Methods and materials for treating chronic heart disorders

Abstract

Methods and materials for treating chronic cardiac disorders are provided herein. For example, methods and materials for using a dual guanylate cyclase A and B activator (e.g., a polypeptide having the sequence set forth in SEQ ID NO:1) to treat hypertrophic cardiomyopathy and other cardiac conditions, such as conditions associated with ventricular hypertrophy, atrial hypertrophy, ventricular remodeling, atrial remodeling, systolic and / or diastolic dysfunction, heart failure (e.g., heart failure with reduced ejection fraction, heart failure with mildly reduced ejection fraction, and heart failure with preserved ejection fraction), and other forms of chronic heart disease (e.g., Fabry disease, Noonan syndrome, Pompe disease, PRKAG2-related cardiomyopathy, Danon disease, Friedrich ataxia cardiomyopathy, amyloidosis, or desminopathy) are provided herein.

Description

[0001] Attorney Docket No. 07039-2327WO1

[0002] 2024-159

[0003] METHODS AND MATERIALS FOR TREATING CHRONIC HEART DISORDERS

[0004] CROSS-REFERENCE TO RELATED APPLICATIONS

[0005] This application claims priority from U.S. Provisional Application Serial No. 63 / 705.163, filed October 9, 2024. The disclosure of the prior application is considered part of (and is incorporated by reference in) the disclosure of this application.

[0006] STATEMENT AS TO FEDERALLY SPONSORED RESEARCH

[0007] This invention was made with government support under HL132854, HL135790, and HL171847 awarded by the National Institutes of Health. The government has certain rights in the invention.

[0008] SEQUENCE LISTING

[0009] This application contains a Sequence Listing that has been submitted electronically as an XML file named 07039-2327WOl_SL_ST26.xml. The XML file, created on August 19, 2025, is 15,496 bytes in size. The material in the XML file is hereby incorporated by reference in its entirety'.

[0010] TECHNICAL FIELD

[0011] This document relates to methods and materials for treating mammals having heart related disorders. For example, this document provides methods and materials for using a dual guanylate cyclase A and B activator to treat mammals having hypertrophic cardiomyopathy (HCM) and / or other chronic heart conditions, such as heart failure (e.g.. heart failure with reduced ejection fraction (HFrEF), heart failure with mildly reduced ejection fraction (HFmrEF), and heart failure with preserved ejection fraction (HFpEF)), conditions associated with ventricular and / or atrial hypertrophy and / or remodeling (e.g., left ventricular hypertrophy and remodeling, right ventricular hypertrophy and remodeling, left atrial hypertrophy and remodeling, and / or right atrial hypertrophy and remodeling), systolic and / or diastolic dysfunction, cardiomyopathy, and / or other heart diseases including HCM phenocopies (e.g., Fabry disease, Noonan syndrome, Pompe disease, PRKAG2-related cardiomyopathy, Danon disease, Friedrich ataxia cardiomyopathy, amyloidosis, and / or desminopathy). Attorney Docket No. 07039-2327WO1

[0012] 2024-159

[0013] BACKGROUND

[0014] Cardiac hypertrophy is defined as an increase in the mass or thickness of the walls and / or chambers of the heart. Cardiac hypertrophy can be due to primary abnormalities in the heart, such as genetic mutations that cause HCM, dilated cardiomyopathy (DCM), and / or ischemic cardiomyopathy (ICM). Cardiac hypertrophy also can occur due to secondary abnormalities, such as hypertension, obesity, diabetes, valvular heart disease (e.g., aortic stenosis, aortic regurgitation, mitral stenosis, or mitral regurgitation), or secondary7to adverse myocardial remodeling following myocardial infarction (Nakamura and Sadoshima, Nat. Rev. Cardiol., 15(7):387-407 (2018)). Although some forms of cardiac hypertrophy may be considered an early adaptive response to stress, cardiac hypertrophy is a known risk factor for adverse cardiovascular outcomes in multiple diseases, including hypertension, aortic stenosis, HCM, DCM, ICM, and congestive heart failure (CHF) (Clarke et al., Eur. Heart. J. 41 :ehaa946-2938 (2020); Gonzales et al., JACC. Cardiovasc. Interv. 13: 1329-1339 (2020); Nakamura et al., Nat. Re. 15:387-407 (2018); Ommen et al., Circul. 149:el239-el 311 (2024); Tsao et al., J. Am. Heart. Assc. 4:e002188 (2024); Verdecchia et al., Circul. 104:2039-2044 (2001)). Therefore, cardiac hypertrophy represents a significant public health burden.

[0015] HCM is the most common inherited cardiac disorder. Hallmark features of HCM include cardiac hypertrophy or thickening of the heart muscle, fibrosis and stiffening of the heart, and abnormal relaxation of the heart. About half of HCM patients develop “obstruction,” which impedes the flow' of blood out of heart during contraction. The most common symptoms in people with HCM are shortness of breath, heart palpitations, and chest discomfort, which result in fatigue and exercise intolerance. Shortness of breath in HCM can be due to congestion caused by retention of salt and water by the kidney. Complications of HCM include heart failure, arrhythmia, stroke, and early death.

[0016] The historical cornerstones of HCM management were controlling the symptoms and preventing sudden death due to arrhythmia. First line therapies for HCM. such as beta blockers and calcium channel blockers, can lower the heart rate and reduce the force of contraction, the degree of obstruction, and the oxygen demand of the heart. However, the use of these drugs is not based on high quality evidence, and they do not directly target the disease itself. If patients remain symptomatic despite these first-line therapies, cardiac myosin inhibitors may be used. Cardiac myosin inhibitors (e.g.. mavacamten) can reduce Attorney Docket No. 07039-2327WO1

[0017] 2024-159 the force of contraction at the cellular level. Further, cardiac myosin inhibitors only target cardiac myocytes, and therefore they do not have direct therapeutic effects in other cells and organs (e.g., cardiac fibroblasts, macrophages, and the kidney) that play a role in patients with symptomatic HCM and HCM disease progression.

[0018] SUMMARY

[0019] This document is based, at least in part, on the identification of dual guanylate cyclase A and B activators that can be used to treat mammals having, or at risk of having, chronic cardiovascular disorders. As demonstrated herein, a dual guanylate cyclase A and B activator (e.g., MV81) can induce a dose dependent increase in cyclic guanosine monophosphate (cGMP) levels (a measure of guanylate cyclase receptor A and / or B target engagement) in human cardiomyocytes, human cardiac fibroblasts, and human THP-1 macrophages. MV81 treatment also can increase plasma and urine cGMP levels and urinary sodium excretion in rats. In addition, administration of MV81 to a mouse model of HCM and a mouse model of heart failure (HF) and cardiac remodeling can modify (e.g., reduce, prevent, or reverse) left ventricular thickening, and administration to normal wild type mice can lead to reduced expression of genes related to hypertrophy and fibrosis.

[0020] This document provides methods and materials for treating mammals having heart related disorders (chronic heart disorders). For example, this document provides methods and materials for using a dual guanylate cyclase A and B receptor activator (e.g., MV81) to treat mammals having chronic heart conditions in which guanylate cyclase A (GC-A) and guanylate cyclase B (GC-B) are present in heart cells of the mammal. In some cases, this document provides methods and materials for using a dual guanylate cyclase A and B receptor activator (e.g., MV81) to treat mammals having HCM or other conditions, such as cardiac disorders associated with ventricular and / or atrial hypertrophy and / or remodeling, systolic and / or diastolic dysfunction, other heart diseases such as HCM phenocopies (e.g., Fabry disease, Noonan syndrome, Pompe disease, PRKAG2-related cardiomyopathy. Danon disease. Friedrich ataxia cardiomyopathy, amyloidosis, and desminopathy), and heart failure (e.g., HFrEF, HFmrEF, and HFpEF).

[0021] In a first aspect, this document provides a method for treating a chronic heart condition. The method can include, consist essentially of, or consist essentially of administering, to a mammal identified as having the chronic heart condition, a Attorney Docket No. 07039-2327WO1

[0022] 2024-159 composition that contains a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five. six. seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat. a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The method can include administering the composition to the mammal in an amount effective to reduce one or more symptoms of the chronic heart condition in the mammal. The one or more symptoms of the chronic heart condition can be selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, or edema. The chronic heart condition can be HCM. The chronic heart condition can be associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g., HFrEF, HFmrEF, and HFpEF), or any combination thereof, or can be a HCM phenocopy. The composition can include a natriuretic peptide containing the amino acid sequence set forth in SEQ ID NO: 1. The composition can include a natriuretic peptide containing the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0023] In another aspect, this document provides a method for reducing inflammation in a mammal having a chronic heart condition. The method can include, consist essentially of, or consist essentially of administering, to a mammal identified as having the chronic heart condition, a composition that contains a natriuretic peptide containing the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, a horse, a cow-, a sheep, a goat, a rabbit, a mouse, or a rat. The chronic heart condition can be HCM. The chronic heart condition can be associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g., HFrEF. Attorney Docket No. 07039-2327WO1

[0024] 2024-159

[0025] HFmrEF. and HFpEF). or any combination thereof, or can be a HCM phenocopy. The composition can include a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1. The composition can include a natriuretic peptide containing the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0026] In another aspect, this document provides a method for reducing TGF-beta levels or TGF-beta activity in a mammal having a chronic heart condition. The method can include, consist essentially of, or consist essentially of administering, to a mammal identified as having the chronic heart condition, a composition that contains a natriuretic peptide containing the amino acid sequence set forth in SEQ ID NO:1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO:1 are maintained and are the only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The chronic heart condition can be HCM. The chronic heart condition can be associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g., HFrEF, HFmrEF. and HFpEF). or any combination thereof, or can be a HCM phenocopy. The composition can include a natriuretic peptide containing the amino acid sequence set forth in SEQ ID NO: 1. The composition can include a natriuretic peptide containing the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0027] In another aspect, this document provides a method for reducing ventricular hypertrophy in a mammal having a chronic heart condition. The method can include, consist essentially of, or consist essentially of administering, to a mammal identified as having the chronic heart condition, a composition that contains a natriuretic peptide containing the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the Attorney Docket No. 07039-2327WO1

[0028] 2024-159 only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The chronic heart condition can be HCM. The chronic heart condition can be associated with left ventricular hypertrophy, left ventricular remodeling, right ventricular hypertrophy, right ventricular remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g.. HFrEF, HFmrEF, and HFpEF), or any combination thereof, or can be a HCM phenocopy. The composition can include a natriuretic peptide containing the amino acid sequence set forth in SEQ ID NO: 1. The composition can include a natriuretic peptide containing the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0029] In another aspect, this document features a method for preventing, attenuating, or reversing diastolic dysfunction in a mammal having a chronic heart condition. The method can include, consist of, or consist essentially of administering, to a mammal identified as having the chronic heart condition, a composition that contains a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO:1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The chronic heart condition can be HCM. The chronic heart condition can be associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, cardiomyopathy, heart failure (e.g., HFrEF. HFmrEF, and HFpEF), or any combination thereof, or can be a HCM phenocopy. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO:1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0030] In another aspect, this document features a method for treating a mammal having a chronic heart condition in which guanylate cyclase A (GC-A) and guanylate cyclase B (GC-B) are present in heart cells of the mammal, where the method includes, consists of. Attorney Docket No. 07039-2327WO1

[0031] 2024-159 or consists essentially of administering, to the mammal, a composition that contains a natriuretic peptide that includes the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The composition can contain the natriuretic peptide in an amount effective to reduce one or more symptoms of the chronic heart condition in the mammal. The one or more symptoms of the chronic heart condition can be selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema. The chronic heart condition can be HCM. The chronic heart condition can be associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g., HFrEF, HFmrEF. and HFpEF), or any combination thereof, or can be a HCM phenocopy. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1. composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1 with one, two. three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0032] In another aspect, this document features the use of a composition containing a natriuretic peptide for treating a chronic heart condition in a mammal, where the natriuretic peptide includes the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, tw o. three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. The use mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The composition can include the natriuretic peptide in an amount effective to reduce one or more symptoms of the chronic heart condition in the mammal. The one or more symptoms of the chronic heart condition can be selected from the group consisting of shortness of breath, heart palpitations, chest Attorney Docket No. 07039-2327WO1

