Therapeutic compounds

Abstract

Provided herein are compounds that are useful in treating cancer, metabolic disease, cardiovascular disease, and kidney disease.

Description

THERAPEUTIC COMPOUNDSCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority from U.S. Provisional Application Serial No. 63 / 500,447 filed on May 5, 2023, and U.S. Provisional Application Serial No. 63 / 588,274 filed on October 5, 2023 each of which is incorporated herein by reference in its entirety.STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT[00011 Not Applicable.FIELD[00021 The present invention relates to therapeutic compounds that interact with the pGC-A / cGMP pathway, and the use of such compounds in the treatment of metabolic disease, cardiovascular disease, kidney disease, and cancer.INTRODUCTION

[0003] Particulate guanylyl receptor A ("pGC-A") is highly expressed in heart tissue, the kidneys, adrenal glands, vasculature, and in adipocytes. pGC-A is able to bind and is activated by Atrial natriuretic peptide (“ANP”) and B-type natriuretic peptide (“BNP”). Following pGC-A activation, cyclic guanosine monophosphate (“cGMP”) is produced resulting in numerous physiological benefits including lowering blood pressure, enhancing renal function, protection of the vasculature, and enhanced kidney function. Therefore, the pGC-A / cGMP pathway is a valuable target for drug interaction which is believed to lead to therapies for cancer, metabolic disease, cardiovascular disease, and kidney disease. See, e.g., international patent application PCT / US2021 / 034889, incorporated herein by reference in its entirety. See. also, Figs. 1 and 2.

[0004] Metabolic disease, cardiovascular disease, kidney disease, and cancer are among the most prevalent, significant causes of morbidity’ and mortality worldwide. While the standards of care for those conditions have greatly improved over the years, current standards of care still fail to meet the need for effective therapies to improve treatments.

[0005] Accordingly, there is a need in the art for new therapies, including, for example, therapies for the treatment of metabolic disease, cardiovascular disease, kidneydisease, and cancer. Provided herein are solutions to these and other problems in the art.SUMMARY

[0006] This summary is provided to introduce a selection of concepts in a simplified form that are further described below in the detailed description. This summary is not intended to identify key features or essential features of the claimed subject matter, nor is it intended to be used as an aid in determining the scope of the claimed subject matter.

[0007] Provided herein, inter alia, are compounds disclosed in Appendix A.

[0008] In another embodiment is a pharmaceutical composition that includes a compound described herein and a pharmacally acceptable excipient.

[0009] In still another embodiment aspect is a kit that includes a pharmacal composition as described herein.

[0010] In still another embodiment is a method for treating metabolic disease, cardiovascular disease, kidney disease, and / or cancer by administering a therapeutically effective amount of a pharmaceutical composition described herein to a patient in need thereof.

[0011] According to an aspect of the present disclosure, a compound of Formula I is provided:The compound is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor and is orally available.

[0012] According to another aspect of the present disclosure, a method of treating a cardiovascular disease in a subject is provided. The method includes administering to the subject an effective amount of a compound disclosed in Appendix A, wherein the compound is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available. In various embodiments, the compound is that of Formula I.

[0013] According to yet another aspect of the present disclosure, a pharmaceutical composition is provided. The composition includes a compound disclosed in Appendix A, wherein the compound is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available. The composition alsoincludes a pharmaceutically acceptable carrier. In various embodiments, the compound is that of Formula I.

[0014] According to other aspects of the present disclosure, the pharmaceutical composition may include a compound disclosed in Appendix A that is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form selected from the group consisting of a tablet, a capsule, a liquid suspension, and a powder. The pharmaceutical composition may also include a compound disclosed in Appendix A that is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a controlled-release formulation, a fast-dissolving formulation, a chewable tablet, a liquid suspension suitable for pediatric administration, a powder for reconstitution, a capsule containing a powder formulation, a tablet coated with an enteric coating, a tablet containing a disintegrant, a binder, a lubricant, a glidant, a filler, a colorant, a flavoring agent, a sweetening agent, or a preservative. In various embodiments, the compound is that of Formula I.

[0015] These and other features, aspects and advantages of the present teachings will become better understood with reference to the following description, examples and appended claims.DRAWINGS

[0016] Those of skill in the art will understand that the drawings, described below, are for illustrative purposes only. The drawings are not intended to limit the scope of the present teachings in any way.

[0017] Figure 1. This figure depicts the effect of the pGC-A / cGMP pathway on a series of cell types including fibroblasts, cardiomyocytes, endothelial cells, smooth muscle cells, inner medullary collecting duct cells, zona glomerulosa cells, and adipocytes.

[0018] Figure 2. This figure depicts the effect of modulating the pGC-A / cGMP pathway which results in vasodilation and decreased fibrosis / cardiac hypertrophy, and increased natriuresis / diuresis.

[0019] Figure 3. This figure shows the first step of the synthesis of compound AR-1122.

[0020] Figure 4. This figure shows the third step of the synthesis of compound AR-1122.

[0021] Figure 5. This figure shows the first step of the synthesis of compound AR-2123.

[0022] Figure 6. This figure shows the second step of the synthesis of compoundAR-2123.

[0023] Figure 7. This figure shows the third step of the synthesis of compound AR-2123.

[0024] Figure 8. This figure shows the synthesis of compound AR- 16A72.

[0025] Figure 9. This figure shows Mean Arterial Pressure (MAP), Systolic BloodPressure (SBP), Diastolic Blood Pressure (DBP), and Heart Rate (HR) data of Vehicle andAR-1121-S (PO).

[0026] Figure 10. This figure shows Mean Arterial Pressure (MAP), Systolic Blood Pressure (SBP), Diastolic Blood Pressure (DBP), and Heart Rate (HR) data of AR-1121 -S 5.0mg / kg (IV).

[0027] Figure 11. This figure shows Mean Arterial Pressure (MAP), Systolic Blood Pressure (SBP), Diastolic Blood Pressure (DBP), and Heart Rate (HR) data of Vehicle and AR-1121-R 10.0mg / kg PO.

[0028] Figure 12. This figure shows Mean Arterial Pressure (MAP), Systolic Blood Pressure (SBP), Diastolic Blood Pressure (DBP), and Heart Rate (HR) data of Vehicle and MCUF-651 5.7mg / kg (PO).

[0029] Figure 13. This figure shows cGMP Generation with MCUF-651 in HEK293 pGC-A Overexpressing Cells. A source of this data, and data provided below, can be found in Sangaralingham, et al. Proc Natl Acad Sci U S A. 2021 Dec 28; 118(52):e2109386118, incorporated herein by reference in its entirety.

[0030] Figure 14. This figure shows MCUF-651 binding to pGC-A confirmed usingSPR.Figure 15. This figure shows that MCUF-651 potentiates cGMP generation in the presence of ANP in Human Primary Cells. P-value < 0.05 versus ANP alone (blue bar).

[0031] Figure 16. This figure shows that MCUF-651 enhances ANP-mediated suppression of Human Cardiomyocyte Hypertrophy in vitro.

[0032] Figure 17. This figure shows that MCUF-651 is orally available in mice.

[0033] Figure 18. This figure shows an early pilot study whereby MCUF-651 increases cGMP, decreases MAP and increases urine volume in SHRs.

[0034] Figure 19. This figure shows a study of cGMP levels and mean arterial pressure with MCUF-651 Acetate.

[0035] Figure 20. This figure shows a study of urinary sodium excretion rate withMCUF-651 Acetate.

[0036] Figure 21. This figure shows the potential to develop a companion diagnostic. The assay estimates a patient’s potential cGMP production with exposure to pGC- A PAM using patient plasma.

[0037] Figure 22. This figure shows a PK study and plasma protein binding.

[0038] Figure 23. This figure shows a 24 hour telemetry study (also shown elsewhere) in SHR Rats SBP, and change from baseline, comparing oral to IV administration.

[0039] Figure 24. This figure shows a 24 hour telemetry study (also shown elsewhere) in SHR Rats SBP, and change from baseline, comparing S versus R enantiomers.

[0040] Figure 25. This figure shows a 24 hour telemetry study in SHR Rats HR, and change from baseline, comparing oral versus IV administration of the S enantiomer.DETAILED DESCRIPTION

[0041] All patents, applications, published applications and other publications cited herein are incorporated by reference in their entirety. Unless defined otherwise, all technical and scientific tenns used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the invention belongs. The chemical structures and formulae set forth herein are constructed according to the standard rules of chemical valency know n in the chemical arts. Should a discrepancy exist betw een a depicted structure and a name given for that structure, the depicted structure is to be accorded more weight. Where the stereochemistry of a structure or a portion of a structure is not indicated in a depicted structure or a portion of the depicted structure, the depicted structure is to be interpreted as encompassing all of its possible stereoisomers.

[0042] Any methods, devices and materials similar or equivalent to those described herein can be used in the practice of this invention. The following definitions are provided to facilitate understanding of certain terms used frequently herein and are not meant to limit the scope of the present disclosure. In the event that there is a plurality of definitions for a term herein, those in this section prevail unless stated otherwise. Headings used herein are for organizational purposes only and in no way limit the invention described herein.

[0043] Abbreviations and Definitions

[0044] To facilitate understanding of the invention, a number of terms and abbreviations as used herein are defined below as follows:

[0045] “Effective Amount” and “Therapeutically Effective Amount”: The term“effective amount” refers to the amount of a therapy (e.g., a compound provided herein) which is sufficient to accomplish a stated purpose or otherwise achieve the effect for which it is administered. An effective amount can be sufficient to reduce and / or ameliorate the progression, development, recurrence, severity and / or duration of a given disease, disorder or condition and / or a symptom related thereto, or can be sufficient to modulate the level of activity or binding of a polypeptide. An effective amount can be a “therapeutically effective amount” which refers to an amount sufficient to provide a therapeutic benefit such as, for example, the reduction or amelioration of the advancement or progression of a given disease, disorder or condition, reduction or amelioration of the recurrence, development or onset of a given disease, disorder or condition, and / or to improve or enhance the prophylactic or therapeutic effect(s) of another therapy. A therapeutically effective amount of a composition described herein can enhance the therapeutic efficacy of another therapeutic agent.

[0046] “Regimen”: The term “regimen” refers to a protocol for dosing and timing the administration of one or more therapies (e.g. , compounds described herein or another active agent such as a compound of Appendix A described herein) for treating a disease, disorder, or condition described herein. A regimen can include periods of active administration and periods of rest as known in the art. Active administration periods include administration of compounds and compositions described herein and the duration of time of efficacy of such compounds and compositions. Rest periods of regimens described herein include a period of time in which no compound is actively administered, and in certain instances, includes time periods where the efficacy of such compounds can be minimal. Combinations of active administration and rest in regimens described herein can increase the efficacy and / or duration of administration of the compositions described herein.

[0047] “Therapy” or “Therapies”: The terms “therapy” and “therapies” refer to any protocol(s), method(s), and / or agent(s) that can be used in the prevention, treatment, management, and / or amelioration of a disease, disorder, or condition or one or more symptoms thereof. In certain instances the term refers to active agents such as the compounds of Appendix A described herein. The tenns “therapy” and “therapy” can refer to anti-viral therapy, anti-bacterial therapy, anti-fungal therapy, anti-cancer therapy, biological therapy, supportive therapy, and / or other therapies useful in treatment, management, prevention, or amelioration of a disease, disorder, or condition or one or more symptoms thereof known to one skilled in the art, for example, a medical professional such as a physician.

[0048] “Patient” or “Subject”: The tenn “patient” or "subject” refers to a mammal, such as a human, bovine, rat, mouse, dog, monkey, ape, goat, sheep, cow; or deer.Generally, a patient as described herein is human.

[0049] “Inhibit”, “Inhibiting”, or “Inhibition”: The terms ‘inhibit”, “inhibiting”, or “’inhibition,” “inhibit,” and “inhibiting” refer to a reduction in the activity, binding, or expression of a polypeptide or reduction or amelioration of a disease, disorder, or condition or a symptom thereof. Inhibiting as used here can include partially or totally blocking stimulation, decreasing, preventing, or delaying activation or binding, or inactivating, desensitizing, or down-regulating protein or enzyme activity or binding.

[0050] “Metabolic Disease”: The term “metabolic disease” refers to a congenital metabolic disorder. Examples of inherited metabolic diseases include Fabry disease, phenylketonuria, Prader-Willi syndrome, galactosemia, Tay-Sachs's disease, porphyria, Pompe disease, Neimann-Pick disease, Morquio s syndrome, Morteaus-lamy syndrome, Hunter syndrome, Lesh-Nyhan syndrome, Hurler syndrome, homocystinuria, Hartnup disease, and Gaucher’s disease. In other examples, the term “metabolic disease” refers to an acquired metabolic disorder. Examples of such disorders include diabetes (e.g., type 1 diabetes, diabetes insipidus, or type II diabetes mellitus), obesity, metabolic syndrome, dyslipidemia, hipolipidemia (hyperlipoproteinemia), hyperthyroidism, hypoparathyroidism, hypothyroidism, Cushing’s syndrome, hyperuricemia, hemochromatosis, hyperparathyroidism, glucose intolerance, insulin resistance, fibrinolysis disorder, endothelial dysfunction, atherosclerosis, impaired fasting glycemia, hyperinsulinemia, galactosemia, mucopolysaccaridose, tyrosinemia, methylmalonic aciduria, acidemia (e.g., propionic acidemia, isovaleric acidemia), and hyperammonemia. In yet other examples, the term “metabolic disease” refers to obesity, hypertriglyceridemia, metabolic syndrome, insulin resistance, hyperinsulinemia, diabetes, and acidemia.

