ceasing)). Optionally, evaluating can also be performed prior to administering (e.g., about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 8 weeks, or about 12 weeks prior to administering).

The evaluating can include comparing the value of the parameter from the subject to a standard (e.g., a standard described herein), and optionally determining if a 0 difference exists between the value of the parameter from the subject and the value from the standard. The evaluating can include determining if one or more of the following are present: the difference between the pancreatic islet mass of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and/or the difference between the pancreatic islet function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreas weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the body weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet triglyceride content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about

25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet lipid content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and the difference between the pancreas function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. In another aspect, the disclosure provides a method for evaluating a subject, e.g., a subject to whom a ghrelin pathway antagonist (e.g., a ghrelin pathway antagonist described herein) is currently being administered. The method includes evaluating one or more of the following parameters of the subject:

5 pancreatic islet mass; pancreatic islet function; pancreas weight; body weight; pancreatic islet triglyceride content; pancreatic islet lipid content; and the pancreas function (e.g., insulin production).

The evaluating can be performed, e.g., about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, o about 24 weeks, about 28 weeks, about 32 weeks, about 36 weeks, about 40 weeks, about 44 weeks, about 48 weeks, about 52 weeks, about 1.5 years, about 2 years, about 2.5 years, about 3 years, about 3.5 years, about 4 years, about 4.5 years, about 5 years, about 5.5 years, about 6 years, about 6.5 years, about 7 years, about 7.5 years, about 8 years, about 9.5 years, about 10 years, about 12 years, about 14 years, about 16 years, about 185 years, or about 20 years after commencing administration of the ghrelin pathway antagonist to the subject (e.g., the evaluating can be performed at one or more of these times, and/or the evalutiang can be performed at weekly intervals, monthly intervals, and so forth). The evaluating can also, or alternatively, be performed at the time administration of the ghrelin pathway antagonist commences (e.g., within 1, 2, 3, 4, or 50 days of commencing). Optionally, evaluating can also be performed prior to administering (e.g., about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 8 weeks, or about 12 weeks prior to administering).

The evaluating can include comparing the value of the parameter from the subject to a standard (e.g., a standard described herein), and optionally determining if a 5 difference exists between the value of the parameter from the subject and the value from the standard.

The evaluating can include determining if one or more of the following are present: the difference between the pancreatic islet mass of the subject and that of a0 standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and/or the difference between the pancreatic islet function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreas weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the body weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet triglyceride content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet lipid content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and the difference between the pancreas function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%.

In another aspect, the disclosure provides a method of selecting a drug for administration to a subject, the method comprising selecting a drug on the basis that the drug can preserve or optimize pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content. Optionally, the method can include providing the drug to the subject, e.g., wherein providing includes administering the drug or transferring the drug to the subject's possession.

In one aspect, the disclosure provides a method of selecting a drug for administration to a subject in need of preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content, the method comprising selecting a drug on the basis that the drug can preserve or optimize pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content. Optionally, the method can include providing the drug to the subject, e.g., wherein providing includes administering the drug or transferring the drug to the subject's possession.

In another aspect, the disclosure provides a method for evaluating a subject, e.g., a subject who has been selected to receive treatment with a ghrelin pathway antagonist (e.g., a ghrelin pathway antagonist described herein). The method includes evaluating one or more of the following parameters of the subject: pancreatic islet mass; pancreatic islet function; pancreas weight; body weight; pancreatic islet triglyceride content; pancreatic islet lipid content; and the pancreas function (e.g., insulin production).

The evaluating can include comparing the value of the parameter from the subject to a standard (e.g., a standard described herein), and optionally determining if a difference exists between the value of the parameter from the subject and the value from the standard.

The evaluating can include determining if one or more of the following are present: the difference between the pancreatic islet mass of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and/or the difference between the pancreatic islet function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreas weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the body weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet triglyceride content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet lipid content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and the difference between the pancreas function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%.

In another aspect, the disclosure provides a method of prescribing a ghrelin pathway antagonist drug, the method comprising: receiving an identifier for the ghrelin pathway antagonist drug, e.g., the chemical structure, chemical name, trade name or generic name of the ghrelin pathway antagonist drug; receiving information that the ghrelin pathway antagonist drug has one or more of the following properties: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content, or is useful for treating one or more of the following subject needs: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content; selecting a subject in need of the ghrelin pathway antagonist drug, e.g., on the basis that the subject is in need of one or more of: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content; and causing the ghrelin pathway antagonist drug to be prescribed, dispensed, or administered to a subject.

In another aspect, the disclosure provides a method of providing a recipient with information about, or with guidelines for, the use of a ghrelin pathway antagonist drug, the method comprising: communicating to the recipient an identifier for the ghrelin pathway antagonist drug, e.g., the chemical structure, chemical name, trade name or generic name of the ghrelin pathway antagonist drug; communicating to the recipient information that the ghrelin pathway antagonist drug has one or more of the following properties: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content, or is useful for treating one or more of the following subject needs: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content; receiving a request from the recipient to purchase the ghrelin pathway antagonist drug; and selling, shipping or transferring the ghrelin pathway antagonist drug to the recipient. In another aspect, the disclosure provides a method of providing a recipient with information about a ghrelin pathway antagonist drug, or with guidelines for, the use of a ghrelin pathway antagonist drug, the method comprising: providing an identifier for the ghrelin pathway antagonist drug, e.g., the chemical structure, chemical name, trade name or generic name of the ghrelin pathway antagonist drug; providing information that the ghrelin pathway antagonist drug has one or more of the following properties: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content, or is useful for treating one or more of the following subject needs: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content; memorializing the identifier and the information; and transferring the memorialization (e.g., the memorialized identifier and information) to the recipient.

In another aspect, the disclosure provides a method of providing a recipient with information about a ghrelin pathway antagonist drug, or with guidelines for the use of a ghrelin pathway antagonist drug, the method comprising: providing an identifier for the ghrelin pathway antagonist drug, e.g., the chemical structure, chemical name, trade name or generic name of the ghrelin pathway antagonist drug; providing information that the ghrelin pathway antagonist drug has one or more of the following properties: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content, or is useful for treating one or more of the following subject needs: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content; associating the identifier with the information, e.g., in a database or by physical association; and transferring the associated identifier and information to the recipient.

In another aspect, the disclosure provides a database, medium, or computer containing or programmed to contain: an identifier for a ghrelin pathway antagonist drug, e.g., the chemical structure, chemical name, trade name or generic name of the ghrelin pathway antagonist drug; information that the ghrelin pathway antagonist drug has one or more of the following properties: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content, or is useful for treating one or more of the following subject needs: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content; and an associative function associating the identifier with the information, e.g., in a database or by physical association.

In another aspect, the disclosure provides a method of making a ghrelin pathway antagonist drug available to a subject, the method comprising: providing to the subject an identifier for the ghrelin pathway antagonist drug, e.g., the chemical structure, chemical name, trade name or generic name of the ghrelin pathway antagonist drug; providing to the subject information that the ghrelin pathway antagonist drug has one or more of the following properties: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content, or is useful for treating one or more of the following subject needs: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content; and placing into commerce, a dose of the ghrelin pathway antagonist drug which can be administered to, provided to, or purchased by the subject.

In another aspect, the disclosure provides a method of causing a subject to request a ghrelin pathway antagonist drug, the method comprising: providing to the subject an identifier for the ghrelin pathway antagonist drug, e.g., the chemical structure, chemical name, trade name or generic name of the ghrelin pathway antagonist drug; providing to the subject information that the ghrelin pathway antagonist drug has one or more of the following properties: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content, or is useful for treating one or more of the following subject needs: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content; and placing into commerce, a dose of the ghrelin pathway antagonist drug which can be administered to, provided to, or purchased by the subject.

In another aspect, the disclosure provides a method for a subject to determine if a ghrelin pathway antagonist drug is appropriate for the subject, the method comprising: receiving an identifier for the ghrelin pathway antagonist drug, e.g., the chemical structure, chemical name, trade name or generic name of the ghrelin pathway antagonist drug; receiving information that the ghrelin pathway antagonist drug has one or more of the following properties: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content, or is useful for treating one or more of the following subject needs: preservation or optimization of pancreatic islet mass, pancreatic islet function, pancreas weight, body weight, pancreatic islet triglyceride content, and/or pancreatic islet lipid content; and contacting a healthcare provider to request treatment with or information about the ghrelin pathway antagonist drug.

In another aspect, the present disclosure provides an article of manufacture comprising: a) a packaging material; b) a ghrelin pathway antagonist; and c) a label or package insert contained within the packaging material indicating that the ghrelin pathway antagonist is effective for treating a subject as described herein. For example, the ghrelin pathway antagonist can be effective for treating a subject to preserve pancreatic islet mass, preserve pancreatic islet function, preserve pancreas weight, preserve body weight, preserve pancreatic islet triglyceride content, preserve pancreatic islet lipid content, and/or spare the pancreas.

In some embodiments, the ghrelin pathway antagonist is ghrelin receptor antagonist [D-Lys³]-GHRP-6. In some embodiments, the ghrelin pathway antagonist is used in combination with another therapeutic agent. In some preferred embodiments, the ghrelin pathway antagonist is used in combination with another ghrelin pathway antagonist, e.g., another ghrelin pathway antagonist described herein. In some embodiments, the ghrelin pathway antagonist is used in combination with a treatment for obesity.

In some aspects, the ghrelin pathway antagonist used in a method described herein is a compound, e.g., a small molecule.

In some embodiments, the compound, including stereoisomers thereof, can be created either singly, in small clusters, or in a combinatorial fashion to give structurally diverse libraries of compounds.

In one aspect, the methods described herein use a compound of formula (I)

formula (I) wherein, R¹ is hydrogen, halo (e.g., fluoro), aryl, heteroaryl, arylalkyl, heteroarylalkyl, cyclyl, cyclylalkyl, heterocyclyl, heterocyclylalkyl, alkyl, alkenyl, alkynyl, or R¹ can be taken together with R² or R³ to form a ring; each of which is optionally substituted with 1-4 R⁶; k' is a bond, O, C(O), C(O)O, OC(O), C(O)NR³, NR³C(O), S, SO, SO₂, CR²=CR², or C≡C; n is 0-6, preferably 1-3;

R is hydrogen, halo (e.g., fluoro), Ci-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl; or R² can be taken together with R¹ to form a ring;

R³ is hydrogen, C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl, or R³ can be taken together with R², R⁴, or R⁵ to form a ring; each of which can be optionally substituted with 1-2 R^6';

A is

_R7a _R7a

-|-(CH₂)_X C (CH₂)_y-|- -<-N— (CH₂)_X C (CH₂)_yγ

_R7b _R8 _R7b -(CH₂)_x M (CH₂)_yγ

x and y are each independently 0-6; M is aryl, heteroaryl, eye IyI, or heterocyclyl, each of which is optionally substituted with 1-4 R⁹;

R⁴ and R⁵ are each independently hydrogen, alkyl, alkenyl, haloalkyl, cyclyl, or heterocyclyl, or R⁴ and R⁵ can be taken together to form a heterocyclic ring, or R⁴ and R⁵ can be taken together to form an azido moiety, or one or both of R⁴ and R⁵ can independently be joined to one or both of R^7a and R^7b to form one or more bridges between the nitrogen to which the R⁴ and R⁵ are attached and R^7a and R^7b, wherein each bridge contains 1 to 5 carbons; or one or both of R⁴ and R⁵ can independently be joined to one or both of R^7a and R^7b to form to form one or more heterocyclic rings including the nitrogen to which the R⁴ and R⁵ are attached, or one or both of R⁴ and R⁵ can independently be joined to R³ to form a ring, or one or both of R⁴ and R⁵ can independently be joined to R⁸ to form a ring; wherein each R⁴ and R⁵ are optionally independently substituted with 1-5 halo, 1-3 hydroxy, 1-3 alkyl, 1-3 alkoxy, 1-3 oxo, 1-3 amino, 1-3 alkylamino, 1-3 dialklyamino, 1-3 nitrile, or 1-3 haloalkyl;

Y is a monocyclic aryl or monocyclic heteroaryl; each of which is optionally substituted with 1-4 R¹⁰; each R⁶ and R⁶ are independently halo, alkyl, alkenyl, alkynyl, cyclyl, heterocyclyl, aryl, heteroaryl, alkoxy, haloalkyl, haloalkyloxy, haloalkylthio, acetyl, cyano, nitro, hydroxy, oxo, C(O)OR², OC(O)R², N(R³)₂, C(O)N(R³)₂, NR³C(O)R², or SR²;

R^7a and R^7b are each independently hydrogen, alkyl, alkenyl, haloalkyl, cyclyl, cyclylalkyl, or heterocyclyl; or one or both of R^7a and R^7b can independently be joined to one or both of R⁴ and R⁵ to form one or more bridges between the nitrogen to which the R⁴ and R⁵ are attached and R^7a and R^7b, wherein each bridge contains 1 to 5 carbons; or one or both of R^7a and R^7b can independently be joined to one or both of R⁴ and R⁵ to form to form one or more heterocyclic rings including the nitrogen to which the R⁴ and R⁵ are attached, or one or both of R^7a and R^7b can independently be joined with R⁸ to form a ring; wherein each R^7a and R^7b can be independently optionally substituted with 1- 5 halo, 1-3 hydroxy, 1-3 alkyl, 1-3 alkoxy, 1-3 amino, 1-3 alkylamino, 1-3 dialklyamino, 1-3 nitrile, or 1-3 haloalkyl;

R⁸ is hydrogen or C₁-C₆ alkyl, or R⁸ can be joined with R⁴, R⁵, R^7a or R^7b to form a ring;

R⁹ is halo, alkyl, cyclyl, heterocyclyl, aryl, heteroaryl, alkoxy, haloalkyl, haloalkyloxy, haloalkylthio, acetyl, cyano, nitro, hydroxy, oxo, C(O)OR², OC(O)R², N(R²)₂, C(O)N(R²)₂, NR²C(O)R², SR²; each R¹⁰ is independently alkyl, alkenyl, alkynyl, halo, cyano, carbonyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, cyclyl, cyclylalkyl, alkoxy, alkoxyalkyl, aryloxy, aryloxyalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, heteroarylalkyl, -OR¹¹, - NR¹¹R^11', -CF₃, -SOR¹², -SO₂R¹², -OC(O)R¹¹, -SO₂NR¹²R^12', -(CH₂)_mR¹⁴ or R¹⁵; each of which is optionally independently substituted with 1-3 R¹⁶;

R¹¹ and R¹¹ are each independently hydrogen, alkyl, alkenyl, alkynyl, cyclyl, heterocyclyl, aryl or heteroaryl; R¹² and R¹² are each independently hydrogen, alkyl, alkenyl, alkynyl, alkylthioalkyl, alkoxyalkyl, aryl, arylalkyl, heterocyclyl, heteroaryl, heteroarylalkyl, heterocycloalkyl or cyclyl, cyclylalkyl, or R¹² and R¹² taken together can be cyclized to form -(CH₂)_qX(CH₂)_s-; wherein each R¹² and R¹² may independently optionally be substituted with 1 to 3 substituents selected from the group consisting of halogen, OR¹¹, alkoxy, heterocycloalkyl, -NR¹¹C(O)NR¹¹R^11', -C(O)NR¹¹R^11', -NR¹¹C(O)R^11', -CN, oxo, -NR¹¹SO₂R^11', -OC(O)R¹¹, -SO₂NR¹¹R^11', -SOR¹³, -S(O)₂R¹³, -COOH and -C(O)OR¹³; each R¹³ is independently alkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl, each of which may optionally be substituted with -(CH₂)_w0H; each R¹⁴ is independently alkoxy, alkoxycarbonyl, -C(O)NR¹²R^12', -NR¹¹R^11', -

C(O)R¹², -NR¹¹C(O)NR¹¹R^11' or -N-heteroaryl; each R¹⁵ is independently -(CH₂)_PN(R¹²)C(O)R^12', -(CH₂)_PCN, - (CH₂)_PN(R¹²)C(O)OR^12', -(CH₂)_PN(R¹²)C(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂R¹², - (CH₂)_PSO₂NR¹²R^12', -(CH₂)_PC(O)NR¹²R^12', -(CH₂)_PC(O)OR¹², -(CH²)_POC(O)OR¹², - (CH₂)_pOC(O)R¹², -(CH₂)_POC(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂NR¹²R^12', -(CH₂)_POR¹², -

(CH₂)_pOC(O)N(R¹²)(CH₂)_mOH, -(CH₂)_PSOR¹², -(CH₂)_PSO₂R¹², -(CH₂)_PNR¹¹R¹¹ or -

(CH₂)_pOCH₂C(O)N(R¹²)(CH₂)_mOH; each R¹⁶ is independently halo, alkyl, alkenyl, alkynyl, alkoxy, - (CH₂)_PNR¹¹C(O)NR¹¹R^11', -(CH₂)_PC(O)NR¹¹R^11', -(CH₂)_PNR¹¹C(O)R^11', -CN, - (CH₂)_PNR¹¹SO₂R^11', -(CH₂)_POC(O)R¹ \ -(CH₂)_PSO₂NR¹¹R^11', -(CH₂)_PSOR¹³, - (CH₂)_pC00H or -(CH₂)_PC(O)OR¹³;

X is CR¹¹R^11', O, S, S(O), S(O)₂, or NR¹¹; m is an integer between 1 and 6; p is an integer from O to 5; q and s are each independently an integer between 1 and 3; and w is an integer between O and 5.

In some embodiments, formula (I), comprises an enriched preparation of formula

(D

foπnula (V). In some embodiments, foπnula (I), comprises an enriched preparation of formula

(I")

formula (I"). In some embodiments, n is 1; k' is a bond or O; and R¹ is aryl, heteroaryl, arylalkyl, or heteroarylalkyl.

In some embodiments, n is 1; k' is O; and R¹ is arylalkyl. For example, R¹ can be phenylmethyl.

In some embodiments, n is 2; k' is a bond; and R¹ is aryl. In some embodiments, n is 0 or 1 ; k' is a bond; and

R¹ is alkyl, for example unsubstituted or substituted with one R⁶. For example, R¹ can be a branched alkyl such as one of the following .

In some embodiments, R² is hydrogen or C₁-C₃ alkyl.

In some embodiments n is 0 and k' is a bond. Exemplary R¹ moieties include methyl, and ethyl. Preferred R¹ moieties include methyl. In some embodiments R¹ is unsubstituted methyl or methyl or ethyl substituted with C(O)N(R³)₂. In some embodiments n is 0 and k' is a bond, and R¹ and R² are both methyl.

In some embodiments, n is 0; k' is a bond; and

R¹ is hydrogen. In some embodiments R³ is hydrogen.

In some embodiments, R¹ and R³ together form a heterocyclic ring such as a pyrrolidine or an azetidine ring. The heterocyclic ring can be unsubstituted or substituted, for example, with 1-2 R⁶.

In some embodiments, R¹ and R² together form a ring. In some embodiments,

A is

_R7a _R7a

4-(CH₂)_x C (CH₂)_y-|- -$-N— (CH₂)_X C (CH₂)_y-|-

R^7b or R⁸ R^7b

For example, A can be

R^7a and R^7b are H; x is 1 ; and y is 0 or 1.

In some embodiments, A is CH₂CH₂ or CH₂CH₂CH₂; and each R⁴ and R⁵ is independently alkyl, or R⁴ and R⁵, when taken together, form a heterocyclic ring. In some embodiments, R^7a and R^7b can each be H.

In some embodiments, at least one of R^7a or R^7b is taken together with at least one of R⁴ or R⁵ to form a heterocyclic ring including the nitrogen to which the R⁴ and R⁵ are attached.

In some embodiments, R^7a and R^7b are each independently alkyl;

R⁴ and R⁵ are each independently hydrogen or alkyl; and x and y are each independently 0 or 1.

In some embodiments,

taken together is

In some embodiments,

, taken together is

In some embodiments,

, taken together is

In some embodiments,

, taken together is

In some embodiments,

taken together is

In some embodiments, Y is a monocyclic heteroaromatic moiety, for example a nitrogen containing heteraromatic moiety such as a nitrogen containing five membered heteraromatic moiety.

In some embodiments, Y is a heterocyclic moiety containing at least two heteroatoms, for example, a five membered heterocyclic moiety containing at least two heteroatoms or at least three heteroatoms. In some embodiments Y is substituted with one R¹⁰. R¹⁰ can be positioned, for example, 1,3 relative to the point of attachment of Y to the adjacent chain carbon or 1,2 relative to the point of attachment of Y to the adjacent chain carbon.

In some embodiments, R¹⁰ is aryl or heteroaryl, for example a monocyclic aryl or monocyclic heteroaryl such as phenyl, pyridyl, oxazolyl, thiazolyl, or thiophenyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy.

In some embodiments, R¹⁰ is a bicyclic heteroaryl, for example indolyl, benzimidazolyl, benzoxazolyl, benzofuranyl, benzothiophenyl, or benzthiazolyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy.

In some embodiments, R¹⁰ is arylalkyl or heteroarylalky, for example a monocyclic or bicyclic arylalkyl or monocyclic or bicyclic heteroary alkyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy.

In some embodiments, R¹⁰ includes an unsaturated or partially unsaturated cyclic moiety, for example a cyclyl or heterocyclyl moiety. The cyclic moiety can either be directly attached to Y or attached via a linker such as an alkylenyl linker. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy. In some embodiments Y is oxadiazole or triazole.

In another aspect, the methods described herein use a compound of formula (II),

formula (II) wherein,

Q\ Q² _J Q^{3 an}d Q⁴ together with the carbon to which they are attached form a heteroaryl moiety, and each Q¹, Q², Q³ and Q⁴ is independently S, O, N, CR², CR¹⁰, NR², or NR 10 In some embodiments, the compound of formula (II), comprises an enriched preparation of formula (H')

formula (IF). In some embodiments, the compound of formula (II), comprises an enriched preparation of formula (H")

formula (H").

In some embodiments, Q¹ and Q⁴ are each independently S, O, N, or NR¹⁰. In some embodiments, Q¹ and Q³ are each independently S, O, N, or NR¹⁰.