[0033] 2024-159 discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema. The chronic heart condition can be HCM. The chronic heart condition can be associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g., HFrEF, HFmrEF, and HFpEF), or any combination thereof, or can be a HCM phenocopy. The composition can include a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1. The composition can include a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cy steine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0034] In still another aspect, this document features the use of a composition containing a natriuretic peptide for reducing inflammation in a mammal having a chronic heart condition, where the natriuretic peptide includes the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO:1 are maintained and are the only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, ahorse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The chronic heart condition can be HCM. The chronic heart condition can be associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g., HFrEF, HFmrEF, and HFpEF), or any combination thereof, or can be a HCM phenocopy. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0035] In another aspect, this document features the use of a composition containing a natriuretic peptide for reducing TGF-beta levels or TGF-beta activity in a mammal having a chronic heart condition, where the natriuretic peptide includes the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two. three, four, five, six, seven, eight, nine, or Attorney Docket No. 07039-2327WO1

[0036] 2024-159 ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The chronic heart condition can be HCM. The chronic heart condition can be associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g., HFrEF, HFrnrEF, and HFpEF), or any combination thereof, or can be a HCM phenocopy. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1 with one. two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0037] In another aspect, this document features the use of a composition containing a natriuretic peptide for reducing ventricular hypertrophy in a mammal having a chronic heart condition, where the natriuretic peptide includes the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, tw o, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The chronic heart condition can be HCM. The chronic heart condition can be associated with left ventricular hypertrophy, left ventricular remodeling, right ventricular hypertrophy, right ventricular remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g., HFrEF, HFrnrEF, and HFpEF), or any combination thereof, or can be a HCM phenocopy. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1 with one. two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. Attorney Docket No. 07039-2327WO1

[0038] 2024-159

[0039] In still another aspect, this document features the use of a composition containing a natriuretic peptide for preventing, attenuating, or reversing diastolic dysfunction in a mammal having a chronic heart condition, where the natriuretic peptide includes the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The chronic heart condition can be HCM. The chronic heart condition can be associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, cardiomyopathy, heart failure (e.g.. HFrEF, HFmrEF, and HFpEF), or any combination thereof, or can be a HCM phenocopy. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0040] In another aspect, this document features the use of a composition containing a natriuretic peptide for treating a mammal having a chronic heart condition in which guanylate cyclase A (GC-A) and guanylate cyclase B (GC-B) are present in heart cells of the mammal, where the natriuretic peptide includes the amino acid sequence set forth in SEQ ID NO: 1 wi th zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. The mammal can be a human. The mammal can be a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. The composition can contain the natriuretic peptide in an amount effective to reduce one or more symptoms of the chronic heart condition in the mammal. The one or more symptoms of the chronic heart condition can be selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema. The chronic heart condition can be HCM. The chronic heart condition can be associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial Attorney Docket No. 07039-2327WO1

[0041] 2024-159 remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g., HFrEF. HFmrEF, and HFpEF), or any combination thereof, or can be aHCM phenocopy. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1. The composition can contain a natriuretic peptide including the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide.

[0042] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Although methods and materials similar or equivalent to those described herein can be used to practice the invention, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting.

[0043] The details of one or more embodiments of the invention are set forth in the accompanying drawings and the description below. Other features, objects, and advantages of the invention will be apparent from the description and drawings, and from the claims.

[0044] DESCRIPTION OF DRAWINGS

[0045] FIG. 1 is a graph plotting cGMP levels in human cardiomyocytes induced by MV81 or vehicle (veh).

[0046] FIG. 2 is a graph plotting cGMP levels in human cardiac fibroblasts induced by vehicle (veh) or equimolar (10'7M) ANP, CNP, or MV81.

[0047] FIG. 3 is a graph plotting cGMP levels in in human THP-1 macrophages induced by MV81 or vehicle (veh).

[0048] FIG. 4 includes a pair of graphs showing the effect of MV 81 or vehicle (veh) on expression of the inflammatory cytokines IL 1 (left) and IL-6 (right) in human THP-1 macrophages that were polarized to the Ml state with LPS+IFNy. Attorney Docket No. 07039-2327WO1

[0049] 2024-159

[0050] FIG. 5 is a graph showing the effect of MV81 or vehicle (veh) on expression of the scavenger receptor CD86 in human THP-1 macrophages that were polarized to the Ml state with LPS+IFNy.

[0051] FIGS. 6A-6B are a pair of graphs plotting plasma cGMP levels (FIG. 6A) and the change in urinary cGMP response (FIG. 6B) after acute intravenous administration of MV81 or vehicle (veh) in rats. **P<0.01, ****p<0.0001 versus baseline within each group.

[0052] FIG. 7 is a graph showing changes in urine sodium excretion after an acute intravenous infusion of MV81 or vehicle (veh) in rats.

[0053] FIG. 8 is a graph showing the plasma cGMP response after chronic subcutaneous administration of MV81(0. 25. 50. or 100 pmol / kg / min) via osmotic minipump for 28 days in normal wild-type mice.

[0054] FIG. 9 is a graph showing the effect on heart w eight / body weight ratio (marker of cardiac hypertrophy) after chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 28 days in normal wild-type mice.

[0055] FIG. 10 is a graph showing the effect on left ventricle w eight / body w eight ratio (marker of left ventricular hypertrophy) after chronic subcutaneous administration of MV81(50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 28 days in normal wild-type mice.

[0056] FIG. 11 is a graph showing the effect on the expression of the indicated genes (which are related to hypertrophy and fibrosis) in the left ventricle after chronic subcutaneous administration of MV81(50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 28 days in normal wild-type mice.

[0057] FIG. 12 is a graph showing the effect on Tgfbl (a pro-hypertrophic and profibrotic cytokine) gene expression in the left ventricle after chronic subcutaneous administration of MV81 (0, 25, 50, or 100 pmol / kg / min) via osmotic minipump for 28 days in normal wild-type mice.

[0058] FIG. 13 is a graph showing the plasma cGMP response after chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 3 months in R403Q (a model of hypertrophic cardiomyopathy) mice. Attorney Docket No. 07039-2327WO1

[0059] 2024-159

[0060] FIG. 14 is a graph showing the effect on LV thickness after chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle via osmotic minipump for 3 months in R403Q (a model of hypertrophic cardiomyopathy) mice.

[0061] FIG. 15 is a graph showing the correlation between plasma cGMP levels and LV thickness after chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 3 months in R403Q (a model of hypertrophic cardiomyopathy) mice.

[0062] FIG. 16 is a graph showing fractional shortening after chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 3 months in R403Q (a model of hypertrophic cardiomyopathy) mice with normal wild-type mice serving as controls.

[0063] FIG. 17 is a graph showing the cardiac output response after chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 3 months in R403Q (a model of hypertrophic cardiomyopathy) mice, with normal wild-type mice serving as a control. **P<0.01 between wild-type and R403Q + Veh at 3 months,#P<0.05 between with R403Q + MV81 versus R403Q + Veh at 3 months using two-way repeated measures ANOVA.

[0064] FIG. 18A is a graph showing E / e' (an echocardiographic marker of diastolic dysfunction and filling pressures) response after chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 3 months in R403Q (a model of hypertrophic cardiomyopathy) mice. Normal wild-type mice (WT) served as a control. FIG. 18B is a graph plotting E / e', and FIG. 18C is a graph plotting e1(a marker of myocardial relaxation) in R403Q mice treated with chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 2 months, with treatment beginning after the onset of disease.

[0065] FIG. 19 is a graph plotting LV thickness after chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 3 weeks in mice with established left ventricular hypertrophy following transverse aortic constriction (TAC). Sham mice served as a control. **P<0.01, ****P<0.0001 versus baseline within TAC + Veh group;#P<0.05 between TAC + Veh versus TAC + MV81 within each time-point.

[0066] FIGS. 20A-20B are graphs showing the effect of chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 3 Attorney Docket No. 07039-2327WO1

[0067] 2024-159 weeks on left ventricular weight / body weight ratio (FIG. 20A, a marker of left ventricular hypertrophy) and left atrial weight / body weight ratio (FIG. 20B) in mice with established left ventricular hypertrophy following transverse aortic constriction (TAC). Sham mice served as a control.

[0068] FIG 21 is a graph plotting plasma cGMP in sham-operated mice and after chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 3 weeks in mice with transverse aortic constriction. *P<0.05, ****p<0.0001 using Welch ANOVA with Tamhane’s T2 multiple comparisons test.

[0069] FIG. 22 is a graph plotting in vitro degradation of ANP, CNP, and MV81 by neprilysin, assessed by quantification of cGMP -generating activity in human cardiac fibroblasts. *P<0.05 using t-test and **P<0.01 using t-test with Welch correction.

[0070] FIG. 23 is a graph plotting myocyte cross sectional area (CSA) in sham-operated mice and in mice after chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 3 weeks in mice with transverse aortic constriction (TAC). ***P<0.01 using ANOVA with Tukey multiple comparison test.

[0071] FIG. 24 shows a gene ontology analysis from left ventricular RNA sequencing to compare TAC mice treated with vehicle versus MV81.

[0072] FIGS. 25A-25D include graphs plotting the expression of genes related to hypertrophy, including Nppa (FIG. 25A), Nppb (FIG. 25B), the transcription factor Nfatc3 (FIG. 25C), and Rcanl (FIG. 25D), which is a sensitive marker of Nfatc3 nuclear translocation, in TAC mice treated with vehicle or MV81.

[0073] FIGS. 26A-26D are graphs plotting the expression of genes related to fibroblast activation, including Fap (FIG. 26A) and Postn (FIG. 26B), and genes related to extracellular matrix deposition, including Col lai (FIG. 26C) and Fnl (FIG. 26D). in TAC mice treated with vehicle or MV81.

[0074] FIGS. 27A-27D are graphs plotting the expression of genes related to inflammation and immunity7, including Ccr2 (FIG. 27A) and II lb (FIG. 27B), and genes encoding transcription factors related to macrophage-fibroblast cross-talk, such as Meoxl (FIG. 27C) and Runxl (FIG. 27D). in TAC mice treated with vehicle or MV81.

[0075] FIGS. 28A-28B show uniform manifold approximation and projection (UMAP) representations of single-nucleus transcriptomic profiles for the GC-A receptor (FIG. 28A) and GC-B receptor (FIG. 28B) in various cell types within non-failing (control) and failing human left ventricular tissue. Attorney Docket No. 07039-2327WO1

[0076] 2024-159

[0077] DETAILED DESCRIPTION

[0078] This document provides methods and materials for treating mammals having a heart-related disorder (e.g., a chronic cardiac disorder), where the methods include administering to the mammals a dual guanylate cyclase A and B activator. As used herein, a “dual guanylate cyclase A and B activator” is a molecule that is capable of activating both guanylate cyclase A and guanylate cyclase B. In some cases, the dual guanylate cyclase A and B activator can be a natriuretic peptide. There are three endogenous natriuretic peptides in humans: atrial natriuretic peptide (ANP, encoded by the NPPA gene in humans), B-type natriuretic peptide (BNP, encoded by the NPPB gene in humans), and C-type natriuretic peptide (CNP, encoded by the NPPC gene in humans). There also are three natriuretic peptide receptors: guanylate cyclase A (GC-A, also referred to as NPR-A or NPR1), guanylate cyclase B (GC-B, also referred to as NPR-B or NPR2), and natriuretic peptide receptor C (NPR-C, also referred to as NPR3). ANP and BNP bind with high affinity to GC-A, whereas CNP binds with high affinity to GC-B. Both GC-A and GC-B contain intracellular domains that catalyze production of cyclic guanosine monophosphate (cGMP). The binding of ligands to GC-A or GC-B results in the production of cGMP, a key second messenger that participates in downstream signaling that results in a variety of biological effects. Excess cGMP produced by GC-A and / or GC- B is egressed out of the cell, which can be measured in plasma. Thus, plasma cGMP is a biomarker of GC-A and / or GC-B target engagement.