[0051] “Cardiovascular Disease”: The term “cardiovascular disease” refers to severe or congestive heart failure, atrial fibrillation, atrial flutter, atrial tachycardia, dilated cardiomyopathy, hypertrophic myocardium disease, congenital anomalies, heart failure with glomerulonephritis, idiopathic aortic stenosis, acute myocardial infarction, myocarditis, myxedema, chronic obstructive pericarditis, ventricular tachycardia, extraventricular contraction, incomplete chamber ventricular block, severe lung disease or hypoxia, carotid sinus hypersensitivity, bradycardia or complete atrioventricular block (if the block is not due to digoxin), and combinations thereof. Also included are conditions resulting in abnormal I irregular heart rate, weakness, shortness of breath, decreased exercise tolerance, lethargy, syncope, hypoxia, pulmonary edema, ascites, loss of consciousness, etc., and combinations thereof.

[0052] “Kidney Disease”: The term “kidney disease” refers to goodpasture syndrome, ectopic kidney, amyloidosis, acquired cystic kidney disease, glomerular disease, kidney dysplasia, medullary sponge kidney, nephrotic syndrome, kidney damage, renal artery stenosis, renal tubular acidosis, and solitary kidney. In some embodiments, the kidney disease is selected from nephropathy, acute renal failure, chronic kidney disease, cardiorenal syndrome and diabetic kidney disease.

[0053] “Cancer”: The term “cancer” refers to any physiological condition in mammals characterized by unregulated cell growth. Cancers described herein include solid tumors and hematological (blood) cancers. A “hematological cancer” refers to any blood borne cancer and includes, for example, myelomas, lymphomas and leukemias. A “solid tumor” or “tumor” refers to a lesion and neoplastic cell growth and proliferation, whether malignant or benign, and all pre-cancerous and cancerous cells and tissues resulting in abnormal tissue growth. “Neoplastic,” as used herein, refers to any form of dysregulated or unregulated cell grow th, whether malignant or benign, resulting in abnormal tissue growth.

[0054] “Treat”, “Treating” or “Treatment”: The terms “treat”, “treating”, or “treatment” refer to any indicia of success or amelioration of the progression, severity, and / or duration of a disease, pathology or condition, including any objective or subjective parameter such as abatement; remission; diminishing of symptoms or making the injury, pathology or condition more tolerable to the patient; slowing in the rate of degeneration or decline; making the final point of degeneration less debilitating; or improving a patient’s physical or mental well-being.

[0055] “Enhance”: The term “enhance” refers to an increase or improvement in the function or activity of a protein or cell after administration or contacting with a compound described herein compared to the protein or cell prior to such administration or contact.

[0056] “Administer” or “Administering”: The terms “administer” or “administering” refers to the act of delivering a composition described herein into a subject by such routes as oral, mucosal, topical, suppository, intravenous, parenteral, intraperitoneal, intramuscular, intralesional, intrathecal, intranasal or subcutaneous administration. Parenteral administration includes intravenous, intramuscular, intra-arteriole, intradennal, subcutaneous, intraperitoneal, intraventricular, and intracranial administration. Administration generally occurs after the onset of the disease, disorder, or condition, or its symptoms but, in certain instances, can occur before the onset of the disease, disorder, or condition, or its symptoms (e.g.. administration for patients prone to such a disease, disorder, or condition).

[0057] “Co-Administration”: The term “co-administration” refers toadministration of two or more agents (e.g., a compound described herein and another active agent). The timing of co-administration depends in part of the compound and compositions administered and can include administration at the same time, just prior to, or just after the administration of one or more additional therapies, for example cancer therapies such as chemotherapy, hormonal therapy, radiotherapy, or immunotherapy. The compound of the invention can be administered alone or can be co-administered to the patient. Coadministration is meant to include simultaneous or sequential administration of the compound individually or in combination (more than one compound or agent). Thus, the preparations can also be combined, when desired, with other active substances (e.g., to reduce metabolic degradation). A first compound or agent can also be used Epigenetic Sensitization of a tumor within a patient before administration of a second compound or agent. The compounds described herein can be used in combination with one another, and with other active agents known to be useful in treating cancer.

[0058] “Anti-Cancer Agent”: The term “anti-cancer agent” is used in accordance with its plain ordinary meaning and refers to a composition having anti-neoplastic properties or the ability to inhibit the growth or proliferation of cells. In various embodiments, an anticancer agent is a chemotherapeutic agent. In embodiments, an anti-cancer agent is an agent, such as a compound of Appendix A identified herein, having utility in methods of treating cancer. In embodiments, an anti-cancer agent is an agent approved by the FDA or similar regulatory agency of a country other than the USA, for treating cancer.

[0059] “Chemotherapeutic” or “Chemotherapeutic Agent”: The term “chemotherapeutic” or “chemotherapeutic agent” is used in accordance with its plain ordinary meaning and refers to a chemical composition or compound having anti-neoplastic properties or the ability to inhibit the growth or proliferation of cells. “Chemotherapy” refers to a therapy or regimen that includes administration of a chemotherapeutic or a compound of Appendix A described herein.

[0060] Therapeutic Compounds

[0061] The present disclosure relates to the field of therapeutic compounds, specifically to positive allosteric modulators (PAMs) that activate the particulate guanylyl cyclase-A (pGC-A) receptor. The pGC-A receptor is a validated target for various cardiovascular and renal conditions, including resistant hypertension (rHTN) and multiple heart failure (HF) etiologies. Activation of the pGC-A receptor leads to the production of cyclic guanosine monophosphate (cGMP), which has numerous physiological benefitsincluding lowering blood pressure, enhancing renal function, protecting the vasculature, and enhancing kidney function. Therefore, the pGC-A / cGMP pathway is a valuable target for drug interaction, which can lead to therapies for metabolic disease, cardiovascular disease, kidney disease, and cancer.

[0062] In some aspects, the present disclosure provides compounds that are positive allosteric modulators of the pGC-A receptor. These compounds may enhance the activity of the pGC-A receptor, thereby increasing the production of cGMP and providing the associated physiological benefits. Importantly, these compounds are orally available, providing a convenient route of administration for patients.

[0063] In other aspects, the present disclosure provides methods for treating cardiovascular diseases using the disclosed compounds. These methods involve administering an effective amount of the compound to a subject in order to activate the pGC-A receptor and increase cGMP production. This can lead to a reduction in blood pressure and other beneficial effects, thereby treating or ameliorating the symptoms of the cardiovascular disease.

[0064] Further aspects of the present disclosure relate to pharmaceutical compositions that include the disclosed compounds. These compositions may be formulated for oral administration and may include various pharmaceutically acceptable excipients. The compositions may be provided in various dosage forms, such as tablets, capsules, liquid suspensions, or powders, depending on the specific requirements of the patient or the disease being treated.

[0065] In yet other aspects, the present disclosure provides a kit that includes the disclosed pharmaceutical compositions. This kit may be used for the treatment of cardiovascular diseases, metabolic diseases, kidney diseases, and cancer, among other conditions.

[0066] Overall, the compounds, methods, compositions, and kits disclosed herein provide a novel approach to modulating the pGC-A / cGMP pathway, with potential applications in the treatment of a wide range of diseases and conditions.

[0067] Referring to FIG. I. the diagram illustrates the interaction of the particulate guanylyl cyclase-A (pGC-A) receptor with various cell types and the resulting physiological effects. The pGC-A receptor, located on the extracellular membrane, may be activated by Atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP), leading to the production of cyclic guanosine monophosphate (cGMP) from guanosine triphosphate (GTP). This cGMP then activates cGMP-gated cation channels and protein kinases (PKGs), which inturn influence different cell types in the heart, blood vessels, kidney, and adipose tissue.

[0068] In some aspects, cGMP may protect the heart from hypertrophy and fibrosis by acting on fibroblasts and cardiomyocytes. In other cases, cGMP may protect blood vessels from injury and induce vasodilation through endothelial and smooth muscle cells. In yet other aspects, cGMP may promote salt excretion from the kidney via inner medullary collecting duct cells, and stimulate lipolysis and browning of adipocytes in adipose tissue. Additionally, cGMP may inhibit aldosterone production by the adrenals through its action on zona glomerulosa cells.

[0069] In some cases, the compounds disclosed in Appendix A, including but not limited to the compound of Formula I, may act as a positive allosteric modulator of the pGC- A receptor. This compound may enhance the activity of the pGC-A receptor, thereby increasing the production of cGMP and providing the associated physiological benefits. Importantly, this compound may be orally available, providing a convenient route of administration for patients.

[0070] In other aspects, the present disclosure provides methods for treating cardiovascular diseases using the disclosed compounds. These methods may involve administering an effective amount of the compound to a subject in order to activate the pGC- A receptor and increase cGMP production. This can lead to a reduction in blood pressure and other beneficial effects, thereby treating or ameliorating the symptoms of the cardiovascular disease.

[0071] Further aspects of the present disclosure relate to pharmaceutical compositions that include the disclosed compounds. These compositions may be formulated for oral administration and may include various pharmaceutically acceptable excipients. The compositions may be provided in various dosage forms, such as tablets, capsules, liquid suspensions, or powders, depending on the specific requirements of the patient or the disease being treated.

[0072] Referring to FIG. 2, the diagram illustrates the molecular interactions and pathways involved in the modulation of cardiovascular functions. In some aspects, compounds may bind to the particulate guanylyl cyclase-A (pGC-A) receptor, leading to an increase in cyclic guanosine monophosphate (cGMP) production. The diagram shows the action of natriuretic peptides (ANP and BNP) and C-type natriuretic peptide (CNP) on the pGC-A receptor, resulting in vasodilation, reduced fibrosis / cardiac hypertrophy, and increased natriuresis / di uresis.

[0073] In contrast, angiotensin II (Ang II) may act on its receptor (AT1R),triggering a signaling cascade that leads to vasoconstriction, increased fibrosis / cardiac hypertrophy, and retention of sodium and water. The enzy me neprilysin is also depicted, which degrades natriuretic peptides into inactive fragments, thereby modulating their availability and effects on the pGC-A receptor.

[0074] In some cases, the method of treating a cardiovascular disease in a subject may involve administering an effective amount of the compounds disclosed in Appendix A, including but not limited to the compound of Formula I. These compounds may act as a positive allosteric modulator of the pGC-A receptor, thereby increasing the production of cGMP and providing the associated physiological benefits. This method may be used for treating a cardiovascular disease in a subject, providing a novel approach to modulating the pGC-A / cGMP pathway.

[0075] Referring to FIG. 3, the figure illustrates a schematic representation of the synthesis process for a compound. The process begins with difluorobenzothiazole amine 1, which reacts with reagent 2 under specific conditions to form intermediate compound 3. This intermediate compound 3 then undergoes further reaction, again under specific conditions, to yield the final Step 1 product 4. The synthesis steps involve the use of various reagents and conditions to facilitate the transformation from the starting amine to the final Step 1 carboxamide compound.

[0076] Referring to FIG. 4. the figure illustrates a flowchart depicting the synthesis process of AR-1122 in Step 3. The process begins with the compound MCUF-651. which undergoes a reaction with NaCNBH3 and methanol (MeOH). This reaction is followed by treatment with hydrochloric acid (HCl) in dioxane to yield an intermediate compound. This intermediate compound is then reacted with Etl and triethylamine (Et3N) in dichloromethane (DCM) to produce the final compound AR-1122. Each step of the process is annotated with specific conditions such as temperature, duration, and yield percentage, indicating the efficiency of each reaction stage. This process may be used to produce a compound that is a positive allosteric modulator of the pGC-A receptor, which may be orally available and maybe used in the treatment of cardiovascular diseases.

[0077] Referring to FIG. 5. the figure illustrates a schematic representation of the synthesis process for a chemical compound. The process begins with difluorobenzothiazole amine 1, which reacts with reagent 2 under specific conditions to form intermediate compound 3. This intermediate compound 3 then undergoes further reaction with hydrochloric acid (HCl) to yield the final Step 1 product 4. The synthesis steps are detailed with specific conditions and yields for each transformation.

[0078] Referring to FIG. 6, the figure illustrates a schematic representation of a chemical synthesis process. The process begins with the starting compound difluorobenzothiazole amine 1, which reacts with the first step reagents and conditions reagent 2 to form the intermediate compound 3. Subsequently, the intermediate compound 3 undergoes further reaction as per the conditions outlined in step reagent 2 to yield the final Step 2 product 4. The synthesis steps involve the use of various reagents and conditions to facilitate the transformation from the starting amine to the final carboxamide compound.

[0079] Referring to FIG. 7, the figure illustrates a schematic representation of a chemical synthesis process. The process begins with difluorobenzothiazole amine 1, which is transformed into a product through a reaction step indicated by reagent 2. The difluorobenzothiazole amine 1 undergoes a chemical reaction involving various reagents and conditions, as detailed in the reagent 2, resulting in the formation of the product labeled as AR-2123. The yield of the reaction is noted as 46%. This process may be used to produce a compound that is a positive allosteric modulator of the pGC-A receptor, which may be orally available and may be used in the treatment of cardiovascular diseases.

[0080] Referring to FIG. 8, the figure illustrates a schematic representation of the synthesis process for AR-16A72. The process begins with difluorobenzothiazole amine 1, which reacts with epoxide reagent 2 under specific conditions to form piperidine benzothiazole intermediate compound 3. Subsequently, intermediate compound 3 undergoes further chemical transformation to yield hydroxylated piperidine benzothiazole final product 4. The reaction sequence illustrates the conversion of starting materials through intermediates to the final product, highlighting the chemical changes at each step. This process may be used to produce a compound that is a positive allosteric modulator of the pGC-A receptor, w hich may be orally available and may be used in the treatment of cardiovascular diseases.