In some embodiments, Q² is CR² or CR¹⁰.

In some embodiments, Q² is S, O, N, or NR¹⁰.

In some embodiments, at least one of Q² or Q³ is CR² or CR¹⁰.

In some embodiments, at least two of Q¹, Q², Q³, or Q⁴ is S, O, N, or NR¹⁰. In some embodiments, Q¹, Q², and Q³ are each independently S, O, N, or NR¹⁰.

In some embodiments, Q¹ is NR¹⁰.

In some embodiments, one of Q², Q³, or Q⁴ is CR²

In some embodiments, Q² is CR¹⁰.

In some embodiments, Q³ is CR²

In some embodiments, Q¹, Q², Q³ and Q⁴ together form

In some embodiments, Q¹ is NR². In some embodiments, Q¹, Q², Q³ and Q⁴ together form

In some embodiments, Q¹ is NR¹⁰.

In another aspect, the methods described herein use a compound of formula (III),

wherein,

Z¹, Z², Z³, Z⁴, and Z⁵ together form an aryl or heteroaryl moiety, and each Z¹, Z², Z³, Z⁴, and Z⁵ is independently N, CR¹⁰, or CR². In some embodiments, the compound of formula (III), comprises an enriched preparation of formula (III')

formula (HF).

In some embodiments, the compound of formula (III), comprises an enriched preparation of formula (III')

formula (HF).

In some embodiments, one of Z¹, Z², Z³, Z⁴, and Z⁵ is N.

In some embodiments, two of Z¹, Z², Z³, Z⁴, and Z⁵ are N. In some embodiments, three of Z¹, Z², Z³, Z⁴, and Z⁵ is N. In some embodiments, two of Z¹ and Z² are N. In some embodiments, two of Z¹ and Z³ are N.

In some embodiments, two of Z¹ and Z⁴ are N.

In some embodiments, two of Z¹, Z³, and Z⁵ are N.

In some embodiments, the compound is a compound of formula (I), wherein Y is substituted with a single substituent R¹⁰. For example, R¹⁰ can be aryl or heteroaryl, optionally substituted with up to three independent R¹⁶.

In some embodiments, R¹⁰ is a bicyclic heteroaryl, for example indolyl, benzimidazolyl, benzoxazolyl, benzofuranyl, benzothiophenyl, or benzthiazolyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy. In some embodiments, R¹⁰ is arylalkyl or heteroarylalky, for example a monocyclic or bicyclic arylalkyl or monocyclic or bicyclic heteroary alkyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy.

In some embodiments, R¹⁰ includes an unsaturated or partially unsaturated cyclic moiety, for example a cyclyl or heterocyclyl moiety. The cyclic moiety can either be directly attached to Y or attached via a linker such as an alkylenyl linker. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy.

In some embodiments, R¹⁰ is R¹⁵. In some embodiments, Y is substituted with a second R¹⁰, for example an alkyl, halo or alkoxy.

In some embodiments, R¹ is aryl, heteroaryl, arylalkyl, or heteroarylalkyl; k' is a bond or O; n is 1 or 2; R² and R³ are both hydrogen; A is

x and y are each independently 0-6;

R⁴ and R⁵ are each independently hydrogen or alkyl; Y is a monocyclic aryl or monocyclic heteroaryl; each of which is optionally substituted with 1-4 R¹⁰; each R¹⁰ is independently alkyl, alkenyl, alkynyl, halo, cyano, carbonyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, cyclyl, cyclylalkyl, alkoxy, alkoxyalkyl, aryloxy, aryloxyalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, heteroarylalkyl, -OR¹¹, - NR¹¹R^11', -CF₃, -SOR¹², -SO₂R¹², -OC(O)R¹¹, -SO₂NR¹²R^12', -(CH₂)_mR¹⁴ or R¹⁵; each of which is optionally independently substituted with 1-3 R¹⁶;

R¹¹ and R¹¹ are each independently hydrogen, alkyl, alkenyl, alkynyl, cyclyl, heterocyclyl, aryl or heteroaryl;

R¹² and R¹² are each independently hydrogen, alkyl, alkenyl, alkynyl, alkylthioalkyl, alkoxyalkyl, aryl, arylalkyl, heterocyclyl, heteroaryl, heteroarylalkyl, heterocycloalkyl or cyclyl, cyclylalkyl, or R¹² and R¹² taken together can be cyclized to form -(CH₂)_qX(CH₂)_s-; wherein each R¹² and R¹² may independently optionally be substituted with 1 to 3 substituents selected from the group consisting of halogen, OR¹¹, alkoxy, heterocycloalkyl, -NR¹¹C(O)NR¹¹R^11', -C(O)NR¹¹R^11', -NR¹¹C(O)R^11', -CN, oxo, -NR¹¹SO₂R^11', -OC(O)R¹¹, -SO₂NR¹¹R^11', -SOR¹³, -S(O)₂R¹³, -COOH and -C(O)OR¹³; each R¹³ is independently alkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl, each of which may optionally be substituted with -(CH₂)_w0H; each R¹⁴ is independently alkoxy, alkoxycarbonyl, -C(O)NR¹²R^12', -NR¹¹R^11', - C(O)R¹², -NR¹¹C(O)NR¹¹R^11' or -N-heteroaryl; each R¹⁵ is independently -(CH₂)_PN(R¹²)C(O)R^12', -(CH₂)_PCN, -

(CH₂)_pN(R¹²)C(O)OR^12', -(CH₂)_pN(R¹²)C(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂R¹², - (CH₂)_PSO₂NR¹²R^12', -(CH₂)_PC(O)NR¹²R^12', -(CH₂)_PC(O)OR¹², -(CH²)_POC(O)OR¹², - (CH₂)_POC(O)R¹², -(CH₂)_POC(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂NR¹²R^12', -(CH₂)_POR¹², - (CH₂)_pOC(O)N(R¹²)(CH₂)_mOH, -(CH₂)_PSOR¹², -(CH₂)_PSO₂R¹², -(CH₂)_PNR¹¹R¹¹ or - (CH₂)_pOCH₂C(O)N(R¹²)(CH₂)_mOH; each R¹⁶ is independently halo, alkyl, alkenyl, alkynyl, alkoxy, - (CH₂)_PNR¹¹C(O)NR¹¹R^11', -(CH₂)_PC(O)NR¹¹R^11', -(CH₂)_PNR¹¹C(O)R^11', -CN, - (CH₂)_PNR¹¹SO₂R^11', -(CH₂)_POC(O)R¹ \ -(CH₂)_PSO₂NR¹¹R^11', -(CH₂)_PSOR¹³, - (CH₂)_pC00H or -(CH₂)_PC(O)OR¹³;

X is CR¹¹R^11', O, S, S(O), S(O)₂, or NR¹¹; m is an integer between 1 and 6; p is an integer from O to 5; q and s are each independently an integer between 1 and 3; and w is an integer between 0 and 5.

For example, in some embodiments, n is 1; k' is a bond or O; and R¹ is aryl, heteroaryl, arylalkyl, or heteroarylalkyl. In some embodiments, n is 1; k' is O; and R¹ is arylalkyl, for example phenylmethyl. In some embodiments, n is 2; k'is a bond; and R¹ is aryl.

For example, in some embodiments, R^7a and R^7b are H; x is 1; and y is O or 1. In some embodiments, A is CH₂CH₂ or CH₂CH₂CH₂.

In some embodiments, each R⁴ and R⁵ is independently alkyl, for example, methyl or ethyl, preferably ethyl. In some embodiments, Y is a monocyclic heteroaromatic moiety, for example a nitrogen containing heteraromatic moiety such as a nitrogen containing five membered heteraromatic moiety.

In some embodiments, Y is a heterocyclic moiety containing at least two heteroatoms, for example, a five membered heterocyclic moiety containing at least two heteroatoms or at least three heteroatoms.

In some embodiments Y is substituted with one R¹⁰. R¹⁰ can be positioned, for example, 1,3 relative to the point of attachment of Y to the adjacent chain carbon or 1,2 relative to the point of attachment of Y to the adjacent chain carbon.

In some embodiments Y is oxadiazole or triazole.

In some embodiments, Y is

, wherein Ql is O or NR², preferably O or NH. In some embodiments, R¹⁰ is aryl, arylalkyl, heteroaryl, or heteroarylalkyl, for example optionally substituted with one or more R¹⁶. In some embodiments, R¹⁰ is substituted with one R¹⁶, such as halo (e.g., fluoro or chloro) or alkoxy. In some embodiments, the compound has a formula (Ia)

formula (Ia)

In some embodiments, R¹ is aryl, heteroaryl, arylalkyl, or heteroarylalkyl; k' is a bond or O; n is 1 or 2; A is CH₂, CH₂CH₂, or CH₂CH₂CH₂; R⁴ and R⁵ are each independently hydrogen or alkyl;

Y is a monocyclic aryl or monocyclic heteroaryl; each of which is optionally substituted with 1-4 R¹⁰; each R¹⁰ is independently alkyl, alkenyl, alkynyl, halo, cyano, carbonyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, cyclyl, cyclylalkyl, alkoxy, alkoxyalkyl, aryloxy, aryloxyalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, heteroarylalkyl, -OR¹¹, - NR¹¹R^11', -CF₃, -SOR¹², -SO₂R¹², -OC(O)R¹¹, -SO₂NR¹²R^12', -(CH₂)_mR¹⁴ or R¹⁵; each of which is optionally independently substituted with 1-3 R¹⁶;

R¹² and R¹² are each independently hydrogen, alkyl, alkenyl, alkynyl, alkylthioalkyl, alkoxyalkyl, aryl, arylalkyl, heterocyclyl, heteroaryl, heteroarylalkyl, heterocycloalkyl or cyclyl, cyclylalkyl, or R¹² and R¹² taken together can be cyclized to form -(CH₂)_qX(CH₂)_s-; wherein each R¹² and R¹² may independently optionally be substituted with 1 to 3 substituents selected from the group consisting of halogen, OR¹¹, alkoxy, heterocycloalkyl, -NR¹¹C(O)NR¹¹R^11', -C(O)NR¹¹R^11', -NR¹¹C(O)R^11', -CN, oxo, -NR¹¹SO₂R^11', -OC(O)R¹¹, -SO₂NR¹¹R^11', -SOR¹³, -S(O)₂R¹³, -COOH and -C(O)OR¹³; each R¹³ is independently alkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl, each of which may optionally be substituted with -(CH₂)_w0H; each R¹⁴ is independently alkoxy, alkoxycarbonyl, -C(O)NR¹²R^12', -NR¹¹R^11', -

C(O)R¹², -NR¹¹C(O)NR¹¹R^11' or -N-heteroaryl; each R¹⁵ is independently -(CH₂)_PN(R¹²)C(O)R^12', -(CH₂)_PCN, - (CH₂)pN(R¹²)C(O)OR^12', -(CH₂)pN(R¹²)C(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂R¹², -

(CH₂)_PSO₂NR¹²R^12', -(CH₂)_PC(O)NR¹²R^12', -(CH₂)_PC(O)OR¹², -(CH²)_POC(O)OR¹², (CH₂)_POC(O)R¹², -(CH₂)_POC(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂NR¹²R^12', -(CH₂)_POR¹², -

(CH₂)_pOC(O)N(R¹²)(CH₂)_mOH, -(CH₂)_PSOR¹², -(CH₂)_PSO₂R¹², -(CH₂)_pNRⁿRⁿ or -

(CH₂)_pOCH₂C(O)N(R¹²)(CH₂)_mOH; each R¹⁶ is independently halo, alkyl, alkenyl, alkynyl, alkoxy, -

(CH₂)_PNR¹¹C(O)NR¹¹R^11', -(CH₂)_PC(O)NR¹¹R^11', -(CH₂)_PNR¹¹C(O)R^11', -CN, - (CH₂)_PNR¹¹SO₂R^11', -(CH₂)_POC(O)R¹ \ -(CH₂)_PSO₂NR¹¹R^11', -(CH₂)_PSOR¹³, -

(CH₂)_pC00H or -(CH₂)_PC(O)OR¹³; X is CR¹¹R^11', O, S, S(O), S(O)₂, or NR¹¹; m is an integer between 1 and 6; p is an integer from 0 to 5; q and s are each independently an integer between 1 and 3; and w is an integer between 0 and 5.

For example, in some embodiments, n is 1; k' is a bond or O; and R¹ is aryl, heteroaryl, arylalkyl, or heteroarylalkyl. In some embodiments, n is 1; k' is O; and R¹ is arylalkyl, for example phenylmethyl. In some embodiments, n is 2; k'is a bond; and R¹ is aryl. In some embodiments, A is CH₂CH₂ or CH₂CH₂CH₂, preferably CH₂CH₂CH₂.

In some embodiments, each R⁴ and R⁵ is independently alkyl, for example, methyl or ethyl, preferably ethyl.

In some embodiments Y is oxadiazole or triazole.

In some embodiments, Y is

, wherein Ql is O or NR², preferably O or NH. In some embodiments, R¹⁰ is aryl, arylalkyl, heteroaryl, or heteroarylalkyl, for example optionally substituted with one or more R¹⁶. In some embodiments, R¹⁰ is substituted with one R¹⁶, such as halo (e.g., fluoro or chloro) or alkoxy. In some embodiments,

R¹ is hydrogen or alkyl, for example unsubstituted or substituted with one R⁶; n is 0 or 1 ; k' is a bond; and

R² and R³ each independently hydrogen or Ci-C₆ alkyl;

A is

x and y are each independently 0-6;

R⁴ and R⁵ are each independently hydrogen or alkyl;

R¹¹ and R¹¹ are each independently hydrogen, alkyl, alkenyl, alkynyl, cyclyl, heterocyclyl, aryl or heteroaryl; R¹² and R¹² are each independently hydrogen, alkyl, alkenyl, alkynyl, alkylthioalkyl, alkoxyalkyl, aryl, arylalkyl, heterocyclyl, heteroaryl, heteroarylalkyl, heterocycloalkyl or cyclyl, cyclylalkyl, or R¹² and R¹² taken together can be cyclized to form -(CH₂)_qX(CH₂)_s-; wherein each R¹² and R¹² may independently optionally be substituted with 1 to 3 substituents selected from the group consisting of halogen, OR¹¹, alkoxy, heterocycloalkyl, -NR¹¹C(O)NR¹¹R^11', -C(O)NR¹¹R^11', -NR¹¹C(O)R^11', -CN, oxo, -NR¹¹SO₂R^11', -OC(O)R¹¹, -SO₂NR¹¹R^11', -SOR¹³, -S(O)₂R¹³, -COOH and -C(O)OR¹³; each R¹³ is independently alkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl, each of which may optionally be substituted with -(CH₂)_w0H; each R¹⁴ is independently alkoxy, alkoxycarbonyl, -C(O)NR¹²R^12', -NR¹¹R^11', - C(O)R¹², -NR¹¹C(O)NR¹¹R^11' or -N-heteroaryl; each R¹⁵ is independently -(CH₂)_PN(R¹²)C(O)R^12', -(CH₂)_PCN, - (CH₂)_PN(R¹²)C(O)OR^12', -(CH₂)_PN(R¹²)C(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂R¹², - (CH₂)_PSO₂NR¹²R^12', -(CH₂)_PC(O)NR¹²R^12', -(CH₂)_PC(O)OR¹², -(CH²)_POC(O)OR¹², - (CH₂)_POC(O)R¹², -(CH₂)_POC(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂NR¹²R^12', -(CH₂)_POR¹², - (CH₂)_pOC(O)N(R¹²)(CH₂)_mOH, -(CH₂)_PSOR¹², -(CH₂)_PSO₂R¹², -(CH₂)_pNRⁿRⁿ or - (CH₂)_pOCH₂C(O)N(R¹²)(CH₂)_mOH; each R¹⁶ is independently halo, alkyl, alkenyl, alkynyl, alkoxy, - (CH₂)_PNR¹¹C(O)NR¹¹R^11', -(CH₂)_PC(O)NR¹¹R^11', -(CH₂)_PNR¹¹C(O)R^11', -CN, -

(CH₂)_PNR¹¹SO₂R^11', -(CH₂)_POC(O)R¹ \ -(CH₂)_PSO₂NR¹¹R^11', -(CH₂)_PSOR¹³, (CH₂)_pC00H or -(CH₂)_PC(O)OR¹³;

In some embodiments, n is 0 or 1 ; k' is a bond; and

R¹ is alkyl, for example unsubstituted or substituted with one R⁶.

In some embodiments n is 0 and k' is a bond. Exemplary R¹ moieties include methyl, and ethyl. Preferred R¹ moieties include methyl. In some embodiments R¹ is unsubstituted methyl or methyl or ethyl substituted with C(O)N(R )₂.

In some embodiments, n is 0 or 1 ; k' is a bond; and R¹ is alkyl, for example unsubstituted or substituted with one R⁶. For example, R¹ can be a branched alkyl such as one of the following .

In some embodiments n is 0 and k' is a bond, and R¹ and R³ are both methyl.

In some embodiments, n is O; k' is a bond; and

R¹ is hydrogen.

In some embodiments, A is CH₂CH₂ or CH₂CH₂CH₂, preferably CH₂CH₂CH₂.

In some embodiments Y is substituted with one R¹⁰. R¹⁰ can be positioned, for example, 1,3 relative to the point of attachment of Y to the adjacent chain carbon or 1,2 relative to the point of attachment of Y to the adjacent chain carbon. In some embodiments, R¹⁰ is aryl or heteroaryl, for example a monocyclic aryl or monocyclic heteroaryl such as phenyl, pyridyl, oxazolyl, thiazolyl, or thiophenyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy. In some embodiments, R¹⁰ is a bicyclic heteroaryl, for example indolyl, benzimidazolyl, benzoxazolyl, benzofuranyl, benzothiophenyl, or benzthiazolyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy.

In some embodiments Y is oxadiazole or triazole.

□ 10

^K V_^N

T Vi-

In some embodiments, Y is ^"Q , wherein Ql is O or NR , preferably O or NH. In some embodiments, R¹⁰ is aryl, arylalkyl, heteroaryl, or heteroarylalkyl, for example optionally substituted with one or more R¹⁶. In some embodiments, R¹⁰ is substituted with one R¹⁶, such as halo (e.g., fluoro or chloro) or alkoxy.

In some embodiments, the compounds has a formula (Ib)

formula (Ib)

In some embodiments, R¹ is hydrogen or alkyl; A is CH₂, CH₂CH₂, or CH₂CH₂CH₂; R² is hydrogen or C₁-C₃ alkyl;

R⁴ and R⁵ are each independently hydrogen or alkyl;

Y is a monocyclic aryl or monocyclic heteroaryl; each of which is optionally substituted with 1-4 R¹⁰; each R¹⁰ is independently alkyl, alkenyl, alkynyl, halo, cyano, carbonyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, cyclyl, cyclylalkyl, alkoxy, alkoxyalkyl, aryloxy, aryloxyalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, heteroarylalkyl, -OR¹¹, - NR¹¹R^11', -CF₃, -SOR¹², -SO₂R¹², -OC(O)R¹¹, -SO₂NR¹²R^12', -(CH₂)_mR¹⁴ or R¹⁵; each of which is optionally independently substituted with 1-3 R¹⁶; R¹¹ and R¹¹ are each independently hydrogen, alkyl, alkenyl, alkynyl, cyclyl, heterocyclyl, aryl or heteroaryl;

(CH₂)_pSO₂NR¹²R^12', -(CH₂)_PC(O)NR¹²R^12', -(CH₂)_PC(O)OR¹², -(CH²)_POC(O)OR¹², - (CH₂)_pOC(O)R¹², -(CH₂)_POC(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂NR¹²R^12', -(CH₂)_POR¹², - (CH₂)_pOC(O)N(R¹²)(CH₂)_mOH, -(CH₂)_PSOR¹², -(CH₂)_PSO₂R¹², -(CH₂)_PNR¹¹R¹¹ or -

(CH₂)_pOCH₂C(O)N(R¹²)(CH₂)_mOH; each R¹⁶ is independently halo, alkyl, alkenyl, alkynyl, alkoxy, - (CH₂)_PNR¹¹C(O)NR¹¹R^11', -(CH₂)_PC(O)NR¹¹R^11', -(CH₂)_PNR¹¹C(O)R^11', -CN, - (CH₂)_PNR¹¹SO₂R^11', -(CH₂)_POC(O)R¹ \ -(CH₂)_PSO₂NR¹¹R^11', -(CH₂)_PSOR¹³, - (CH₂)_PCOOH or -(CH₂)_PC(O)OR¹³;

In some embodiments Y is substituted with one R¹⁰. R¹⁰ can be positioned, for example, 1,3 relative to the point of attachment of Y to the adjacent chain carbon or 1,2 relative to the point of attachment of Y to the adjacent chain carbon. In some embodiments, R¹⁰ is aryl or heteroaryl, for example a monocyclic aryl or monocyclic heteroaryl such as phenyl, pyridyl, oxazolyl, thiazolyl, or thiophenyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy.

In some embodiments, R¹⁰ is arylalkyl or heteroarylalky, for example a monocyclic or bicyclic arylalkyl or monocyclic or bicyclic heteroary alkyl. In some eemmbbooddiimmeennttss,, RR¹¹⁰⁰ iiss ssuubbssttiittuutteedd wwiitthh 11--33 RR¹¹⁶⁶.. IInn ssoommee eemmbbooddiimmeernts, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy. In some embodiments, R¹⁰ includes an unsaturated or partially unsaturated cyclic moiety, for example a cyclyl or heterocyclyl moiety. The cyclic moiety can either be directly attached to Y or attached via a linker such as an alkylenyl linker. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy.

In some embodiments Y is oxadiazole or triazole.

In some embodiments, Y is

, wherein Ql is O or NR², preferably O or NH. In some embodiments, R¹⁰ is aryl, arylalkyl, heteroaryl, or heteroarylalkyl, for example optionally substituted with one or more R¹⁶. In some embodiments, R¹⁰ is substituted with one R¹⁶, such as halo (e.g., fluoro or chloro) or alkoxy.