[0079] In some cases, methods and materials provided herein can be used to treat mammals having heart conditions (e.g., chronic heart conditions) in which guanylate cyclase A (GC-A) and guanylate cyclase B (GC-B) are present in heart cells of the mammal. Examples of such heart conditions can include, without limitation, heart failure (e.g., HFrEF, HFmrEF, or HFpEF), conditions associated with ventricular and / or atrial hypertrophy and / or remodeling (e.g., left ventricular hypertrophy and remodeling, right ventricular hypertrophy and remodeling, left atrial hypertrophy and remodeling, and / or right atrial hypertrophy and remodeling), and systolic and / or diastolic dysfunction, as well as other heart diseases.

[0080] In some embodiments, methods and materials provided herein can be used to reduce inflammation in a mammal having a chronic heart condition. In some embodiments, methods and materials provided herein can be used to reduce transforming Attorney Docket No. 07039-2327WO1

[0081] 2024-159 growth factor-beta (TGF-beta) levels in a mammal having a chronic heart condition. In some embodiments, methods and materials provided herein can be used to reduce ventricular hypertrophy (e.g., left ventricular thickening or right ventricular thickening) in a mammal.

[0082] Any appropriate chronic cardiac disorder can be treated using the methods and materials described herein. A non-limiting example of a chronic cardiac disorder that can be treated as described herein is HCM. For example, this document provides methods and materials for using a dual guanylate cyclase A and B activator to treat HCM. In some cases, a dual guanylate cyclase A and B activator can be a natriuretic peptide. For example, a natriuretic peptide such as MV81 (SEQ ID NO: 1) can be used to treat a chronic cardiac condition as described herein. In some cases, MV81 (SEQ ID NO: 1) can be used to treat mammals having a chronic heart condition in which GC-A and GC-B are present in heart cells of the mammal. In some cases, MV81 (SEQ ID NO: 1) can be used to treat mammals (e.g., humans) having HCM and / or another chronic cardiac condition, such as a condition associated with hypertrophy and / or remodeling (e.g., left ventricular hypertrophy, left ventricular remodeling, left atrial hypertrophy, left atrial remodeling, right ventricular hypertrophy, right ventricular remodeling, right atrial hypertrophy, and / or right atrial remodeling), systolic and / or diastolic dysfunction, or another chronic heart disease. For example, MV81 (SEQ ID NO: 1) can be used to treat mammals having a chronic heart disease that is a HCM phenocopy, such as Fabry disease, Noonan syndrome, Pompe disease, PRKAG2-related cardiomyopathy, Danon disease, Friedrich ataxia cardiomyopathy, amyloidosis, and / or desminopathy. The methods and materials provided herein can be used to treat a chronic cardiac disorder (e.g., HCM and / or HF) in any appropriate mammal including, without limitation, humans, monkeys, dogs, cats, horses, cows, sheep, goats, rabbits, mice, and rats.

[0083] The methods and materials provided herein can include using a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or having the amino acid sequence set forth in SEQ ID NO: 1 with one to ten (e.g., one, two, three, four, five, six, seven, eight, nine, or ten) amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide. Examples of such natriuretic peptides include, without limitation, those set forth in TABLE 1. Peptides having the sequences set forth in Attorney Docket No. 07039-2327WO1

[0084] 2024-159

[0085] TABLE 1 can form ring structures due to disulfide bonds between the cysteine residues, which are underlined.

[0086] TABLE 1. Exemplary7natriuretic peptides

[0087] In some cases, an amino acid substitution can be a conservative amino acid substitution. For example, an amino acid residue of SEQ ID NO: 1 can be selected and replaced with an amino acid residue that does not differ significantly in its effect on maintaining (a) the structure of the peptide backbone in the area of the substitution, (b) Attorney Docket No. 07039-2327WO1

[0088] 2024-159 the charge or hydrophobicity of the molecule at the target site, or (c) the bulk of the side chain. For example, naturally occurring residues can be divided into groups based on side-chain properties: (1) hydrophobic amino acids (norleucine, methionine, alanine, valine, leucine, and isoleucine); (2) neutral hydrophilic amino acids (cysteine, serine, and threonine); (3) acidic amino acids (aspartic acid and glutamic acid); (4) basic amino acids (asparagine, glutamine, histidine, lysine, and arginine); (5) amino acids that influence chain onentation (glycine and proline); and (6) aromatic amino acids (tryptophan, tyrosine, and phenylalanine). Substitutions made within these groups can be considered conservative substitutions. Non-limiting examples of conservative substitutions include, without limitation, substitution of valine for alanine, lysine for arginine, glutamine for asparagine, glutamic acid for aspartic acid, serine for cysteine, asparagine for glutamine, aspartic acid for glutamic acid, proline for glycine, arginine for histidine, leucine for isoleucine, isoleucine for leucine, arginine for lysine, leucine for methionine, leucine for phenyalanine, glycine for proline, threonine for serine, serine for threonine, ty rosine for tryptophan, phenylalanine for tyrosine, and / or leucine for valine. Further examples of conservative substitutions that can be made within S EQ ID NO: 1 are set forth in TABLE

[0089] 2.

[0090] TABLE 2. Examples of conservative amino acid substitutions Attorney Docket No. 07039-2327WO1

[0091] 2024-159

[0092] In some cases, an amino acid substitution can be anon-conservative amino acid substitution. For example, an amino acid residue of SEQ ID NO: 1 can be selected and replaced with an amino acid residue from a different class of amino acids as compared with the selected amino acid. For example, an acidic amino acid (aspartic acid or glutamic acid) present in SEQ ID NO: 1 can be selected and replaced with a basic amino acid (asparagine, glutamine, histidine, lysine, or arginine).

[0093] A polypeptide described herein (e.g., a polypeptide having the amino acid sequence set forth in SEQ ID NO: 1, or having the amino acid sequence set forth in SEQ ID NO: 1 with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide) can be obtained using any appropriate technique including, without limitation, solid phase polypeptide synthesis techniques such as those involving the use of an Applied BioSystems (Foster City. CA) Peptide Synthesizer or a Biosearch Inc. (San Rafael, CA) automatic peptide synthesizer. Disulfide bonds between cysteine residues can be introduced by mild oxidation of the linear polypeptides using KCN as described elsewhere (see, e.g., U.S. Patent No. 4,757,048). In some cases, a polypeptide described herein (e.g., a polypeptide having the amino acid sequence set forth in SEQ ID NO: 1) can be obtained recombinantly. For example, a polypeptide described herein (e.g., a polypeptide having the amino acid sequence set forth in SEQ ID NO: 1, or having the amino acid sequence set forth in SEQ ID NO: 1 with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide) can be obtained by expressing a recombinant nucleic acid encoding the polypeptide such as an expression vector encoding a polypeptide described herein within host cells. The resulting polypeptide then can be purified using, for example, affinity chromatographic techniques and HPLC. The extent of purification can be measured using any appropriate method including, without limitation, column chromatography, polyacrylamide gel electrophoresis, or high-performance liquid chromatography. In some cases, a polypeptide described herein (e.g., a polypeptide having the amino acid sequence set forth in SEQ ID NO: 1, or having the amino acid sequence set forth in SEQ ID NO: 1 with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the Attorney Docket No. 07039-2327WO1

[0094] 2024-159 natriuretic peptide) can be designed or engineered to contain a tag sequence that allows the polypeptide to be purified (e.g., captured onto an affinity matrix) and / or an enzyme or label sequence that allows the polypeptide to be detected. For example, a tag such as c- myc, hemagglutinin, polyhistidine, or FLAG™ tag (Kodak) can be used to aid polypeptide purification. Examples of enzy me and label sequences that can be used to aid polypeptide detection include, without limitation, alkaline phosphatase. GFP. RFP. luciferases, and Cas polypeptides. Such tags, enzymes, and labels can be inserted anywhere within the polypeptide, including at either the carboxyl terminus or the amino terminus. In some cases, the polypeptide described herein can have the structure and can be obtained as described elsewhere (see, WO 2021 / 183928).

[0095] Any appropriate method can be used to confirm that a polypeptide useful in the methods provided herein (e g., a polypeptide having the amino acid sequence set forth in SEQ ID NO: 1, or having the amino acid sequence set forth in SEQ ID NO: 1 with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide) has one or more biological (e.g., natriuretic) activities. For example, a polypeptide described herein can be confirmed as having a biological (e.g., anti-fibrotic, anti-hypertrophic, and / or anti-remodeling) activity in vitro by measuring its effect on intracellular cGMP levels in human cardiomyocytes, human cardiac fibroblast cells, and / or human THP-1 macrophages. A polypeptide described herein can be confirmed as having a biological (e.g., anti-fibrotic, anti-hypertrophic, and / or anti-remodeling) activity7in vivo by, for example, testing its effects on factors such as plasma and / or urinary cGMP generation, heart weight, altered gene expression, plasma and / or urinary7sodium excretion, and / or left ventricular thickness in mammals.

[0096] In some cases, a polypeptide described herein (e.g., a polypeptide having the amino acid sequence set forth in SEQ ID NO: 1, or having the amino acid sequence set forth in SEQ ID NO:1 with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of the natriuretic peptide) can be formulated as a pharmaceutical composition by admixture with one or more pharmaceutically7acceptable non-toxic excipients or carriers. Pharmaceutical compositions can be prepared for parenteral administration, for example, in the form of liquid solutions or suspensions in aqueous physiological buffer solutions; for oral administration, for example, in the form of tablets Attorney Docket No. 07039-2327WO1

[0097] 2024-159 or capsules; or for intranasal administration, for example in the form of powders, nasal drops, or aerosols. Compositions for other routes of administration can be prepared as desired using appropriate methods.

[0098] Formulations for parenteral administration can include as common excipients, sterile water, saline, polyalkylene glycols such as polyethylene glycol, oils of vegetable origin, hydrogenated naphthalenes, and combinations thereof. In some cases, biocompatible, biodegradable lactide polymer, lactide / glycolide copolymer, polyoxethylene-polyoxypropylene copolymers, or combinations thereof can be used as excipients for controlling the release of the polypeptide in vivo. Other suitable parenteral delivery systems that can be used include, without limitation, ethylene-vinyl acetate copolymer particles, osmotic pumps, implantable infusion systems, liposomes, and combinations thereof. Formulations for inhalation administration can include excipients such as lactose. Inhalation formulations can be aqueous solutions containing, for example, polyoxyethylene-9-lauryl ether, glycocholate, deoxy cholate, or combinations thereof, or they can be oily solutions for administration in the form of nasal drops. If desired, a composition containing a polypeptide described herein (e.g., a polypeptide having the amino acid sequence set forth in SEQ ID NO: 1) can be formulated as a gel to be applied intranasally. Formulations for parenteral administration can include glycocholate for buccal administration.

[0099] For oral administration, tablets or capsules can be prepared using appropriate methods with pharmaceutically acceptable excipients such as binding agents (e.g., pregelatinized maize starch, polyvinylpyrrolidone, or hydroxypropyl methylcellulose); fillers (e.g.. lactose, microcrystalline cellulose or calcium hydrogen phosphate); lubricants (e.g. magnesium stearate, talc or silica); disintegrants (e.g., potato starch or sodium starch glycolate); or wetting agents (e.g., sodium lauryl sulfate). Tablets can be coated using appropriate methods. Preparations for oral administration can be formulated to give controlled release of the polypeptide.

[0100] Nasal preparations can be presented in a liquid form or as a dry product. Nebulised aqueous suspensions or solutions can include carriers or excipients to adjust pH and / or tonicity.