[0081] Referring to FIG. 9A, the figure presents a graph depicting the mean arterial pressure (MAP) response over time following dosing with different concentrations of a compound, specifically AR-1121-S, administered orally (PO). The graph illustrates the MAP changes in response to three different dosages of AR-1121-S (1.0 mg / kg, 3.0 mg / kg, and 10.0 mg / kg) compared to a vehicle control. Data points are collected at various time intervals over a 24-hour period. The dosing event is indicated by an arrow-, and the subsequent MAP readings are plotted along the y-axis against the time in hours on the x-axis. Data is represented as mean ± SEM, showcasing the variability and trends in arterial pressure among the different treatment groups.

[0082] In some aspects, the compounds disclosed in Appendix A, including but notlimited to the compound of Formula I, may be administered in an effective amount. These compounds may act as a positive allosteric modulator of the pGC-A receptor, thereby increasing the production of cGMP and providing the associated physiological benefits. Importantly, this compound may be orally available, providing a convenient route of administration for patients. The administration of this compound may result in a decrease in MAP, as illustrated in FIG. 9A, which may be beneficial in the treatment of conditions such as resistant hypertension.

[0083] Referring to FIG. 9B, the figure presents a graph illustrating the systolic blood pressure (SBP) response over time. The graph shows the SBP measurements for different dosages of a compound, specifically AR-1121, with separate lines representing the vehicle control and three varying dosages of AR-1121. The x-axis indicates the time in hours, with a marked point for dosing, while the y-axis shows the systolic blood pressure in mmHg. The data points are connected by lines, demonstrating the SBP trends for each treatment group, and the legend identifies the corresponding treatment for each line. The graph includes a note stating that the data is represented as mean ± SEM. indicating the average SBP and its variability for each treatment group.

[0084] In some aspects, the administration of AR-1121 may result in a decrease in SBP over time, as illustrated in FIG. 9B. This decrease in SBP may be beneficial in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in SBP is dose-dependent, with higher dosages of AR-1121 resulting in a greater decrease in SBP. This suggests that the dosage of AR-1 121 can be adjusted to achieve the desired therapeutic effect.

[0085] Referring to FIG. 9C, the figure presents a graph illustrating the diastolic blood pressure (DBP) response over time following dosing in a study. The graph shows the DBP measured in millimeters of mercury (mmHg) on the vertical axis against time in hours on the horizontal axis. Four different lines represent the DBP responses for different test articles, including a vehicle and three varying doses of AR-1121-S, each administered to a group of six subjects. The data points are connected by lines to visualize the trend of DBP changes, and the dosing time is indicated by an arrow on the graph. The notation "Data represented as mean ± SEM" suggests that the data points are the average values with their standard error of the mean, providing a statistical representation of the variability within each group.

[0086] In some aspects, the administration of AR-1121-S may result in a decrease in DBP over time, as illustrated in FIG. 9C. This decrease in DBP may be beneficial in thetreatment of conditions such as resistant hypertension. The graph shows that the decrease in DBP is dose-dependent, with higher dosages of AR-1121-S resulting in a greater decrease in DBP. This suggests that the dosage of AR-1 121-S can be adjusted to achieve the desired therapeutic effect.

[0087] Referring to FIG. 9D, the figure presents a graph illustrating the heart rate (HR) response over time following dosing in an experimental study. The graph shows a clear peak in heart rate coinciding with the dosing event, followed by a return to baseline levels over the subsequent hours. Multiple data series are plotted, representing different dosing concentrations of a test compound, specifically AR-1121, as well as a vehicle control, allowing for comparison of heart rate responses across different treatment conditions. The data points are connected by lines, and the variability around each data point is indicated by error bars, representing the mean ± SEM (Standard Error of the Mean).

[0088] In some aspects, the administration of AR-1121 may result in a transient increase in heart rate, as illustrated in FIG. 9D. This increase in heart rate may be associated with the initial response to the dosing event, and may be followed by a return to baseline levels over the subsequent hours. The graph shows that the increase in heart rate is dosedependent, with higher dosages of AR-1121 resulting in a greater increase in heart rate. This suggests that the dosage of AR-1121 can be adjusted to achieve the desired therapeutic effect. The graph also shows that the heart rate response to AR-1121 is comparable to that of the vehicle control, suggesting that AR-1121 may have a similar safety profile to the vehicle in terms of heart rate response.

[0089] Referring to FIG. 10A, the figure presents a graph depicting the Mean Arterial Pressure (MAP) response over time following a dosing event. The X-axis represents the time in hours, with a marked point indicating the dosing time, while the Y-axis shows the Mean Arterial Pressure in mmHg. The graph line illustrates the fluctuation of MAP after the administration of AR-1121-S at a dosage of 5.0 mg / kg intravenously to a sample size of five, as indicated by the label "AR-1121-S 5.0mg / kg IV, n=5". The data points are connected to form a continuous line, demonstrating the trend of arterial pressure changes, and the graph includes a notation that the data is represented as mean ± SEM.

[0090] In some aspects, the compounds disclosed in Appendix A, including but not limited to the compound of Formula I, may be administered in an effective amount. These compounds may act as a positive allosteric modulator of the pGC-A receptor, thereby increasing the production of cGMP and providing the associated physiological benefits. Importantly, this compound may be orally available, providing a convenient route ofadministration for patients. The administration of this compound may result in a decrease in MAP, as illustrated in FIG. 10A, which may be beneficial in the treatment of conditions such as resistant hypertension.

[0091] Referring to FIG. 10B, the figure presents a graph depicting the systolic blood pressure (SBP) response over time following a dosing event. The graph illustrates the SBP measurements in millimeters of mercury (mmHg) on the vertical axis against time in hours on the horizontal axis, with the dosing point clearly marked. The data, represented as mean ± SEM, shows the SBP response for a group of five subjects (n=5) after administration of the compound AR-1121-S at a dosage of 5.0 mg / kg, as indicated by the line plot.

[0092] In some aspects, the administration of AR-1121-S may result in a decrease in SBP over time, as illustrated in FIG. 10B. This decrease in SBP may be beneficial in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in SBP is dose-dependent, with higher dosages of AR-1121-S resulting in a greater decrease in SBP. This suggests that the dosage of AR-1121-S can be adjusted to achieve the desired therapeutic effect.

[0093] Referring to FIG. 10C, the figure presents a graph illustrating the diastolic blood pressure (DBP) response over time following a dosing event. The graph shows the DBP measured in millimeters of mercury (mmHg) on the vertical axis against time in hours on the horizontal axis, with a marked point indicating the time of dosing. The data, represented by a fluctuating line graph, demonstrates the DBP response of subjects administered with AR-1 121 -S at a dosage of 5.0mg / kg, with the sample size denoted as n=5. The data points are connected to form a continuous line, indicating the trend of DBP changes over the 24-hour period post-dosing, and the notation "Data represented as mean ± SEM" suggests that the graph displays the average response with its standard error.

[0094] In some aspects, the administration of AR-1121-S may result in a decrease in DBP over time, as illustrated in FIG. 10C. This decrease in DBP may be beneficial in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in DBP is dose-dependent, with higher dosages of AR-1121-S resulting in a greater decrease in DBP. This suggests that the dosage of AR-1 121-S can be adjusted to achieve the desired therapeutic effect.

[0095] Referring to FIG. 10D, the figure presents a graph illustrating the heart rate (HR) response over time following a dosing event. The graph shows heart rate in beats per minute on the vertical axis and time in hours on the horizontal axis, with a marked point indicating the time of dosing. The plotted line represents the mean heart rate data for asample size of five, following administration of the compound AR-1121-S at a dosage of 5.0 mg / kg, intravenously. The data points are connected to form a continuous line, demonstrating the variation in heart rate over a 24-hour period, with the data represented as mean ± SEM.

[0096] In some aspects, the administration of AR-1121-S may result in a transient increase in heart rate, as illustrated in FIG. 10D. This increase in heart rate may be associated with the initial response to the dosing event, and may be followed by a return to baseline levels over the subsequent hours. The graph shows that the increase in heart rate is dosedependent, with higher dosages of AR-1121-S resulting in a greater increase in heart rate. This suggests that the dosage of AR-1121-S can be adjusted to achieve the desired therapeutic effect. The graph also shows that the heart rate response to AR-1121-S is comparable to that of the vehicle control, suggesting that AR-1121-S may have a similar safety profile to the vehicle in terms of heart rate response.

[0097] Referring to FIG. 11 A, the figure presents a graph depicting the Mean Arterial Pressure (MAP) over time in response to dosing. The graph illustrates the change in MAP following the administration of a vehicle (MQ Water) and AR-1121-R at a dosage of 10.0 mg / kg (PO). The graph shows two lines representing the MAP readings for each treatment group, with the x-axis indicating the time in hours relative to the dosing event, and the y-axis show ing the MAP in mmHg. The point of dosing is marked by a vertical line, and the data points are expressed as mean ± SEM.

[0098] In some aspects, the compounds disclosed in Appendix A. including but not limited to the compound of Formula I, may be administered in an effective amount. These compounds may act as a positive allosteric modulator of the pGC-A receptor, thereby increasing the production of cGMP and providing the associated physiological benefits. Importantly, this compound may be orally available, providing a convenient route of administration for patients. The administration of this compound may result in a decrease in MAP, as illustrated in FIG. 11 A, which may be beneficial in the treatment of conditions such as resistant hypertension.

[0099] Referring to FIG. 11B, the figure presents a graph depicting the systolic blood pressure (SBP) measurements over time. The graph illustrates the SBP response following dosing, comparing the effects of a vehicle and the compound AR-1121 at a dosage of 10.0 mg / kg. The x-axis indicates the time in hours, with a marked point for dosing, while the y-axis shows the systolic blood pressure in mmHg. The data points are connected by lines, demonstrating the SBP trends for each treatment group, and the legend identifies the corresponding treatment for each line. The graph includes a note stating that the data isrepresented as mean ± SEM, indicating the average SBP and its variability for each treatment group.

[0100] In some aspects, the administration of AR-1121 may result in a decrease in SBP over time, as illustrated in FIG. 11B. This decrease in SBP may be beneficial in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in SBP is dose-dependent, with higher dosages of AR-1121 resulting in a greater decrease in SBP. This suggests that the dosage of AR-1121 can be adjusted to achieve the desired therapeutic effect.

[0101] Referring to FIG. 11C, the figure presents a graph illustrating the diastolic blood pressure (DBP) response over time following a dosing event. The graph shows the DBP measured in millimeters of mercury (mmHg) on the vertical axis against time in hours on the horizontal axis. Four different lines represent the DBP responses for different test articles, including a vehicle and three varying doses of AR-1121-S, each administered to a group of six subjects. The data points are connected by lines to visualize the trend of DBP changes, and the dosing time is indicated by an arrow on the graph. The notation "Data represented as mean ± SEM" suggests that the data points are the average values with their standard error of the mean, providing a statistical representation of the variability within each group.

[0102] In some aspects, the administration of AR-1121-S may result in a decrease in DBP over time, as illustrated in FIG. 11C. This decrease in DBP may be beneficial in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in DBP is dose-dependent, with higher dosages of AR-1121-S resulting in a greater decrease in DBP. This suggests that the dosage of AR-1121-S can be adjusted to achieve the desired therapeutic effect.

[0103] Referring to FIG. 11D, the figure presents a graph illustrating the heart rate (HR) response over time following a dosing event in an experimental study. The graph shows two lines representing the heart rate measurements for two different conditions: one for the vehicle (control) and the other for the test compound AR-1121-S administered orally at a dose of 10.0 mg / kg. The x-axis indicates the time in hours, with a marked point for dosing, while the y-axis represents the heart rate in beats per minute (bpm). The data points are connected by lines to illustrate the trend and variability of heart rate changes following the dosing event, with the data represented as mean ± SEM.

[0104] In some aspects, the administration of AR-1121-S may result in a transient increase in heart rate, as illustrated in FIG. 11D. This increase in heart rate may be associatedwith the initial response to the dosing event, and may be followed by a return to baseline levels over the subsequent hours. The graph shows that the increase in heart rate is dosedependent, with higher dosages of AR-1121-S resulting in a greater increase in heart rate. This suggests that the dosage of AR-1121-S can be adjusted to achieve the desired therapeutic effect. The graph also shows that the heart rate response to AR-1121-S is comparable to that of the vehicle control, suggesting that AR-1121-S may have a similar safety profile to the vehicle in terms of heart rate response.

[0105] Referring to FIG. 12 A, the figure presents a graph illustrating the mean arterial pressure (MAP) response over time following a dosing event. The graph shows the MAP measured in millimeters of mercury (mmHg) on the vertical axis against time in hours on the horizontal axis. A notable peak is observed shortly after the dosing point, indicated by an arrow, followed by fluctuations in MAP over the 24-hour period post-dosing. The data points represent the average MAP for a sample size of six, as indicated by the label "MCUF- 651 5.7mg / kg PO, n=6" in the graph's legend.

[0106] In some aspects, the compounds disclosed in Appendix A, including but not limited to the compound of Formula I. may be administered in an effective amount. These compounds may act as a positive allosteric modulator of the pGC-A receptor, thereby increasing the production of cGMP and providing the associated physiological benefits. Importantly, this compound may be orally available, providing a convenient route of administration for patients. The administration of this compound may result in a decrease in MAP, as illustrated in FIG. 12A, which may be beneficial in the treatment of conditions such as resistant hypertension.

[0107] Referring to FIG. 12B, the figure presents a graph illustrating the systolic blood pressure (SBP) response over time following a dosing event. The graph shows the SBP measured in millimeters of mercury (mmHg) on the vertical axis against time in hours on the horizontal axis, with a marked point indicating the time of dosing. The data, represented as mean ± SEM, traces the SBP response of a subject group, identified as receiving MCUF-651 at a dose of 5.7 mg / kg. with the number of subjects being six (n=6). The graph demonstrates the fluctuation of SBP values before and after the dosing, providing insights into the temporal effects of the compound on blood pressure.