In some embodiments, R¹ and R³ together form a heterocyclic ring such as a pyrrolidine or an azetidine ring (The heterocyclic ring can be unsubstituted or substituted, for example, with 1-2 R⁶.); n is 0 or 1 ; k' is a bond;

R² hydrogen or Ci-C₆ alkyl, preferably hydrogen; A is

x and y are each independently 0-6; R⁴ and R⁵ are each independently hydrogen or alkyl;

Y is a monocyclic aryl or monocyclic heteroaryl; each of which is optionally substituted with 1-4 R¹⁰; each R¹⁰ is independently alkyl, alkenyl, alkynyl, halo, cyano, carbonyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, cyclyl, cyclylalkyl, alkoxy, alkoxyalkyl, aryloxy, aryloxyalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, heteroarylalkyl, -OR¹¹, -

NR¹¹R^11', -CF₃, -SOR¹², -SO₂R¹², -OC(O)R¹¹, -SO₂NR¹²R^12', -(CH₂)_mR¹⁴ or R¹⁵; each of which is optionally independently substituted with 1-3 R¹⁶; R¹¹ and R¹¹ are each independently hydrogen, alkyl, alkenyl, alkynyl, cyclyl, heterocyclyl, aryl or heteroaryl;

R¹² and R¹² are each independently hydrogen, alkyl, alkenyl, alkynyl, alkylthioalkyl, alkoxyalkyl, aryl, arylalkyl, heterocyclyl, heteroaryl, heteroarylalkyl, heterocycloalkyl or cyclyl, cyclylalkyl, or R¹² and R¹² taken together can be cyclized to form -(CH₂)_qX(CH₂)_s-; wherein each R¹² and R¹² may independently optionally be substituted with 1 to 3 substituents selected from the group consisting of halogen, OR¹¹, alkoxy, heterocycloalkyl, -NR¹¹C(O)NR¹¹R^11', -C(O)NR¹¹R^11', -NR¹¹C(O)R^11', -CN, oxo, -NR¹¹SO₂R^11', -OC(O)R¹¹, -SO₂NR¹¹R^11', -SOR¹³, -S(O)₂R¹³, -COOH and -C(O)OR¹³; each R¹³ is independently alkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl, each of which may optionally be substituted with -(CH₂)_w0H; each R¹⁴ is independently alkoxy, alkoxycarbonyl, -C(O)NR¹²R^12', -NR¹¹R^11', - C(O)R¹², -NR¹¹C(O)NR¹¹R^11' or -N-heteroaryl; each R¹⁵ is independently -(CH₂)_PN(R¹²)C(O)R^12', -(CH₂)_PCN, - (CH₂)pN(R¹²)C(O)OR^12', -(CH₂)pN(R¹²)C(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂R¹², -

(CH₂)_PSO₂NR¹²R^12', -(CH₂)_PC(O)NR¹²R^12', -(CH₂)_PC(O)OR¹², -(CH²)_POC(O)OR¹²,

(CH₂)_POC(O)R¹², -(CH₂)_POC(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂NR¹²R^12', -(CH₂)_POR¹², - (CH₂)_pOC(O)N(R¹²)(CH₂)_mOH, -(CH₂)_PSOR¹², -(CH₂)_PSO₂R¹², -(CH₂)_pNRⁿRⁿ or - (CH₂)_pOCH₂C(O)N(R¹²)(CH₂)_mOH; each R¹⁶ is independently halo, alkyl, alkenyl, alkynyl, alkoxy, -

(CH₂)_PNR¹¹C(O)NR¹¹R^11', -(CH₂)_PC(O)NR¹¹R^11', -(CH₂)_PNR¹¹C(O)R^11', -CN, - (CH₂)_PNR¹¹SO₂R^11', -(CH₂)_POC(O)R¹ \ -(CH₂)_PSO₂NR¹¹R^11', -(CH₂)_PSOR¹³, - (CH₂)_pC00H or -(CH₂)_PC(O)OR¹³;

In some embodiments, A is CH₂CH₂ or CH₂CH₂CH₂, preferably CH₂CH₂CH₂. In some embodiments, each R⁴ and R⁵ is independently alkyl, for example, methyl or ethyl, preferably ethyl. In some embodiments, Y is a monocyclic heteroaromatic moiety, for example a nitrogen containing heteraromatic moiety such as a nitrogen containing five membered heteraromatic moiety.

In some embodiments Y is oxadiazole or triazole.

In some embodiments, Y is

, wherein Ql is O or NR², preferably O or

NH. In some embodiments, R¹⁰ is aryl, arylalkyl, heteroaryl, or heteroarylalkyl, for example optionally substituted with one or more R¹⁶. In some embodiments, R¹⁰ is substituted with one R¹⁶, such as halo (e.g., fluoro or chloro) or alkoxy. In some embodiments, the compounds has a formula (Ic)

formula (Ic) n is 0, 1, 2, 3, or 4; preferably 1 or 2; A is CH₂, CH₂CH₂, or CH₂CH₂CH₂;

R⁴ and R⁵ are each independently hydrogen or alkyl;

C(O)R¹², -NR¹¹C(O)NR¹¹R^11' or -N-heteroaryl; each R¹⁵ is independently -(CH₂)_PN(R¹²)C(O)R^12', -(CH₂)_PCN, - (CH₂)_PN(R¹²)C(O)OR^12', -(CH₂)_PN(R¹²)C(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂R¹², - (CH₂)_PSO₂NR¹²R^12', -(CH₂)_PC(O)NR¹²R^12', -(CH₂)_PC(O)OR¹², -(CH²)_POC(O)OR¹², - (CH₂) _pP0OCC((0O))RR¹¹²²,, --((CCHH₂₂))_PPOOCC((OO))NNRR¹¹²²RR^1122'',, --((CCHH₂₂))_pPrN(R¹²)SO₂NR¹²R^12', -(CH₂)_POR¹² (CH₂)pOC(O)N(R¹²)(CH₂)_mOH, -(CH₂)_PSOR¹², -(CH₂)_PSO₂R¹², -(CH₂)_PNR¹¹R¹¹ or - (CH₂)_pOCH₂C(O)N(R¹²)(CH₂)_mOH; each R¹⁶ is independently halo, alkyl, alkenyl, alkynyl, alkoxy, - (CH₂)_PNR¹¹C(O)NR¹¹R^11', -(CH₂)_PC(O)NR¹¹R^11', -(CH₂)_PNR¹¹C(O)R^11', -CN, - (CH₂)_PNR¹¹SO₂R^11', -(CH₂)_POC(O)R¹ \ -(CH₂)_PSO₂NR¹¹R^11', -(CH₂)_PSOR¹³, - (CH₂)_pC00H or -(CH₂)_PC(O)OR¹³;

In some embodiments, R¹⁰ is a bicyclic heteroaryl, for example indolyl, benzimidazolyl, benzoxazolyl, benzofuranyl, benzothiophenyl, or benzthiazolyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy. In some embodiments, R¹⁰ is arylalkyl or heteroarylalky, for example a monocyclic or bicyclic arylalkyl or monocyclic or bicyclic heteroaryalkyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy. In some embodiments, R¹⁰ includes an unsaturated or partially unsaturated cyclic moiety, for example a cyclyl or heterocyclyl moiety. The cyclic moiety can either be directly attached to Y or attached via a linker such as an alkylenyl linker. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, cyano, or methoxy. In some embodiments Y is oxadiazole or triazole.

□ 10

^K V_^N

T Vi-

In some embodiments, Y is ^"Q , wherein Ql is O or NR , preferably O or NH. In some embodiments, R¹⁰ is aryl, arylalkyl, heteroaryl, or heteroarylalkyl, for example optionally substituted with one or more R¹⁶. In some embodiments, R¹⁰ is substituted with one R¹⁶, such as halo (e.g., fluoro or chloro) or alkoxy. In another aspect, the methods described herein use a compound of formula (IV)

formula (IV) wherein,

R¹ is hydrogen, aryl, heteroaryl, arylalkyl, heteroarylalkyl, cyclyl, cyclylalkyl, heterocyclyl, heterocyclylalkyl, alkyl, alkenyl, alkynyl, or R¹ can be taken together with

R² or R³ to form a ring; each of which is optionally substituted with 1-4 R⁶; k' is a bond, O, C(O), C(O)O, OC(O), C(O)NR³, NR³C(O), S, SO, SO₂, CR²=CR², or C≡C; n is 0-6, preferably 1-3;

R is hydrogen, Ci-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl;

A' is heterocyclyl; optionally substituted with 1-3 R⁹; Y is a monocyclic aryl or monocyclic heteroaryl; each of which is optionally substituted with 1-4 R¹⁰; each R⁶ is independently halo, alkyl, alkenyl, alkynyl, cyclyl, heterocyclyl, aryl, heteroaryl, alkoxy, haloalkyl, haloalkyloxy, haloalkylthio, acetyl, cyano, nitro, hydroxy, oxo, C(O)OR², OC(O)R², N(R³)₂, C(O)N(R³)₂, NR³C(O)R², or SR²;

R⁹ is halo, alkyl, cyclyl, heterocyclyl, aryl, heteroaryl, alkoxy, haloalkyl, haloalkyloxy, haloalkylthio, acetyl, cyano, nitro, hydroxy, oxo, C(O)OR², OC(O)R², N(R²)₂, C(O)N(R²)₂, NR²C(O)R², SR²; each R¹⁰ is independently alkyl, alkenyl, alkynyl, halo, cyano, carbonyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, cyclyl, cyclylalkyl, alkoxy, alkoxyalkyl, aryloxy, aryloxyalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, heteroarylalkyl, -OR¹¹, - NR¹¹R^11', -CF₃, -SO₂R¹², -OC(O)R¹¹, -SO₂NR¹²R^12', -(CH₂)_mR¹⁴ or R¹⁵; each of which is optionally independently substituted with 1-3 R¹⁶;

C(O)R¹², -NR¹¹C(O)NR¹¹R^11' or -N-heteroaryl; each R¹⁵ is independently heterocycloalkyl, heteroaryl, -CN, - (CH₂)_PN(R¹²)C(O)R^12', -(CH₂)_PCN, -(CH₂)_PN(R¹²)C(O)OR^12', - (CH₂)_pN(R¹²)C(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂R¹², -(CH₂)_PSO₂NR¹²R^12', - (CH₂)_PC(O)NR¹²R^12', -(CH₂)_PC(O)OR¹², -(CH²)_POC(O)OR¹², -(CH₂)_POC(O)R¹², - (CH₂)_POC(O)NR¹²R^12', -(CH₂)_pN(R¹²)SO₂NR¹²R¹²¹, -(CH₂)_POR¹², - (CH₂)_pOC(O)N(R¹²)(CH₂)_mOH, -(CH₂)_PSOR¹² or -(CH₂)_pOCH₂C(O)N(R¹²)(CH₂)_mOH; each R¹⁶ is independently halo, alkyl, alkenyl, alkynyl, alkoxy, - (CH₂)_PNR¹¹C(O)NR¹¹R^11', -(CH₂)_PC(O)NR¹¹R^11', -(CH₂)_PNR¹¹C(O)R^11', -CN, - (CH₂)_PNR¹¹SO₂R^11', -(CH₂)_POC(O)R¹ \ -(CH₂)_PSO₂NR¹¹R^11', -(CH₂)_PSOR¹³, - (CH₂)_pC00H or -(CH₂)_PC(O)OR¹³;

X is CR¹¹R^11', O, S, S(O), S(O)₂, or NR¹¹; m is an integer between 1 and 6; p is an integer from O to 5. q and s are each independently an integer between 1 and 3; and w is an integer between 0 and 5.

In some embodiments, the compound of formula (IV), comprises an enriched preparation of formula (IV)

formula (IV).

In some embodiments, the compound of formula (IV), comprises an enriched preparation of formula (IV")

R¹

formula (IV"). In some embodiments, A' is a 5 or 6 membered heterocyclyl. In some embodiments, the 5 or 6 membered heterocyclyl includes at least two nitrogen atoms.

In some embodiments, A' is

In some embodiments, A' is substituted with one R⁹, for example, N(R²)₂.

In some embodiments, n is 1; k' is a bond or O; and R¹ is aryl, heteroaryl, arylalkyl, or heteroarylalkyl.

In some embodiments, n is 2; k' is a bond; and R¹ is aryl. In some embodiments, Y is a monocyclic heteroaromatic moiety, for example, a nitrogen containing heteraromatic moiety, such as a nitrogen containing 5 membered heteraromatic moiety.

In some embodiments, Y is a heterocyclic moiety containing at least two heteroatoms, for example, a 5 membered heterocyclic moiety containing at least two heteroatoms or a heterocyclic moiety containing at least 3 heteroatoms.

In some embodiments, Y is substituted with 1 R¹⁰. The R¹⁰ can be positioned, for example, 1,3 relative to the point of attachment of Y to the adjacent chain carbon or can be positioned, for example, 1,2 relative to the point of attachment of Y to the adjacent chain carbon. In some embodiments, R¹⁰ is aryl or heteroaryl, for example a monocyclic aryl or monocyclic heteroaryl such as phenyl, pyridyl, or thiophenyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, or methoxy. In some embodiments, R¹⁰ is a bicyclic heteroaryl, for example indolyl, imidazolyl, benzoxazolyl, or benzthiazolyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, or methoxy.

In some embodiments, Y is oxadiazole or triazole.

In another aspect, the methods described herein use a compound of formula (V),

formula (V) wherein,

Q\ Q² _J Q^{3 an}d Q⁴ together with the carbon to which they are attached form a heteroaryl moiety, and each Q¹, Q², Q³ and Q⁴ is independently S, O, N, CR², CR¹⁰, NR², or NR¹⁰.

In some embodiments, the compound of formula (V), comprises an enriched preparation of formula (V)

formula (V).

In some embodiments, the compound of formula (V), comprises an enriched preparation of formula (V")

formula (V").

In some embodiments, Q² is CR² or CR¹⁰. In some embodiments, Q² is S, O, N, or NR¹⁰. In some embodiments, at least one of Q² or Q³ is CR² or CR¹⁰. In some embodiments, at least two of Q¹, Q², Q³, or Q⁴ is S, O, N, or NR IO In some embodiments, Q¹, Q², and Q³ are each independently S, O, N, or NR IO

In some embodiments, Q¹ is NR¹⁰.

In some embodiments, one of Q², Q³, or Q⁴ is CR²

In some embodiments, Q² is CR¹⁰.

In some embodiments, Q³ is CR²

In some embodiments, Q¹, Q², Q³ and Q⁴ together form

In some embodiments, Q¹ is NR².

In some embodiments, Q¹, Q², Q³ and Q⁴ together form

^■

In some embodiments, Q¹ is NR¹⁰. In another aspect, the methods described herein use a compound of formula (VI),

formula (VI) wherein

Z¹, Z², Z³, Z⁴, and Z⁵ together form an aryl or heteroaryl moiety, and each Z¹, Z², Z³, Z⁴, and Z⁵ is independently N, CR¹⁰, or CR². In some embodiments, the compound of formula (VI), comprises an enriched preparation of a compound of formula (VF).

R¹

formula (VI').

In some embodiments, the compound of formula (VI), comprises an enriched preparation of a compound of formula (VI").

R¹

formula (VI").

In some embodiments, one of Z¹, Z², Z³, Z⁴, and Z⁵ is N. In some embodiments, two of Z¹, Z², Z³, Z⁴, and Z⁵ are N.

In some embodiments, three of Z¹, Z², Z³, Z⁴, and Z⁵ is N. In some embodiments, two of Z¹ and Z² are N. In some embodiments, two of Z¹ and Z³ are N. In some embodiments, two of Z¹ and Z⁴ are N.

In some embodiments, two of Z¹, Z³, and Z⁵ are N.

In some embodiments, the compound is a compound of formula (IV), wherein Y is substituted with a single substituent R¹⁰. For example, R¹⁰ can be aryl or heteroaryl, optionally substituted with up to three independent R¹⁶.

In some embodiments, R¹⁰ is aryl or heteroaryl, for example a monocyclic aryl or monocyclic heteroaryl such as phenyl, pyridyl, or thiophenyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, or methoxy. In some embodiments, R¹⁰ is a bicyclic heteroaryl, for example indolyl, imidazolyl, benzoxazolyl, or benzthiazolyl. In some embodiments, R¹⁰ is substituted with 1-3 R¹⁶. In some embodiments, R¹⁶ is halo, alkyl, or alkoxy, for example chloro, fluoro, methyl, or methoxy.

The compounds of formulae (I), (II), (III), (IV), (V), and (VI) are also described in US Patent Apps. 11/451,086 and 11/564,815, and PCT Pub. Nos. WO 2006/135860 and WO 2007/146914. In another aspect, the methods described herein use a pharmaceutically acceptable salt comprising a compound of any of the formulae described herein.

In another aspect, a composition comprising a compound of any of the formulae described herein and a pharmaceutically acceptable carrier is used in a method of the invention. In one aspect, a compound that has a structure of a formula described herein, and that competes with ghrelin for binding to GHS-R is used in a method of the invention.

In another aspect, a compound listed in Table 1 is used in a method of the invention.

In one embodiment, the compound is an enantiomerically enriched isomer of a stereoisomer described herein. For example, the compound has an enantiomeric excess of at least about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%. Enantiomer, when used herein, refers to either of a pair of chemical compounds whose molecular structures have a mirror-image relationship to each other.

In one embodiment, a preparation of a compound disclosed herein is enriched for an isomer of the compound having a selected stereochemistry, e.g., R or S, corresponding to a selected stereocenter, e.g., the position corresponding to the carbon alpha to the sulfonamide nitrogen in formula (I). Exemplary R/S configurations can be those provided in an example described herein, e.g, those described in the Table below, or the configuration of the majority or minority species in a synthetic scheme described herein. For example, the compound has a purity corresponding to a compound having a selected stereochemistry of a selected stereocenter of at least about 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%.

In one embodiment, a compound described herein includes a preparation of a compound disclosed herein that is enriched for a structure or structures having a selected stereochemistry, e.g., R or S, at a selected stereocenter, e.g., the carbon alpha to the sulfonamide nitrogen of a formula described herein e.g., formula (I), (II), (III), (IV), (V), or (VI).

Exemplary R/S configurations can be those provided in an example described herein, e.g, those described in the Table below, or the configuration of the majority or minority species in a synthetic scheme described herein. For example, the compound has a purity corresponding to a compound having a selected stereochemistry of a selected stereocenter of at least about 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%.

An "enriched preparation," as used herein, is enriched for a selected stereoconfiguration of one, two, three or more selected stereocenters within the subject compound. Exemplary selected stereocenters and exemplary stereoconfigurations thereof can be selected from those provided, herein, e.g., in an example described herein, e.g., those described in the Table below. By enriched is meant at least 60%, e.g., of the molecules of compound in the preparation have a selected stereochemistry of a selected stereocenter. In preferred embodiments it is at least 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%. Enriched refers to the level of a subject molecule(s) and does not connote a process limitation unless specified.

In one embodiment, a preparation of a compound disclosed herein, is enriched for isomers (subject isomers) which are diastereromers of a compound described herein. For example, a compound having a selected stereochemistry, e.g., R or S, corresponding to a selected stereocenter, e.g., the position corresponding to the carbon alpha to the sulfonamide nitrogen of a formula described herein e.g., formula (I), (II), (III), (IV), (V), or (VI). Exemplary R/S configurations can be those provided in an example described herein, e.g., those described in the table below, or the configuration of the majority or minority species in a synthetic scheme described herein. For example, the compound has a purity corresponding to a compound having a selected stereochemistry of a selected stereocenter of at least about 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%. Diastereromer, when used herein, refers to a stereoisomer of a compound having two or more chiral centers that is not a mirror image of another stereoisomer of the same compound.

In another aspect, the methods described herein use an organic compound that modulates (e.g., antagonizes, agonizes, or inversely agonizes) GHS-R activity, the compound having a molecular weight of less than 700 Daltons, and having fewer than four L- or D- amino acids (e.g., and any salt thereof) for use in a method of the invention. For example, the compound may, in certain embodiments, bind or otherwise include a metal cation.

In one embodiment, the compound has a molecular weight less than [D-Lys-3]- GHRP-6 or H(2)N-D-arg-Pro-Lys-Pro-d-Phe-Gln-d-Trp-Phe- d-Trp-Leu-Leu-NH(2) (L 756,867) or within 2, 1.5, 1.4, 1.2, 1.1, 0.8, 0.6, or 0.5 fold that of [D-Lys-3]-GHRP-6 or L 756,867.

In another aspect, a pharmaceutical composition that includes a compound described herein, e.g., a compound listed in Table 1 or described above, and a pharmaceutically acceptable carrier is used in a method of the invention.

The term "halo" refers to any radical of fluorine, chlorine, bromine or iodine. The term "alkyl" refers to a hydrocarbon chain that may be a straight chain or branched chain, containing the indicated number of carbon atoms. For example, Cl-ClO indicates that the group may have from 1 to 10 (inclusive) carbon atoms in it. The term "lower alkyl" refers to a C1-C8 alkyl chain. In the absence of any numerical designation, "alkyl" is a chain (straight or branched) having 1 to 10 (inclusive) carbon atoms in it. The term "alkoxy" refers to an -O-alkyl radical. The term "alkylene" refers to a divalent alkyl (i.e., -R-). The term "aminoalkyl" refers to an alkyl substituted with an amino. The term

"mercapto" refers to an -SH radical. The term "thioalkoxy" refers to an -S-alkyl radical. The term "alkenyl" refers to a hydrocarbon chain that may be a straight chain or branched chain having one or more carbon-carbon double bonds. The alkenyl moiety contains the indicated number of carbon atoms. For example, C2-C10 indicates that the group may have from 2 to 10 (inclusive) carbon atoms in it. The term "lower alkenyl" refers to a C2-C8 alkenyl chain. In the absence of any numerical designation, "alkenyl" is a chain (straight or branched) having 2 to 10 (inclusive) carbon atoms in it.