[0101] This document also provides methods and materials for treating a mammal (e.g., a human) that has a chronic heart disease (e.g., HCM and / or HF), is developing a chronic heart disease (e.g., HCM and / or HF), is likely to develop more severe effects of a chronic Attorney Docket No. 07039-2327WO1

[0102] 2024-159 heart disease (e.g., HCM and / or HF), and / or is likely to experience a poor outcome from a chronic heart disease (e.g.. HCM and / or HF). For example, a mammal can be treated with one or more natriuretic peptides to reduce a symptom of HCM or another chronic heart disorder, to counteract a symptom of HCM or another chronic heart disorder, and / or to reduce the likelihood of a poor outcome from HCM or another chronic heart disorder. Symptoms of HCM and other chronic heart disorders can include, without limitation, shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness (e.g., especially wi th exertion or upon standing quickly), syncope (fainting) or presyncope (e.g., during or after physical activity ), orthopnea, and paroxysmal nocturnal dyspnea, lightheadedness, especially with or after activity or exercise, abnormal heart rhythms (arrhythmias), heart murmurs, edema (e.g., swelling in the legs and / or feet), or any combination thereof. Examples of natriuretic peptides that can be used alone or in combination to treat a mammal as described herein include, without limitation, a polypeptide having the sequence set forth in SEQ ID NO: 1 and a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten (e.g., one, two, three, four, five, six, seven, eight, nine, or ten) amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide.

[0103] In some cases, one or more additional therapeutic agents also can be administered to a mammal (e.g., a human) in addition to one or more natriuretic peptides, in order to reduce a symptom of chronic heart disease (e.g.. HCM and / or HF), to counteract a symptom of chronic heart disease (e.g., HCM and / or HF), and / or to reduce the likelihood of a poor outcome from chronic heart disease (e.g., HCM and / or HF). Examples of such therapeutic agents include, without limitation, cardiac myosin inhibitors (e.g., mavacamten and aficamten). ACE inhibitors (ACEi’s). angiotensin II receptor blockers (ARBs; e.g., ARBs neprilysin inhibition such as sacubitril-valsartan), aldosterone receptor antagonists, aldosterone synthase inhibitors, diuretics (e.g., furosemide, bumetanide, ethacrynic acid, torsemide, acetazolamide, dorzolamide, amiloride, spironolactone, eplerenone, triamterene, potassium canrenoate, bendroflumethiazide, or hydrochlorothiazide), beta-blockers, statins, myosin inhibitors (e.g.. mavacamten, aficamten, or BMS-98645 / MYK-224), myosin modulators (e.g., EDG-7500 or MYK- 581), and cardiac mitotropes (e.g., ninerafaxstat, perhexiline, trimetazidine, or elamipretide). The one or more natriuretic peptides and the one or more additional Attorney Docket No. 07039-2327WO1

[0104] 2024-159 therapeutic agents can be administered simultaneously (e.g., in the same composition or in separate compositions that are administered at the same time), or sequentially.

[0105] Any appropriate route of administration can be used to administer a composition containing one or more natriuretic peptides described herein (or one or more natriuretic peptides described herein in combination with one or more additional therapeutic agents) to treat a mammal (e.g., a human) that has chronic heart disease (e.g.. HCM and / or HF), is developing chronic heart disease (e.g., HCM and / or HF), is likely to develop more severe effects of chronic heart disease (e.g., HCM and / or HF), and / or is likely to experience a poor outcome from chronic heart disease (e.g., HCM and / or HF). For example, parenteral administration (e.g.. by subcutaneous, intrathecal, intraventricular, intramuscular, or intraperitoneal injection, or by intravenous drip) can be used to administer a composition containing one or more natriuretic peptides described herein (or one or more natriuretic peptides described herein in combination with one or more additional therapeutic agents) to a mammal (e.g., a human). In some cases, the administration can be rapid (e.g., by injection) or can occur over a period of time (e.g., by slow infusion or administration of slow release formulations). In some cases, a composition containing one or more natriuretic peptides described herein (or one or more natriuretic peptides described herein in combination with one or more additional therapeutic agents) can be administered to a mammal (e.g., a human) via topical administration (e.g., transdermal, sublingual, ophthalmic, or intranasal), pulmonary administration (e.g., by inhalation or insufflation of powders or aerosols), or oral administration.

[0106] In some cases, a mammal having chronic heart disease (e.g.. HCM and / or HF) can be instructed to self-administer one or more natriuretic peptides described herein (and, in some cases, one or more additional therapeutic agents).

[0107] A composition containing one or more natriuretic peptides, such as a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide, and / or containing one or more such natriuretic peptides in combination with one or more additional therapeutic agents, can be administered to a mammal (e.g., a human) in an appropriate amount, at an appropriate frequency, and for an appropriate duration effective to achieve a desired Attorney Docket No. 07039-2327WO1

[0108] 2024-159 outcome. In some cases, a composition provided herein can be administered to a mammal (e.g., a human) in an appropriate amount, at an appropriate frequency, and for an appropriate duration effective to reduce the severity of a symptom of a chronic heart disorder such as HCM. In some cases, a composition provided herein can be administered to a mammal (e.g., a human) in an appropriate amount, at an appropriate frequency, and for an appropriate duration effective to reduce, reverse, reduce the likelihood of, or prevent adverse myocardial remodeling, such as hypertrophy or fibrosis. In some cases, a composition provided herein can be administered to a mammal (e.g., a human) in an appropriate amount, at an appropriate frequency, and for an appropriate duration effective to reduce, reverse, reduce the likelihood of, or prevent systolic and / or diastolic dysfunction. In some cases, a composition provided herein can be administered to a mammal (e.g., a human) in an appropriate amount, at an appropriate frequency, and for an appropriate duration effective to reduce, reduce the likelihood of, or prevent adverse cardiovascular events, such as heart failure, hospitalization, arrhythmia, or death.

[0109] In some cases, a composition containing one or more natriuretic peptides, such as a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide, and / or containing one or more such natriuretic peptides in combination with one or more additional therapeutic agents, can be administered to a mammal identified as having chronic heart disease (e.g., HCM and / or HF) to reduce the severity of a symptom of the chronic heart disease (e.g., HCM and / or HF) by, for example, at least 5 percent (e.g., at least 10 percent, at least 25 percent, at least 50 percent, at least 75 percent, or 100 percent). Any appropriate method can be used to determine whether or not a mammal (e.g., a human) has experienced a reduction in the severity of a symptom of a chronic heart disease (e.g., HCM and / or HF).

[0110] An effective amount of a composition containing one or more natriuretic peptides, such as a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide, and / or one or more such natriuretic peptides in combination with one or more additional therapeutic Attorney Docket No. 07039-2327WO1

[0111] 2024-159 agents, can be any amount that reduces, reverses, or prevents development of a symptom of chronic heart disease (e.g., HCM and / or HF) without producing significant toxicity to the mammal. In some cases, an effective amount of a composition containing one or more natriuretic peptides, such as a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1. or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide, and / or one or more such natriuretic peptides in combination with one or more additional therapeutic agents, can be any amount that reduces, reverses, reduces the likelihood of, or prevents adverse myocardial remodeling (e.g.. hypertrophy or fibrosis), any amount that reduces, reverses, reduces the likelihood of, or prevents systolic and / or diastolic dysfunction, and / or any amount that reduces, reduces the likelihood of, or prevents adverse cardiovascular events (e.g., heart failure, hospitalization, arrhythmia, or death). An effective amount can vary' depending on the relative potency of the peptide and / or the route of administration, and can generally be estimated based on ECso found to be effective in in vitro and in vivo animal models.

[0112] In some cases, an effective amount of a natriuretic peptide described herein (e.g., a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO:1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only tw o cysteine residues of the natriuretic peptide) that is subcutaneously administered can be from about 0.001 pg / kg to about 500 pg / kg of body weight. For example, an effective amount of a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1 can be from about 0.001 pg / kg to about 0.01 pg / kg, from about 0. 1 pg / kg to about 50 pg / kg, from about 0.01 pg / kg to about 0. 1 pg / kg, from about 0. 1 pg / kg to about 0.5 pg / kg, from about 0.5 pg / kg to about 1 pg / kg, from about 1 pg / kg to about 10 pg / kg, from about 10 pg / kg to about 50 pg / kg, from about 50 pg / kg to about 100 pg / kg, or from about 100 pg / kg to about 500 pg / kg.

[0113] In some embodiments, an effective amount of a natriuretic peptide described herein (e.g., a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide) that is Attorney Docket No. 07039-2327WO1

[0114] 2024-159 subcutaneously administered can be from about 0.05 pg to about 10 mg. For example, an effective amount of a subcutaneously administered natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1 can be from about 0.5 pg to about 1 mg, from about 0.5 pg to about 5 pg, from about 5 pg to about 10 pg, from about 10 pg to about 50 pg, from about 50 pg to about 100 pg, from about 100 pg to about 500 pg, from about 500 pg to about 1 mg, from about 1 mg to about 2.5 mg, from about 2.5 mg to about 5 mg, or from about 5 mg to about 10 mg.

[0115] In some embodiments, an effective amount of a natriuretic peptide described herein (e.g., a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide) that is orally administered can be from about 0.05 mg to about 1000 mg. For example, an effective amount of an orally administered natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1 can be from about 0.05 mg to about 750 mg, from about 0.5 mg to about 100 mg, from about 0.5 mg to about 1 mg, from about 1 mg to about 5 mg, from about 5 mg to about 10 mg, from about 10 mg to about 50 mg, from about 50 mg to about 100 mg, from about 100 mg to about 200 mg, from about 200 mg to about 500 mg, or from about 500 mg to about 1000 mg.

[0116] In some embodiments, an effective amount of a natriuretic peptide described herein (e.g., a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide) that is intravenously administered can be from about 0.05 ng / kg / min to about 500 ng / kg / min. For example, an effective amount of an intravenously administered natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1 can be from about 0.05 ng / kg / min to about 500 ng / kg / min, from about 0. 1 ng / kg / min to about 250 ng / kg / min, from about 0.5 ng / kg / min to about 200 ng / kg / min. from about 0.5 ng / kg / min to about 50 ng / kg / min, from about 1 ng / kg / min to about 150 ng / kg / min, from about 5 ng / kg / min to about 100 ng / kg / min, from about 10 ng / kg / min to about 50 ng / kg / min, from about 0.05 ng / kg / min to about 0.5 ng / kg / min, from about 0.5 ng / kg / min to about 1 ng / kg / min, from about 1 ng / kg / min to about 10 ng / kg / min, from about 10 ng / kg / min to about 50 Attorney Docket No. 07039-2327WO1

[0117] 2024-159 ng / kg / min, from about 50 ng / kg / min to about 100 ng / kg / min, or from about 100 ng / kg / min to about 500 ng / kg / min.

[0118] If a particular mammal fails to respond to a particular amount, then the amount can be increased by, for example, two-fold. After receiving this higher amount, the subject can be monitored for both responsiveness to the treatment and toxicity' symptoms, and adjustments made accordingly. The effective amount can remain constant or can be adjusted as a sliding scale or variable dose depending on the mammal’s response to treatment. Various factors can influence the actual effective amount used for a particular application. For example, the frequency of administration, duration of treatment, use of multiple treatment agents, route of administration, and severity of the condition (e.g.. HCM and / or HF) may require an increase or decrease in the actual effective amount administered.

[0119] A composition containing one or more natriuretic peptides, such as a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide, and / or one or more natriuretic peptides in combination with one or more additional therapeutic agents can be administered once or more than once, at any appropriate frequency and for any appropriate duration. In some cases, an effective frequency for administering a composition containing one or more natriuretic peptides, such as a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide, and / or one or more natriuretic peptides in combination with one or more additional therapeutic agents can be any frequency that reduces the severity' of a symptom of chronic heart disease (e.g., HCM and / or HF), any frequency that reduces, reverses, reduces the likelihood of, or prevents adverse myocardial remodeling, any frequency that reduces, reverses, reduces the likelihood of, or prevents systolic and / or diastolic dysfunction, and / or any frequency that reduces, reverses, reduces the likelihood of, or prevents the occurrence of adverse cardiovascular events, without producing significant toxicity to the mammal. For example, the frequency of administration can be once or more daily (e.g., once, twice, three times, or four times Attorney Docket No. 07039-2327WO1

[0120] 2024-159 daily), biweekly, weekly, monthly, or even less frequently. The frequency of administration can remain constant or can be variable during the duration of treatment. In some cases, a course of treatment can include rest periods. For example, a composition containing one or more natriuretic peptides, such as a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide, and / or one or more natriuretic peptides in combination with one or more additional therapeutic agents can be administered over a two month period followed by a month rest period, and then repeated. As with the effective amount, various factors can influence the actual frequency of administration. For example, the effective amount, duration of treatment, use of multiple treatment agents, route of administration, and severity of the condition (e.g., HCM and / or HF) may require an increase or decrease in administration frequency.