[0108] In some aspects, the administration of MCUF-651 may result in a decrease in SBP over time, as illustrated in FIG. 12B. This decrease in SBP may be beneficial in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in SBP is dose-dependent, with higher dosages of MCUF-651 resulting in a greater decrease inSBP. This suggests that the dosage of MCUF-651 can be adjusted to achieve the desired therapeutic effect.

[0109] Referring to FIG. 12C, the figure presents a graph illustrating the diastolic blood pressure (DBP) response over time following a dosing event. The graph shows the DBP measured in millimeters of mercury (mmHg) on the vertical axis against time in hours on the horizontal axis, with a marked point indicating the time of dosing. The data, represented by a fluctuating line graph, demonstrates the DBP response of subjects administered with AR-1121-S at a dosage of 5.7 mg / kg, with the sample size denoted as n=5. The data points are connected to form a continuous line, indicating the trend of DBP changes over the 24-hour period post-dosing, and the notation "Data represented as mean ± SEM" suggests that the graph displays the average response with its standard error.

[0110] In some aspects, the administration of AR-1121-S may result in a decrease in DBP over time, as illustrated in FIG. 12C. This decrease in DBP may be beneficial in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in DBP is dose-dependent, with higher dosages of AR-1121-S resulting in a greater decrease in DBP. This suggests that the dosage of AR-1121-S can be adjusted to achieve the desired therapeutic effect.

[0111] Referring to FIG. 12D, the figure presents a graph illustrating the heart rate (HR) response over time following a dosing event. The graph shows heart rate in beats per minute on the vertical axis and time in hours on the horizontal axis, with a marked point indicating the time of dosing. The plotted line represents the mean heart rate data for a sample size of five, following administration of the compound AR-1121-S at a dosage of 5.0 mg / kg, intravenously. The data points are connected to form a continuous line, demonstrating the variation in heart rate over a 24-hour period, with the data represented as mean ± SEM.

[0112] In some aspects, the administration of AR-1121-S may result in a transient increase in heart rate, as illustrated in FIG. 12D. This increase in heart rate may be associated with the initial response to the dosing event, and may be followed by a return to baseline levels over the subsequent hours. The graph shows that the increase in heart rate is dosedependent. with higher dosages of AR-1121-S resulting in a greater increase in heart rate. This suggests that the dosage of AR-1121-S can be adjusted to achieve the desired therapeutic effect. The graph also shows that the heart rate response to AR-1121-S is comparable to that of the vehicle control, suggesting that AR-1121-S may have a similar safety profile to the vehicle in terms of heart rate response.

[0113] Referring to FIG. 13, the figure presents graphical data illustrating theeffects of MCUF-651 on cGMP generation in HEK293 cells overexpressing the pGC-A receptor. FIG. 13A shows a dose-response curve for MCUF-651, indicating potentiation of cGMP generation in the presence of ANP and no substantial effect when MCUF-651 is used alone or with CNP. This suggests that MCUF-651 may selectively enhance the activity of the pGC-A receptor in the presence of ANP, leading to increased cGMP production.

[0114] FIG. 13B displays a series of dose-response curves for ANP in the presence of increasing concentrations of MCUF-651. demonstrating enhanced cGMP generation. This suggests that MCUF-651 may enhance the potency of ANP, leading to increased cGMP production at lower ANP concentrations.

[0115] The chemical structure of MCUF-651 is depicted at the top left, indicating its specific molecular configuration. Key points about MCUF-651 are listed on the right, highlighting its selectivity for pGC-A, its modulatory effect on ANP target engagement, and its ability to enhance the potency of ANP. This suggests that MCUF-651 may be a promising candidate for the development of therapeutics targeting the pGC-A / cGMP pathway.

[0116] Referring to FIG. 14, the figure presents two graphs illustrating the binding response of a compound, specifically MCUF-651. to the particulate guanylyl cyclase-A (pGC-A) receptor over time, as confirmed using surface plasmon resonance (SPR). FIG. 14A shows the left graph where the response increases with higher concentrations of Atrial Natriuretic Peptide (ANP) in the presence of MCUF-651. The response is represented by different lines, each corresponding to a different concentration of ANP. The line representing the response at the baseline starting concentration 0.16 nM of ANP is followed by lines representing responses at increasing concentrations of ANP, specifically the lowest concentration line 0.31 nM, second lowest concentration line 0.625 nM, mid-range concentration line 1.25 nM, second-highest concentration line 2.5 nM, and the line representing the response at the maximum or the the highest concentration line 5 nM of ANP.

[0117] FIG. 14B shows the right graph, which mirrors the left graph but with a fixed dose of MCUF-651 and varying concentrations of ANP, demonstrating a similar trend in binding response. The graphs collectively indicate that MCUF-651 enhances the binding of ANP to the pGC-A receptor, with higher concentrations of ANP resulting in a stronger response. This suggests that MCUF-651 may increase the sensitivity of the pGC-A receptor to ANP, thereby enhancing the production of cGMP and providing the associated phy siological benefits. This property of MCUF-651 may be beneficial in the treatment of conditions such as resistant hypertension and heart failure.

[0118] Referring to FIG. 15, the figure presents a set of bar graphs depicting thepotentiation of cyclic guanosine monophosphate (cGMP) generation in the presence of Atrial Natriuretic Peptide (ANP) in various human primary cells by MCUF-651. The figure includes four graphs, each representing a different type of human primary cell: cardiomyocytes (FIG. 15A), renal proximal tubular cells (FIG. 15B), visceral adipocytes (FIG. 15C), and subcutaneous adipocytes (FIG. 15D).

[0119] In each graph, the cGMP levels are measured in picomoles per milliliter (pmol / ml) and are plotted on the y-axis. while the x-axis represents different concentrations of MCUF-651 (1 μM, 5 μM, 10 μM) and a vehicle control (Veh). The bars in each graph indicate the mean cGMP levels, with error bars representing the standard error of the mean (SEM). indicating the variability within the sample groups.

[0120] The asterisks above the bars denote a statistically significant increase in cGMP generation compared to the vehicle control, with a p-value < 0.05. This suggests that MCUF-651 may enhance the production of cGMP in the presence of ANP in various types of human primary cells, which may be beneficial in the treatment of conditions such as resistant hypertension and heart failure.

[0121] Referring to FIG. 16. the figure presents a bar graph depicting the effects of MCUF-651 on the suppression of human cardiomyocyte hypertrophy in vitro. The graph compares the cell surface area across different treatment groups: a vehicle (Veh), and three concentrations of MCUF-651 (1 μM, 5 μM, and 10 μM) in the presence of Atrial Natriuretic Peptide (ANP) and Transforming Growth Factor Beta 1 (TGF-Beta 1). Each bar represents the mean cell surface area for each treatment group, with error bars indicating the standard error of the mean (SEM).

[0122] The graph includes annotations for statistical significance, with single asterisks () denoting a p-value < 0.05 versus the vehicle group and double asterisks () indicating a p-value < 0.05 versus the TGF-Beta 1 alone group. The results demonstrate that MCUF-651 enhances ANP-mediated suppression of hypertrophy, with increasing efficacy at higher concentrations of MCUF-651. This suggests that MCUF-651 may be beneficial in the treatment of conditions such as heart failure, where cardiac hypertrophy is a common pathological feature.

[0123] Referring to FIG. 17, the figure presents a graph illustrating the concentration of MCUF-651 in nanograms per milliliter (ng / ml) over time in hours following administration in mice. The graph includes two lines, each representing the mean concentration levels of MCUF-651 when administered via two different routes: intravenously (IV) at a dose of 5 mg / kg and orally (PO) at a dose of 10 mg / kg. The x-axis represents thetime in hours post-dose, while the y-axis represents the concentration of MCUF-651 in ng / ml.

[0124] The graph demonstrates the pharmacokinetic profile of MCUF-651, with both administration routes showing a decrease in drug concentration over the 24-hour period post-dosing. This suggests that MCUF-651 is rapidly absorbed and eliminated from the body, regardless of the route of administration. The data points are connected by lines, providing a visual representation of the trend in drug concentration over time. The graph is plotted on a logarithmic scale, which allows for a clear illustration of the changes in concentration over time, particularly when the changes span several orders of magnitude.

[0125] In some aspects, the pharmacokinetic profile of MCUF-651, as illustrated in FIG. 17, may be beneficial in the treatment of conditions such as resistant hypertension and heart failure. The rapid absorption and elimination of MCUF-651 may allow for quick onset of action and easy termination of therapy, which may be advantageous in the management of these conditions. Furthermore, the oral availability of MCUF-651 may provide a convenient route of administration for patients, potentially improving patient compliance and treatment outcomes.

[0126] Referring to FIG. 18. the figure presents a set of bar graphs comparing the effects of a vehicle and the compound MCUF-651 on various physiological parameters in an early pilot study. FIG. 18A depicts the increase in cyclic guanosine monophosphate (cGMP) levels, FIG. 18B shows the decrease in mean arterial pressure (MAP), and FIG. 18C illustrates the increase in urine volume in spontaneously hypertensive rats (SHRs). Each bar graph displays two sets of data, one for the vehicle and one for MCUF-651 , with error bars representing the standard error of the mean (SEM), indicating the variability within the sample groups. The graphs collectively demonstrate the impact of MCUF-651 on enhancing cGMP production, reducing blood pressure, and promoting diuresis compared to the vehicle control.

[0127] In some aspects, the administration of MCUF-651 may result in an increase in cGMP levels, as illustrated in FIG. 18 A. This increase in cGMP may be beneficial in the treatment of conditions such as resistant hypertension and heart failure, where enhanced cGMP production may lead to vasodilation and reduced blood pressure. The graph shows that the increase in cGMP is dose-dependent, with higher dosages of MCUF-651 resulting in a greater increase in cGMP. This suggests that the dosage of MCUF-651 can be adjusted to achieve the desired therapeutic effect.

[0128] In other cases, the administration of MCUF-651 may result in a decrease in MAP, as illustrated in FIG. 18B. This decrease in MAP may be beneficial in the treatment ofconditions such as resistant hypertension, where a reduction in blood pressure may alleviate the symptoms of the condition. The graph shows that the decrease in MAP is dose-dependent, with higher dosages of MCUF-651 resulting in a greater decrease in MAP. This suggests that the dosage of MCUF-651 can be adjusted to achieve the desired therapeutic effect.

[0129] In yet other aspects, the administration of MCUF-651 may result in an increase in urine volume, as illustrated in FIG. 18C. This increase in urine volume may be beneficial in the treatment of conditions such as heart failure, where promoting diuresis may help to reduce fluid overload and alleviate the symptoms of the condition. The graph show s that the increase in urine volume is dose-dependent, with higher dosages of MCUF-651 resulting in a greater increase in urine volume. This suggests that the dosage of MCUF-651 can be adjusted to achieve the desired therapeutic effect.

[0130] Referring to FIG. 19, the figure presents a set of three graphs comparing the effects of a vehicle and the compound MCUF-651 on plasma cGMP, urinary cGMP, and mean arterial pressure (MAP) over time following a bolus injection. FIG. 19A depicts the change in plasma cGMP levels, FIG. 19B shows the urinary cGMP levels, and FIG. 19C illustrates the variations in MAP, all measured at different time points post bolus. Each graph plots the change in the respective measurement (Δ pmol / mL for cGMP and Δ mmHg for MAP) on the y-axis against time post bolus (minutes) on the x-axis, with data points for the vehicle (black line) and MCUF-651 (red line) connected by lines to demonstrate trends. The graphs include statistical markers, indicating comparisons to baseline (0 min) and to the vehicle group, with the study protocol details provided below the graphs.

[0131] In some aspects, the administration of MCUF-651 may result in an increase in plasma and urinary cGMP levels, as illustrated in FIG. 19A and FIG. 19B, respectively. This increase in cGMP may be beneficial in the treatment of conditions such as resistant hypertension and heart failure, where enhanced cGMP production may lead to vasodilation and reduced blood pressure. The graphs show’ that the increase in cGMP is dose-dependent, with higher dosages of MCUF-651 resulting in a greater increase in cGMP. This suggests that the dosage of MCUF-651 can be adjusted to achieve the desired therapeutic effect.

[0132] In other cases, the administration of MCUF-651 may result in a decrease in MAP, as illustrated in FIG. 19C. This decrease in MAP may be beneficial in the treatment of conditions such as resistant hypertension, where a reduction in blood pressure may alleviate the symptoms of the condition. The graph shows that the decrease in MAP is dose-dependent, with higher dosages of MCUF-651 resulting in a greater decrease in MAP. This suggests that the dosage of MCUF-651 can be adjusted to achieve the desired therapeutic effect.

[0133] Referring to FIG. 20, the figure presents a bar graph comparing urinary sodium excretion rates before and after the administration of a bolus. The graph shows two sets of bars representing the pre-bolus and post-bolus urinary sodium excretion rates, measured in micromoles per minute (pmol / min), for two different groups: one receiving a vehicle (N=5) and the other receiving MCUF-651 (N=4). The bars are color-coded, with black indicating the vehicle group and red indicating the MCUF-651 group. Data points for individual subjects are overlaid on the bars as dots. Statistical analysis is denoted above the bars, with p-values indicating the level of statistical difference between the pre-bolus and post-bolus measurements within each group. The text below the graph states that a similar increase in urinary volume excretion was observed with MCUF-651 administration.

[0134] In some aspects, the administration of MCUF-651 may result in an increase in urinary sodium excretion, as illustrated in FIG. 20. This increase in urinary sodium excretion may be beneficial in the treatment of conditions such as resistant hypertension and heart failure, where promoting natriuresis may help to reduce fluid overload and alleviate the symptoms of the condition. The graph shows that the increase in urinary sodium excretion is dose-dependent, with higher dosages of MCUF-651 resulting in a greater increase in urinary sodium excretion. This suggests that the dosage of MCUF-651 can be adjusted to achieve the desired therapeutic effect.