The term "alkynyl" refers to a hydrocarbon chain that may be a straight chain or branched chain having one or more carbon-carbon triple bonds. The alkynyl moiety contains the indicated number of carbon atoms. For example, C2-C10 indicates that the group may have from 2 to 10 (inclusive) carbon atoms in it. The term "lower alkynyl" refers to a C2-C8 alkynyl chain. In the absence of any numerical designation, "alkynyl" is a chain (straight or branched) having 2 to 10 (inclusive) carbon atoms in it.

The term "aryl" refers to a 6-carbon monocyclic, 10-carbon bicyclic, or 14-carbon tricyclic aromatic ring system wherein 0, 1, 2, 3, or 4 atoms of each ring may be substituted by a substituent. Examples of aryl groups include phenyl, naphthyl and the like. The term "arylalkyl" or the term "aralkyl" refers to alkyl substituted with an aryl. The term "arylalkenyl" refers to an alkenyl substituted with an aryl. The term "arylalkynyl" refers to an alkynyl substituted with an aryl. The term "arylalkoxy" refers to an alkoxy substituted with aryl.

The terms "cycloalkyl" or "cyclyl" as employed herein includes saturated and partially unsaturated cyclic hydrocarbon groups having 3 to 12 carbons, preferably 3 to 8 carbons, and more preferably 3 to 6 carbons, wherein the cycloalkyl group may be optionally substituted. Preferred cycloalkyl groups include, without limitation, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, and cyclooctyl. The term "heteroaryl" refers to an aromatic 5-8 membered monocyclic, 8-12 membered bicyclic, or 11-14 membered tricyclic ring system having 1-3 heteroatoms if monocyclic, 1-6 heteroatoms if bicyclic, or 1-9 heteroatoms if tricyclic, said heteroatoms selected from O, N, or S (e.g., carbon atoms and 1-3, 1-6, or 1-9 heteroatoms of N, O, or S if monocyclic, bicyclic, or tricyclic, respectively), wherein 0, 1, 2, 3, or 4 atoms of each ring may be substituted by a substituent. Examples of heteroaryl groups include pyridyl, furyl or furanyl, imidazolyl, benzimidazolyl, pyrimidinyl, thiophenyl or thienyl, quinolinyl, indolyl, thiazolyl, and the like. The term "heteroarylalkyl" or the term "heteroaralkyl" refers to an alkyl substituted with a heteroaryl. The term "heteroarylalkenyl" refers to an alkenyl substituted with a heteroaryl. The term "heteroarylalkynyl" refers to an alkynyl substituted with a heteroaryl. The term "heteroarylalkoxy" refers to an alkoxy substituted with heteroaryl.

The term "heterocyclyl" or "heterocyclylalkyl"refers to a nonaromatic 5-8 membered monocyclic, 5-12 membered bicyclic, or 11-14 membered tricyclic ring system having 1-3 heteroatoms if monocyclic, 1-6 heteroatoms if bicyclic, or 1-9 heteroatoms if tricyclic, said heteroatoms selected from O, N, or S (e.g., carbon atoms and 1-3, 1-6, or 1-9 heteroatoms of N, O, or S if monocyclic, bicyclic, or tricyclic, respectively), wherein 0, 1, 2 or 3 atoms of each ring may be substituted by a substituent. Examples of heterocyclyl groups include piperazinyl, pyrrolidinyl, dioxanyl, morpholinyl, tetrahydrofuranyl, and include both bridged and fused ring systems. The term "heterocyclylalkyl" refers to an alkyl substituted with a heterocyclyl.

The term "sulfonyl" refers to a sulfur attached to two oxygen atoms through double bonds. An "alkylsulfonyl" refers to an alkyl substituted with a sulfonyl. The term "amino acid" refers to a molecule containing both an amino group and a carboyxl group. Suitable amino acids include, without limitation, both the D- and L- isomers of the 20 naturally occurring amino acids found in peptides (e.g., A, R, N, C, D, Q, E, G, H, I, L, K, M, F, P, S, T, W, Y, V (as known by the one letter abbreviations)) as well as unnaturally occurring amino acids prepared by organic synthesis or other metabolic routes. The term "amino acid side chain" refers to any one of the twenty groups attached to the α-carbon in naturally occurring amino acids. For example, the amino acid side chain for alanine is methyl, the amino acid side chain for phenylalanine is phenylmethyl, the amino acid side chain for cysteine is thiomethyl, the amino acid side chain for aspartate is carboxymethyl, the amino acid side chain for tyrosine is A- hydroxyphenylmethyl, etc. The term "substituents" refers to a group "substituted" on an alkyl, cycloalkyl, aryl, heterocyclyl, or heteroaryl group at any atom of that group. Any moiety described herein can be further substituted with a substituent. Suitable substituents include, without limitation, halo, hydroxy, mercapto, oxo, nitro, haloalkyl, alkyl, aryl, aralkyl, alkoxy, thioalkoxy, aryloxy, amino, alkoxycarbonyl, amido, carboxy, alkanesulfonyl, alkylcarbonyl, and cyano groups.

An "antagonist" of a particular protein includes compounds that, at the protein level, directly bind or modify the subject component such that an activity of the subject component is decreased, e.g., by competitive or non-competitive inhibition, destabilization, destruction, clearance, or otherwise. For example, the decreased activity can include reduced ability to respond to an endogenous ligand. For example, an antagonist of GHS-R can reduce the ability of GHS-R to respond to ghrelin.

An "agonist" of a particular protein includes compounds that, at the protein level, directly bind or modify the subject component such that an activity of the subject component is increased, e.g., by activation, stabilization, altered distribution, or otherwise.

An "inverse agonist" of a particular protein includes a compound that, at the protein level, causes an inhibition of the constitutive activity of the protein (e.g., a receptor), with a negative intrinsic activity, for example by binding to and/or stabilizing an inactive form of the protein, which pushes the equilibrium away from formation of an active conformation of the protein.

Generally, a receptor exists in an active (Ra) and an inactive (Ri) conformation. Certain compounds that affect the receptor can alter the ratio of Ra to Ri (Ra/Ri). For example, a full agonist increases the ratio of Ra/Ri and can cause a "maximal", saturating effect. A partial agonist, when bound to the receptor, gives a response that is lower than that elicited by a full agonist (e.g., an endogenous agonist). Thus, the Ra/Ri for a partial agonist is less than for a full agonist. However, the potency of a partial agonist may be greater or less than that of the full agonist.

Certain compounds that agonize GHS-R to a lesser extent than ghrelin can function in assays as antagonists as well as agonists. These compounds antagonize activation of GHS-R by ghrelin because they prevent the full effect of ghrelin-receptor interaction. However, the compounds also, on their own, activate some receptor activity, typically less than a corresponding amount of ghrelin. Such compounds may be referred to as "partial agonists of GHS-R".

A subject with "normal" GH levels is one who would return a normal result using the glucose tolerance test in which glucose is ingested and blood levels of GH are measured by enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA) or polyclonal immunoassay. A normal result for this test is characterized by less than 1 ng/mL of GH within 1 to 2 hours of an oral glucose load. However, GH levels of a subject with excessive GH, as in one with acromegaly may not decrease below 1 ng/mL after ingesting glucose. Because GH levels oscillate every twenty to thirty minutes and varies in level according to the time of day, stress level, exercise, etc., a standard means of determining if GH levels are excessive is to administer glucose. This approach normalizes GH and is less affected by the pulsatility of GH, age, gender, or other factors. Alternatively or as a confirmation, since IGF-I levels are invariably increased in acromegalic individuals, IGF-I levels can be measured and compared to age and gender matched normal controls.

The term "an indicator of GH/IGF-1 axis activity" refers to a detectable property of the GH/IGF-1 axis that is indicative of activity of the axis. Exemplary properties include circulating GH concentration, circulating IGF-I concentration, frequency of GH pulses, amplitude of GH pulses, GH concentration in response to glucose, IGF-I receptor phosphorylation, and IGF-I receptor substrate phosphorylation. A compound that modulates activity of GHS-R can alter one or more indicators of GH/IGF-1 axis activity.

In some aspects, the disclosure provides the use of a composition described herein (e.g., a ghrelin pathway antagonist, alone or in combination with another agent described herein) for the preparation of a medicament, e.g., for the treatment indication described herein, e.g., for preserving pancreatic islet mass. In some aspects, the disclosure provides the use of a ghrelin pathway antagonist, alone or in combination with another agent described herein, for use in treatment.

All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In the case of conflict, the present specification, including definitions, controls. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting.

The details of one or more embodiments of the invention are set forth in the accompanying drawings and the description below.

DESCRIPTION OF THE DRAWINGS

FIGS. IA- 1C are bar graphs showing measurements taken for ghrelin receptor (GhrR) wild type (WT) (shaded bars) and knock out (KO) (open bars) mice that were fed a high fat diet. FIG. IA: body weight measurements (g). FIG. IB: islet cell mass measurements (mg). FIG 1C: pancreas weight measurements (mg). FIG. 2 is bar graphs showing the islet triglyceride content (ng/islet) for GhrR WT and KO mice fed a normal chow (NC; open bars) or a high fat (HF; shaded bars) diet. ANOVA results are also shown.

FIGS. 3 A and 3B are bar graphs showing measurements taken for mice treated with EX- 1350 ghrelin pathway antagonist (shaded bars) or with a vehicle control (open bars). FIG. 3A: pancreas weight measurements (mg). FIG. 3B: body weight measurements (g).

FIG. 4 is bar graphs showing islet cell mass measurements (mg) taken for mice treated with EX- 1350 ghrelin pathway antagonist (shaded bars) or with a vehicle control (open bars).

DETAILED DESCRIPTION

The present invention is based, in part, on the discovery that administering a ghrelin pathway antagonist (optionally in combination with an additional treatment) can preserve pancreatic islet mass in a subject. The preservation of pancreatic islet mass can preserve islet function and/or delay the development (or progression) of prediabetes or diabetes in a subject at risk thereof, or in a subject having a risk factor or precursor stage therefore, e.g., obesity, insulin resistance, or prediabetes. A ghrelin pathway antagonist can lead to pancreas sparing in a subject.

Increased dietary fat intake is associated with obesity and insulin resistance. As insulin resistance develops (e.g., in a subject with prediabetes), the pancreatic islet attempts to compensate for increased blood glucose concentrations by increasing the rate of insulin secretion. The islet achieves this by increasing the number of beta cells and thus increasing its mass. However, the pancreatic islet mass may increase due to increased lipid content, as dietary fat intake increases and obesity develops. Increased islet lipid content can cause an impairment of islet cell function, including: reduced insulin secretion, a decreased response to glucose-stimulated insulin secretion, and/or increased metabolism of glucose and/or free fatty acids (FFAs) causing glucolipotoxicity, lipotoxicity, oxidative stress, and/or dysregulation of triglyceride and/or FFA cycling. This can eventually lead to β-cell exhaustion and endoplasmic reticulum (ER) stress, which can lead to islet dysfunction and type 2 diabetes. Eventually, the cells are less able or no longer able to compensate for the impaired islet cell function and islet cell mass is reduced by increased apoptosis and/or necrosis. By preserving islet mass, e.g., by preventing or delaying an increase in islet mass, e.g., in prediabetes, e.g., by preventing an increase in islet lipid content or decreasing islet lipid content, the impairment of islet cell function is prevented, decreased (e.g., minimized) and/or delayed. Further, by preserving islet mass, e.g., by preventing or delaying decrease in islet mass e.g., in diabetes (e.g., due to β-cell exhaustion, endoplasmic reticulum (ER) stress, apoptosis, and/or necrosis), the impairment of islet cell function is prevented, decreased (e.g., minimized) and/or delayed. Ghrelin pathway antagonists can be used to prevent an increase in pancreatic islet lipid content and/or to decrease islet lipid content. This can delay or prevent the development or progression of prediabetes or diabetes. Ghrelin pathway antagonists can be used to preserve pancreatic islet mass (e.g., the mass of individual islet cells and/or the number of cells in the islet). Ghrelin pathway antagonists can be used to prevent impairment, delay impairment, and/or decrease the extent of impairment of islet cell function. The antagonists can prevent or delay the progression or incidence of prediabetes or diabetes.

Pancreatic Islets The endocrine cells of the pancreas are grouped in the islets of Langerhans

("islets"). The islets constitute approximately 1 to 2% of the mass of the pancreas. There are about one million islets in a healthy adult human pancreas, which are distributed evenly throughout the organ; their combined weight is about 1 to 1.5 grams.

Hormones produced in the islets are secreted directly into the blood flow by (at least) five different types of cells:

Alpha (α) cells producing glucagon (15-20% of total islet cells) Beta (β) cells producing insulin and amylin (65-80%) Delta (δ) cells producing somatostatin (3-10%) PP cells producing pancreatic polypeptide (3-5%) and Epsilon (ε) cells producing ghrelin (<1%).

The tight coupling between insulin and glucagon secretion in relation to blood nutrients is possible because of the anatomical arrangement of cells in the islets and the direction of the blood flow in each islet. Each islet contains a central core of insulin- secreting β-cells and a mantle of glucagon-secreting α- and/or somatostatin-releasing δ- cells or a mantle of δ- and PP cells, so called because they release pancreatic polypeptide. The δ-cells are branched and send processes into the core of the islet.

Each islet is highly vascularized with small arterioles entering its core. These break up into a network of capillaries that form venules carrying blood to the mantle. Such an arrangement of blood flow allows high concentrations of insulin to bathe the α-, δ- and PP cells. Additionally, the capillaries within the islets are fenestrated, facilitating peptide entry into the blood stream.

Insulin is secreted in response to increases in glucose concentration in extracellular fluid. This metabolic signal requires metabolism of glucose to pyruvate and appears to be detected by the activity of the enzyme glucokinase that catalyzes production of glucose-6-phosphate. This initiates an insulin-releasing signal involving a rise in ATP, closure of a K⁺ channel, depolarization and opening of a Ca²⁺ channel. This process is very rapid and secretion of insulin occurs within one minute of exposure to glucose. Detection of changes in glucose concentration is facilitated by the presence of canaliculi containing interstitial fluid along the lateral surfaces of neighboring β-cells between the arterioles and venules. Concentrated in the microvilli of these canaliculi are the specific GLUT2 glucose transporters enabling the intracellular concentration of glucose in the β-cells to be essentially the same as that of the interstitial fluid.

Pancreatic Islets and Diabetes. It has been proposed that there are five stages in the progression of diabetes, each of which is marked by important changes in β -cell mass, phenotype, and function (Weir and Bonner-Weir, Diabetes 53:S16-S21 (2004)). Stage 1 can be described as compensation: insulin secretion increases to maintain normal glucose levels in the face of insulin resistance resulting from obesity, physical inactivity, and genetic predisposition. Stage 2 occurs when glucose levels rise to levels of -5.0-6.5 mmol/1 (89-116 mg/dl) — a stable state of β-cell adaptation. Stage 3 is an unstable period of early decompensation in which glucose levels rise relatively rapidly to stage 4, which is characterized as stable decompensation. Finally, there is the severe decompensation of stage 5 that represents profound β -cell failure with progression to ketosis. Movement across stages 1-4 can be in either direction. For example, individuals with type 2 diabetes who undergo gastric reduction surgery can move from stage 4 to stage 1. Even conventional treatments with diet, exercise, and oral agents often return people to stage 2. For type 1 diabetes, as remission develops, progression from stage 4 to stage 2 is typically found.

The most common example of compensation is found with the insulin resistance due to obesity, which is accompanied by higher overall rates of insulin secretion and increased acute glucose-stimulated insulin secretion (GSIS) following an intravenous glucose challenge. Much of the increase in insulin secretion undoubtedly results from an increase in β-cell mass, as has been found in autopsy studies in humans and numerous rodent models. β-Cell mass is normally tightly maintained through a balance of β-cell birth β -cell replication and islet neogenesis and β-cell death through apoptosis. Most of the increase in β-cell mass with insulin resistance is probably due to increased β-cell number, but β -cell hypertrophy may also contribute. It is not yet clear if the higher plasma insulin levels can be entirely explained by the larger β -cell mass or whether there is also increased secretion per given unit of β-cell mass.

Measurements of Pancreas Weight and Pancreas Parameters Pancreatic islet mass can be measured according to any such technique known in the art. For example, positron emission tomography (PET) can be used to estimate islet mass. Souza et al. (J. Clin. Invest. 116:1506-1513 (2006)) describe the use of a labeled ligand that binds to β-cells and longitudinal noninvasive PET-based measurements to determine pancreatic β-cell mass. As described therein, the β-cells of the pancreas and neurons share expression of a large number of genes and display some functional similarities. Vesicular monoamine transporter type 2 (VMAT2) is expressed at dopamine nerve terminals in the CNS and by β-cells, but is absent from the exocrine pancreas and many other intra-abdominal tissues. A specific ligand for VMAT2, dihydrotetrabenazine (DTBZ), is in clinical use for PET imaging of the CNS. Souza et al. studied the binding of [³H]DTBZ to total membrane fractions prepared from purified human islets and purified exocrine pancreas tissue. [³H]DTBZ bound specifically to islet membranes, but not to membranes from the exocrine pancreas. VMAT2 was not present in the other endocrine cells of the islets or the exocrine pancreas. The successful use of [¹¹C]DTBZ to image the endocrine pancreas in vivo, and to estimate β-cell mass, and changes in β- cell mass, was demonstrated.

Total pancreatic volume can be measured by computed tomography (CT), for example, from the contour of the pancreas in a CT image (see, e.g., Saisho et al., Clin. Anat. 20:933-942 (2007)). Likewise, pancreas parencymal volume, pancreas fat volume, and a fat/parenchyma ratio can be determined by CT density (Id.). Additional assays to image the pancreas and determine pancreatic islet mass include:

- a β-cell-specific anti-IC2 mAb, modified with a radioisotope chelator, can be used in radioimmuno scintigraphy to image and measure β-cells;

- the in vivo uptake of 6-deoxy-6-[¹²⁵I]iodo-d-glucose by islets and acinar tissue can be measured; and - pancreatic uptake of [2-(14)C]alloxan, which is preferentially taken up by endocrine cells, particularly β -cells, and which targets the GLUT2 transporter, can be measured.

In addition, pancreatic islet function and impairment of function can be measured indirectly, for example, by:

Performing an oral glucose tolerance test (OGTT),

Measurements of fasting blood glucose concentrations, and

Measurements of blood Hbalc.

Further, although islet lipid content cannot be measured directly, an indirect way of measuring an individual' s plasma triglyceride content can give an indication of lipid toxicity.

To determine if a treatment is, e.g., preserving a parameter of the pancreas, the parameter can be measured before, during, and/or after treatment (e.g., administration of a ghrelin pathway antagonist described herein). For example, islet mass can be measured prior to commencing treatment, at regular intervals during treatment, and/or upon cessation of treatment or upon switching treatments and/or treatment schedule, and/or treatment dose.

Ghrelin Pathway Ghrelin is an appetite-stimulating hormone produced by P/Dl cells lining the fundus of the human stomach and epsilon cells of the pancreas. Ghrelin levels increase before meals and decrease after meals. It is considered the counterpart of the hormone leptin, produced by adipose tissue, which induces satiation when present at higher levels. Ghrelin is also produced in the hypothalamic arcuate nucleus where it stimulates the secretion of growth hormone from the anterior pituitary gland.

Ghrelin is also made by a small population of neurons in the arcuate nucleus. Ghrelin plays a significant role in neurotrophy, particularly in the hippocampus, and is essential for cognitive adaptation to changing environments and the process of learning.

Receptors for ghrelin are expressed by neurons in the arcuate nucleus and the ventromedial hypothalamus. The ghrelin receptor (GhrR) is a G protein-coupled receptor, also known as the GHS receptor (growth hormone secretagogue receptor; GHS-

R).

GHS-R can regulate the secretion of growth hormone (GH). GH itself is a regulator of IGF-I production. Thus, compounds, e.g., compounds described herein, that modulate GHS-R activity can be used to modulate (e.g., increase or decrease) activity of the GH/IGF-1 axis. For example, agonists of GHS-R can be used to increase GH activity and/or IGF-I activity. Antagonists of GHS-R can be used to decrease GH activity and/or IGF-I activity. See also US Pat. App. No. 10/656,530, the contents of which include uses for which a compound described herein may be used, e.g., as a modulator of the GH/IGF- 1 axis.

The GH/IGF-1 axis includes a series of extracellular and intracellular signaling components that have the transcription factor Forkhead as a downstream target. Major components of the GH/IGF-1 axis can be divided into three categories: pre-IGF-1, IGF- 1, and post IGF-I components. "Pre IGF-I components" include GH, GH-R, ghrelin, GHS-R, GHRH, GHRH-R, SST, and SST-R. "Post-IGF-1 components" include IGF-I-R and intracellular signaling components including PI(3) kinase, PTEN phosphatase, PI(3,4)P2, 14-3-3 protein, and PI(3,4,5)P3 phosphatidyl inositol kinases, AKT serine/threonine kinase (e.g., AKT-I, AKT-2, or AKT-3), or a Forkhead transcription factor (such as FOXO-I, FOXO-3, or FOXO-4). A "somatotroph axis signaling pathway component" refers to a protein that is one of the following: (i) a protein that is located in a somatotroph and that regulates GH release by the somatotroph, or (ii) a protein that directly binds to a protein in class (i). Exemplary somatotroph axis signaling pathway components of class (i) include cell surface receptors such as GHS-R, GHRH-R, and SST-R. Exemplary somatotroph axis signaling pathway components of class (ii) include GHRH, ghrelin, and SST.

A compound that modulates GH levels, e.g., by altering GHS-R activity can have downstream effects. For example, the compound can alter (e.g., increase or decrease) the levels or activity of an IGF-I receptor signaling pathway effector. A "IGF-I Receptor signaling pathway effector" refers a protein or other biologic whose levels are directly regulated by a Forkhead transcription factor in response to IGF-I. For example, expression of the gene encoding the protein can be directly regulated by a Forkhead transcription factor such as FOXO-I, F0X0-3a, or F0X0-4. Exemplary IGF-I Receptor signaling pathway effector can include: GADD45, PA26, Selenoprotein P, Whipl, cyclin G2, and NIP3.