[0121] An effective duration for administering a composition containing one or more natriuretic peptides, such as a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide, and / or one or more natriuretic peptides in combination with one or more additional therapeutic agents can be any duration that reduces the severity of a symptom of chronic heart disease (e.g., HCM and / or HF), any duration that reduces, reverses, reduces the likelihood of. or prevents adverse myocardial remodeling, any duration that reduces, reverses, reduces the likelihood of. or prevents systolic and / or diastolic dysfunction, and / or any duration that reduces, reverses, reduces the likelihood of, or prevents the occurrence of adverse cardiovascular events, without producing significant toxicity to the mammal. In some cases, the effective duration can vary from 15 minutes to several days (e.g., 15 to 30 minutes. 30 to 60 minutes, 1 to 3 hours. 3 to 6 hours, 6 to 12 hours, 12 to 24 hours. 24 to 36 hours. 36 to 48 hours. 48 to 72 hours. 3 to 5 days, or 5 to 7 days), or even weeks or months. In general, an effective duration for administering a composition containing one or more natriuretic peptides, such as a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, Attorney Docket No. 07039-2327WO1

[0122] 2024-159 provided that the two cysteine residues are maintained and are the only two cysteine residues of the natriuretic peptide, and / or one or more natriuretic peptides in combination with one or more additional therapeutic agents can range from about 1 month to about 10 years or more (e.g., from about 1 month to about 6 months, from about 6 months to about 12 months, from about 1 year to about 2 years, from about 2 years to about 5 years, from about 5 years to about 10 years, or more than 10 years). In some cases, an effective duration can be for as long as the individual mammal (e.g., a human) is alive. Multiple factors can influence the actual effective duration used for a particular treatment. For example, an effective duration can vary7with the frequency of administration, effective amount, use of multiple treatment agents, route of administration, and severity of the condition (e.g.. HCM and / or HF).

[0123] After administering a composition containing one or more natriuretic peptides, such as a natriuretic peptide having the amino acid sequence set forth in SEQ ID NO: 1, or a polypeptide having the sequence set forth in SEQ ID NO: 1 but with one to ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues are maintained and are the only tw o cysteine residues of the natriuretic peptide to a mammal (e.g., a human) having chronic heart disease (e.g., HCM and / or HF), the mammal can be monitored to determine whether or not the treatment w as effective against the heart disease (e.g., HCM and / or HF). For example, a mammal (e.g., a human) can be assessed after treatment to determine whether cardiac structure and / or function was improved. Any appropriate method can be used to assess cardiac structure and / or function. For example, shortness of breath, edema, and / or lung congestion can be monitored to determine whether a treatment described herein is effective. In some cases, echocardiography (transthoracic or transesophageal), cardiac MRI. chest x-ray, cardiac CT, EKG. nuclear testing, treadmill stress testing, pharmacologic stress testing, right and / or left heart catheterization, and / or cardiopulmonary exercise testing, and / or biomarkers (such as BNP, NT-proBNP, troponin, creatinine) and / or scores such as the NYHA classification, KCCQ score, and / or MLHF score can be used to monitor an effective response to a treatment described herein. In some cases, plasma cGMP can be measured in order to guide dosing or as a marker of efficacy. For example, an increase in plasma cGMP levels after administration of a natriuretic peptide described herein can indicate that the administered dose was appropriate and / or that the treatment w as effective. Attorney Docket No. 07039-2327WO1

[0124] 2024-159

[0125] Exemplary Embodiments

[0126] Embodiment 1 is a method for treating a chronic heart condition, wherein said method comprises administering, to a mammal identified as having said chronic heart condition, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0127] Embodiment 2 is the method of embodiment 1, wherein said mammal is a human.

[0128] Embodiment 3 is the method of embodiment 1, wherein said mammal is a monkey, a dog. a cat. a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

[0129] Embodiment 4 is the method of any one of embodiments 1 to 3, wherein said method comprises administering said composition to said mammal in an amount effective to reduce one or more symptoms of said chronic heart condition in said mammal.

[0130] Embodiment 5 is the method of embodiment 4, wherein said one or more symptoms of said chronic heart condition are selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema.

[0131] Embodiment 6 is the method of any one of embodiments 1 to 5. wherein said chronic heart condition is HCM.

[0132] Embodiment 7 is the method of any one of embodiments 1 to 5, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure (e.g., HFrEF, HFmrEF, and HFpEF), or any combination thereof, or is a HCM phenocopy.

[0133] Embodiment 8 is the method of any one of embodiments 1 to 7, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ 1D NO: 1.

[0134] Embodiment 9 is the method of any one of embodiments 1 to 7, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid Attorney Docket No. 07039-2327WO1

[0135] 2024-159 additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0136] Embodiment 10 is a method for reducing inflammation in a mammal having a chronic heart condition wherein said method comprises administering, to a mammal identified as having said chronic heart condition, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0137] Embodiment 11 is the method of embodiment 10, wherein said mammal is a human.

[0138] Embodiment 12 is the method of embodiment 10, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

[0139] Embodiment 13 is the method of any one of embodiments 10 to 12, wherein said chronic heart condition is HCM.

[0140] Embodiment 14 is the method of any one of embodiments 10 to 13, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

[0141] Embodiment 15 is the method of any one of embodiments 10 to 14, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

[0142] Embodiment 16 is the method of any one of embodiments 10 to 14, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0143] Embodiment 17 is a method for reducing TGF-beta levels or TGF-beta activity in a mammal having a chronic heart condition wherein said method comprises administering, to a mammal identified as having said chronic heart condition, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six. seven, eight, nine, or ten Attorney Docket No. 07039-2327WO1

[0144] 2024-159 amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0145] Embodiment 18 is the method of embodiment 17, wherein said mammal is a human.

[0146] Embodiment 19 is the method of embodiment 17, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

[0147] Embodiment 20 is the method of any one of embodiments 17 to 19, wherein said chronic heart condition is HCM.

[0148] Embodiment 21 is the method of any one of embodiments 17 to 20, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

[0149] Embodiment 22 is the method of any one of embodiments 17 to 21, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

[0150] Embodiment 23 is the method of any one of embodiments 17 to 21, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one. two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0151] Embodiment 24 is a method for reducing ventricular hypertrophy in a mammal having a chronic heart condition, wherein said method comprises administering, to a mammal identified as having said chronic heart condition, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0152] Embodiment 25 is the method of embodiment 24, wherein said mammal is a human.

[0153] Embodiment 26 is the method of embodiment 24, wherein said mammal is a monkey, a dog. a cat. a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat. Attorney Docket No. 07039-2327WO1

[0154] 2024-159

[0155] Embodiment 27 is the method of any one of embodiments 24 to 26, wherein said chronic heart condition is HCM.

[0156] Embodiment 28 is the method of any one of embodiments 24 to 27, wherein said chronic heart condition is associated with left ventricular hypertrophy, left ventricular remodeling, right ventricular hypertrophy, right ventricular remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

[0157] Embodiment 29 is the method of any one of embodiments 24 to 28, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO:1.

[0158] Embodiment 30 is the method of any one of embodiments 24 to 28. wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0159] Embodiment 31 is the use of a composition comprising a natriuretic peptide for treating a chronic heart condition in a mammal, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0160] Embodiment 32 is the use of embodiment 31, wherein said mammal is a human.

[0161] Embodiment 33 is the use of embodiment 31, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

[0162] Embodiment 34 is the use of any one of embodiments 31 to 33, wherein said composition comprises said natriuretic peptide in an amount effective to reduce one or more symptoms of said chronic heart condition in said mammal.

[0163] Embodiment 35 is the use of embodiment 34, wherein said one or more symptoms of said chronic heart condition are selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema. Attorney Docket No. 07039-2327WO1

[0164] 2024-159

[0165] Embodiment 36 is the use of any one of embodiments 31 to 35, wherein said chronic heart condition is HCM.

[0166] Embodiment 37 is the use of any one of embodiments 31 to 35, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

[0167] Embodiment 38 is the use of any one of embodiments 31 to 37, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO:1.

[0168] Embodiment 39 is the use of any one of embodiments 31 to 37, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, tw o. three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0169] Embodiment 40 is the use of a composition comprising a natriuretic peptide for reducing inflammation in a mammal having a chronic heart condition, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO:1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only tw o cysteine residues of said natriuretic peptide.

[0170] Embodiment 41 is the use of embodiment 40, wherein said mammal is a human.

[0171] Embodiment 42 is the use of embodiment 40, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

[0172] Embodiment 43 is the use of any one of embodiments 40 to 42, wherein said chronic heart condition is HCM.

[0173] Embodiment 44 is the use of any one of embodiments 40 to 43, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy. Attorney Docket No. 07039-2327WO1

[0174] 2024-159

[0175] Embodiment 45 is the use of any one of embodiments 40 to 44, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

[0176] Embodiment 46 is the use of any one of embodiments 40 to 44, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0177] Embodiment 47 is the use of a composition comprising a natriuretic peptide for reducing TGF-beta levels or TGF-beta activity in a mammal having a chronic heart condition, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0178] Embodiment 48 is the use of embodiment 47, wherein said mammal is a human.

[0179] Embodiment 49 is the use of embodiment 47, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

[0180] Embodiment 50 is the use of any one of embodiments 47 to 49, wherein said chronic heart condition is HCM.

[0181] Embodiment 51 is the use of any one of embodiments 47 to 50, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

[0182] Embodiment 52 is the use of any one of embodiments 47 to 51, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

[0183] Embodiment 53 is the use of any one of embodiments 47 to 51, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide. Attorney Docket No. 07039-2327WO1

[0184] 2024-159

[0185] Embodiment 54 is the use of a composition comprising a natriuretic peptide for reducing ventricular hypertrophy in a mammal having a chronic heart condition, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0186] Embodiment 55 is the use of embodiment 54, wherein said mammal is a human.

[0187] Embodiment 56 is the use of embodiment 54, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

[0188] Embodiment 57 is the use of any one of embodiments 54 to 56, wherein said chronic heart condition is HCM.

[0189] Embodiment 58 is the use of any one of embodiments 54 to 57, wherein said chronic heart condition is associated with left ventricular hypertrophy, left ventricular remodeling, right ventricular hypertrophy, right ventricular remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

[0190] Embodiment 59 is the use of any one of embodiments 54 to 58, w herein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO:1.

[0191] Embodiment 60 is the use of any one of embodiments 54 to 58, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, tw o. three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0192] Embodiment 61 is a method for preventing, attenuating, or reversing diastolic dysfunction in a mammal having a chronic heart condition, wherein said method comprises administering, to a mammal identified as having said chronic heart condition, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five. six. seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only tw o cysteine residues of said natriuretic peptide. Attorney Docket No. 07039-2327WO1

[0193] 2024-159

[0194] Embodiment 62 is the method of embodiment 61, wherein said mammal is a human.

[0195] Embodiment 63 is the method of embodiment 61, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

[0196] Embodiment 64 is the method of any one of embodiments 61 to 63, wherein said chronic heart condition is HCM.

[0197] Embodiment 65 is the method of any one of embodiments 61 to 64, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

[0198] Embodiment 66 is the method of any one of embodiments 61 to 65. wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

[0199] Embodiment 67 is the method of any one of embodiments 61 to 65, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0200] Embodiment 68 is the use of a composition comprising a natriuretic peptide for preventing, attenuating, or reversing diastolic dysfunction in a mammal having a chronic heart condition, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two. three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0201] Embodiment 69 is the use of embodiment 68, wherein said mammal is a human.

[0202] Embodiment 70 is the use of embodiment 68, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

[0203] Embodiment 71 is the use of any one of embodiments 68 to 70, wherein said chronic heart condition is HCM.