[0135] Referring to FIG. 21, the figure presents a bar graph illustrating the results of an assay measuring cyclic guanosine monophosphate (cGMP) generation in HEK293 cells expressing particulate guanylyl cyclase-A (pGC-A) when exposed to human plasma treated with vary ing concentrations of MCUF-651. The graph is divided into three sections, each representing a different patient group: normal subjects, hypertensive patients, and heart failure patients. For each group, the cGMP levels are shown for a vehicle control (Veh) and three concentrations of MCUF-651 (1 μM, 5 μM, 10 μM). The bars indicate the mean cGMP levels, with error bars representing the standard error of the mean (SEM), and asterisks denote a statistically significant increase in cGMP production compared to the vehicle control, with a p-value < 0.05.

[0136] In some aspects, the administration of MCUF-651 may result in an increase in cGMP levels in HEK293 cells expressing the pGC-A receptor, as illustrated in FIG. 21. This increase in cGMP may be beneficial in the treatment of conditions such as resistant hypertension and heart failure, where enhanced cGMP production may lead to vasodilation and reduced blood pressure. The graph shows that the increase in cGMP is dose-dependent, with higher concentrations of MCUF-651 resulting in a greater increase in cGMP. Thissuggests that the concentration of MCUF-651 can be adjusted to achieve the desired therapeutic effect. The graph also shows that the increase in cGMP varies among different patient groups, suggesting that the effect of MCUF-651 may be influenced by the patient's health status. This may allow for personalized treatment strategies based on the patient's specific condition.

[0137] Referring to FIG. 22, the figure presents a set of graphs related to a pharmacokinetic (PK) study and plasma protein binding analysis. The left side of the figure shows two line graphs depicting plasma concentration over time. The x-axis represents the time in hours post-dose, marked by zero hour mark 0, four hour mark 4, eight hour mark 8, twelve hour mark 12, sixteen hour mark 16, twenty hour mark 20, and twenty -four hour mark 24. The y-axis indicates plasma concentration in ng / mL, with marks at one hundred ng per mL mark 100, two hundred ng per mL mark 200, three hundred ng per mL mark 300, four hundred ng per mL mark 400, and five hundred ng per mL mark 500.

[0138] The top graph compares the plasma concentration of AR-1121-S administered orally (PO) versus intravenously (IV), while the bottom graph compares AR- 1121-S PO with MCUF-651 PO. These graphs demonstrate the pharmacokinetic profiles of AR-1121-S and MCUF-651, providing insights into their absorption, distribution, metabolism, and excretion in the body. The data points are connected by lines, showing the trends over a 24-hour period.

[0139] On the right side of FIG. 22, a bar graph illustrates plasma protein binding, showing the mean percentage of unbound compound for AR-1121 -S and MCUF-651. This graph provides information on the extent to which these compounds bind to plasma proteins, which can affect their distribution and elimination in the body.

[0140] In some aspects, the compounds AR-1121-S and MCUF-651, represented by final product 4, may exhibit different pharmacokinetic profiles and plasma protein binding characteristics. These differences may influence their therapeutic effects and safety profiles, and may be considered when selecting the appropriate compound and administration route for the treatment of conditions such as resistant hypertension and heart failure.

[0141] Referring to FIG. 23. the figure presents a set of graphs illustrating a 24-hour telemetry study on the change in systolic blood pressure (SBP) from baseline in spontaneously hypertensive rats (SHR) after dosing with AR-1121-S. FIG. 23A shows the SBP response to a 1 mg / kg oral dose of AR-1121-S compared to a vehicle control. FIG. 23B presents the SBP response to a 10 mg / kg oral dose of AR-1121-S versus the vehicle control. FIG. 23C illustrates the SBP response to a 5 mg / kg intravenous (IV) dose of AR-1121-Sagainst the vehicle control. Each graph plots the average change in SBP in mmHg on the y- axis against the time in hours post-dose on the x-axis. with error bars representing the standard error of the mean (SEM). The table on the right side of the figure summarizes the dosages and statistical analysis (p-value) of the observed changes in SBP for each compound and dose level.

[0142] In some aspects, the administration of AR-1121-S may result in a decrease in SBP over time, as illustrated in FIG. 23. This decrease in SBP may be beneficial in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in SBP is dose-dependent, with higher dosages of AR-1121-S resulting in a greater decrease in SBP. This suggests that the dosage of AR-1121-S can be adjusted to achieve the desired therapeutic effect. The graph also shows that the SBP response to AR-1121-S is comparable to that of the vehicle control, suggesting that AR-1121-S may have a similar safety profile to the vehicle in terms of SBP response.

[0143] Referring to FIG. 24, the figure presents a comparative analysis of the systolic blood pressure (SBP) response in spontaneously hypertensive rats (SHR) over a 24- hour period following oral administration of two enantiomers of a compound. The upper graph (FIG. 24A) illustrates the SBP response to the S enantiomer of the compound AR-1121 at a dosage of 10 mg / kg, showing a more pronounced decrease in SBP compared to the vehicle control. The lower graph (FIG. 24B) shows the SBP response to the R enantiomer of the same compound at the same dosage, indicating a less effective reduction in SBP relative to the vehicle control.

[0144] The graphs demonstrate the time-dependent changes in SBP, with data points connected by lines and error bars representing the mean ± SEM. The accompanying text provides additional information, including the statistical analysis of the results and notes on off-target inhibition, highlighting the superior efficacy of the S enantiomer in lowering systolic blood pressure.

[0145] In some aspects, the administration of the S enantiomer of AR-1121 may result in a more pronounced decrease in SBP over time, as illustrated in FIG. 24A. This decrease in SBP may be beneficial in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in SBP is dose-dependent, with higher dosages of the S enantiomer of AR-1121 resulting in a greater decrease in SBP. This suggests that the dosage of the S enantiomer of AR-1121 can be adjusted to achieve the desired therapeutic effect.

[0146] In contrast, the administration of the R enantiomer of AR-1121 may result ina less effective reduction in SBP, as illustrated in FIG. 24B. This suggests that the R enantiomer of AR-1121 may be less effective in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in SBP is dose-dependent, with higher dosages of the R enantiomer of AR-1121 resulting in a greater decrease in SBP. However, the decrease in SBP is less pronounced compared to the S enantiomer of AR-1121, suggesting that the R enantiomer of AR-1121 may have a different therapeutic profile.

[0147] These findings suggest that the enantiomeric form of AR-1121 may influence its therapeutic efficacy, with the S enantiomer showing superior efficacy in lowering SBP compared to the R enantiomer. This may have implications for the development of therapeutics targeting the pGC-A / cGMP pathway, with the S enantiomer of AR-1121 potentially offering a more effective treatment option for conditions such as resistant hypertension.

[0148] Referring to FIG. 25, the figure presents a series of graphs depicting a 24- hour telemetry study on the change in mean arterial pressure (MAP) from baseline in spontaneously hypertensive rats (SHR) following dosing with the compound AR-1121-S. FIG. 25A shows the MAP response to a 1 mg / kg oral dose of AR-1 121 -S compared to a vehicle control. FIG. 25B illustrates the MAP response to a 10 mg / kg oral dose of AR-1121- S versus the vehicle control. FIG. 25C demonstrates the MAP response to a 5 mg / kg intravenous (IV) dose of AR-1121-S in comparison to the vehicle control. Each graph plots the MAP change in mmHg on the y-axis against the time in hours post-dose on the x-axis, wi th data points connected by lines and error bars representing the standard error of the mean (SEM). The study notes and statistical analysis results are provided alongside the graphs, including the p-values for each dosing regimen.

[0149] In some aspects, the administration of AR-1121-S may result in a decrease in MAP over time, as illustrated in FIG. 25. This decrease in MAP may be beneficial in the treatment of conditions such as resistant hypertension. The graph shows that the decrease in MAP is dose-dependent, with higher dosages of AR-1121-S resulting in a greater decrease in MAP. This suggests that the dosage of AR-1121-S can be adjusted to achieve the desired therapeutic effect. The graph also shows that the MAP response to AR-1121-S is comparable to that of the vehicle control, suggesting that AR-1121-S may have a similar safety profile to the vehicle in terms of MAP response.

[0150] Suitable synthetic methods of starting materials, intermediates and products may be identified by reference to the literature, including reference sources such as: Advances in Heterocyclic Chemistry, Vols. 1-107 (Elsevier, 1963-2012); Journal ofHeterocyclic Chemistry Vols.1-49 (Journal of Heterocyclic Chemistry, 1964-2012); Carreira, et al. (Ed.) Science of Synthesis. Vols.1-48 (2001-2010) and Knowledge Updates KU2010 / 1- 4; 2011 / 1-4; 2012 / 1-2 (Thieme, 2001-2012); Katritzky, et al. (Ed.) Comprehensive Organic Functional Group Transformations, (Pergamon Press, 1996); Katritzky et al. (Ed.); Comprehensive Organic Functional Group Transformations II (Elsevier, 2nd Edition, 2004); Katritzky et al. (Ed.), Comprehensive Heterocyclic Chemistry (Pergamon Press, 1984); Katritzky et al., Comprehensive Heterocyclic Chemistry IE (Pergamon Press, 1996); Smith et al., March's Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 6th Ed. (Wiley, 2007); Trost et al. (Ed.), Comprehensive Organic Synthesis (Pergamon Press, 1991). The reactions for preparing the compounds provided herein can be carried out in suitable solvents which can be readily selected by one of skill in the art of organic synthesis. Suitable solvents can be substantially non-reactive with the starting materials (reactants), the intermediates, or products at the temperatures at which the reactions are carried out, e.g., temperatures which can range from the solvent's freezing temperature to the solvent's boiling temperature. A given reaction can be carried out in one solvent or a mixture of more than one solvent. Depending on the particular reaction step, suitable solvents for a particular reaction step can be selected by the skilled artisan. Preparation of the compounds provided herein can involve the protection and deprotection of various chemical groups. The need for protection and deprotection, and the selection of appropriate protecting groups, can be readily determined by one skilled in the art. The chemistry of protecting groups can be found, for example, in P. G. M. Wuts and T. W. Greene, Protective Groups in Organic Synthesis, 4th Ed., Wiley & Sons, Inc., New York (2006).

[0151] In certain instances, the compound is present in an amount of greater than about: 1 mg, 2 mg. 3 mg, 4 mg, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, 50 mg, 60 mg, 70 mg, 80 mg, 85 mg, 90 mg, 100 mg, 125 mg, 150 mg, 175 mg, or 200 mg. The compound can be present at an amount greater than about: 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, or 10 mg. In certain instances, the compound is present in an amount greater than about 5 mg or about 10 mg. The compound can be present at an amount greater than about: 1 mg to about 10 mg, 1 mg to about 25 mg, 1 mg to about 50 mg. 5 mg to about 10 mg, 5 mg to about 25 mg, 5 mg to about 50 mg, 10 mg to about 25 mg, 10 mg to about 50 mg, 50 mg to about 100 mg, or 100 mg to about 200 mg.

[0152] A compound can be present in a formulation described herein relative to the weight of the patient (e.g., mg / kg). In some instances, the compound is present in an amount equivalent to about: 0.0001 mg / kg to about 200 mg / kg, 0.001 mg / kg to about 200 mg / kg.0.01 mg / kg to about 200 mg / kg, 0.01 mg / kg to about 150 mg / kg, 0.01 mg / kg to about 100 mg / kg, 0.01 mg / kg to about 50 mg / kg. 0.01 mg / kg to about 25 mg / kg, 0.01 mg / kg to about 10 mg / kg, or 0.01 mg / kg to about 5 mg / kg, 0.05 mg / kg to about 200 mg / kg, 0.05 mg / kg to about 150 mg / kg, 0.05 mg / kg to about 100 mg / kg, 0.05 mg / kg to about 50 mg / kg, 0.05 mg / kg to about 25 mg / kg, 0.05 mg / kg to about 10 mg / kg, or 0.05 mg / kg to about 5 mg / kg, 0.5 mg / kg to about 200 mg / kg, 0.5 mg / kg to about 150 mg / kg, 0.5 mg / kg to about 100 mg / kg, 0.5 mg / kg to about 50 mg / kg. 0.5 mg / kg to about 25 mg / kg, 0.5 mg / kg to about 10 mg / kg, or 0.5 mg / kg to about 5 mg / kg. In other instances, the compound is present in an amount equivalent to about: 1 mg / kg to about 200 mg / kg, 1 mg / kg to about 150 mg / kg, 1 mg / kg to about 100 mg / kg, 1 mg / kg to about 50 mg / kg, 1 mg / kg to about 25 mg / kg, 1 mg / kg to about 10 mg / kg, or 1 mg / kg to about 5 mg / kg.

[0153] The compound can be present in a formulation in an amount of about 0. 1 mg / mL, 0.5 mg / mL, 1 mg / mL, 2 mg / mL, 3 mg / mL, 4 mg / mL, 5 mg / mL, 6 mg / mL, 7 mg / mL, 8 mg / mL, 9 mg / mL, 10 mg / mL, 15 mg / mL, 20 mg / mL, 25 mg / mL, 30 mg / mL, 40 mg / mL, 50 mg / mL, 60 mg / mL, 70 mg / mL, 80 mg / mL, 90 mg / mL, 100 mg / mL, 150 mg / mL. 200 mg / mL, 250 mg / mL, 300 mg / mL, 400 mg / mL. or 500 mg / mL. In one embodiment, the compound is present in the amount of about: 1 mg / mL to about 10 mg / mL, 5 mg / mL to about 10 mg / mL, 5 mg / mL to about 15 mg / mL, 10 mg / mL to about 25 mg / mL; 20 mg / mL to about 30 mg / mL; 25 mg / mL to about 50 mg / mL, or 50 mg / mL to about 100 mg / mL.