As used herein, "activity of the GH/IGF-1 axis" refers to the net effect of the axis components with respect to ability to stimulate GH secretion, increase IGF-I levels, or increase IGF-I receptor signaling. Accordingly, "downregulating the GH/IGF-1 axis" refers to modulating one or more components such that one or more of the following is reduced, e.g., decreased GH, decreased IGF-I, or decreased IGF-I receptor signaling. For example, in some instances, GH levels are maintained but its action is inhibited; thus IGF-I levels are decreased without decreasing GH levels. In some instances, both GH and IGF-I levels are decreased.

Ghrelin Pathway Antagonists

Ghrelin pathway antagonists that can be used in methods to preserve pancreatic islet mass or islet lipid content can be a compound, e.g., a small molecule (e.g., small organic molecule) (e.g., less than 7 kDa in molecular weight, e.g., 6, 5, 4, 3, 2, 1, 0.7, 0.6, 0.5, 0.4, 0.3, or 0.2 kDa) or macromolecule. The antagonist can be a peptide, polypeptide, protein, antibody, antibody fragment (e.g., Fc fragment), peptidomimetic, peptoid, nucleic acid, or other chemical compound or a combination thereof. One or more ghrelin pathway antagonists can be used alone or in combination with another agent, e.g., another antagonist described herein. In the case of a combination therapy, the amounts and times of administration can be those that provide, e.g., a synergistic therapeutic effect, or an additive therapeutic effect. The ghrelin pathway antagonist and other agent can be administered together or in sequence. The ghrelin pathway antagonist and other agent can be in the same or in separate compositions, e.g., in the same or separate dosage forms. Further, the administration of the ghrelin pathway antagonist (with or without the additional antagonist) can be used as a primary, e.g., first line treatment, or as a secondary treatment, e.g., for subjects who have an inadequate response to a previously administered therapy (i.e., a therapy other than one with a ghrelin pathway antagonist). In some embodiments, a ghrelin pathway antagonist can be used in combination with a standard treatment for a disorder or disease, e.g., obesity or diabetes.

Examples of antagonists at the protein level include antibodies, proteins, and peptides (e.g., fragments of naturally occurring ligands, random or semi-randomly generated binding peptides, and so forth), non-proteinaceous molecules include, e.g., molecules less than 3000, 2000, 1000, 700, 500, 400, 300, or 200 in molecular weight. Examples of antagonists at the nucleic acid level include nucleic acid aptamers RNAi, antisense RNAs, a ribozyme molecule, and a molecule which binds to a regulatory element for the subject component (e.g., an artificial transcription factor, e.g., a zinc finger protein).

The antagonists can target any component of the ghrelin pathway, e.g., ghrelin or the ghrelin receptor (GHS-R). For example, the antagonists include: anti-ghrelin antibodies, anti-ghrelin receptor antibodies, soluble forms of ghrelin or the ghrelin receptor (e.g., a fusion protein containing the extracellular domain of the ghrelin receptor, e.g., an Fc fusion), and small molecule compounds, described below.

An exemplary antagonist is the ghrelin receptor antagonist [D-Lys³]-GHRP-6, also referred to as antagonist for Growth Hormone Releasing Peptide 6 (see also His-D- Trp-D-Lys-Trp-D-Phe-Lys-NH2; Sigma- Aldrich Product No. G4535). Other antagonists include antibodies to ghrelin and GHS-R. See, e.g., Nakazato et al. (2001) Nature 409:194.

Antibodies to ghrelin and GHS-R are commercially available, e.g., from AbCam, Abnova, Acris Antibodies, Santa Cruz Biotechnology, and Vincibiochem.

Ghrelin and/or GHS-R siRNA is commercially available, e.g., from Santa Cruz Biotechnology, Qiagen Inc., Dharmacon, Inc., Invitrogen. Exemplary ghrelin pathway antagonists include: TZP-301 (Tranzyme Pharma); ghrelin antagonists from 7TM Pharma; ghrelin receptor antagonists from Novo Nordisk; ghrelin antagonists JMV-2866 and JMV-2844 (Zentaris); ghrelin antagonist peptide, GIy- Ser-Ser(octanoyl)-Phe (Ardana); JMV2959; and NOX-BI l (Noxxon), a spiegelmer- based ghrelin antagonist. Spiegelmers are L-RNAs that are able to bind to L-peptides. NOX-BI l was identified using Noxxon's SELEX technology, which allows selection of a D-RNA aptamer which binds to the D-peptide with high affinity. The L-RNA that will then bind to the L-peptide is synthesized from the D-RNA sequence.

Additional ghrelin pathway antagonists are described., e.g., in PCT Pub. Nos. WO 02/08250; WO 01/87335; WO 2007/038678; WO 2004/084943; WO 2005/112903; WO 2006/137974; and US Pub. App. Nos. US 2006-257867; US 2005-272648; US 2005- 201938; US 2003-211967.

Ghrelin and GHS-R Sequences.

Exemplary ghrelin sequences include:

Exemplary GHS-R sequences include:

Antibodies. Immunoglobulins that bind to a component of the ghrelin pathway, e.g., ghrelin or the ghrelin receptor (GHS-R) and, for example, that reduce ghrelin pathway activity, can also be produced. For example, an immunoglobulin can bind to a receptor and modulate receptor activity or the ability of a ligand to interact or modulate the receptor. For example, an immunoglobulin can bind to GHS-R and prevent ghrelin binding, without itself activating the receptor. Similarly, an immunoglobulin can bind to a secreted pathway component, e.g., a GH Secretagogue (GHS) (e.g., ghrelin), and e.g., prevent the component from binding to and/or activating its receptor (e.g., GHS-R). In a preferred embodiment, the immunoglobulin is human, humanized, deimmunized, or otherwise non-antigenic in the subject.

An immunoglobulin can be, for example, an antibody or an antigen-binding fragment thereof. As used herein, the term "immunoglobulin" refers to a protein consisting of one or more polypeptides that include one or more immunoglobulin variable domain sequences. A typical immunoglobulin includes at least a heavy chain immunoglobulin variable domain and a light chain immunoglobulin variable domain. An immunoglobulin protein can be encoded by immunoglobulin genes. The recognized human immunoglobulin genes include the kappa, lambda, alpha (IgAl and IgA2), gamma (IgGl, IgG2, IgG3, IgG4), delta, epsilon and mu constant region genes, as well as the myriad immunoglobulin variable region genes. Full-length immunoglobulin "light chains" (about 25 KDa or 214 amino acids) are encoded by a variable region gene at the NH2-terminus (about 110 amino acids) and a kappa or lambda constant region gene at the COOH- terminus. Full-length immunoglobulin "heavy chains" (about 50 KDa or 446 amino acids), are similarly encoded by a variable region gene (about 116 amino acids) and one of the other aforementioned constant region genes, e.g., gamma (encoding about 330 amino acids). The term "antigen-binding fragment" of an antibody (or simply

"antibody portion," or "fragment"), as used herein, refers to one or more fragments of a full-length antibody that retain the ability to specifically bind to the antigen. Examples of antigen-binding fragments include: (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CL and CHl domains; (ii) a F(ab')₂ fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; (iii) a Fd fragment consisting of the VH and CHl domains; (iv) a Fv fragment consisting of the VL and VH domains of a single arm of an antibody, (v) a dAb fragment (Ward et al., (1989) Nature 341:544-546), which consists of a VH domain; and (vi) an isolated complementarity determining region (CDR). Furthermore, although the two domains of the Fv fragment, VL and VH, are coded for by separate genes, they can be joined, using recombinant methods, by a synthetic linker that enables them to be made as a single protein chain in which the VL and VH regions pair to form monovalent molecules (known as single chain Fv (scFv); see e.g., Bird et al. (1988) Science 242:423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883). Such single chain antibodies are also encompassed within the term "antigen-binding fragment" of an antibody. These antibody fragments are obtained using conventional techniques known to those with skill in the art, and the fragments are screened for utility in the same manner as are intact antibodies.

In one embodiment, the antibody against the ghrelin pathway component is a fully human antibody (e.g., an antibody made in a mouse which has been genetically engineered to produce an antibody from a human immunoglobulin sequence), or a non- human antibody, e.g., a rodent (mouse or rat), goat, primate (e.g., monkey). Preferably, the non-human antibody is a rodent (mouse or rat antibody). Methods of producing rodent antibodies are known in the art. Human monoclonal antibodies can be generated using transgenic mice carrying the human immunoglobulin genes rather than the mouse system (see, e.g., PCT Pub. Nos. WO 91/00906 and WO 92/03918). Other methods for generating immunoglobulin ligands include phage display (e.g., as described in US Pat. No. 5,223,409 and PCT Pub. No. WO 92/20791).

RNAi. It is also possible to attenuate ghrelin pathway activity using a double- stranded RNA (dsRNA) that mediates RNA interference (RNAi). The dsRNA can be delivered to cells or to an organism. Endogenous components of the cell or organism can trigger RNA interference (RNAi) which silences expression of genes that include the target sequence. dsRNA can be produced by transcribing a cassette in both directions, for example, by including a T7 promoter on either side of the cassette. The insert in the cassette is selected so that it includes a sequence from a ghrelin pathway component to be attenuated. The sequence need not be full length, for example, an exon, or at least 50 nucleotides, preferably from the 5' half of the transcript, e.g., within 300 nucleotides of the ATG. See also, the HISCRIBE™ RNAi Transcription Kit (New England Biolabs,

MA) and Fire, A. (1999) Trends Genet. 15, 358-363. dsRNA can be digested into smaller fragments. See, e.g., US Pub. App. No. 2002-0086356. dsRNAs can be used to silence gene expression in mammalian cells. See, e.g., Clemens, J. C. et al. (2000) Proc. Natl. Sci. USA 97, 6499-6503; Billy, E. et al. (2001)

Proc. Natl. Sci. USA 98, 14428-14433; Elbashir et al. (2001) Nature. 411(6836):494-8;

Yang, D. et al. (2002) Proc. Natl. Acad. Sci. USA 99, 9942-9947.

For example, double stranded RNA molecules complementary to a nucleic acid encoding a GHS (e.g., ghrelin) or GHS-R, can be used to attenuate activity of the ghrelin pathway.

In one embodiment, an siRNA is used. siRNAs are small double stranded RNAs

(dsRNAs) that optionally include overhangs. For example, the duplex region is about 18 to 25 nucleotides in length, e.g., about 19, 20, 21, 22, 23, or 24 nucleotides in length.

Typically the siRNA sequences are exactly complementary to the target mRNA. dsRNAs and siRNAs in particular can be used to silence gene expression in mammalian cells (e.g., human cells). See, e.g., Clemens, J. C. et al. (2000) Proc. Natl. Sci. USA 97, 6499-6503;

Billy, E. et al. (2001) Proc. Natl. Sci. USA 98, 14428-14433; Elbashir et al. (2001)

Nature. 411(6836):494-8; Yang, D. et al. (2002) Proc. Natl. Acad. Sci. USA 99, 9942-

9947; US Pub. App. Nos. 2003-0166282 and 2003-0143204. Small Molecules. The small molecules described herein can be used for a variety of purposes, e.g., as ghrelin pathway modulators, e.g., antagonists. The compounds can, e.g., antagonize GHS-R activity and can be used to reduce GHS-R activity, e.g., in a subject.

Representative compounds that can be used in practicing the methods of the invention are depicted below in Table 1. Other exemplary compounds are within the scope set forth in the Summary and/or are described in US Patent App. 11/451,086 and

PCT Pub. No. WO 2006/135860. Methods of preparing GHS-R modulating compounds are described, e.g., in US Patent Apps. 11/451,086 and 11/564,815, and PCT Pub. Nos.

WO 2006/135860 and WO 2007/146914. Table 1: Exemplary GHS-R Modulating Compounds

* A refers to a compound having antagonist activity with a Ki < 100 nM in a cell based assay.

B refers to a compound having antagonist activity with a Ki between 100 nM and 500 nM in a cell based assay.

C refers to a compound having antagonist activity with a Ki between 500 nM and 1000 nM in a cell based assay.

D refers to a compound having antagonist activity with Ki, < 1000 nM in a cell-based assay.

E refers to other exemplary compounds. Representative compounds that modulate GHS-R include the compounds of formulas (I), (II), (III), (IV), (V), and (VI) below, where all variables are as described herein.

formula (I) formula (II) formula (III)

formula (IV) formula (V) formula (VI).

In some preferred embodiments, Y is a 5 membered heteroaromatic moiety substituted with 1 or 2 substituents as described herein. Exemplary Y moieties are reproduced below.

In each instance, any atom, including the hydrogens depicted on the nitrogen atoms, can be substituted with R . 10. In some preferred embodiments, the heteroaryl moiety includes 1 or 2 R . 10 substituents. In some preferred embodiments, R 10 is aryl, arylalkyl, or R¹⁵. When two R¹⁰ substituents are included, in some embodiments, one R¹⁰ is R¹⁵ and the second R . 10 is a different substituent, such as alkyl, alkoxy, halo, etc. In certain instances, R¹ is an aryl moietiy such as a phenyl moiety, for example unsubstituted or substituted aryl moiety. In some instances, R¹ is a heteroaryl moiety such as an indole moiety. In many instances where R¹ is aryl or heteroaryl (or other lipophilic moiety such as alkyl), K is an oxygen or a bond.

5 A and R⁴ and R⁵ can be chosen to vary the compound's type of interaction with GHS-

R. For example, in some instances where R⁴ and R⁵ are both hydrogen, the compound is an agonist of GHS-R. In other instances where R⁴ and R⁵ are both independently alkyl, the compound is an antagonist of GHS-R.

Other aspects of this invention relate to a composition having a compound of any of o the formulae described herein and a pharmaceutically acceptable carrier; or a compound of any of the formulae described herein, an additional therapeutic compound (e.g., an anti-hypertensive compound or a cholesterol lowering compound), and a pharmaceutically acceptable carrier; or a compound of any of the formulae described herein, an additional therapeutic compound, and a pharmaceutically acceptable carrier.5 Combinations of substituents and variables envisioned by this invention are only those that result in the formation of stable compounds. The term "stable", as used herein, refers to compounds which possess stability sufficient to allow manufacture and which maintains the integrity of the compound for a sufficient period of time to be useful for the purposes detailed herein (e.g., therapeutic or prophylactic administration to a subject).0

Synthesis of Ghrelin Receptor-Modulating Compounds

The compounds described herein can be made using a variety of synthetic techniques.

In some embodiments, a Y moiety, or other ring corresponding to a Y moiety, can5 be synthesized onto an amino acid or amino acid type starting material as depicted in schemes A and B below.

Scheme A

In the schemes provided herein, all variables are defined as herein and PG is a nitrogen protecting group. The nitrogen protected amino acid is reacted with a N- hydroxy imidamide (amidoxime) moiety (which is prepared by reacting a cyano containing moiety with hydroxylamine) to produce an oxadiazole containing moiety. The resulting compound can be further manipulated to form a compound of formula (I) by removing the nitrogen protecting group and reacting the resulting moiety with an activated sulfone, such as a sulfonyl chloride as depicted below.

Scheme B below depicts the formation of a triazole containing moiety which can be further reacted in a manner similar to the oxadiazole containing moiety to form a compound of formula (I).

The triazole precursor moiety can be prepared in a variety of manners, for example, by reacting a cyano containing moiety with a hydrazine hydrate (to form the intermediate amidrazone). In other embodiments, a compound of formula (I) can be prepared by first reacting an activated sulfone moiety (e.g., a sulfonyl chloride) with an amino acid moiety or protected amino acid, as depicted in Scheme C below.

Scheme C.

The free carboxyl moiety can then be further manipulated to produce a compound of formula (I). For example, the free carboxyl moiety can be reacted with a compound of formula (X) or (XI) above to form an oxadiazole or triazole containing compound of formula (I) in a manner similar to that described in schemes A and B above. As can be appreciated by the skilled artisan, further methods of synthesizing the compounds of the formulae herein will be evident to those of ordinary skill in the art. Additionally, the various synthetic steps may be performed in an alternate sequence or order to give the desired compounds. Synthetic chemistry transformations and protecting group methodologies (protection and deprotection) useful in synthesizing the compounds described herein are known in the art and include, for example, those such as described in R. Larock, Comprehensive Organic Transformations, VCH Publishers (1989); T. W. Greene and P. GM. Wuts, Protective Groups in Organic Synthesis, 2d. Ed., John Wiley and Sons (1991); L. Fieser and M. Fieser, Fieser and Fieser's Reagents for Organic Synthesis, John Wiley and Sons (1994); and L. Paquette, ed., Encyclopedia of Reagents or Organic Synthesis, John Wiley and Sons (1995), and subsequent editions thereof.

Additionally, the compounds disclosed herein can be prepared on a solid support or using a solid phase peptide synthesis.

The term "solid support" refers a material to which a compound is attached to facilitate identification, isolation, purification, or chemical reaction selectivity of the compound. Such materials are known in the art and include, for example, beads, pellets, disks, fibers, gels, or particles such as cellulose beads, pore-glass beads, silica gels, polystyrene beads optionally cross-linked with divinylbenzene and optionally grafted with polyethylene glycol, poly-acrylamide beads, latex beads, dimethylacrylamide beads optionally cross-linked with N,N'-bis-acryloyl ethylene diamine, glass particles coated with hydrophobic polymer, and material having a rigid or semi-rigid surface. The solid supports optionally have functional groups such as amino, hydroxy, carboxy, or halo groups, (see, Obrecht, D. and Villalgrodo, J. M., Solid-Supported Combinatorial and Parallel Synthesis of Small-Molecular-Weight Compound Libraries, Pergamon-Elsevier Science Limited (1998)), and include those useful in techniques such as the "split and pool" or "parallel" synthesis techniques, solid-phase and solution-phase techniques, and encoding techniques (see, for example, Czarnik, A. W., Curr. Opin. Chem. Bio., (1997) 1, 60).

The term "solid phase peptide" refers to an amino acid, which is chemically bonded to a resin (e.g., a solid support). Resins are generally commercially available (e.g., from SigmaAldrich). Some examples of resins include Rink-resins, Tentagel S RAM, MBHA, and BHA-resins.

The compounds of this invention may contain one or more asymmetric centers and thus occur as racemates and racemic mixtures, single enantiomers and enantiometric mixtures, individual diastereomers and diastereomeric mixtures. All such isomeric forms of these compounds are expressly included in the present invention. The compounds of this invention may also be represented in multiple tautomeric forms, in such instances, the invention expressly includes all tautomeric forms of the compounds described herein (e.g., alkylation of a ring system may result in alkylation at multiple sites, the invention expressly includes all such reaction products). All such isomeric forms of such compounds are expressly included in the present invention. All crystal forms of the compounds described herein are expressly included in the present invention.

As used herein, the compounds of this invention, including the compounds of formulae described herein, are defined to include pharmaceutically acceptable derivatives or prodrugs thereof. A "pharmaceutically acceptable derivative or prodrug" means any pharmaceutically acceptable salt, ester, salt of an ester, or other derivative of a compound of this invention (for example an imidate ester of an amide), which, upon administration to a recipient, is capable of providing (directly or indirectly) a compound of this invention. Particularly favored derivatives and prodrugs are those that increase the bioavailability of the compounds of this invention when such compounds are administered to a mammal (e.g., by allowing an orally administered compound to be more readily absorbed into the blood) or which enhance delivery of the parent compound to a biological compartment (e.g., the brain or lymphatic system) relative to the parent species. Preferred prodrugs include derivatives where a group which enhances aqueous solubility or active transport through the gut membrane is appended to the structure of formulae described herein. The compounds of this invention may be modified by appending appropriate functionalities to enhance selective biological properties. Such modifications are known in the art and include those which increase biological penetration into a given biological compartment (e.g., blood, lymphatic system, central nervous system), increase oral availability, increase solubility to allow administration by injection, alter metabolism and alter rate of excretion.

Pharmaceutically acceptable salts of the compounds of this invention include those derived from pharmaceutically acceptable inorganic and organic acids and bases. Examples of suitable acid salts include acetate, adipate, benzoate, benzenesulfonate, butyrate, citrate, digluconate, dodecylsulfate, formate, fumarate, glycolate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, lactate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, palmoate, phosphate, picrate, pivalate, propionate, salicylate, succinate, sulfate, tartrate, tosylate and undecanoate. Salts derived from appropriate bases include alkali metal (e.g., sodium), alkaline earth metal (e.g., magnesium), ammonium and N-(alkyl)₄ ⁺ salts. This invention also envisions the quaternization of any basic nitrogen-containing groups of the compounds disclosed herein. Water or oil-soluble or dispersible products may be obtained by such quaternization.

Evaluating Compounds

A variety of methods can be used to evaluate a compound for ability to modulate (e.g., antagonize) the ghrelin pathway (e.g., ghrelin or GHS-R activity). Evaluation methods include in vitro binding assays, in vitro cell-based signaling assays, and in vivo methods. For example, the evaluation methods can evaluate binding activity, or an activity downstream of GHS-R, e.g., a signaling activity downstream of GHS-R such as inositol phosphate production, Ca²⁺ mobilization, or gene transcription (e.g., CREB- mediated gene transcription). The effects of a compound in pancreatic islet mass or islet lipid content can be evaluated as described herein, e.g., in the Examples.

Binding assays. Generally, the compounds can be evaluated to determine if they antagonize the ghrelin pathway, e.g., evaluated for their ability to bind to a ghrelin pathway component (e.g., ghrelin or GHS-R), and/or to determine if they compete with one or more known compounds that interact with a ghrelin pathway component (e.g., ghrelin or GHS-R), and the extent of such interactions. For example, the compounds can be evaluated to determine if they compete with ghrelin, ipamorelin, L-692,400 or L- 692,492 for binding to GHS-R. One exemplary binding assay is as follows: GHS-R expressing COS-7 cells cultured at a density of 1 x 10⁵ cells per well so that binding is assayed in the range of about 5 - 8 % binding of the radioactive ligand. For example, the cells can express an endogenous nucleic acid encoding GHS-R or an exogenous nucleic acid encoding GHS- R. Cells transfected with an exogenous nucleic acid encoding GHS-R can be used, e.g., two days, after transfection. Competition binding experiments are performed for 3 hours at 4°C using 25 pM of ¹²⁵I-ghrelin in 0.5 ml of 50 mM HEPES buffer, pH 7.4, supplemented with 1 mM CaCl₂, 5 mM MgCl₂, and 0.1 % (w/v) bovine serum albumin, 40 mg/ml bacitracin. Non-specific binding can be determined as the binding in the presence of 1 mM of unlabeled ghrelin. Cells are washed twice in 0.5 ml of ice-cold buffer and then lysed with 0.5-1 ml of lysis buffer (8 M Urea, 2 % NP40 in 3 M acetic acid). After washing and lysis, the bound radioactivity is counted. Assays can be run in duplicate or triplicate, e.g., to provide statistical power.