[0204] Embodiment 72 is the use of any one of embodiments 68 to 71, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular Attorney Docket No. 07039-2327WO1

[0205] 2024-159 remodeling, atrial hypertrophy, atrial remodeling, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

[0206] Embodiment 73 is the use of any one of embodiments 68 to 72, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

[0207] Embodiment 74 is the use of any one of embodiments 68 to 72, wherein said composition comprises a natriuretic peptide compnsing the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0208] Embodiment 75 is a use of a composition comprising a natriuretic peptide for treating a mammal having a chronic heart condition in which guanylate cyclase A (GC-A) and guanylate cyclase B (GC-B) are present in heart cells of the mammal, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only tw o cysteine residues of said natriuretic peptide.

[0209] Embodiment 76 is the use of embodiment 75, wherein said mammal is a human.

[0210] Embodiment 77 is the use of embodiment 75, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

[0211] Embodiment 78 is the use of any one of embodiments 75 to 77, wherein said composition comprises said natriuretic peptide in an amount effective to reduce one or more symptoms of said chronic heart condition in said mammal.

[0212] Embodiment 79 is the use of embodiment 78, wherein said one or more symptoms of said chronic heart condition are selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema.

[0213] Embodiment 80 is the use of any one of embodiments 75 to 79, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

[0214] Embodiment 81 is the use of any one of embodiments 75 to 79, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic Attorney Docket No. 07039-2327WO1

[0215] 2024-159 dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

[0216] Embodiment 82 is the use of any one of embodiments 75 to 81, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

[0217] Embodiment 83 is the use of any one of embodiments 75 to 81, wherein said composition comprises a natriuretic peptide compnsing the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0218] Embodiment 84 is a method for treating a mammal having a chronic heart condition in which guanylate cyclase A (GC-A) and guanylate cyclase B (GC-B) are present in heart cells of the mammal, wherein said method comprises administering, to said mammal, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0219] Embodiment 85 is the method of embodiment 84, wherein said mammal is a human.

[0220] Embodiment 86 is the method of embodiment 84, w erein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow; a sheep, a goat, a rabbit, a mouse, or a rat.

[0221] Embodiment 87 is the method of any one of embodiments 84 to 86, wherein said composition comprises said natriuretic peptide in an amount effective to reduce one or more symptoms of said chronic heart condition in said mammal.

[0222] Embodiment 88 is the method of embodiment 87, wherein said one or more symptoms of said chronic heart condition are selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema.

[0223] Embodiment 89 is the method of any one of embodiments 84 to 88, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM). Attorney Docket No. 07039-2327WO1

[0224] 2024-159

[0225] Embodiment 90 is the method of any one of embodiments 84 to 88, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

[0226] Embodiment 91 is the method of any one of embodiments 84 to 90, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

[0227] Embodiment 92 is the method of any one of embodiments 84 to 90, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one. two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

[0228] The invention will be further described in the following examples, which do not limit the scope of the invention described in the claims.

[0229] EXAMPLES

[0230] Example 1 : Effects of MV81 in vitro and in vivo MA TERIALS AND METHODS

[0231] Human cardiomyocyte (hCM) cGMP analysis: Human cardiomyocytes (hCMs) were purchased from ScienCell (Catalog #6200) and were grown in cardiac myocyte medium (Cat#6101, ScienCell) in 6-well plates until they reached 80-90% confluency. The treatment buffer used in all experiments contained Hank's Balanced Salt Solution (HBSS), 0.1% BSA, 2 mM HEPES, and 0.5 mM 3-isobutyl-l-methylzanthine (IBMX, a non-specific phosphodiesterase inhibitor) (Sigma, St. Louis, MO). To test the samples, hCMs were incubated for 10 minutes in treatment buffer only (vehicle) or treatment buffer with MV81 at doses of 10'8M or 10'7M. After treatment for 10 minutes, cells were washed with phosphate buffered solution (PBS) once, lysed with 0.1 M HC1, and sonicated for 10 minutes. Intracellular cGMP levels in the lysates were measured using a commercial cGMP ELISA kit (Enzo Life Sciences, Farmingdale, NY) as instructed by the manufacturer.

[0232] Primary human cardiac fibroblasts (hCFs): Primary human cardiac fibroblasts (hCFs). isolated from a healthy donor without known cardiovascular disease, were Attorney Docket No. 07039-2327WO1

[0233] 2024-159 procured from Cell Biologies Inc. and maintained in Fibroblast Media 2 (PromoCell) supplemented with 10% fetal bovine serum. FGF2, and insulin according to the manufacturer’s instructions. Cells from passages 4-6 were cultured to 80% confluence in 6 well plates, washed with PBS, and incubated with treatment buffer (HBSS, 2mM HEPES buffer, 0.1% bovine serum albumin. 0.5mM IBMX) for 10 minutes. The treatment buffer was aspirated, and the cells were treated with treatment buffer containing vehicle (5% dextrose in water) or 10‘7M peptide (ANP, CNP, or MV81) for 15 minutes. The buffer was aspirated, and the cells were well washed with PBS before lysing them with 0. IM HC1. The cells were scraped wi th a spatula and placed at -80°C for downstream analysis. Intracellular cGMP levels were quantified using a commercially available ELISA (Enzo) according to the manufacturer’s instructions. The cGMP values were normalized to total protein content using the Pierce BCA Protein Assay Kit (Thermo Fisher).

[0234] Human THP-1 Macrophages: THP-1 monocytes (a human leukemia monocytic cell line) were differentiated into M0 macrophages incubation with 150nM phorbol 12- myristate 13-acetate for 24 hours, followed by another 24 hours of incubation in RPMI medium supplemented with 10% fetal bovine serum and 1% pen / strep. M0 macrophages were washed and pre-incubated with treatment buffer (HBSS, 2mM HEPES buffer, 0.1% bovine serum albumin. 0.5mM IBMX) for 10 minutes. The treatment buffer was aspirated, and the cells were treated with treatment buffer containing vehicle (5% dextrose in water) or 10'7M MV81 for 15 minutes. The buffer was aspirated, and the cells were w ashed with PBS before lysing the cells with 0. IM HC1. The cells were scraped with a spatula and placed at -80°C for downstream analysis. Intracellular cGMP levels were quantified using a commercially available ELISA (Enzo) according to the manufacturer’s instructions. M0 macrophages were pre-treated with vehicle or MV81 (10‘7M) for 24 hours and then polarized to Ml macrophages by incubating with 10 pg / mL of lipopolysaccharide (LPS) and 20 ng / mL of interferon gamma (IFN-y). Additionally, at the time of polarization, the cells were supplied with fresh MV81 (10'7M). Following 24 hours of polanzation, the cells were washed with PBS and then stored in TR1ZOL® at - 80°C for downstream analysis.

[0235] Acute Infusion In Vivo Rat Experiments: A total of 13 Sprague Dawley Rats (male, 250-375 g) were randomly treated with one of the following: (i) vehicle (0.9% saline, n=5), (ii) MV 81 (100 pmol / kg / min, n=4), or (iii) MV81 (600 pmol / kg / min, n=4). Attorney Docket No. 07039-2327WO1

[0236] 2024-159

[0237] Investigators who conducted the in vivo study were not blinded to the treatments, but biochemical analysis was performed by a different investigator who was blinded to the treatment.

[0238] On the day of the experiment, the rats were anesthetized with isoflurane and kept on a heating pad at 37°C to maintain a normal body temperature for the entire study. Vascular and bladder cannulation procedures were conducted. Briefly, a polyethylene-50 (PE-50) tube catheter was placed into a jugular vein for saline / peptide infusion and another PE-50 tube was placed into the carotid artery for blood sampling. Urine samples were passively collected from the bladder through a PE-90 tube catheter. A 15-minute equilibrium period was followed by a 30-minute pre-infusion period to collect baseline urine samples. The pre-infusion period was followed by a 45-minute continuous infusion of vehicle or MV81 (15-minute lead-in drug infusion and 30-minute clearance during drug infusion). Immediately after peptide infusion ended, another 30-minute washout clearance period was started before termination and sacrifice. Two additional urine samples were collected during the 30-minute infusion clearance and during the 30-minute washout clearance periods, respectively. Blood samples were collected and placed in tubes containing EDTA on ice at baseline, at the end of 30-minute drug infusion, at the middle of the washout clearance period, and at the end of the washout clearance period, to determine the levels of circulating cGMP. Blood was centrifuged at 2500 rpm at 4°C for 10 minutes, and the plasma was aliquoted. All plasma and urine were stored at -80°C until being assayed. For biochemical analysis, plasma cGMP levels were measured with a cGMP ELISA kit (Enzo Life Sciences, Farmingdale, NY). Urinary sodium concentrations were determined with an electrocyte analyzer (Diamond Diagnostics Inc., Holliston, MA).

[0239] Normal Wild-Type Mouse Studies: Male mice between 4-5 month-old male mice of 129Sv / Ev background were used to assess the dose response of chronic subcutaneous infusion of MV81 via osmotic minipump (Alzet) for 28 days according to the manufacturer’s instructions. For the dosing study, mice (N=3 per group) were randomly treated with vehicle (5% dextrose in water) or MV81 (25 pmol / kg / min. 50 pmol / kg / min, or 100 pmol / kg / min). Following 28 days of infusion, mice were euthanized, and blood samples were obtained via right ventricular puncture using EDTA-washed syringes. Samples were immediately placed on ice. Plasma was isolated by centrifugation at 4°C and stored at -80°C until analysis. The heart and left ventricle were harvested, dissected, Attorney Docket No. 07039-2327WO1

[0240] 2024-159 weighed, snap frozen in liquid nitrogen, and stored at -80°C. Plasma concentrations of cGMP were measured using a commercially available assay (Complete cGMP Assay Kit, Enzo Life Sciences), according to the manufacturer’s instructions.

[0241] Gene Expression in Human Macrophages arid Normal Wild-Type Mice: Gene expression was analyzed following RNA isolation from human macrophages or from previously frozen mouse left ventricle tissue using TRIZOL® and the Direct-zol RNA MiniPrep Kit (Zymo Research). Reverse transcription was performed using the SuperScrip IV First-Strand Synthesis System (Thermo Fisher). Quantitative real-time polymerase chain reactions were performed using IQ™ SYBR* GREEN Supermix (Bio Rad). Mouse-specific primer sequences were obtained from pga.mgh.harvard.edu / primerbank / . Gene expression was calculated using the 2'AACtmethod. For human macrophages, 18S rRNA was used as a housekeeping gene and for mouse tissue, Gapdh was used as a housekeeping gene. Target genes included Nppb (an established marker of wall-stress and hypertrophy), Collal (a component of fibrillar collagen, which is a major component of the extracellular matrix), Postn (a marker of fibroblast activation and component of the extracellular matrix). Tgfbl (a pro- hypertrophic, pro-fibrotic, and pro-inflammatory cytokine), Serpinel (a marker of intracellular TGFP activation), Fap (fibroblast activation protein), and Runxl (a pro- fibrotic and pro-inflammatory transcription factor).

[0242] R4030 Mouse Model of Hypertrophic Cardiomyopathy: The R403Q mouse is a model of hypertrophic cardiomyopathy. R403Q mice aged 9-17 weeks were randomly treated with vehicle (5% dextrose in water) or MV81 (50 pmol / kg / min) for 3 months. Minipumps were exchanged approximately every 28 days. Echocardiography (echo; VisualSonics) was performed at baseline and repeated before osmotic pump exchange and the terminal point of the study (after about 3 months of treatment). Thickness of the inferior wall of the LV at end-diastole w as measured using short axis M-mode images. Diastolic function was measured with apical four chamber view as the ratio of early peak mitral filling velocity (E) to the velocity of the septal mitral annulus during diastole (e'). Cardiac output was measured in a standard manner using a trace of the LV endocardial borders during systole and diastole. In separate experiments, mice began treatment at approximately 6-8 months of age, after the onset of LV hypertrophy (which w as confirmed by echo) and treated with MV81 (50 pmol / kg / min) for 2 months. At the terminal timepoint of the study, the mice were euthanized. Blood samples were obtained Attorney Docket No. 07039-2327WO1

[0243] 2024-159 via right ventricular puncture using EDTA-washed syringes. Samples were immediately placed on ice. Plasma was isolated by centrifugation at 4°C and stored at -80°C until analysis. The heart and left ventricle were dissected, weighed, snap frozen in liquid nitrogen, and stored at -80°C. Plasma concentrations of cGMP were measured using a commercially available assay (Complete cGMP Assay Kit, Enzo Life Sciences), according to the manufacturer’s instructions.