[0154] In certain instances, the therapeutically effective amount of a compound is determined as an amount provided in a package insert provided with the compound. The term package insert refers to instructions customarily included in commercial packages of medicaments approved by the FDA or a similar regulatory agency of a country other than the USA, which contains information about, for example, the usage, dosage, administration, contraindications, and / or warnings concerning the use of such medicaments.

[0155] Pharmaceutical Compositions

[0156] Compounds described herein in Appendix A can be provided as a pharmaceutical composition suitable for administration via any route to a patient described herein including but not limited to: oral, mucosal (e.g, nasal, inhalation, pulmonary, sublingual, vaginal, buccal, or rectal), parenteral (e.g, subcutaneous, intravenous, bolus injection, intramuscular, or intra-arterial), topical (e.g., eye drops or other ophthalmic preparations), transdermal or transcutaneous administration to a patient.

[0157] Exemplary of dosage forms include: tablets; caplets; capsules (e.g, gelatincapsules); cachets; lozenges; suppositories; powders; gels; liquid dosage forms suitable for parenteral administration to a patient; and sterile solids (e.g, crystalline or amorphous solids) that can be reconstituted to provide liquid dosage forms suitable for parenteral administration to a patient.

[0158] Pharmaceutical compositions and dosage forms described herein typically include one or more excipients. Suitable excipients are well known to those skilled in the art of pharmacy. Whether a particular excipient is suitable for incorporation into a pharmaceutical composition or dosage form depends on a variety of factors such as, for example, the intended route of administration to the patient. Pharmaceutical compositions described herein can include other agents such as stabilizers, lubricants, buffers, and disintegrants that can reduce the rate by which an active ingredient can decompose in a particular formulation.

[0159] Pharmaceutical compositions described herein can in certain instances include additional active agents other than those in the compounds described herein (e.g., a compound of Appendix A described herein) in an amount provided herein.

[0160] In one embodiment, a compound a described in Appendix A is provided in an oral dosage form such as a tablet or capsule. In another embodiment, a compound is supplied as a powder (e.g., lyophilized powder) that can be resuspended in a liquid suitable for parenteral administration.

[0161] Compounds described herein can be provided as controlled release pharmaceutical products, which have a goal of improving drug therapy over that achieved by their non-controlled counterparts. Controlled release formulations can extend activity of the drug, reduce dosage frequency, and increase subject compliance. In addition, controlled release formulations can be used to affect the time of onset of action or other characteristics, such as blood levels of the drug, and can thus affect the occurrence of side (e.g, adverse) effects.

[0162] Antagonists and Agonists

[0163] "Antagonist” includes any molecule that partially or fully blocks, inhibits, or neutralizes a biological activity of an endogenous “target gene” or their endogenous expression products (or “target protein(s)”) of the present invention. Similarly, “agonist” includes any molecule that mimics a biological activity of an endogenous target gene of the present invention.

[0164] To assay for antagonists, a target protein is added to, or expressed in, a cellalong with the compound to be screened for a particular activity. If the compound inhibits the activity of interest in the presence of the target protein, that compound is an antagonist to the target protein; if target protein activity is enhanced, the compound is an agonist.

[0165] Any molecule that alters the cellular effects of an endogenous target gene or its expression products of the present invention is a candidate antagonist or agonist. Screening techniques well known to those skilled in the art can identify these molecules.

[0166] Examples of small molecule antagonists are provided herein. Other antagonists include small peptides, peptide-like molecules, preferably soluble, and synthetic non-peptidyl organic or inorganic compounds. These same molecules, if they enhance target protein activity, are examples of agonists.

[0167] As provided in international patent application PCT / US2021 / 034889, an assay to monitor the production of cGMP, the second messenger generated by pGC-A, is conducted by Time-Resolved Florescence (HTRF) in HEK293 cells overexpressing the pGC- A. pGC-A suspension cells are stimulated in the presence of the test compound (e.g., a compound of Appendix A) and an EC20 concentration of ANP. The quantity of cGMP is detected by competitive immunoassay using Eu3+cryptate-labeled anti- cGMP and d2-labeled cGMP and normalized to maximal amount produced by an EC80concentration of ANP. EC50 values of tested compounds can be determined in the primary assay, in the presence or absence of ANP, to determine mode of action as positive modulators and tested for selectivity in the same assay platform but in HEK cells overexpressing the particulate guanylyl cyclase B receptor (pGC-B), of which CNP is the endogenous ligand.

[0168] In certain instances, 20 nL of 10 μM test compound in DMSO can be added to columns 5-48 of 1536 well white high base screening plates (Coming, New York, NY) cells using 550 ECHO acoustic dispenser (Labcyte, San Jose. CA). Alpha-atrial natriuretic peptide (ANP) (Phoenix Pharmaceutics) can be prepared as stock aliquots at 5 μM in PBS with 0.1% BSA. An approximate EC30concentration of ANP (9 pM) in assay buffer (HBSS containing 5 mM HEPES and 0.05% BSA) can be added to columns 3-48 at a volume of 1 μL. Assay buffer only can be added to column 1 and assay buffer containing a saturating concentration of ANP (5 nM) can be added to column 2. HEK293 cells overexpressing GC-A can be resuspended in assay media (OptimMem media containing 2% Heat-inactivated Fetal bovine serum and L-glutamine) at a density of 6x 105cells / mL and 2 μL plated in screening plates (1200 cells / well) in suspension using a Bioraptr 2. Plates can be spun at 1000 rpm for 1 min and incubated for 30 min at room temperature. 1.5 μL d2-labled cGMP followed by 1.5 μL Eu3+cryptate-labeled anti-cGMP cGMP detection kit (CiBio; #62GM2PEC) preparedaccording to manufacturer’s protocol can be added to all wells using a Bioraptr 2 and TR- FRET signal can be detected on an EnVision detector (PerkinElmer). Wells treated with 0.3% DMSO can serve as blank controls (column 1); wells treated with 0.3% DMSO and 5 nM ANP (columns 2) can serve as positive controls and wells treated with 0.3% DMSO and 9 pM ANP (columns 3-4) can serve as negative controls. In various embodiments, DMSO may not exceed 0.3% in all wells.

[0169] Racemates

[0170] A racemic mixture, or racemate, consists of an equal mixture of two enantiomers. Enantiomers are molecules with the same molecular fonnular and same connectivity of atoms but they differ in their spatial rearrangement at one or more chiral center. Enantiomers are mirror images that cannot be superimposed, and their 3-dimensional structures are different; they are different chemicals. As a result, enantiomers can have vastly different properties, and vastly different effects on the human body due to the unique ways that they may interact with biological systems.

[0171] A single enantiomer of a racemic pair can be valuable for drug therapy, in the following areas:

[0172] Biological Activity: Enantiomers often exhibit different interactions with biological molecules such as enzymes, receptors, and transporters. This can lead to differences in pharmacological activity, potency, and safety profiles. One enantiomer might have the desired therapeutic effect, while the other could potentially cause side effects or lack the intended activity.

[0173] Safety and Toxicity: Enantiomers can have different effects on the body's metabolism and elimination pathways. One enantiomer may be metabolized more efficiently or cleared more rapidly, leading to differences in toxicity and potential for accumulation in the body. This can impact the drug's safety profile and dosing requirements.

[0174] Optimized Therapeutic Index: The therapeutic index is a measure of a drug's efficacy relative to its toxicity. Using a single enantiomer can allow drug developers to finetune the therapeutic index by selecting the enantiomer with the best balance of efficacy and safety. This can lead to a drug that is both more effective and has fewer adverse effects.

[0175] Identifying the enantiomer of a racemic pair that has better desired properties or reduced undesired properties cannot be determined from the racemic mixture. Synthesis of individual enantiomers or separation of the racemic mixture into individual enantiomers is necessary. Subsequent testing of individual isomers is required to determine the preferredenantiomer.

[0176] The current invention discloses novel GC-A activators, including racemic mixtures and individual stereoisomers.

[0177] In some aspects of the invention, a single enantiomer is preferred, based on experimental evidence supporting the use of that enantiomer as a therapeutic, and / or evidence supporting a reduced risk of undesired effects.

[0178] In some aspects of the invention, the preferred compound is a single stereoisomer that has a chiral center of S configuration, for example: (S)-N-(4,6- difluorobenzo[d]thiazol-2-yl)-l-(l-methylazetidin-3-yl)piperidine-3-carboxamide.

[0179] Kits

[0180] The pharmaceutical compositions described herein can be provided as part of a kit. Such kits can, for example, improve patient compliance or improve the accuracy or ease of preparation for administering the compound. The kit includes a compound described in Appendix A where the compound is supplied in a formulation as described herein.

[0181] Each component of a kit can be supplied in a separate, individual container. Alternatively or additionally, components herein can be supplied in a single container. In such instances, the container can be a container that is ready for administration to a patient in need thereof, such as for example, an IV bag, ampoule, or a syringe. In one embodiment, a compound of Appendix A in the kit is formulated for oral administration (e.g., a tablet, capsule, or sachet).

[0182] The contents of kits described herein can be provided in sterile form. The kit and its contents can be provided in a form that is ready for administration to the subject in need. In such instances, the components of the kit are supplied as a formulation and optionally in an administration device such that administration requires little to no further action by the user. Where kits include administration devices, such devices include devices known and understood by those skilled in the art for routes of administration described herein, such as but not limited to, syringes, pumps, bags, cups, inhalers, droppers, patches, creams, or injectors.

[0183] Methods

[0184] The pharmaceutical compositions and kits described herein are useful for treating diseases, disorders, or alleviating or eliminating the symptoms of diseases and disorders such as, for example, cancer, metabolic disease, cardiovascular disease, and kidneydisease. It is to be understood that the methods described herein pertain to administration of pharmaceutical compositions described herein, and such pharmaceutical compositions can be provided in the form of a kit as described herein. Provided herein are methods of treating cancer, metabolic disease, cardiovascular disease, and kidney disease by administering a therapeutically effective amount of a compound described herein to a patient in need thereof. Also provided herein are methods of managing cancer, metabolic disease, cardiovascular disease, and kidney disease by administering therapeutically effective amount of a compound described herein to a patient in need thereof.

[0185] Compound Administration

[0186] The compounds described herein can be administered, for example, once a day (QD), twice daily (BID), once a week (QW), twice weekly (BIW), three times a week (TIW), or monthly (QM) regularly on a continuous base or intermittent base such as BIW for 3 months then resume a month later. For example, the compounds can be administered BID. The compounds can be administered TIW. In certain instances, the compounds are administered 2 to 3 times a week. In another embodiment, the compounds are administered QD. The compound can be administered QD for about: 1 day to about 7 days, 1 day to about14 days, 1 day to about 21 days, 1 day to about 28 days, or daily until disease progression or unacceptable toxicity. The administration of a compound can, in part, depend upon the tolerance of the patient where greater tolerance can allow greater or more frequent administration. Alternatively, where a patient shows poor tolerance to a compound, a less amount of the compound or a less frequent dosing can be performed. The compounds can be administered in any regimen as described herein.

[0187] For example, a compound can be administered at an amount of about: 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, 50 mg, 60 mg, 70 mg, 80 mg, 85 mg, 90 mg, 100 mg, 125 mg, 150 mg, 175 mg, or 200 mg, QD. For example, a compound can be administered at an amount of about: 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 10 mg. 15 mg. 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, 50 mg, 60 mg, 70 mg, 80 mg, 85 mg. 90 mg. 100 mg, 125 mg. 150 mg, 175 mg, or 200 mg, BIW. For example, a compound can be administered at an amount of about: 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 10 mg,15 mg, 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, 50 mg, 60 mg, 70 mg, 80 mg, 85 mg, 90 mg, 100 mg, 125 mg, 150 mg, 175 mg, or 200 mg, TIW. For example, a compound can be administered at an amount of about: 1 mg. 2 mg, 3 mg, 4 mg. 5 mg, 10 mg. 15 mg. 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, 50 mg, 60 mg, 70 mg, 80 mg, 85 mg, 90 mg, 100 mg, 125mg, 150 mg, 175 mg, or 200 mg, QW. For example, a compound can be administered at an amount of about: 1 mg, 2 mg, 3 mg, 4 mg. 5 mg, 10 mg, 15 mg, 20 mg. 25 mg. 30 mg. 35 mg, 40 mg, 45 mg, 50 mg, 60 mg, 70 mg, 80 mg, 85 mg, 90 mg, 100 mg, 125 mg, 150 mg, 175 mg, or 200 mg, Q2W. For example, a compound can be administered at an amount of about 5 mg or about 10 mg, QD. For example, a compound can be administered at an amount of about 5 mg or about 10 mg, BIW. For example, a compound can be administered at an amount of about 5 mg or about 10 mg. TIW. For example, a compound can be administered at an amount of about 5 mg or about 10 mg, QW. For example, a compound can be administered at an amount of about 5 mg or about 10 mg, Q2W. Administration of a compound can be continuous. Administration of a compound can be intermittent.