Values of the dissociation and inhibition constants (K_d and K₁) can be estimated from competition binding experiments using the equation: K_d = IC₅₀-L and K₁ = IC₅₀ / (1 + L / K_d), where L is the concentration of radioactive ligand. B_max values can be estimated from competition binding experiments using the equation B_max = B₀ ICso/tligand], where B₀ is the specifically bound radioligand. Cell-Based Activity Assays. For example, the ability of the compound to modulate accumulation of a second messenger signaling component downstream of GHS-R can be evaluated. For example, inositol phosphates (IP), as a result of Gq signaling in a mammalian cell, e.g., COS-7 cells. Other tissue culture cells, Xenopus oocytes, and primary cells can also be used.

Phosphatidylinositol turnover assay. An example of an assay is as follows. One day after transfection, COS-7 cells are incubated for 24 hours with 5 μCi of [³H]-myo- 5 inositol in 1 ml medium supplemented with 10% fetal calf serum, 2 mM glutamine and 0.01 mg/ml gentamicin per well. Cells are then washed twice in buffer, 20 mM HEPES, pH 7.4, supplemented with 140 mM NaCl, 5 mM KCl, 1 mM MgSO₄, 1 mM CaCl₂, 10 mM glucose, 0.05 % (w/v) bovine serum; and incubated in 0.5 ml buffer supplemented with 10 mM LiCl at 37°C for 30 min. For some assays, it is also useful to incubate the o cells with adenosine deaminase ADA (200U/mg, Boehringer Mannheim, Germany) for 30 min in a concentration of lU/ml .

After incubation with the compound of interest for 45 min at 37°C, cells are extracted with 10 % ice-cold perchloric acid and placed on ice for 30 min. The resulting supernatants are neutralized with KOH in HEPES buffer, and [³H]-inositol phosphate is5 purified on Bio-Rad AG 1-X8 anion exchange resin as described. Assays can be run in duplicate, triplicate, etc.

Other second messenger assays. Another second messenger that can be evaluated is Ca²⁺. Ca²⁺ mobilization can be evaluated using a calcium sensitive detector, such as aequorin protein or a dye, e.g., FURA-2. In an exemplary assay, calcium mobilization is0 evaluated in a recombinant cell that expresses GHS-R and aequorin.

Gene expression assay. HEK293 cells (30 000 cells/well) seeded in 96-well plates are transiently transfected with a mixture of pFA2-CREB and pFR-Luc reporter plasmid (PATHDETECT™ CREB trans-Reporting System, Stratagene) and nucleic acid encoding GHS. One day after transfection, cells are treated with the compound of interest in an5 assay volume of 100 μl medium for 5 hrs. After treatment, cells are cultured in low serum (2.5%). After the incubation period, the assay is ended by washing the cells twice with PBS and adding lOOμl luciferase assay reagent (LUCLITE™, Packard Bioscience). Luminescence is measured (e.g., as relative light units (RLU)) using in a luminometer such as the TOPCOUNTER™ (Packard Bioscience) for 5 sec. 0 Other transcription based assays can include evaluating transcription of GHS-R regulated genes in primary cells that express GHS-R (e.g., cells from pituitary, brain, spinal cord, uterus, spleen, pancreas, kidney, adrenal gland, skeletal muscle, thyroid, liver, small intestine, and heart) or in recombinant cells that express GHS-R. rriRNA levels can be evaluated by any method, e.g., microarray analysis, Northern blotting, or RT-PCR. Exemplary genes that are directly or indirectly regulated by GHS-R activity include leptin, resistin, and adiponectin. GHS-R activity may also affect insulin, IGF-I, and leptin levels in circulation.

IC₅₀ and EC₅₀ values can be determined by nonlinear regression, e.g., using the Prism 3.0 software (GraphPad Software, San Diego).

In vivo assays. Exemplary in vivo assays include the fast-refeeding assay, e.g., as follows.

Prior to compound administration, mice are weighed and sorted into groups based on comparable body weight. Food is removed at 6pm for an overnight (- 16 hour) fast. Beginning at 10 am on the next morning, mice are administered with either vehicle (e.g., saline + acetic acid, pH=5) or the compound of interest. Mice are then returned to their home cages and pre- weighed food (approximately 90 grams) is immediately returned to the food hoppers in each cage. The weight of the food remaining in the food hoppers is measured at 30 minutes, 1 hour, 2 hours, and 4 hours post compound/vehicle administration. Final body weights are then recorded for the mice.

The compound of interest can also be evaluated in other experiments. For example, the compound can be administered to lean or obese mice (e.g., (ob/ob)

C57BL/6J mice), or other experimental animals. The compound can be administered intraperitoneally or intracerebroventricularly. After administration, the animal is evaluated, e.g., for feeding behavior, anxiety, or one or more physiological parameters, e.g., a metabolic parameter. ICV Administration. For intra-third cerebroventricular (ICV) administration, each drug can be diluted in 4 Tl of artificial cerebrospinal fluid for injection. For ICV injection, mice are anaesthetised with sodium pentobarbital (80-85 mg/kg intraperitoneally) and placed in a stereotaxic instrument seven days before the experiments. A hole is made in each skull using a needle inserted 0.9 mm lateral to the central suture and 0.9 mm posterior to the bregma. A 24 gauge cannula bevelled at one end over a distance of 3 mm is implanted into the third cerebral ventricle for ICV injection.

Gastric emptying assessment. Another test for food consumption after administration of a compound of interest is the gastric emptying assessment. Before the gastric emptying assessment, mice are food deprived for 16 hours with free access to water. Fasted mice are given free access to preweighed pellets for one hour and then administered the compound of interest. The mice are again deprived of food for one or two hours after the compound administration. Food intake is measured by weighing uneaten pellets. Mice are killed by cervical dislocation two or three hours after the compound administration. Immediately after the stomach was exposed by laparotomy, quickly ligated at both the pylorus and cardia, removed, and the dry content is weighed. Gastric emptying is calculated according to the following formula: gastric emptying (%) = (1 - (dry weight of food recovered from the stomach/weight of food intake)) x 100.

Anxiety tests. Anxiety can be assessed in the standard elevated plus maze, 50 cm above the ground. The four arms can be made 27 cm long and 6 cm wide. Two opposing arms are enclosed by walls 15 cm high (closed arms) while the other arms are devoid of walls (open arms). Each mouse is placed in the center of the maze facing one of the enclosed arms 10 minutes after injection with a compound. The cumulative time spent in each arm and the number of entries into the open or closed arms is recorded during a five minute test session. The time spent in the open arms is expressed as a percentage of total entry time (100-open/ open+closed) and the number of entries in the open arms is expressed as a percentage of the total number of entries (100-open/total entries).

Parameter analysis. Mice or other animals provided with the test compound can be analyzed for one or more biological parameters, e.g., metabolic parameters. For mice, serum is obtained from blood from the orbital sinus under ether anaesthesia at the end of a treatment (e.g., eight hours after removal of food and the final intraperitoneal injection). Mice are killed by cervical dislocation. Immediately after, the epididymal fat pad mass can be assessed based on removal and weighing of the white adipose tissue (WAT) and the gastrocnemius muscle. Blood glucose can be measured by the glucose oxidase method. Serum insulin and free fatty acids (FFA) can be measured by enzyme immunoassay and an enzymatic method (Eiken Chemical Co., Ltd, Tokyo, Japan), respectively. Serum triglycerides and total cholesterol can be measured by an enzymatic method (Wako Pure Chemical Industries, Ltd, Tokyo, Japan). mRNA analysis. RNA is isolated from the stomach, epididymal fat or other relevant tissues using the RNeasy Mini Kit (Qiagen, Tokyo, Japan). Total RNA is denatured with formaldehyde, electrophoresed in 1% agarose gel, and blotted onto a

Hybond N+ membrane. The membranes are hybridized with a labeled cDNA probe (e.g., radioactively, chemically, or fluorescently labeled) for the gene of interest. The total integrated densities of hybridization signals can be determined by densitometry. Data can be normalized to a glyceraldehyde 3-phosphate dehydrogenase mRNA abundance or to actin mRNA abundance and expressed as a percentage of controls. Exemplary genes that can be evaluated include ghrelin, leptin, resistin, and adiponectin. It is also possible to use a transgenic animal that includes a reporter construct with a regulatory region from the gene of interest or to use a recombinant cell with such a construct.

A compound described herein can have a K₁ (as an antagonist) of less than 200, 100, 80, 70, 60, or 50 nM, in one or more of the described assays. A compound described herein can have a K_D as an agonist of greater than 20, 40, 50, 100, 200, 300, or 500 nM, in one or more of the described assays.

A compound described herein can also specifically interact with GHS-R, e.g., relative to other cell surface receptors. The motilin receptor, for example, is a homolog of GHS-R. A disclosed compound may preferentially interact with GHS-R relative to the motilin receptor, e.g., at least a 2, 5, 10, 20, 50, or 100 preference. In another embodiment, the disclosed compound may also interact with motilin receptor, and, e.g., alter motilin receptor activity.

In one embodiment, the compound may alter an intracellular signaling activity downstream of GHS-R, e.g., Gq signaling, phospholipase C signaling, and cAMP response element (CRE) driven gene transcription.

Compounds may also be evaluated for their therapeutic activity with respect to any disorder, e.g., a disorder described herein. Animal models for many disorders are well known in the art. Administration of Compounds and Formulations Thereof

The ghrelin pathway antagonists described herein can, for example, be administered by injection, intravenously, intraarterially, subdermally, intraperitoneally, intramuscularly, or subcutaneously; or orally, buccally, nasally, transmucosally, topically, in an ophthalmic preparation, or by inhalation, with a dosage ranging from about 0.001 to about 100 mg/kg of body weight, e.g., between 0.001-lmg/kg, 1-lOOmg/kg, or 0.01- 5mg/kg, every 4 to 120 hours, e.g., about every 6, 8, 12, 24, 48, or 72 hours, or according to the requirements of the particular compound. The methods herein contemplate administration of an effective amount of compound or compound composition to achieve the desired or stated effect. Typically, the pharmaceutical compositions of this invention will be administered from about 1 to about 6 times per day. Alternatively, the compounds can be administered as a continuous infusion. Such administration can be used as a chronic or acute therapy. The amount of active ingredient that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. A typical preparation will contain from about 5% to about 95% active compound (w/w). Alternatively, such preparations contain from about 20% to about 80% active compound.

Lower or higher doses than those recited above may be required. Specific dosage and treatment regimens for any particular subject will depend upon a variety of factors, including the activity of the specific compound employed, the age, body weight, general health status, sex, diet, time of administration, rate of excretion, drug combination, the severity and course of the disease, condition or symptoms, the subject's disposition to the disease, condition or symptoms, and the judgment of the treating physician.

Upon improvement of a subject's condition, a maintenance dose of a compound, composition or combination of this invention may be administered, if necessary.

Subsequently, the dosage or frequency of administration, or both, may be reduced, as a function of the symptoms, to a level at which the improved condition is retained. Subjects may, however, require intermittent treatment on a long-term basis upon any recurrence of disease symptoms. Pharmaceutical compositions of this invention comprise a compound of the formulae described herein or a pharmaceutically acceptable salt thereof; an additional compound including for example, a steroid or an analgesic; and any pharmaceutically acceptable carrier, adjuvant or vehicle. Alternate compositions of this invention comprise a compound of the formulae described herein or a pharmaceutically acceptable salt thereof; and a pharmaceutically acceptable carrier, adjuvant or vehicle. The compositions delineated herein include the compounds of the formulae delineated herein, as well as additional therapeutic compounds if present, in amounts effective for achieving a modulation of disease or disease symptoms, including kinase mediated disorders or symptoms thereof. The compositions are made by methods including the steps of combining one or more compounds delineated herein with one or more carriers and, optionally, one or more additional therapeutic compounds delineated herein.

The pharmaceutical compositions of this invention may be orally administered in any orally acceptable dosage form including, but not limited to, capsules, tablets, emulsions and aqueous suspensions, dispersions and solutions. In the case of tablets for oral use, carriers which are commonly used include lactose and corn starch. Lubricating agents, such as magnesium stearate, are also typically added. For oral administration in a capsule form, useful diluents include lactose and dried corn starch. When aqueous suspensions and/or emulsions are administered orally, the active ingredient may be suspended or dissolved in an oily phase which can be combined with emulsifying and/or suspending agents. If desired, certain sweetening and/or flavoring and/or coloring agents may be added.

The pharmaceutical compositions may be in the form of a sterile injectable preparation, for example, as a sterile injectable aqueous or oleaginous suspension. This suspension may be formulated according to techniques known in the art using suitable dispersing or wetting agents (such as, for example, Tween 80) and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3- butanediol. Among the acceptable vehicles and solvents that may be employed are mannitol, water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any bland fixed oil may be employed including synthetic mono- or diglycerides. Fatty acids, such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically-acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions. These oil solutions or suspensions may also contain a long-chain alcohol diluent or dispersant, or carboxymethyl cellulose or similar dispersing agents which are commonly used in the formulation of pharmaceutically acceptable dosage forms such as emulsions and or suspensions. Other commonly used surfactants such as Tweens or Spans and/or other similar emulsifying agents or bioavailability enhancers which are commonly used in the manufacture of pharmaceutically acceptable solid, liquid, or other dosage forms may also be used for the purposes of formulation. The pharmaceutical compositions of this invention may also be administered in the form of suppositories for rectal administration. These compositions can be prepared by mixing a compound of this invention with a suitable non-irritating excipient which is solid at room temperature but liquid at the rectal temperature and therefore will melt in the rectum to release the active components. Such materials include, but are not limited to, cocoa butter, beeswax and polyethylene glycols.

Pharmaceutically acceptable carriers, adjuvants and vehicles that may be used in the pharmaceutical compositions of this invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, self-emulsifying drug delivery systems (SEDDS) such as d-α-tocopherol polyethyleneglycol 1000 succinate, surfactants used in pharmaceutical dosage forms such as Tweens or other similar polymeric delivery matrices, serum proteins, such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol and wool fat. Cyclodextrins such as α -, β-, and γ-cyclodextrin, may also be advantageously used to enhance delivery of compounds of the formulae described herein. In some cases, the pH of the formulation may be adjusted with pharmaceutically acceptable acids, bases or buffers to enhance the stability of the formulated compound or its delivery form.

The term parenteral as used herein includes subcutaneous, intracutaneous, intravenous, intramuscular, intraarticular, intraarterial, intrasynovial, intrasternal, intrathecal, intralesional and intracranial injection or infusion techniques.

The pharmaceutical compositions of this invention may be administered by nasal aerosol or inhalation. Such compositions are prepared according to techniques well- known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and/or other solubilizing or dispersing agents known in the art.

When the compositions of this invention comprise a combination of a compound of the formulae described herein and one or more additional therapeutic or prophylactic agents, both the compound and the additional compound should be present at dosage levels of between about 1 to 100%, and more preferably between about 5 to 95% of the dosage normally administered in a monotherapy regimen. Additionally, combinations of a plurality of compounds described herein are also envisioned. The additional compounds may be administered separately, as part of a multiple dose regimen, from the compounds of this invention. Alternatively, those compounds may be part of a single dosage form, mixed together with the compounds of this invention in a single composition. The additional agent can optionally be present in a composition in combination with a ghrelin pathway antagonist. Alternatively, the additional agent can be administered in combination with the ghrelin pathway antagonist but present in a separate composition. The routes, doses, and/or dosing schedule (e.g., when and/or how often) of administration for the agents, and treatment duration for the agents can be the same or can differ.

Pharmaceutical Compositions

A ghrelin pathway antagonist can be incorporated into a pharmaceutical composition for administration to a subject, e.g., a human, a non-human animal, e.g., an animal patient (e.g., pet or agricultural animal) or an animal model (e.g., an animal model for aging or a metabolic disorder (e.g., a disorder of the GH/IGF-1 axis or a pancreatic or insulin related disorder). Such compositions typically include the antagonist (e.g., a small molecule that is a ghrelin pathway antagonist, nucleic acid molecule, protein, or antibody) and a pharmaceutically acceptable carrier.

As used herein, the language "pharmaceutically acceptable carrier or adjuvant" includes solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration. Supplementary active compounds can also be incorporated into the compositions. The term "pharmaceutically acceptable carrier or adjuvant" refers to a carrier or adjuvant that may be administered to a subject, together with a compound of this invention, and which does not destroy the pharmacological activity thereof and is nontoxic when administered in doses sufficient to deliver a therapeutic amount of the compound. A pharmaceutical composition is formulated to be compatible with its intended route of administration. Examples of routes of administration include parenteral, e.g., intravenous, intradermal, subcutaneous, oral (e.g., inhalation), transdermal (topical), transmucosal, and rectal administration. Solutions or suspensions used for parenteral, intradermal, or subcutaneous application can include the following components: a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose. pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide. The parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic.

Pharmaceutical compositions suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion. For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor EL™ (BASF, Parsippany, NJ) or phosphate buffered saline (PBS). In all cases, the composition must be sterile and should be fluid to the extent that easy syringability exists. It should be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyetheylene glycol, and the like), and suitable mixtures thereof. The proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants. Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars, polyalcohols such as manitol, sorbitol, sodium chloride in the composition. Prolonged absorption of the injectable compositions can be brought about by including in the composition an agent which delays absorption, for example, aluminum monostearate and gelatin.

Sterile injectable solutions can be prepared by incorporating the active compound in the required amount in an appropriate solvent with one or a combination of ingredients enumerated above, as required, followed by filtered sterilization. Generally, dispersions are prepared by incorporating the active compound into a sterile vehicle which contains a basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation are vacuum drying and freeze-drying which yields a powder of the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof.

Oral compositions generally include an inert diluent or an edible carrier. For the purpose of oral therapeutic administration, the active compound can be incorporated with excipients and used in the form of tablets, troches, or capsules, e.g., gelatin capsules. Oral compositions can also be prepared using a fluid carrier for use as a mouthwash. Pharmaceutically compatible binding agents, and/or adjuvant materials can be included as part of the composition. The tablets, pills, capsules, troches and the like can contain any of the following ingredients, or compounds of a similar nature: a binder such as microcrystalline cellulose, gum tragacanth or gelatin; an excipient such as starch or lactose, a disintegrating agent such as alginic acid, Primogel, or corn starch; a lubricant such as magnesium stearate or Sterotes; a glidant such as colloidal silicon dioxide; a sweetening agent such as sucrose or saccharin; or a flavoring agent such as peppermint, methyl salicylate, or orange flavoring.

For administration by inhalation, the compounds are delivered in the form of an aerosol spray from pressured container or dispenser which contains a suitable propellant, e.g., a gas such as carbon dioxide, or a nebulizer. Systemic administration can also be by transmucosal or transdermal means. For transmucosal or transdermal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art, and include, for example, for transmucosal administration, detergents, bile salts, and fusidic acid derivatives. Transmucosal administration can be accomplished through the use of nasal sprays or suppositories. For transdermal administration, the active compounds are formulated into ointments, salves, gels, or creams as generally known in the art.

The compounds can also be prepared in the form of suppositories (e.g., with conventional suppository bases such as cocoa butter and other glycerides) or retention enemas for rectal delivery.

In one embodiment, the active compounds are prepared with carriers that will protect the compound against rapid elimination from the body, such as a controlled release formulation, including implants and microencapsulated delivery systems. Biodegradable, biocompatible polymers can be used, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid.

Methods for preparation of such formulations will be apparent to those skilled in the art. The materials can also be obtained commercially from Alza Corporation and Nova Pharmaceuticals, Inc. Liposomal suspensions (including liposomes targeted to particular cells, e.g., a pituitary cell) can also be used as pharmaceutically acceptable carriers. These can be prepared according to methods known to those skilled in the art, for example, as described in US Pat. No. 4,522,811. It is advantageous to formulate oral or parenteral compositions in dosage unit form for ease of administration and uniformity of dosage. Dosage unit form as used herein refers to physically discrete units suited as unitary dosages for the subject to be treated; each unit containing a predetermined quantity of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier.

Toxicity and therapeutic efficacy of the antagonists (or pharmaceutical compositions thereof) can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., for determining the LD50 (the dose lethal to 50% of the population) and the ED50 (the dose therapeutically effective in 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index and it can be expressed as the ratio LD50/ED50. Compounds which exhibit high therapeutic indices are preferred. While compounds that exhibit toxic side effects may be used, care should be taken to design a delivery system that targets such compounds to the site of affected tissue in order to minimize potential damage to uninfected cells and, thereby, reduce side effects.

The data obtained from the cell culture assays and animal studies can be used in formulating a range of dosage for use in humans. The dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration utilized. For any compound used in the method of the invention, the therapeutically effective dose can be estimated initially from cell culture assays. A dose may be formulated in animal models to achieve a circulating plasma concentration range that includes the IC50 (i.e., the concentration of the test compound which achieves a half-maximal inhibition of symptoms) as determined in cell culture. Such information can be used to more accurately determine useful doses in humans. Levels in plasma may be measured, for example, by high performance liquid chromatography.