[0244] Transverse Aortic Constriction in Mice: Transverse aortic constriction (TAC) is a well-established model of pressure overload heart failure, which can induce cardiac hypertrophy and dysfunction. Male 8-10 week old C57BL / 6 mice underwent baseline echo prior to TAC using a 25 gauge needle or a sham procedure. Mice underwent echo once weekly, and at three weeks it was apparent that mice in the TAC group had progressive left ventricular (LV) hypertrophy based on echocardiographic measures of LV wall thickness, as described above. TAC mice were then randomized to vehicle (5% dextrose in water) or MV81 (50 pmol / kg / min) via subcutaneous osmotic minipump. Mice were followed for another three weeks and were euthanized, and the heart, left ventricle, and left atrium were dissected, w eighed, snap frozen in liquid nitrogen and stored at - 80°C.

[0245] RESULTS cGMP in human cardiomyocytes (hCMs), human cardiac fibroblasts (hCFs), and human THP-1 macrophages'. MV81 induced a dose dependent increase in cGMP levels in hCMs as compared to vehicle (FIG. 1). MV81 also induced higher cGMP levels in hCFs compared to ANP and CNP at 10'7M (FIG. 2). In addition, MV81 induced an increase in cGMP levels in human THP-1 macrophages as compared to vehicle (FIG. 3). Furthermore. MV81 attenuated the expression of the pro-inflammatory cytokines IL 1 and IL-6 (FIG. 4), as well as the expression of scavenger receptor CD86 (FIG. 5) in THP-1 macrophages polarized to Ml state with LPS+IFNy. Collectively, these data indicated that MV81 can activate GC-A and / or GC-B in multiple cell ty pes as shown by the generation of the second messenger of GC-A and GC-B, cGMP. This activation supported the potential for multiple mechanisms of therapeutic benefit involving pathways that are known to contribute to the development or progression of heart disease (e.g., HCM and / or HF). For example, these studies suggested that targeting hCFs with a peptide such as MV81 can favorably modulate extracellular remodeling and fibrosis, and suggested that targeting hCMs with a peptide such as MV81 can beneficially influence Attorney Docket No. 07039-2327WO1

[0246] 2024-159 cellular mechanics (e.g., systolic and / or diastolic function), myocyte metabolism, and / or myocyte electrophysiology, and suggested that targeting macrophages or other immune cells with a peptide such as MV81 can modulate inflammation and / or cross-talk with other cells, including but not limited to hCFs and / or hCMs.

[0247] Plasma and Urinary cGMP and Urinary Sodium Excretion in Normal Rats: MV81 increased plasma cGMP (FIG. 6A) and urinary’ cGMP (FIG. 6B) levels in a dose dependent manner in normal rats. In particular, both 100 pmol / kg / min and 600 pmol / kg / min doses of MV81 resulted in a sustained elevation in plasma cGMP levels during the first fifteen minutes of the washout period compared to the vehicle. Furthermore, compared to the 100 pmol / kg / min of MV81 and the vehicle, the 600 pmol / kg / min of MV81 resulted in a sustained elevation in plasma cGMP levels 30 minutes into the washout period. MV81 was more effective at increasing urinary sodium excretion compared to the vehicle, both at the 100 pmol / kg / min dose and at the 600 pmol / kg / min dose (FIG. 7). In normal rats, the 600 pmol / kg / min of MV81 was more effective than 100 pmol / kg / min dose. Overall. MV81 increased urinary sodium excretion compared to vehicle, and the increase in urine sodium observed during the infusion was sustained during the washout period (FIG. 7).

[0248] Plasma cGMP Generation, Heart Weight, and Altered Gene Expression in Normal Wild-Type Mice: Chronic subcutaneous infusion of MV81 in normal wild type mice for 28 days induced a dose-dependent increase in plasma cGMP level (FIG. 8). The elevation of plasma cGMP w ith chronic subcutaneous infusion of MV81 in normal wild ty pe mice resulted in a significantly lower heart weight / body weight ratio (FIG. 9) and left ventricle / body weight ratio (FIG. 10). Chronic subcutaneous infusion of MV81 for 28 days also significantly reduced the expression of genes related to hypertrophy’ and fibrosis in the left ventricle, as determined by quantitative polymerase chain reaction (FIG. 11). Nppb is a marker of wall stress and cardiac hypertrophy, and Collal, Postn, Tgfbl, Serpinel, Fap, and Runxl are related to fibroblast activation, and all of these genes are known to be elevated in humans with hypertrophic cardiomyopathy and in the R403Q mouse model of hypertrophic cardiomyopathy (Geisterfer-Lowrance et al.. Science, 272:731-734 (1996)). Chronic subcutaneous infusion of MV81 for 28 days in normal wild-ty pe mice caused a dose-dependent decrease in LV expression of Tgfbl (FIG. 12). Attorney Docket No. 07039-2327WO1

[0249] 2024-159

[0250] Surprisingly, these data indicated MV81 was effective at favorably modifying myocardial gene expression and structure in the absence of overt heart disease.

[0251] Plasma cGMP Generation and LV Structure and Function in R O 30 Mice: To test the effects of MV81 on hypertrophic cardiomyopathy, R403Q mice were treated with chronic subcutaneous infusion of MV81 for three months. Chronic subcutaneous infusion of MV81 for three months increased plasma cGMP levels in the R403Q mice (FIG. 13), indicating that hypertrophic cardiomyopathy is a state of natriuretic peptide / cGMP deficiency. R403Q mice treated with chronic subcutaneous infusion of vehicle for three months resulted in a thicker or hypertrophied LV wall compared to the R403Q mice treated with chronic subcutaneous infusion of MV81for three months (FIG. 14). In addition, a negative correlation was observed between LV thickness and plasma cGMP levels in R403Q mice treated with vehicle or MV81 for three months (FIG. 15). In addition, these studies demonstrated that chronic subcutaneous infusion with MV81 did not modify fractional shortening (a marker of contractility) in the R403Q mice and was comparable to normal wild-type mice (FIG. 16). Chronic subcutaneous infusion of MV81 for three months also maintained normal cardiac output in R403Q mice and was similar to normal wildtype mice (FIG. 17). Further, MV81, compared to vehicle treatment, was associated with lower E / e', which indicated more favorable LV filling pressures and diastolic function, in the R403Q mice (FIG. 18A). Collectively, these data indicated that MV81 can favorably modify adverse myocardial structure and function in mouse HCM model. In further studies, R403Q mice were provided with chronic subcutaneous administration of MV81 (50 pmol / kg / min) or vehicle (Veh) via osmotic minipump for 2 months, with treatment beginning after the onset of disease. MV81 appeared to low er E / e1(FIG. 18B) and resulted in higher e' (FIG. 18C). Normal wild-type mice (WT) served as a control. These studies demonstrated reversal of diastolic dysfunction by MV81.

[0252] MV81 reversed established LV hypertrophy in mice following TAG: Mice developed progressive LV hypertrophy following three weeks of TAC. Mice were then randomized to vehicle or MV81 treatment groups. Treatment with MV81 for three weeks resulted in reversal of LV hypertrophy, whereas mice treated with vehicle showed progression of LV hypertrophy (FIG. 19). Among the TAC mice treated with vehicle, there was a significant increase in left ventricle / body w eight ratio, indicating further progression of LV hypertrophy. Moreover, TAC mice treated with MV81 resulted in normalization of left ventricle / body weight ratio and was similar to sham operated mice Attorney Docket No. 07039-2327WO1

[0253] 2024-159

[0254] (FIG. 20A) TAC mice treated with MV81 resulted in normalization of the left atrial / body weight ratio and were similar to sham operated mice (FIG. 20B). Collectively, these data indicated that MV81 can reverse established LV hypertrophy and normalize LV and LA mass due to pressure overload (as seen in obstructive HCM, HF, aortic stenosis, or hypertension).

[0255] Example 2 - Effects of MV81 in vivo, including myocyte cross-sectional area and transcriptomic signatures MA TERIALS AND METHODS

[0256] Blood collection and plasma cGMP measurement: Blood samples were obtained via right ventricular puncture using EDTA-washed syringes. Samples were immediately placed on ice. Plasma was isolated by centrifugation at 4°C and stored at -80°C until analysis. Plasma concentrations of cGMP w ere measured using a commercially available assay (Complete cGMP Assay Kit, Enzo Life Sciences), according to the manufacturer’s instructions.

[0257] In vitro neprilysin resistance assay: Human cardiac fibroblasts (Lonza) were cultured according to the manufacturer’s instructions. Intracellular cGMP levels were quantified following incubation of the peptide with either vehicle or recombinant human neprilysin (R&D Systems), as described elsewhere (Chen et al., J. Mol. Cell. Cardiol., 130: 140-150 (2019)), with slight modifications. The incubation buffer consisted of HBSS supplemented with 0. 1% BSA and 10 pM zinc chloride. Peptide (400 pL of IO6M) was incubated with either vehicle (saline) or neprilysin (1,000 ng in 100 pL) for 30 minutes at 37°C. Cells were pre-treated with buffer (incubation buffer plus 0.5 mM IBMX), washed, and then treated with either vehicle or peptide (10'7M) in buffer for 15 minutes. Following treatment, cells were washed and then lysed with 0. 1 M HO, and lysates were stored at -80°C. Intracellular cGMP was quantified using the Complete cGMP ELISA Kit (Enzo Life Sciences).

[0258] Histology and myocyte cross sectional area (CSA) quantification: Following euthanasia and exsanguination, the LV was rapidly excised and submerged in KC1 solution to arrest the myocardium in diastole. The tissue w as fixed in 10% formalin for 24 hours and subsequently transferred to 70% ethanol. The LV w as paraffin-embedded, and 5-pm thick sections were mounted onto glass slides. Sections were stained with wheat germ agglutinin (WGA; Invitrogen) to delineate cell borders. Slides were digitally Attorney Docket No. 07039-2327WO1

[0259] 2024-159 scanned at 20* magnification. Myocyte CSA was quantified by manually tracing 50 randomly selected short-axis myocytes per section using ImageJ software. Image acquisition magnification was kept constant, and all analyses were performed in a blinded manner.

[0260] RNA isolation and sequencing: Following euthanasia and exsanguination, the LV was rapidly excised and snap-frozen in liquid nitrogen. Total RNA was extracted using the Direct-zol RNA Miniprep Kit (Zymo Research) following the manufacturer’s protocol. Purified RNA samples were submitted to Novogene (Sacramento, CA) for high- throughput sequencing using paired-end 150 bp reads on an Illumina platform. A nominal read depth of 20 million read pairs per sample was targeted. Raw sequencing data were processed and analyzed using the Partek Genomics Suite. Gene expression counts were normalized to counts per million (CPM) to account for differences in sequencing depth across samples.

[0261] RESULTS

[0262] MV81 increases plasma cGMP in mice with transverse aortic constriction: After six w eeks of sham or TAC surgery', plasma concentrations of cGMP were significantly elevated in mice treated with vehicle for three weeks compared to sham-operated controls. Administration of MV 81 for three weeks resulted in a further increase in plasma cGMP levels relative to both vehicle-treated TAC mice and sham-operated mice (FIG. 21).

[0263] MV81 is resistant to degradation by neprilysin: ANP, CNP, and MV81 each increased intracellular cGMP levels in cultured human cardiac fibroblasts. Incubation with recombinant NEP markedly reduced cGMP generation induced by ANP and CNP (FIG. 22), indicating susceptibility' to enzymatic degradation. In contrast, cGMP generation induced by MV81 was not reduced by NEP (FIG. 22), suggesting that MV81 is resistant to NEP-medicated degradation.

[0264] MV81 normalizes myocyte CSA after transverse aortic constriction: After six weeks of sham or TAC surgery, mice treated with vehicle beginning three weeks after TAC for a duration of three weeks exhibited a significant increase in cardiac myocyte cross-sectional area (CSA) compared to sham-operated controls (FIG. 23). This increase was consistent with pathological hypertrophic remodeling. In contrast, mice treated with MV81 beginning three weeks after TAC for a duration of three weeks showed a Attorney Docket No. 07039-2327WO1

[0265] 2024-159 significantly reduced myocyte CSA relative to vehicle-treated TAC mice (FIG. 23). Myocyte CSA in MV81 -treated mice was not significantly different from that of sham- operated controls, indicating effective reversal of pathological hypertrophy.