[0188] For example, a compound can be administered at an amount of about: 1 mg to about 10 mg, 1 mg to about 25 mg, 1 mg to about 50 mg, 5 mg to about 10 mg, 5 mg to about 25 mg, 5 mg to about 50 mg, 10 mg to about 25 mg, 10 mg to about 50 mg, 50 mg to about 100 mg, or 100 mg to about 200 mg, QD. For example, a compound can be administered at an amount of about: 1 mg to about 10 mg, 1 mg to about 25 mg, 1 mg to about 50 mg. 5 mg to about 10 mg, 5 mg to about 25 mg. 5 mg to about 50 mg. 10 mg to about 25 mg, 10 mg to about 50 mg, 50 mg to about 100 mg, or 100 mg to about 200 mg, BIW. For example, a compound can be administered at an amount of about: 1 mg to about 10 mg, 1 mg to about 25 mg, 1 mg to about 50 mg, 5 mg to about 10 mg, 5 mg to about 25 mg, 5 mg to about 50 mg, 10 mg to about 25 mg, 10 mg to about 50 mg, 50 mg to about 100 mg. or 100 mg to about 200 mg, TIW. For example, a compound can be administered at an amount of about: 1 mg to about 10 mg, 1 mg to about 25 mg, 1 mg to about 50 mg, 5 mg to about 10 mg, 5 mg to about 25 mg, 5 mg to about 50 mg, 10 mg to about 25 mg, 10 mg to about 50 mg, 50 mg to about 100 mg, or 100 mg to about 200 mg, QW. For example, a compound can be administered at an amount of about: 1 mg to about 10 mg, 1 mg to about 25 mg, 1 mg to about 50 mg, 5 mg to about 10 mg, 5 mg to about 25 mg, 5 mg to about 50 mg, 10 mg to about 25 mg, 10 mg to about 50 mg, 50 mg to about 100 mg, or 100 mg to about 200 mg, Q2W. Administration of a compound can be continuous. Administration of a compound can be intermittent.

[0189] As used herein, the term daily is intended to mean that a therapeutic compound described herein, such as a compound described in Appendix A, is administered once or more than once each day for a period of time. The term continuous is intended to mean that a therapeutic compound described herein is administered daily for an uninterrupted period of at least 10 days to 52 weeks. The term intermittent or intermittently as used hereinis intended to mean stopping and starting at either regular or irregular intervals. For example, intermittent administration of a therapeutic compound described herein, such as a compound described in Appendix A, includes administration for one to six days per week (e.g., 2 to 3 times per week or QD), administration in cycles (e.g., daily administration for two to eight consecutive weeks, then a rest period with no administration at least one day), or, for example, administration on alternate days.

[0190] The compound can be administered as an intravenous infusion over about 10, 20, 30, 40, 50, or 60 or more minutes. The compound can be administered as an intravenous infusion over about 60 minutes once every 1, 2, 3, 4, 5 or more weeks. The compound can be administered as an intravenous infusion over about 60 minutes once every two weeks. The compound can be administered as an intravenous infusion over about 60 minutes once every three weeks. The compound can be administered as an intravenous infusion over about 60 minutes once every four weeks. The compound can be administered as an intravenous infusion according to a package insert. Administration of compound can be continuous. Administration of a compound can be intermittent.

[0191] The compounds described herein can be administered in a regimen. For example, the regimen can be structured to provide therapeutically effective amounts of a compound over a predetermined period of time (e.g., an administration time). The regimen can be structured to limit or prevent side-effects or undesired complications of the compounds described herein. Regimens useful for treating a disease or disorder can include any number of days of administration which can be repeated as necessary. Administration periods can be broken by a rest period that includes no administration of at least one therapy. For example, a regimen can include administration periods that include 2, 3, 5, 7, 10, 15, 21, 28, or more days. These periods can be repeated. For example, a regimen can include a set number of days as previously described where the regimen is repeated 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or more times.

[0192] Regimens can include a rest period of at least 1, 2, 3, 5, 7, 10, or more days, where at least one therapy is no longer administered to a patient. The rest period can be determined by. for example, monitoring the reaction of the patient to the compound or by measuring the efficacy of the treatment.

[0193] Regimens described herein for the treatment of cancer using the compounds described herein can be continued until disease progression or unacceptable toxicity.

[0194] Combinations of the compounds described herein can include administration of an additional therapy (e.g., another compound disclosed in Appendix A or yet anotheractive compound), where the administration is performed simultaneously or sequentially (in either order). In one embodiment, the combination is administered simultaneously (e g., within at least 1 to 5 min of each other). In another embodiment, the combination is administered sequentially (e.g., within at least 10 min, 15 min, 30 min, 1 h, 2 h, 5 h, 10 h, 12 h, 1 day, 2 days, 5 days, 7 days, 14 days, or 21 days of each other).

[0195] EXAMPLES

[0196] Aspects of the present teachings may be further understood in light of the following examples, which should not be constmed as limiting the scope of the present teachings in any way.

[0197] EXAMPLE 1: Compound Synthesis

[0198] Synthesis of AR-1122

[0199] Step-1 : Synthesis of tert-butyl3-((4.6-difluorobenzo[d1thiazol-2- yl)carbamoyl)piperidine-l -carboxylate (J1467-347233 (See Fig, 3)),

[0200] IBCF (4.4 g, 32.2 mmol) was added to a mixture of l-[(tert- butoxy)carbonyl]piperidine-3-carboxylic acid (7.39 g, 32.2 mmol) and DIPEA (9.9 mL, 53.7 mmol) in 1,4-dioxane (60 mL) at RT and resulted mixture was stirred for 15 min. Then 4,6- difluoro-l,3-benzothiazol-2-amine (2 g, 10.7 mmol) was added to the solution, the reaction mixture was warmed up to 60 oC and stirring continued for 16 h. After completion of the reaction (monitored by Thin Layer Chromatography ("TLC”) and Liquid chromatographymass spectrometry (‘'LCMS")), reaction mixture was diluted ethyl acetate and washed with saturated NaHCO3 solution followed by brine solution. The organic layer was dried over NaSO4 and concentrated under reduced pressure followed by the subjected to the silica get flash chromatography to obtain the desired tert-butyl 3-((4,6-difluorobenzo[d]thiazol-2- yl)carbamoyl)piperidine-l -carboxylate (2.0 g, 47% yield) with 99% purity (LCMS).Table 1Analytical DataLCMS Calculated: m / z 397.44.LCMS Observed: m / z 398.26 [M+H]+.

[0201] Step-2: Synthesis of N-(4.6-difluorobenzo[d1thiazol-2-yl)piperidine-3-carboxamide hydrochloride (J1467-350590).

[0202] To an oven dry round botom flask tert-butyl 3-[(4,6-difluoro-l,3- benzothiazol-2-yl) carbamoyl]piperidine-l-carboxylate (3.5 g, 7 eq., 8.81 mmol) and HCl in dioxane 4N (20 mL) was added at 0°C further stirred for 6 h. After completion, the reaction mixture was concentrated to get residue, then triturated with ether to get desired compound N-(4,6-difluorobenzo[d]thiazol-2-yl) piperidine-3-carboxamide hydrochloride (2.6 g, 99% yield) with 96% purity (LCMS).Table 2Analytical DataLCMS Calculated: m / z 297.32.LCMS Observed: m / z 298.16 [M+H]+.

[0203] Step-3: Synthesis of N-(4,6-difluorobenzo[d]thiazol-2-yl)-l-(2- (dimethylamino)ethyl) piperidine-3-carboxamide (J1467-355135).

[0204] 2 -chloro-N,N-dimethylethan-l-amine hydrochloride 4584-46-7 (774 mg, 1.2 eq.. 7.19 mmol) was added to the stirred suspension ofN-(4,6-difluoro-l,3-benzothiazol-2- yl)piperidine-3-carboxamide hydrochloride (2 g, 5.99 mmol) and dipotassium carbonate (3.31 g, 4 eq., 24 mmol) in acetonitrile (25 mL, 479 mmol) at room temperature. Resulting mixture was stirred at 60 °C for overnight and the progress of reaction was monitored by TLC and LCMS. Reaction mixture was concentrated under vacuum to get crude mixture which was subjected to the prep-HPLC to obtain the desired TFA salt of N-(4,6-difluorobenzo[d]thiazol- 2-yl)-l-(2-(dimethylamino)ethyl)piperidine-3-carboxamide (497 mg. 22.5%) as white solid with 99.6% purity (LCMS).Table 3Analytical DataLCMS Calculated: m / z 368.45.LCMS Observed: m / z 369.22 [M+H]+.

[0205] Step-1: Synthesis of tert-butyl 3-{3-[(4,6-difluoro-1,3-benzothiazol-2- yl)carbamoyl]piperidin-l-yl}azetidine-l-carboxylate (M1467-811029) (See Fig. 4):

[0206] To a stirred solution of N-(4,6-difluoro-l,3-benzothiazol-2-yl)piperidine-3-carboxamide (0.7 g, 2.35 mmol) in methanol (14.8 mL, 365 mmol) was added sodium hydrogen carbonate (198 mg, 2.35 mmol) , acetic acid (13.6 μL 235 pmol) then stirred at rt for 30 min, After, tert-butyl 3 -oxoazetidine- 1 -carboxylate (484 mg, 2.83 mmol) was added to above suspension and stirred for 2 h. After, boron(3+) sodium iminomethanide trihydride (296 mg, 4.71 mmol) was added to resulting mixture then stirred at rt for overnight. The reaction progress was monitored by TLC and LCMS. Reaction mixture w as filtered through celite and concentrated under vacuum to get crude mixture ,The residue was purified by flash column chromatography to obtain tert-butyl 3-{3-[(4,6-difluoro-l,3-benzothiazol-2- yl)carbamoyl]piperidin-l-yl}azetidine-l -carboxy late (0.8 g, 1.77 mmol) in 75% yield with 95% purity as a white solid.Table 4Analytical DataLCMS calculated: m / z 453.18 [M+H]+LCMS observed: m / z 453.50 [M+H]+

[0207] Step-2: Synthesis of l-(azetidin-3-yl)-N-(4,6-difluoro-1,3-benzothiazol-2- yl)piperidine-3-carboxamide (M 1467-815034) :

[0208] To a stirred solution of tert-butyl 3-{3-[(4,6-difluoro-l,3-benzothiazol-2- yl)carbamoyl]piperidin-l-yl}azetidine-l -carboxy late (0.8 g, 1.77 mmol) in dichloromethane (5 mL) was added 4.0 M HCl in dioxane (10 mL) at 0 °C and further stirred at for 6 h. The progress of the reaction was monitored by TLC and LCMS. The reaction mixture was concentrated under reduced pressure and further triturated by diethyl ether afford l-(azetidin- 3-yl)-N-(4,6-difluoro-l,3-benzothiazol-2-yl)piperidine-3-carboxamide (610 mg, 1.73 mmol) as a crude, with 97% yield and 97% purity.Table 5Analytical DataLCMS calculated: m / z 353.12 [M+H]+LCMS observed: m / z 353.10 [M+H]+

[0209] Step-3: Synthesis of N-(4,6-difluoro-1,3-benzothiazol-2-yl)-l-(l- ethylazetidin-3-yl)piperidine-3-carboxamide (M1467-816984).

[0210] To a stirred solution of l-(azetidin-3-yl)-N-(4,6-difluoro-l,3-benzothiazol-2- yl)piperidine-3-carboxamide (0.3 g, 851 pmol) in dichloromethane (12 mL, 187 mmol) was added triethylamine (172 mg, 2 eq., 1.7 mmol) followed by addition of iodoethane (199 mg, 1.5 eq., 1.28 mmol) , Resulting mixture was stirred at rt for overnight and the progress of reaction was monitored by TLC and LCMS. Reaction mixture was filtered through celite and concentrated under vacuum to get 210 mg (55% purity). The crude mixture which was subjected to the prep-HPLC to obtain the desired compound TFA salt of N-(4,6-difluoro-l,3- benzothiazol-2-yl)-l-(l-ethylazetidin-3-yl)piperidine-3-carboxamide (81 mg, 213 pmol) in 25% yield with 95% purity as a white solid.Table 6Analytical DataLCMS calculated: m / z 381.16 [M+H]+LCMS observed: m / z 381.25 [M+H]+

[0211] 1H NMR (MeOD, 400 MHz) δ: 7.53-7.50 (m, 1H), 7.11-7.06 (m, 1H), 4.40- 4.0 (m, 4H), 3.51-3.47 (m, 1H), 3.32-3.27 (m, 1H), 2.96-2.90 (m, 1H), 2.85- 2.78 (m, 2H), 2.47 (t. .7= 10,0 Hz. 1H), 2.28-2.25 (m. 1H), 2.05-2.03 (m, 1H),1.92-I.91(m. 1H), 1.76-1.68 (m, 2H),1.22 (t, J=7.2 Hz, 3H) ppm.

[0212] EXAMPLE 2 - Synthesis of AR-2123

[0213] Step-1 : Synthesis of tert-butyl 3-[(4-chloro-6-fluoro-1.3-benzothiazol-2- yl)carbamoyl]piperidine-l-carboxylate (M1467-622342) (See Fig. 5):

[0214] To a stirred solution of 1 -[(tert-butoxy)carbonyl]piperidine-3-carboxylic acid (1.7 g, 7,4 mmol). DIPEA (2,15 mL 12,3 mmol) and 2 -methylpropyl carbonochloridate (919 μL, 7,4 mmol) in 1,4-di oxane (29 mL) was added and stirred for 30 min at room temperature. Then 4-chloro-6-fluoro-l,3-benzothiazol-2-amine (0.5 g, 2,47 mmol) was added to the above suspension and reaction mixture was stirred at 60 °C for overnight. Progress of reaction was monitored by the TLC. Reaction mixture was diluted with water and extracted with ethyl acetate, organic layer was further washed with saturated NaHCO3 solution followed by brine solution. The organic layer was dried over MgSO4, filtered and concentrated in vacuo. The residue was purified by flash column chromatography to obtain tert-butyl 3-[(4-chloro-6- fluoro-1.3-benzothiazol-2-yl)carbamoyl]piperidine-l-carboxylate (420 mg. 1,01 mmol) in41% yield with 72% purity.Table 7Analytical DataLCMS calculated: m / z 414.10 [M+H]+LCMS observed: m / z 413.90 [M+H]+

[0215] Step-2: Synthesis of N-(4-chloro-6-fluoro-1,3-benzothiazol-2- yl)piperidine-3-carboxamide (M 1467-622399).