In some implementations, a therapeutically effective amount of protein or polypeptide (i.e., an effective dosage) ranges from about 0.001 to 30 mg/kg body weight, preferably about 0.01 to 25 mg/kg body weight, more preferably about 0.1 to 20 mg/kg body weight, and even more preferably about 1 to 10 mg/kg, 2 to 9 mg/kg, 3 to 8 mg/kg, 4 to 7 mg/kg, or 5 to 6 mg/kg body weight. The protein or polypeptide can be administered one time per week for between about 1 to 10 weeks, preferably between 2 to 8 weeks, more preferably between about 3 to 7 weeks, and even more preferably for about 4, 5, or 6 weeks. The skilled artisan will appreciate that certain factors may influence the dosage and timing required to effectively treat a subject, including but not limited to the severity of the disease or disorder, previous treatments, the general health and/or age of the subject, and other diseases present. Moreover, treatment of a subject with a therapeutically effective amount of a compound can include a single treatment or, preferably, can include a series of treatments.

For antibody compounds, one preferred dosage is 0.1 mg/kg of body weight (generally 10 mg/kg to 20 mg/kg). If the antibody is to act in the brain, a dosage of 50 mg/kg to 100 mg/kg is usually appropriate. Generally, partially human antibodies and fully human antibodies have a longer half-life within the human body than other antibodies. Accordingly, lower dosages and less frequent administration is often possible. Modifications such as lipidation can be used to stabilize antibodies and to enhance uptake and tissue penetration (e.g., into the brain). A method for lipidation of antibodies is described by Cruikshank et al. ((1997) J. Acquired Immune Deficiency Syndromes and Human Retrovirology 14:193). The present invention encompasses agents which modulate expression or activity.

An agent may, for example, be a small molecule. For example, such small molecules include, but are not limited to, peptides, peptidomimetics (e.g., peptoids), amino acids, amino acid analogs, polynucleotides, polynucleotide analogs, nucleotides, nucleotide analogs, organic or inorganic compounds (i.e., including heteroorganic and organometallic compounds) having a molecular weight less than about 10,000 grams per mole, organic or inorganic compounds having a molecular weight less than about 5,000 grams per mole, organic or inorganic compounds having a molecular weight less than about 1,000 grams per mole, organic or inorganic compounds having a molecular weight less than about 500 grams per mole, and salts, esters, and other pharmaceutically acceptable forms of such compounds. Exemplary doses include milligram or microgram amounts of the small molecule per kilogram of subject or sample weight (e.g., about 1 microgram per kilogram to about 500 milligrams per kilogram, about 100 micrograms per kilogram to about 5 milligrams per kilogram, or about 1 microgram per kilogram to about 50 micrograms per kilogram. It is furthermore understood that appropriate doses of a small molecule may depend upon a number of factors, such as the potency of the small molecule with respect to the expression or activity to be modulated (e.g., affinity for target compound and efficacy) and pharmacokinetic properties. When one or more of these small molecules is to be administered to an animal (e.g., a human) in order to modulate expression or activity of a polypeptide or nucleic acid of the invention, a physician, veterinarian, or researcher may, for example, prescribe a relatively low dose at first, subsequently increasing the dose until an appropriate response is obtained. In addition, it is understood that the specific dose level for any particular animal subject will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, gender, and diet of the subject, the time of administration, the route of administration, the rate of excretion, any drug combination, and the degree of expression or activity to be modulated.

The nucleic acid molecules that modulate the ghrelin pathway can be inserted into vectors and used as gene therapy vectors. Gene therapy vectors can be delivered to a subject by, for example, intravenous injection, local administration (see US Pat.

No. 5,328,470) or by stereotactic injection (see e.g., Chen et al. Proc. Natl. Acad. Sci. USA 91:3054-3057, 1994). The pharmaceutical preparation of the gene therapy vector can include the gene therapy vector in an acceptable diluent, or can comprise a slow release matrix in which the gene delivery vehicle is imbedded. Alternatively, where the complete gene delivery vector can be produced intact from recombinant cells, e.g., retroviral vectors, the pharmaceutical preparation can include one or more cells which produce the gene delivery system.

The pharmaceutical compositions can be included in a container, pack, or dispenser together with instructions for administration. A ghrelin pathway antagonist can be provided in a kit. The kit includes (a) the antagonist, e.g., a composition that includes the antagonist, and (b) informational material. The informational material can be descriptive, instructional, marketing or other material that relates to the methods described herein and/or the use of the antagonist for the methods described herein. For example, the informational material describes methods for administering the antagonist to alter lifespan regulation or at least one symptom of aging or an age related disease.

In one embodiment, the informational material can include instructions to administer the antagonist in a suitable manner, e.g., in a suitable dose, dosage form, or mode of administration (e.g., a dose, dosage form, or mode of administration described herein). In another embodiment, the informational material can include instructions for identifying a suitable subject, e.g., a human, e.g., an adult human. The informational material of the kits is not limited in its form. In many cases, the informational material, e.g., instructions, is provided in printed matter, e.g., a printed text, drawing, and/or photograph, e.g., a label or printed sheet. However, the informational material can also be provided in other formats, such as Braille, computer readable material, video recording, or audio recording. In another embodiment, the informational material of the kit is a link or contact information, e.g., a physical address, email address, hyperlink, website, or telephone number, where a user of the kit can obtain substantive information about the modulator and/or its use in the methods described herein. Of course, the informational material can also be provided in any combination of formats. In addition to the antagonist, the composition of the kit can include other ingredients, such as a solvent or buffer, a stabilizer or a preservative, and/or a second agent for treating a condition or disorder described herein, e.g. increased pancreatic islet mass. Alternatively, the other ingredients can be included in the kit, but in different compositions or containers than the antagonist. In such embodiments, the kit can include instructions for admixing the antagonist and the other ingredients, or for using the modulator together with the other ingredients.

The antagonist can be provided in any form, e.g., liquid, dried or lyophilized form. It is preferred that the antagonist be substantially pure and/or sterile. When the antagonist is provided in a liquid solution, the liquid solution preferably is an aqueous solution, with a sterile aqueous solution being preferred. When the antagonist is provided as a dried form, reconstitution generally is by the addition of a suitable solvent. The solvent, e.g., sterile water or buffer, can optionally be provided in the kit.

The kit can include one or more containers for the composition containing the antagonist. In some embodiments, the kit contains separate containers, dividers or compartments for the antagonist (e.g., in a composition) and informational material. For example, the antagonist (e.g., in a composition) can be contained in a bottle, vial, or syringe, and the informational material can be contained in a plastic sleeve or packet. In other embodiments, the separate elements of the kit are contained within a single, undivided container. For example, the antagonist (e.g., in a composition) is contained in a bottle, vial or syringe that has attached thereto the informational material in the form of a label. In some embodiments, the kit includes a plurality (e.g., a pack) of individual containers, each containing one or more unit dosage forms (e.g., a dosage form described herein) of the antagonist (e.g., in a composition). For example, the kit includes a plurality of syringes, ampules, foil packets, or blister packs, each containing a single unit dose of the antagonist. The containers of the kits can be air tight and/or waterproof.

The antagonist (e.g., in a composition) can be administered to a subject, e.g., an adult subject, e.g., a subject in need ofpreserved pancreatic islet mass. The method can include evaluating a subject, e.g., to evaluate pancreatic islet mass, and thereby identifying a subject as having increased islet mass or being pre-disposed it.

Treatments

The compounds described herein can be administered to cells in culture, e.g. in vitro or ex vivo, or to a subject, e.g., in vivo, to treat, prevent, and/or diagnose a variety of disorders, including those described herein below.

As used herein, the term "treat" or "treatment" is defined as the application or administration of a compound, alone or in combination with, a second compound to a subject, e.g., a patient, or application or administration of the compound to an isolated tissue or cell, e.g., cell line, from a subject, e.g., a patient, who has a disorder (e.g., a disorder as described herein), a symptom of a disorder, or a predisposition toward a disorder, with the purpose to cure, heal, alleviate, relieve, alter, remedy, ameliorate, improve or affect the disorder, one or more symptoms of the disorder or the predisposition toward the disorder (e.g., to prevent at least one symptom of the disorder or to delay onset of at least one symptom of the disorder).

As used herein, an amount of a compound effective to treat a disorder, or a "therapeutically effective amount" refers to an amount of the compound which is effective, upon single or multiple dose administration to a subject, in treating a cell, or in curing, alleviating, relieving or improving a subject with a disorder beyond that expected in the absence of such treatment.

As used herein, an amount of a compound effective to prevent a disorder, or a "prophylactically effective amount" of the compound refers to an amount effective, upon single- or multiple-dose administration to the subject, in preventing or delaying the occurrence of the onset or recurrence of a disorder or a symptom of the disorder.

As used herein, the term "subject" is intended to include human and non-human animals. Exemplary human subjects include a human patient having a disorder, e.g., a disorder described herein or a normal subject. The term "non-human animals" of the invention includes all vertebrates, e.g., non-mammals (such as chickens, amphibians, reptiles) and mammals, such as non-human primates, domesticated and/or agriculturally useful animals, e.g., sheep, dog, cat, cow, pig, etc.

Prediabetes

In one aspect, a ghrelin pathway antagonist described herein can be used to preserve pancreatic islet mass in a subject that has, or is predisposed to, prediabetes, and/or complications thereof. Administration of a ghrelin pathway antagonist that preserves pancreatic islet mass may preserve islet function, and thereby maintain or increase insulin sensitivity and/or maintain or decrease insulin levels in the subject. A subject in need of such a treatment may be a subject who has insulin resistance or other symptom of prediabetes .

Prediabetes (e.g., a pre-diabetic stage) is a condition in which blood sugar (glucose) levels are higher than normal, but not high enough to be classified as type 2 diabetes. In a subject with prediabetes, the long-term damage of diabetes may already be starting. In the United States, 54 million adults have prediabetes, according to the American Diabetes Association. Without intervention, prediabetes is likely to become type 2 diabetes in as little as 10 years.

In prediabetes, the pancreatic islet attempts to compensate for increased blood glucose concentrations by increasing the rate of insulin secretion. The islet achieves this by increasing the number of beta (β) cells and thus increasing its mass.

Symptoms. Often, prediabetes has no signs or symptoms, but a subject may have one or more symptoms of type 2 diabetes such as: increased thirst, frequent urination, extreme hunger, unexplained weight loss, fatigue, blurred vision, slow-healing sores or frequent infections. Causes. In prediabetes and diabetes, the pancreas fails to make enough insulin or cells of the body become resistant to the action of insulin, or both. Exactly why this happens is uncertain, although excess fat — especially abdominal fat — and inactivity seem to be important factors.

Risk factors. The same factors that increase the risk of developing type 2 diabetes increase the risk of developing prediabetes, including:

Weight: Being overweight is a primary risk factor for prediabetes. The more fatty tissue a subject has, especially around the abdomen, the more resistant cells become to insulin.

Inactivity: The less active a subject, the greater the risk of prediabetes. Physical activity helps control weight, use glucose as energy, and make cells more sensitive to insulin.

Age: The risk of prediabetes increases with age, especially after age 45. But diabetes is also increasing dramatically among children, adolescents and younger adults.

Family history: The risk of prediabetes increases if a parent or sibling has type 2 diabetes.

Race: Although it's unclear why, people of certain races (including blacks, Hispanics, American Indians and Asian- Americans) are more likely to develop prediabetes.

Gestational diabetes: If a subject developed gestational diabetes when pregnant, the risk of later developing diabetes increases. If a subject gave birth to a baby who weighed more than 9 pounds, the subject is also at increased risk of diabetes. Polycystic ovary syndrome: For women, having polycystic ovary syndrome increases the risk of diabetes.

Other conditions associated with diabetes include: high blood pressure, high levels of low-density (LDL), or "bad," cholesterol, low levels of high-density lipoprotein (HDL), or "good," cholesterol, and high levels of triglycerides, another fat in the blood. When these conditions- high blood pressure, high blood sugar and abnormal blood fats- occur together with obesity, they are associated with resistance to insulin. This is often referred to as metabolic syndrome. Diagnosis. Fasting blood sugar test: A blood sample will be taken after a subject fasts for at least eight hours or overnight. With this test, a blood sugar level lower than 100 milligrams per deciliter (mg/dL) is normal. A blood sugar level from 100 to 125 mg/dL is considered prediabetes. This is sometimes referred to as impaired fasting glucose (IFG). A blood sugar level of 126 mg/dL or higher may indicate diabetes. Oral glucose tolerance test. A blood sample will be taken after a subject fasts for at least eight hours or overnight. The subject then drinks a sugary solution, and their blood sugar level is measured again after two hours. A blood sugar level less than 140 mg/dL is normal. A blood sugar level from 140 to 199 mg/dL is considered prediabetes. This is sometimes referred to as impaired glucose tolerance (IGT). A blood sugar level of 200 mg/dL or higher may indicate diabetes.

Treatment. Treatments for prediabetes include: eating healthy foods, increased physical activity, weight loss, and medication. Medications include one or more ghrelin pathway antagonists described herein, alone or in combination with another treatment, e.g., a treatment used to treat prediabetes or diabetes (e.g., see below). Examples of treatments include: metformin (Glucophage) and acarbose (Precose).

Metabolic Disorders and Diabetes

In one aspect, a ghrelin pathway antagonist described herein can be used to preserve pancreatic islet mass in a subject that has, or is predisposed to, a metabolic disorder, such as insulin-resistance, a pre-diabetic state, type 2 diabetes, and/or complications thereof. Administration of a ghrelin pathway antagonist that preserves pancreatic islet mass may preserve islet function, and thereby maintain or increase insulin sensitivity and/or maintain or decrease insulin levels in the subject. A subject in need of such a treatment may be a subject who has insulin resistance or other precursor symptom of type 2 diabetes, who has type 2 diabetes, or who is likely to develop any of these conditions. For example, the subject may be a subject having insulin resistance, e.g., having high circulating levels of insulin and/or associated conditions, such as hyperlipidemia, dyslipogenesis, hypercholesterolemia, impaired glucose tolerance, high blood glucose sugar level, other manifestations of syndrome X, hypertension, atherosclerosis and lipodystrophy. In an exemplary embodiment, a ghrelin pathway antagonist may be administered as a combination therapy for treating or preventing a metabolic disorder or type 2 diabetes. For example, one or more ghrelin pathway antagonists may be administered in combination with one or more anti-diabetic agents. Exemplary antidiabetic agents include, for example, an aldose reductase inhibitor, a glycogen phosphorylase inhibitor, a sorbitol dehydrogenase inhibitor, a protein tyrosine phosphatase IB inhibitor, a dipeptidyl protease inhibitor, insulin (including orally bioavailable insulin preparations), an insulin mimetic, metformin, acarbose, a peroxisome proliferator- activated receptor- [gamma] (PPAR- [gamma]) ligand such as troglitazone, rosaglitazone, pioglitazone or GW- 1929, a sulfonylurea, glipazide, glyburide, or chlorpropamide wherein the amounts of the first and second compounds result in a therapeutic effect. Other anti-diabetic agents include a glucosidase inhibitor, a glucagon-like peptide- 1 (GLP-I), insulin, a PPAR [alpha]/[gamma] dual agonist, a meglitimide and an [alpha] P2 inhibitor. In an exemplary embodiment, an anti-diabetic agent may be a dipeptidyl peptidase IV (DP-IV or DPP-IV) inhibitor, such as, for example LAF237 from Novartis (NVP DPP728; 1 -[[[2-[(5- cyanopyridin-2-yl)amino] ethyl] amino] acetyl] -2- cyano-(S)- pyrrolidine) or MK-04301 from Merck.

Diabetes. Type 2 diabetes, once known as adult-onset or noninsulin-dependent diabetes, is a chronic condition that affects the way a subject's body metabolizes glucose. Type 2 diabetes also includes "non-obese type 2" and "obese type 2." Type 2 diabetes is often preventable, but the condition is on the rise, fueled largely by the current obesity epidemic.

In type 2 diabetes, a subject's body is resistant to the effects of insulin, or the body produces some, but not enough, insulin to maintain a normal glucose level. Left uncontrolled, the consequences of type 2 diabetes can be life-threatening.

As a subject progresses through a pre-diabetic stage and towards developing type 2 diabetes, the islet cells are less able, or no longer able, to compensate for the impaired islet cell function, and islet cell mass is reduced by increased apoptosis and/or necrosis. Type 1 diabetes is a similar, although much less common, condition in which the pancreas produces little or no insulin.

Management of diabetes includes eating healthy foods, engaging in regular physical activity, and maintaining a healthy weight. If diet and exercise aren't enough, diabetes medications or insulin therapy may be needed. Symptoms. Type 2 diabetes symptoms include: increased thirst and frequent urination, extreme hunger, weight loss, fatigue, blurred vision, slow-healing sores or frequent infections, and acanthosis nigricans- is a sign of insulin resistance.

Causes. In prediabetes and diabetes, the pancreas fails to make enough insulin or cells of the body become resistant to the action of insulin, or both. Exactly why this happens is uncertain, although excess fat — especially abdominal fat — and inactivity seem to be important factors.

Risk factors.

Weight: Being overweight is a primary risk factor for type 2 diabetes. The more fatty tissue you have, the more resistant your cells become to insulin. Inactivity: The less active a subject, the greater the risk of type 2 diabetes.

Physical activity helps control weight, uses glucose as energy and makes cells more sensitive to insulin.

Family history: The risk of type 2 diabetes increases if a parent or sibling has type 2 diabetes. Race: Although it's unclear why, people of certain races- including blacks, Hispanics, American Indians and Asian Americans- are more likely to develop type 2 diabetes.

Age: The risk of type 2 diabetes increases with age, especially after age 45. But type 2 diabetes is increasing dramatically among children, adolescents and younger adults.

Prediabetes: Prediabetes is a condition in which your blood sugar level is higher than normal, but not high enough to be classified as type 2 diabetes. Left untreated, prediabetes often progresses to type 2 diabetes. Gestational diabetes: If a subject developed gestational diabetes when pregnant, the risk of developing type 2 diabetes increases. If a subject gave birth to a baby weighing more than 9 pounds, she is also at risk of type 2 diabetes.

Diagnosis.

Random blood sugar test. A blood sample will be taken at a random time. Regardless of when the subject last ate, a random blood sugar level of 200 milligrams per deciliter (mg/dL) or higher suggests diabetes.

Fasting blood sugar test. A blood sample will be taken after an overnight fast. A fasting blood sugar level between 70 and 100 mg/dL is normal. A fasting blood sugar level from 100 to 125 mg/dL is considered prediabetes, which indicates a high risk of developing diabetes. If it's 126 mg/dL or higher on two separate tests, the subject is diagnosed with diabetes.

Further, type 2 diabetes can be characterized by (1) reduced pancreatic -beta-islet- cell secretion of insulin such that less than necessary amounts of insulin are produced to keep blood glucose levels in balance and/or (2) "insulin resistance," wherein the body fails to respond normally to insulin. (See US Pat. Nos. 5,266,561 and 6,518,069). For example, glucose-stimulated insulin levels typically fail to rise above 4.0 nmol/L. (US Pat. No. 5,266,561)

Molecular indications of type 2 diabetes include islet amyloid deposition in the pancreases. Treatments for diabetes include: Sulfonylureas. These medications lower blood glucose by stimulating the pancreas to release more insulin. This class includes Dymelor, Diabinese, Orinase, Tolinase, Glucotrol, Glucotrol XL, DiaBeta, Micronase, Glynase PresTab and Amaryl.

Biguanides. These medications improve insulin's ability to move glucose into cells especially into the muscle cells. They also prevent the liver from releasing stored glucose. Examples include metformin (Glucophage, Glucophage XR, Riomet, Fortamet and Glumetza).

Thiazolidinediones. These medications improve insulin's effectiveness in muscle and in fat tissue. They lower the amount of glucose released by the liver and make fat cells more sensitive to the effects of insulin. Actos and Avandia are the two drugs of this class.

Alpha-glucosidase inhibitors. These drugs block enzymes that help digest starches, slowing the rise in blood glucose. This class includes Precose and Glyset.

Meglitinides. These diabetes medicines lower blood glucose by stimulating the pancreas to release more insulin. This class includes Prandin (rapaglinide) and Starlix (nateglinide).

Dipeptidyl peptidase IV (DPP-IV) inhibitors. The DPP-IV inhibitors work to lower blood sugar in patients with type 2 diabetes by increasing insulin secretion from the pancreas and reducing sugar production. The medication may be taken alone or with other medications such as metformin. This class includes Januvia.

Combination therapy. There are several combination diabetes pills that combine two medications into one tablet. One example of this is Glucovance, which combines glyburide (a sulfonylurea) and metformin. Others include Metaglip, which combines glipizide (a sulfonylurea) and metformin, and Avandamet which utilizes both metformin and rosiglitazone (Avandia) in one pill. An exemplary example is PRAND IMET™, a Replaglinide and Metformin Fixed-Dose Combination Tablet.

One or more of these medications can be used with a ghrelin pathway antagonist described herein. Obesity

In one aspect, a ghrelin pathway antagonist described herein can be used to decrease pancreatic islet mass in a subject that has, or is predisposed to, obesity. Administration of a ghrelin pathway antagonist that decreases pancreatic islet mass may improve islet function, and thereby limit or prevent complications associates with obesity in the subject. A subject in need of such a treatment may be a subject who has obesity or a precursor of obesity (e.g., the subject is overweight and/or poor diet and/or sedentary lifestyle), or who is likely to develop obesity (e.g., the subject is overweight and/or poor diet and/or sedentary lifestyle, and/or has a genetic predisposition to obesity). Obesity is a disease in which excess body fat has accumulated to such an extent that health may be negatively affected. It is commonly defined as a body mass index (weight divided by height squared) of 30 kg/m or higher. This distinguishes it from being overweight as defined by a BMI of between 25-29.9. Many studies show an association between excessive body weight and various diseases, particularly cardiovascular diseases, diabetes mellitus type 2, sleep apnea, cancer and osteoarthritis. As a result, obesity has been found to reduce life expectancy.