[0266] In mice with pressure overload, MV81 modifies transcriptomic signatures associated with adverse cardiac remodeling: RNA sequencing of LV tissue from mice subjected to TAC and treated with either vehicle or MV81 revealed significant transcriptional changes. Gene ontology (GO) enrichment analysis identified several biological processes that were differentially regulated between the two groups. The most significantly overrepresented ontology' in vehicle-treated TAC mice was related to collagen-containing extracellular matrix (FIG. 24), consistent with pathological remodeling and fibrosis. Additional enriched pathways included those associated with cardiac hypertrophy, fibrotic signaling, immune activation, inflammatory response, and metabolic dysregulation (FIG. 24), all of which are hallmarks of pressure overload- induced cardiac dysfunction. For example, in mice subjected to TAC, MV81 reduced the expression of genes associated with hypertrophy, including Nppa (FIG. 25A), Nppb (FIG. 25B), the transcription factor Nfatc3 (FIG. 25C), and Rcanl (FIG. 25D), compared to vehicle treatment. MV81 also suppressed genes related to fibroblast activation, such as Fap (FIG. 26A) and Postn (FIG. 26B), and extracellular matrix deposition, including Col lai (FIG. 26C) and Fnl (FIG. 26D), compared to vehicle treatment. Further, MV81 attenuated the expression of genes involved in inflammation and immunity, such as Ccr2 (FIG. 27A) and II lb (FIG. 27B), as well as transcription factors implicated in macrophage-fibroblast crosstalk, including Meoxl (FIG. 27C) and Runxl (FIG. 27D), compared to vehicle treatment. These findings suggested that MV81 modulates the cardiac transcriptome in a manner that counteracts maladaptive remodeling, supporting its therapeutic role. All enriched ontologies had a false discovery rate (FDR) < 0.05, confirming statistical significance.

[0267] Example 3 - Single-nuclear transcriptomics of particulate guanylyl cyclase receptors in the human heart

[0268] MA TERIALS AND METHODS

[0269] Human heart samples and single nucleus RNA sequencing (snRNA-seq) '. Data from a study described elsewhere (Chaffin et al., Nature, 608(7921):174-180 (2022)) were accessed at the Broad Institute Single Cell Portal under project ID SCP1303 Attorney Docket No. 07039-2327WO1

[0270] 2024-159

[0271] (singlecell.broadinstitute.org / single_cell / study / SCP1303). Briefly, single-nuclei RNA sequencing (snRNAseq) was performed using nearly 600.000 nuclei isolated from failing human left ventricles, including samples from 11 individuals with DCM, 15 with HCM, and 16 non-failing control hearts. UMAPs for GC-A (gene name NPR1) and GC-B (gene name NPR2) were created according to cell type. Results were combined for DCM, HCM, and controls because there was no difference in the pattern of expression of GC-A or GC-B among these three groups.

[0272] RESULTS

[0273] Spatial and cell-specific patterns ofNP receptors gene expression in the human heart'. To resolve cell-specific NP receptor expression dynamics, snRNA-seq datasets obtained as described elsewhere (Chaffin et al., supra) were analyzed. These data offered insights into the cellular expression patterns of NP receptors, thereby informing opportunities for targeted pharmacological intervention. Expression of the GC-A receptor (FIG. 28A) was predominantly detected in endothelial cells, with comparatively less expression in cardiac myocytes, cardiac fibroblasts, vascular smooth muscle cells, pericytes and macrophages. While expression of the GC-B receptor (FIG. 28B) was ubiquitously detected throughout the heart, with higher expression in cardiac myocytes, cardiac fibroblasts, vascular smooth muscle cells, pericytes, and macrophages, but lower expression in endothelial cells compared to GC-A.

[0274] OTHER EMBODIMENTS

[0275] It is to be understood that while the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

Claims

1. Attorney Docket No: 07039-2327WO12024-159WHAT IS CLAIMED IS:

1. Use of a composition comprising a natriuretic peptide for treating a chronic heart condition in a mammal, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

2. The use of claim 1, wherein said mammal is a human.

3. The use of claim 1, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a co w. a sheep, a goat, a rabbit, a mouse, or a rat.

4. The use of claim 1, wherein said composition comprises said natriuretic peptide in an amount effective to reduce one or more symptoms of said chronic heart condition in said mammal.

5. The use of claim 4, wherein said one or more symptoms of said chronic heart condition are selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema.

6. The use of claim 1. wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

7. The use of claim 1, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

8. The use of claim 1. wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.Attorney Docket No: 07039-2327WO12024-1599. The use of claim 1, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 wi th one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

10. Use of a composition comprising a natriuretic peptide for reducing inflammation in a mammal having a chronic heart condition, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO:1 are maintained and are the only two cysteine residues of said natriuretic peptide.1 1. The use of claim 10, wherein said mammal is a human.

12. The use of claim 10, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow. a sheep, a goat, a rabbit, a mouse, or a rat.

13. The use of claim 10, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

14. The use of claim 10, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

15. The use of claim 10, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NOT.

16. The use of claim 10, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.Attorney Docket No: 07039-2327WO12024-15917. Use of a composition comprising a natriuretic peptide for reducing TGF-beta levels or TGF-beta activity in a mammal having a chronic heart condition, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

18. The use of claim 17, wherein said mammal is a human.

19. The use of claim 17, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

20. The use of claim 17, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

21. The use of claim 17, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

22. The use of claim 17, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

23. The use of claim 17, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

24. Use of a composition comprising a natriuretic peptide for reducing ventricular hypertrophy in a mammal having a chronic heart condition, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, orAttorney Docket No: 07039-2327WO12024-159 substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

25. The use of claim 24, wherein said mammal is a human.

26. The use of claim 24, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

27. The use of claim 24, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

28. The use of claim 24, wherein said chronic heart condition is associated with left ventricular hypertrophy, left ventricular remodeling, right ventricular hypertrophy, right ventricular remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

29. The use of claim 24, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

30. The use of claim 24, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

31. Use of a composition comprising a natriuretic peptide for preventing, attenuating, or reversing diastolic dysfunction in a mammal having a chronic heart condition, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

32. The use of claim 31 , wherein said mammal is a human.Attorney Docket No: 07039-2327WO12024-15933. The use of claim 31, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

34. The use of claim 31, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

35. The use of claim 31, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

36. The use of claim 31, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NOT.

37. The use of claim 31, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

38. Use of a composition comprising a natriuretic peptide for treating a mammal having a chronic heart condition in which guanylate cyclase A (GC-A) and guanylate cyclase B (GC-B) are present in heart cells of the mammal, wherein said natriuretic peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only tw o cysteine residues of said natriuretic peptide.

39. The use of claim 38, wherein said mammal is a human.

40. The use of claim 38, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

41. The use of claim 38, wherein said composition comprises said natriuretic peptide in an amount effective to reduce one or more symptoms of said chronic heart condition in said mammal.Attorney Docket No: 07039-2327WO12024-15942. The use of claim 41, wherein said one or more symptoms of said chronic heart condition are selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema.

43. The use of claim 38, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

44. The use of claim 38, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

45. The use of claim 38, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO:1.

46. The use of claim 38, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

47. A method for treating a chronic heart condition, wherein said method comprises administering, to a mammal identified as having said chronic heart condition, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

48. The method of claim 47, wherein said mammal is a human.

49. The method of claim 47, wherein said mammal is a monkey, a dog, a cat, a pig. a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.Attorney Docket No: 07039-2327WO12024-15950. The method of claim 47, wherein said method comprises administering said composition to said mammal in an amount effective to reduce one or more symptoms of said chronic heart condition in said mammal.51 . The method of claim 50, wherein said one or more symptoms of said chronic heart condition are selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema.

52. The method of claim 47, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

53. The method of claim 47, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

54. The method of claim 47, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO:1.

55. The method of claim 47. wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

56. A method for reducing inflammation in a mammal having a chronic heart condition wherein said method comprises administering, to a mammal identified as having said chronic heart condition, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.Attorney Docket No: 07039-2327WO12024-15957. The method of claim 56, wherein said mammal is a human.

58. The method of claim 56, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow. a sheep, a goat, a rabbit, a mouse, or a rat.

59. The method of claim 56, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

60. The method of claim 56, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

61. The method of claim 56, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NOT.

62. The method of claim 56, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

63. A method for reducing TGF-beta levels or TGF-beta activity' in a mammal having a chronic heart condition wherein said method comprises administering, to a mammal identified as having said chronic heart condition, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NOT with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

64. The method of claim 63, wherein said mammal is a human.

65. The method of claim 63, wherein said mammal is a monkey, a dog, a cat, a pig. a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.Attorney Docket No: 07039-2327WO12024-15966. The method of claim 63, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

67. The method of claim 63, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

68. The method of claim 63, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1.

69. The method of claim 63, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

70. A method for reducing ventricular hypertrophy in a mammal having a chronic heart condition, wherein said method comprises administering, to a mammal identified as having said chronic heart condition, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO with zero, one, two. three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO are maintained and are the only two cysteine residues of said natriuretic peptide.

71. The method of claim 70, wherein said mammal is a human.

72. The method of claim 70, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow. a sheep, a goat, a rabbit, a mouse, or a rat.

73. The method of claim 70, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

74. The method of claim 70, wherein said chronic heart condition is associated with left ventricular hypertrophy, left ventricular remodeling, right ventricular hypertrophy,Attorney Docket No: 07039-2327WO12024-159 right ventricular remodeling, systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

75. The method of claim 70, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 .

76. The method of claim 70, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

77. A method for preventing, attenuating, or reversing diastolic dysfunction in a mammal having a chronic heart condition, wherein said method comprises administering, to a mammal identified as having said chronic heart condition, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with zero, one. two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO:1 are maintained and are the only two cysteine residues of said natriuretic peptide.

78. The method of claim 77, wherein said mammal is a human.

79. The method of claim 77, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

80. The method of claim 77, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

81. The method of claim 77, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

82. The method of claim 77, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NOT.Attorney Docket No: 07039-2327WO12024-15983. The method of claim 77, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 wi th one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

84. A method for treating a mammal having a chronic heart condition in which guanylate cyclase A (GC-A) and guanylate cyclase B (GC-B) are present in heart cells of the mammal, wherein said method comprises administering, to said mammal, a composition that comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with zero, one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues within SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

85. The method of claim 84, wherein said mammal is a human.

86. The method of claim 84, wherein said mammal is a monkey, a dog, a cat, a pig, a horse, a cow, a sheep, a goat, a rabbit, a mouse, or a rat.

87. The method of claim 84, wherein said composition comprises said natriuretic peptide in an amount effective to reduce one or more symptoms of said chronic heart condition in said mammal.

88. The method of claim 87, wherein said one or more symptoms of said chronic heart condition are selected from the group consisting of shortness of breath, heart palpitations, chest discomfort, fatigue, dizziness, syncope, presyncope, orthopnea, paroxysmal nocturnal dyspnea, lightheadedness, arrhythmia, heart murmurs, and edema.

89. The method of claim 84, wherein said chronic heart condition is hypertrophic cardiomyopathy (HCM).

90. The method of claim 84, wherein said chronic heart condition is associated with ventricular hypertrophy, ventricular remodeling, atrial hypertrophy, atrial remodeling,Attorney Docket No: 07039-2327WO1 2024-159 systolic dysfunction, diastolic dysfunction, cardiomyopathy, heart failure, or any combination thereof, or is a HCM phenocopy.

91. The method of claim 84, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 .

92. The method of claim 84, wherein said composition comprises a natriuretic peptide comprising the amino acid sequence set forth in SEQ ID NO: 1 with one, two, three, four, five, six, seven, eight, nine, or ten amino acid additions, deletions, or substitutions, provided that the two cysteine residues in SEQ ID NO: 1 are maintained and are the only two cysteine residues of said natriuretic peptide.

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