Treatments include diet and exercise. Medications may also be used. Anti- obesity drugs can operate through one or more of the following mechanisms:

Suppression of the appetite. Epilepsy medications and catecholamines and their derivatives (such as amphetamine-based drugs) are the main tools used for this. Drugs blocking the cannabinoid receptors may also be used; as well as stimulants (e.g., dexedrine, digoxin);

Increase of the body's metabolism;

Interference with the body's ability to absorb specific nutrients in food. For example, Orlistat blocks fat breakdown and thereby prevents fat absorption; fiber supplements such as glucomannan and guar gum have been used for the purpose of inhibiting digestion and lowering caloric absorption;

Specific medications include: Orlistat, Sibutramine, Metformin, Byetta, Symlin, Rimonabant. One or more of these medications can be used with a ghrelin pathway antagonist described herein. Patient Selection

The methods described herein include methods of administering a ghrelin pathway antagonist (e.g., a ghrelin pathway antagonist described herein) to a subject in need of such treatment, e.g., wherein the subject has prediabetes, diabetes, other metabolic disorder, obesity, insulin resistance or is developing insulin resistance characterized as described herein.

The methods described herein include administering a ghrelin pathway antagonist to a subject in need of such treatment, e.g., wherein the subject has prediabetes, diabetes, other metabolic disorder, obesity, insulin resistance or is developing insulin resistance characterized as described herein. The method includes evaluating a subject to determine if the subject is in need of such treatment, e.g., wherein the subject has prediabetes, diabetes, other metabolic disorder, obesity, insulin resistance or is developing insulin resistance characterized as described herein, and if the subject is in need of such treatment then treating or instructing to treat the subject with a ghrelin pathway antagonist.

Methods are described herein for selecting a subject on the basis that the subject is in need of such treatment, e.g., wherein the subject has prediabetes, diabetes, other metabolic disorder, obesity, insulin resistance or is developing insulin resistance characterized as described herein, and administering a ghrelin pathway antagonist to that subject. Methods are also described for selecting a pharmaceutical agent (e.g., a drug) for treating a subject suffering from one of these conditions, for example, prediabetes, diabetes, other metabolic disorder, obesity, insulin resistance or is developing insulin resistance characterized as described herein. The method includes evaluating a subject to determine if the subject suffering from prediabetes, diabetes, other metabolic disorder, obesity, insulin resistance or is developing insulin resistance characterized as described herein, and if the subject is suffering from one of these conditions, selecting a ghrelin pathway antagonist to treat the subject (e.g., selecting a ghrelin pathway antagonist on the basis that the subject is suffering from one of these conditions). Exemplary methods of determining whether the subject is suffering from one of these conditions (e.g., methods for diagnosing) are provided herein. The methods described herein include methods of administering a ghrelin pathway antagonist (e.g., a ghrelin pathway antagonist described herein) to a subject in need of such treatment, e.g., wherein one or more of: the difference between the pancreatic islet mass of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and/or the difference between the pancreatic islet function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreas weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the body weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet triglyceride content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet lipid content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and the difference between the pancreas function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. The methods described herein include administering a ghrelin pathway antagonist to a subject in need of such treatment, e.g., the subject is in need of such treatment if one or more of the following are present: the difference between the pancreatic islet mass of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and/or the difference between the pancreatic islet function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreas weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the body weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet triglyceride content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet lipid content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and the difference between the pancreas function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%. The method includes evaluating a subject to determine if the subject is in need of such treatment, e.g., determining if one or more of the above differences exists, and if the subject is in need of such treatment, then treating or instructing to treat the subject with a ghrelin pathway antagonist.

Methods are described herein for selecting a subject on the basis that the subject is in need of such treatment, e.g., if one or more of the following are present: the difference between the pancreatic islet mass of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and/or the difference between the pancreatic islet function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreas weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the body weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet triglyceride content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet lipid content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and the difference between the pancreas function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%, and if one or more are present, administering a ghrelin pathway antagonist to that subject. Methods are also described for selecting a pharmaceutical agent (e.g., a drug) for treating a subject suffering from one of these conditions. The method optionally includes evaluating a subject to determine if one or more of the following is present: the difference between the pancreatic islet mass of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and/or the difference between the pancreatic islet function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreas weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the body weight of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet triglyceride content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about

25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; the difference between the pancreatic islet lipid content of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%; and the difference between the pancreas function of the subject and that of a standard is greater than about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or about 90%, and if one or more are present, selecting a ghrelin pathway antagonist to treat the subject (e.g., selecting a ghrelin pathway antagonist on the basis that the subject is suffering from one of these conditions). Exemplary methods of determining whether the subject is suffering from one of these conditions are provided herein.

EXAMPLES

Experiments were performed to evaluate the role of the ghrelin pathway in regulating pancreatic islet mass and islet lipid content.

It was hypothesized that when given a high fat diet, ghrelin receptor (GhrR) nullizygous (e.g., knock out (KO)) mice have a lower pancreatic islet cell mass and improved pancreatic islet function compared with wild type (WT) mice on the same diet, and that improved islet function is a result from lower islet lipid content.

Example 1 Methods. Ghrelin Receptor Nullizygous Mouse. The GhrR KO mouse is a model of ghrelin pathway antagonism. The GhrR KO mice were bred from a founder mouse that was heterozygous for GhrR on a C57BL/6 background. The mouse was obtained from Charles River Laboratories (Wilmington, MA). See Longo et al., Regulatory Pept. "Improved insulin sensitivity and metabolic flexibility in ghrelin receptor knockout mice", e-published on March 30, 2008; doi:10.1016/j.regpep.2008.03.011.

Islet Cell Mass. Islet cell mass measurements were performed as follows. Mice were weighed, euthanized, and their pancreas was dissected out and weighed. The pancreas was fixed in formalin and embedded in paraffin. Levels were cut through the pancreas at 4μm intervals. Every 50^th level was placed on a slide and stained with hematoxylin and eosin (H & E). Up to fourteen levels were taken from a single pancreas. Each level is placed on a grid system. Points which cross over an islet are marked. The number of points per level is used for the islet cell mass calculation.

Total Islet Cell Mass = islet points x pancreas weight (mg) total number of reference points

See also, Bock et al., Diabetes 52:1716-1722 (2003).

Method for isolating islets from the pancreas. Mice were euthanized by CO₂ asphyxiation and the pancreas removed. Pancreatic islets were isolated by collagenase digestion at 37⁰C using a physiological saline solution supplemented with ImM CaCl₂,

2mM glucose and equilibrated with CO₂:O₂(5%:95%), pH7.4. Islets were hand-picked using a binocular microscope, washed and microdissected to remove residual acinar material. Batches of 100 islets were placed into 1.5ml Eppendorf microfuge tubes, frozen on dry ice, and stored at -8O⁰C until use. Triglyceride Content. Triglycerides were measured using a Serum Triglyceride

Determination Kit (Sigma, TROlOO). Lipids were extracted from islets using hexane/isopropanol.

Triglyceride Assay Enzymatic Reactions: Lipoprotein Lipase

Triglycerides > Glycerol + Fatty Acids

GK Glycerol + ATP > G- 1 -P +ADP

GPO G- 1 -P + O₂ > DAP + H₂O₂

POD H₂O₂ + 4-AAP +ESPA > Quinoneimine dye + H₂O

G-I-P = glycerol- 1 -phosphate GK = glycerol kinase GPO = glycerol phosphate oxidase DAP = dihydroxyacetone phosphate

4-AAP = 4 aminoantipyrine ESPA = sodium N-ethyl-N-(3-sulfopropyl) m-anisidine POD = peroxidase

Absorbance at 540nm is directly proportional to the triglyceride concentration in the sample.

Preserved Pancreas and Islet Cell Mass in GhrR KO Mice. GhrR WT and KO mice were fed a high fat diet ad libitum for 20 weeks. The body weights, islet cell masses, and pancreas weights for both WT and KO mice were measured. The results are shown in FIGS. 1A-1C. The P values are also shown in each graph. Analysis of covariance (ANCOVA) was performed:

Pancreas Weight

Genotype: F(l-19)=4.32; P=0.051

Body Weight: F(l-19)=2.90; P=0.104

Islet Cell Mass

Genotype: F₍₁_₁₉₎=5.15; P=0.035

Body Weight: F_(M9)=14.27; P=OOOl

As shown in the figure, the average islet cell mass and average pancreas weight differed significantly between the GhrR WT and KO mice, both weights being lower in the GhrR KO mice. Histological analysis demonstrated that islets from KO mice had seemingly normal morphology, as compared to the islets from WT mice, where altered islet morphology was detected (data not shown). Without being bound by theory, it is possible that, in the GhrR KO mice, due to improved insulin sensitivity in the periphery, no expansion of β-cells or increased insulin production is needed; thereby protecting the pancreas.

Decreased Islet Triglyceride Content in GhrR KO Mice. Groups of ten GhrR WT and ten GhrR KO mice were fed a normal chow (NC) or high fat (HF) diet for 24 weeks. The triglyceride content of the islets from both groups of mice was measured. One hundred islets were isolated from each of five mice. This was performed twice (n=2) such that 100 islets were examined from each of the ten mice in each treatment group. The results are shown in FIG. 2. As shown in the figure, GhrR KO mice fed a high fat diet have lower average islet triglyceride content than WT mice fed a high fat diet. Also, average islet triglyceride content was much higher in WT mice fed a high fat diet than WT mice fed normal chow. In contrast, KO mice fed a high fat diet has only slightly higher average triglyceride content than KO mice fed normal chow. Two-way analysis of variance (ANOVA) was also performed, as shown in FIG. 2.

Example 2 Effects of a Ghrelin Pathway Antagonist on Pancreas Weight, Body Weight, and Islet Cell Mass. GhrR WT mice were orally dosed with EX- 1350, a ghrelin pathway antagonist, twice a day for 56 days at a dose of 60 mg per kilogram (mpk). As a negative control, a group of mice were dosed with vehicle alone. Both sets of mice were fed a high fat diet. Body weight and pancreas weight measurements were taken. The results are shown in FIGS. 3 A and 3B. As shown in FIG. 3 A, the difference in average pancreas weight between the two groups was not statistically significant. FIG. 3B shows that the average body weight of the EX-1350 treated group was lower than the control group. The difference was statistically significant (*p<0.035).

Islet cell mass was also measured for both sets of mice. The results are shown in FIG. 4. As shown in the figure, mice treated with EX-1350 had lower islet cell mass than vehicle-treated controls (p= 0.073).

Histological analysis demonstrated that islets from EX-1350-treated mice were smaller than islets of vehicle-treated mice, and the islets from EX-1350-treated mice contained less fat than the islets of vehicle-treated mice (data not shown).

The structure of EX-1350 is:

A number of embodiments of the invention have been described. Nevertheless, it will be understood that various modifications may be made without departing from the spirit and scope of the invention. Accordingly, other embodiments are within the scope of the following claims.

Claims

WHAT IS CLAIMED IS:

1. A method for preserving pancreatic islet mass in a subject, the method comprising: administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves pancreatic islet mass.

2. The method of claim 1 further comprising: evaluating pancreatic islet mass in the subject prior to, during, or after administration of the ghrelin pathway antagonist.

3. The method of claim 1, wherein the ghrelin pathway antagonist prevents or delays an increase in pancreatic islet mass in a subject with prediabetes.

4. The method of claim 1, wherein the ghrelin pathway antagonist prevents or delays a decrease in pancreatic islet mass in a subject with diabetes.

5. The method of claim 1, wherein the subject has insulin resistance.

6. The method of claim 1, wherein the subject has prediabetes or a risk factor thereof.

7. The method of claim 1, wherein the subject has diabetes or a risk factor thereof.

8. The method of claim 1, wherein the subject has obesity or a risk factor thereof.

9. A method for preserving pancreatic islet function in a subject, the method comprising: administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves pancreatic islet function.

10. The method of claim 9 further comprising: evaluating pancreatic islet function in the subject prior to, during, or after administration of the ghrelin pathway antagonist.

11. The method of claim 9, wherein the method decreases or delays glucolipotoxicity, lipotoxicity, oxidative stress, or dysregulation of triglyceride or FFA cycling.

12. The method of claim 9, wherein the subject has insulin resistance.

13. The method of claim 9, wherein the subject has prediabetes or a risk factor thereof.

14. The method of claim 9, wherein the subject has diabetes or a risk factor thereof.

15. The method of claim 9, wherein the subject has obesity or a risk factor thereof.

16. A method for preserving pancreas weight in a subject, the method comprising: administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves pancreas weight.

17. The method of claim 16 further comprising: evaluating pancreas weight in the subject prior to, during, or after administration of the ghrelin pathway antagonist.

18. The method of claim 16, wherein the subject has insulin resistance.

19. The method of claim 16, wherein the subject has prediabetes or a risk factor thereof.

20. The method of claim 16, wherein the subject has diabetes or a risk factor thereof.

21. The method of claim 16, wherein the subject has obesity or a risk factor thereof.

22. A method for preserving body weight in a subject, the method comprising: administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves body weight.

23. The method of claim 22 further comprising: evaluating body weight in the subject prior to, during, or after administration of the ghrelin pathway antagonist.

24. The method of claim 22, wherein the subject has insulin resistance.

25. The method of claim 22, wherein the subject has prediabetes or a risk factor thereof.

26. The method of claim 22, wherein the subject has diabetes or a risk factor thereof.

27. The method of claim 22, wherein the subject has obesity or a risk factor thereof.

28. A method for preserving pancreatic islet triglyceride (TG) content in a subject, the method comprising: administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves pancreatic islet TG content.

29. The method of claim 28 further comprising: evaluating pancreatic islet TG content in the subject prior to, during, or after administration of the ghrelin pathway antagonist.

30. The method of claim 28, wherein the subject has insulin resistance.

31. The method of claim 28, wherein the subject has prediabetes or a risk factor thereof.

32. The method of claim 28, wherein the subject has diabetes or a risk factor thereof.

33. The method of claim 28, wherein the subject has obesity or a risk factor thereof.

34. A method for preserving pancreatic islet lipid content in a subject, the method comprising: administering a ghrelin pathway antagonist to the subject, wherein the ghrelin pathway antagonist preserves pancreatic islet lipid content.

35. The method of claim 34 further comprising: evaluating pancreatic islet lipid content in the subject prior to, during, or after administration of the ghrelin pathway antagonist.

36. The method of claim 34, wherein the ghrelin pathway antagonist prevents or delays an increase in pancreatic islet lipid content in a subject with prediabetes.

37. The method of claim 34, wherein the ghrelin pathway antagonist prevents or delays an increase in pancreatic islet lipid content in a subject with diabetes.

38. The method of claim 34, wherein the subject has insulin resistance.

39. The method of claim 34, wherein the subject has prediabetes or a risk factor thereof.

40. The method of claim 34, wherein the subject has diabetes or a risk factor thereof.

41. The method of claim 34, wherein the subject has obesity or a risk factor thereof.

42. The method of claim any one of claims 1-41, wherein the ghrelin pathway antagonist comprises a non-proteinaceous molecule.

43. The method of any one of claims 1-41, wherein the ghrelin pathway antagonist comprises a peptide, polypeptide, protein, antibody, antibody fragment, peptidomimetic, peptoid, nucleic acid, or other chemical compound or a combination thereof.

44. The method of claim 43, wherein the ghrelin pathway antagonist comprises a protein.

45. The method of claim 44, wherein the protein is an antibody, a peptide, or a molecule which binds to a regulatory element for the subject component.

46. The method of claim 43, wherein the ghrelin pathway antagonist comprises a nucleic acid.

47. The method of claim 46, wherein the nucleic acid comprises a nucleic acid aptamer, RNAi, an antisense RNA, or a ribozyme molecule.

48. The method of any one of claims 1-41, wherein the ghrelin pathway antagonist binds to ghrelin or the ghrelin receptor (GHS-R).

49. The method of any one of claims 1-41, wherein the ghrelin pathway antagonist is ghrelin receptor antagonist [D-Lys ]-GHRP-6.

50. The method of any one of claims 1-41, wherein the ghrelin pathway antagonist is used in combination with another therapeutic agent.

51. The method of any one of claims 1-41, wherein the ghrelin pathway antagonist is used in combination with another ghrelin pathway antagonist.

52. The method of any one of claims 1-41, wherein the ghrelin pathway antagonist is used in combination with a treatment for obesity.

53. The method of any one of claims 1-41, wherein the ghrelin pathway antagonist is used in combination with a treatment for metabolic syndrome.

54. The method of any one of claims 1-41, wherein the ghrelin pathway antagonist is used in combination with a treatment for prediabetes.

55. The method of any one of claims 1-41, wherein the ghrelin pathway antagonist is used in combination with a treatment for diabetes.

56. The method of any one of claims 1-41, wherein the ghrelin pathway antagonist is a compound of formula (I)

formula (I) wherein,

R¹ is hydrogen, aryl, heteroaryl, arylalkyl, heteroarylalkyl, cyclyl, cyclylalkyl, heterocyclyl, heterocyclylalkyl, alkyl, alkenyl, alkynyl, or R¹ can be taken together with R² or R³ to form a ring; each of which is optionally substituted with 1-4 R⁶; k' is a bond, O, C(O), C(O)O, OC(O), C(O)NR³, NR³C(O), S, SO, SO₂, CR²=CR², or C≡C; n is 0-6, preferably 1-3;

R² is hydrogen, Ci-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl; or R² can be taken together with R¹ to form a ring;

A is

_R7a _R7a

-|-(CH₂)_X C (CH₂)_y-|- -|— N— (CH₂)_X C (CH₂)_y-<-

_R7b _R8 _R7b

x and y are each independently 0-6;

M is aryl, heteroaryl, cyclyl, or heterocyclyl, each of which is optionally substituted with 1-4 R⁹;

Y is a monocyclic aryl or monocyclic heteroaryl; each of which is optionally substituted with 1-4 R¹⁰; each R⁶ and R⁶ are independently halo, alkyl, alkenyl, alkynyl, cyclyl, heterocyclyl, aryl, heteroaryl, alkoxy, haloalkyl, haloalkyloxy, haloalkylthio, acetyl, ccyyaannco, nitro, hydroxy, oxo, C(O)OR², OC(O)R², N(R³)₂, C(O)N(R³)₂, NR³C(O)R², or

SR²;

R⁸ is hydrogen or Ci-C₆ alkyl, or R⁸ can be joined with R⁴, R⁵, R^7a or R^7b to form a ring;

R⁹ is halo, alkyl, cyclyl, heterocyclyl, aryl, heteroaryl, alkoxy, haloalkyl, haloalkyloxy, haloalkylthio, acetyl, cyano, nitro, hydroxy, oxo, C(O)OR², OC(O)R², N(R²)₂, C(O)N(R²)₂, NR²C(O)R², SR²; each R¹⁰ is independently alkyl, alkenyl, alkynyl, halo, cyano, carbonyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, cyclyl, cyclylalkyl, alkoxy, alkoxyalkyl, aryloxy, aryloxyalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, heteroarylalkyl, -OR¹¹, - NR¹¹R^11', -CF₃, -SOR¹², -SO₂R¹², -OC(O)R¹¹, -SO₂NR¹²R^12', -(CH₂)_mR¹⁴ or R¹⁵; each of which is optionally independently substituted with 1-3 R¹⁶; R¹¹ and R¹¹ are each independently hydrogen, alkyl, alkenyl, alkynyl, cyclyl, heterocyclyl, aryl or heteroaryl;

R¹² and R¹² are each independently hydrogen, alkyl, alkenyl, alkynyl, alkylthioalkyl, alkoxyalkyl, aryl, arylalkyl, heterocyclyl, heteroaryl, heteroarylalkyl, heterocycloalkyl or cyclyl, cyclylalkyl, or R¹² and R¹² taken together can be cyclized to form -(CH₂)_qX(CH₂)_s-; wherein each R¹² and R¹² may independently optionally be substituted with 1 to 3 substituents selected from the group consisting of halogen, OR¹¹, alkoxy, heterocycloalkyl, -NR¹¹C(O)NR¹¹R^11', -C(O)NR¹¹R^11', -NR¹¹C(O)R^11', -CN, oxo, - NR¹¹SO₂R^11', -OC(O)R¹¹, -SO₂NR¹¹R^11', -SOR¹³, -S(O)₂R¹³, -COOH and -C(O)OR¹³; each R¹³ is independently alkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl, each of which may optionally be substituted with -(CH₂)_w0H; each R¹⁴ is independently alkoxy, alkoxycarbonyl, -C(O)NR¹²R^12', -NR¹¹R^11', - C(O)R¹², -NR¹¹C(O)NR¹¹R^11' or -N-heteroaryl; each R¹⁵ is independently -(CH₂)_PN(R¹²)C(O)R^12', -(CH₂)_PCN, - (CH₂)pN(R¹²)C(O)OR^12', -(CH₂)pN(R¹²)C(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂R¹², - (CH₂)_PSO₂NR¹²R^12', -(CH₂)_PC(O)NR¹²R^12', -(CH₂)_PC(O)OR¹², -(CH²)_POC(O)OR¹², - (CH₂)_POC(O)R¹², -(CH₂)_POC(O)NR¹²R^12', -(CH₂)_PN(R¹²)SO₂NR¹²R^12', -(CH₂)_POR¹², - (CH₂)_pOC(O)N(R¹²)(CH₂)_mOH, -(CH₂)_PSOR¹², -(CH₂)_PSO₂R¹², -(CH₂)_pNR^πRⁿ or - (CH₂)_pOCH₂C(O)N(R¹²)(CH₂)_mOH; each R¹⁶ is independently halo, alkyl, alkenyl, alkynyl, alkoxy, - (CH₂)_PNR¹¹C(O)NR¹¹R^11', -(CH₂)_PC(O)NR¹¹R^11', -(CH₂)_PNR¹¹C(O)R^11', -CN, - (CH₂)pNRⁿSO₂R^n', -(CH₂)_POC(O)R¹ \ -(CH₂)_PSO₂NR¹¹R^11', -(CH₂)_PSOR¹³, - (CH₂)_pC00H or -(CH₂)_PC(O)OR¹³;

X is CR¹¹R^11', O, S, S(O), S(O)₂, or NR¹¹; m is an integer between 1 and 6; p is an integer from O to 5. q and s are each independently an integer between 1 and 3; and w is an integer between O and 5.