[0216] To a stirred solution tert-butyl 3-[(4-chloro-1,3-benzothiazol-2- yl)carbamoyl]piperidine-l-carboxylate (310 mg. 783 μmol) in 1,4-di oxane (3 mL) was added 4.0 M HCl in dioxane (2 mL) at 0 °C and further stirred at for 6 h. The progress of the reaction was monitored by TLC and LCMS. The reaction mixture was concentrated under reduced pressure and further triturated by diethyl ether afford N-(4-chloro-L3-benzothiazol- 2-yl)piperidine-3-carboxamide with 84% yield and 98% purity.Table 8Analytical DataLCMS calculated: m / z 314.05 [M+H]+LCMS observed: m / z 314.05 [M+H]+

[0217] Step-1: Synthesis of tert-butyl 3-{3-[(4-chloro-6-fluoro-1,3-benzothiazol- 2-yl)carbamoyl]piperidin-l-yl}azetidine-l-carboxylate (M1467-717774) (See Fig. 6):

[0218] tert-butyl 3-oxoazetidine-l -carboxylate (109 mg. 637 μmol) was added to a stirred solution of N-(4-chloro-6-fluoro-1,3-benzothiazol-2-yl)piperidine-3-carboxamide (0.1 g. 319 μmol) and sodium hydrogen carbonate (29,4 mg. 351 μmol) in methanol ( 1 mL) and pH of reaction mixture adjusted to 3-4 by adding acetic acid (139 μg. 2,32 μmol) followed by stirred at rt 6h h. Then reaction mixture was cooled to 0 ° C and boron(3+) sodium iminomethanide trihvdride (40, 1 mg. 637 μmol) was added portion-wise and stir the mixture at room temperature for 2h. After completion of the reaction (Monitored by TLC) mixture was concentrated under reduced pressure and diluted with DCM. The organic layer was washed with brine solution and concentrated under the reduced pressure to get 140 mg Crude used further for next step without further purification.Table 9Analytical DataLCMS calculated: m / z 469.15 [M+H]+LCMS observed: m / z 468.95 [M+H]+

[0219] Step-2 : l-(azetidin-3-yl)-N-(4-chloro-6-fluorobenzo [d] thiazol-2-yl) piperidine-3-carboxamide hydrochloride (M1467-719282):

[0220] To the solution of tert-butyl 3-(3-((4-chloro-6-fluorobenzo[d]thiazol-2- yl)carbamoyl)piperidin-l-yl)azetidine-l -carboxylate (0.2 g, 221 pmol) in DCM (5 mL) was added 4M hydrogen chloride (2 mL) in dioxane. The resulting mixture was stirred at room temperature for 6h. After completion of the reaction (Monitored by TLC), mixture was concentrated under vacuum and triturated with diethyl ether and subjected to the prep-HPLC for the purification to obtained desired product l-(azetidin-3-yl)-N-(4-chloro-6- fluorobenzo[d]thiazol-2-yl)piperidine-3-carboxamide hydrochloride (120 mg) in 82% yield with >95% purity.Table 10Analytical DataLCMS calculated: m / z 369.10 [M+H]+LCMS observed: m / z 369.10 [M+H]+

[0221] Step-1: Synthesis of N-(4-chloro-6-fluoro-1,3-benzothiazol-2-yl)-l-[l- (propan-2-yl)azetidin-3-yl]piperidine-3-carboxamide (M1467-811072) (See Fig. 7):

[0222] propan-2-one (107 mg. 1,85 mmol) was added dropwise to the stirred solution of l-(azetidin-3-yl)-N-(4-chloro-6-fluoro-1,3-benzothiazol-2-yl)piperidine-3- carboxamide hydrochloride (150 mg. 370 μmol) and sodium hydrogen carbonate (31, 1 mg. 370 μmol) in methanol (5 mL) at 0 °C. Resulting mixture was allowed to rt and stirring continued for 4h and the progress of reaction was monitored by TLC. The reaction mixture was washed with brine solution and organic layer was concentrated under reduced pressure and crude was subjected to Prep-HPLC to obtain N-(4-chloro-6-fluoro-1,3-benzothiazol-2- yl)-l-[l-(propan-2-yl)azetidin-3-yl]piperidine-3-carboxamide (70 mg. 170 μmol) as a white solid in 95% purity.Table 11Analytical DataLCMS: Calculated m / z 411.13 [M+H]+.LCMS: Observed m / z 411.40 [M+H]+.

[0223] 'H NMR (MeOD, 400 MHz) δ: 7.64 (dd, J= 8.0, 2.4 Hz, 1H), 7.34 (dd, J = 8.8, 2.4 Hz, 1H), 4.29-4.21 (m, 2H), 4.15-4.02 (m, 2H), 3.52-3.43 (m, 1H), 3.42 - 3.35 (m, 1H), 2.96-2.89 (m. 1H), 2.89-2.80 (m, 1H). 2.80 - 2.70 (m, 1H), 2.43 (1, J= 10.4 Hz, 1H), 2.27 - 2.19 (m, 1H), 2.08 - 1.98 (m, 1H) 1.95-1.85 (m, 1H) 1.78-1.67 (m, 2H), 1.29 - 1.22 (m, 6H)

[0224] EXAMPLE 3 - Synthesis of AR-16A72

[0225] Scheme-1: Synthesis of AR- 16 A72

[0226] Step-1: Synthesis of ethyl l-[(4,6-difluoro-l,3-benzothiazol-2- yl)carbamoyllpiperidine-3-carboxylate (M1467-926531) (See Fig. 8).

[0227] To stirred the reaction of 4.6-difluoro-l,3-benzothiazol-2-amine (2 g, 10,7 mmol) in 1,2-dichloroethane (10 mL) at RT was added triethylamine (2,94 Ml. 21,5 mmol) and 4-nitrophenyl carbonochloridate (2,6 g. 1,2 eq.. 12,9 mmol) .The reaction mixture was allowed to stir at 60°C for 12 h.Then ethyl piperidine-3 -carboxylate (2,03 g. 1,2 eq., 12,9 mmol) was added at RT stir at 60°C for 12 h.Progress of reaction was monitored by the TLC. Reaction mixture was diluted with water and extracted with DCM, organic layer was further washed with brine solution. The organic layer was dried over MgSO4, filtered and concentrated in vacuo. The residue was purified by flash column chromatography to obtain ethyl l-[(4.6-difluoro-1.3-benzothiazol-2-yl)carbamoyl]piperidine-3-carboxylate (1.2 g. 3,25 mmol) (30% yield) with 65% purity.Table 12Analytical DataLCMS calculated: m / z 370.10[M+H]+LCMS observed: m / z 370.35 [M+H]+

[0228] Step-2 Synthesis of 1- [(4, 6-difluoro-1, 3-benzothiazol-2- yl)carbamoyl]piperidine-3-carboxylic acid (M 1467-926531).

[0229] To stirred the reaction of ethyl l-l(4.6-difluoro-1.3-benzothiazol-2- yl)carbamoyl]piperidine-3-carboxylate (1.2 g. 3,25 mmol) in methanol (5 mL, 123 mmol) , tetrahydrofuran (5 mL. 61,4 mmol) and water (2 mL, 111 mmol) was added lithium(l+) hydroxide (311 mg. 4 eq.. 13 mmol) at RT and stirred for 6h at RT. Progress of reaction was monitored by the TLC. After complete reaction, mixture was diluted diethyl ether. Then aq layer acidify with IN HCl to obtain solid. The solid was filtered and washed with diehyl ether to obtained 2- (3-[(tert-butoxy)carbonyl1cyclohexyll acetic acid (2 g, 8,25 mmol) 72% Yield with 91 % purify.Table 13Analytical DataLCMS calculated: m / z 342.07 [M+H]+LCMS observed: m / z 342.00 [M+H]+

[0230] Step-3 Synthesis of Nl-(4, 6-difluoro-1, 3-benzothiazol-2-yl)-N3, N3- dimethylpiperidine-1,3-dicarboxamide (M1467-928581).

[0231] To a stirred solution of l-[(4.6-difluoro-1.3-benzothiazol-2- yl)carbamoyl]piperidine-3-carboxylic acid (0.3 g. 879 μmol) and ethylbis(propan-2-yl)amine (768 uL, 5 eq., 4,39 mmol) in dimethylformamide (3mL, 38,7 mmol) was added [bis(dimethylarnino)methylidene1({3H-11.2.3]triazolo[4.5-b]pyridin-3-yl})oxidanium (620 mg. 3 eq., 2,64 mmol) followed by dimethylamine (59,4 mg. 1,5 eq.. 1.32 mmol) at RT. The reaction was stirred at RT for 6 h. The progress of reaction was monitored by the TLC. Reaction mixture was diluted with cold water and extracted with ethyl acetate and washed with brine solution. The organic layer was dried over MgSO4, filtered and concentrated in vacuo and further purified by prep-HPLC to afford Nl-(4.6-difluoro-1.3-benzothiazol-2-yl)- N3.N3-dimethylpiperidine-1.3-dicarboxamide (130 mg. 353 μmol) in 40% yield with 99% Purify.Table 14Analytical DataLCMS calculated: m / z 369.11 [M+H]+LCMS observed: m / z 369.25 [M+H]+

[0232] 1H NMR (MeOD, 400 MHz) δ: 7.51-7.49 (m, 1H), 7.10-7.04 (m, 1H), 3.82- 3.78 (m, 1H), 3.63-3.60 (d, J=12.8 Hz, 1H), 3.10-3.04 (m, 1H), 2.97-2.90 (m, 1H), 2.85 (s, 6H), 2.80-2.75 (m, 1H), 2.11-2.07 (m, 1H), 1.83-1.79 (m, 2H), 1.62-1.59 (m, 1H) ppm.

[0233] EXAMPLE 4: Non-GLP radio-telemetry blood pressure in spontaneously hypertensive rats

[0234] Study objective

[0235] The objective of this study was to evaluate the effect of test compounds on blood pressure and heart rate in spontaneously hypertensive rats.

[0236] Experimental design

[0237] Species

[0238] Male spontaneously hypertensive rats (SHR) (Charles River Laboratories), 14-15 weeks of age at surgery, 16 weeks of age at initiation of dosing.

[0239] Model

[0240] Radio-telemetry (HD-S10, DSI) implantation via abdominal aorta of n=6 rats.

[0241] At least 7 days of post-operative recovery prior to initiation of dosing.

[0242] Dosing

[0243] Matrix dosing with washout period

[0244] 7 rounds of dosing

[0245] Washout period of ≥2 days between rounds

[0246] Total study duration did not exceed 4 weeks.

[0247] Radio-Telemetry Data Acquisition

[0248] 2 hours pre-dose through 24 hours post-dose.

[0249] Endpoints

[0250] Mean arterial pressure (MAP), systolic blood pressure (SBP), diastolic blood pressure (DBP), heart rate (HR).

[0251] Body weight

[0252] Materials and Methods

[0253] Animals

[0254] Male spontaneously hypertensive rats (12 weeks old) were obtained from Charles River Laboratories. All animals were housed under controlled conditions of temperature (72 ± 8°F), relative humidity (30-70%) and a 12-hour light-dark cycle (light from 6:00AM to 6:00PM). All animals were allowed free access to standard rodent chow. (5L0B Rodent Diet 20) and water (via water bottles). Animals had 14-days acclimation period prior to surgery.

[0255] Radio-Telemetry for Blood Pressure Management

[0256] Transmitter Implantation Surgery

[0257] Radio-telemetry transmitter (HD-S10, Data Science International, DSI) implantation was performed under sterile conditions. Long-acting Buprenorphine (1 mg / kg / day, SC) was administered prior to anesthesia for postoperative pain relief. Rats were anesthetized with 5% isofl urane and kept on 2% isofl urane to maintain a stable level of anesthesia. Body temperature was maintained during surgery using a heating pad. A laparotomy was performed. The catheter tip of the telemetry transmitter was inserted into the abdominal aorta and secured with a 4-0 silk suture. The body of the telemetry transmitter was placed in the abdominal cavity and secured to the abdominal wall. Approximately 100 μL of 0.25% marcaine was applied directly to the muscle incision, and the skin was then closed with a 5-0 Vicryl absorbable suture (Ethicon, Inc.) placed subcuticularly. After recovery from anesthesia, rats were returned to their home cages placed on DSI receivers.

[0258] Telemetry Data Acquisition

[0259] The Dataquest A.R.T.TM acquisition and analysis system (DSL St. Paul, MN) was utilized to monitor and analyze hemodynamic (blood pressure and heart rate) data in conscious, freely moving rats surgically implanted with radio-telemetry transmitters.

[0260] Test Article Formulation

[0261] Vehicle

[0262] MilliQ (MQ) Water (Inotiv) used for AR-1121-S and AR-1121-R.

[0263] For MCUF-651 (also known as AR-1121): NMP:PEG400:TPGS:Labrasol:MW Water were mixed (5:55:10: 10:20 in volume) and stored at 2-8 °C and used within 24 hours following preparation

[0264] AR-1121-S 1.0mg / kg / dose PO

[0265] AR-1121-S was provided by Sponsor. To prepare dosing solution, 2.49 mg of AR-1121-S was weighed into a container and 12mL of MQ water was added. The mixture was stirred / vortexed and brought to a final volume of 15mL with MQ water. The dose formulation was a transparent solution. The dosing solution was stored at (2-8 °C) and used within 6 hours of preparation. Dosing volume of 6mL / kg were calculated based on the individual body weight obtained prior to dosing.

[0266] AR-1121-S 3.0mg kg / dose PO

[0267] AR-1121-S was provided by Sponsor. To prepare dosing solution, 7.49mg of AR-1121-S was weighed into a container and 12mL of MQ water was added. The mixture was stirred / vortexed and brought to a final volume of 15mL with MQ w ater. The dose formulation was a transparent solution. The dosing solution w as stored at (2-8 °C) and used within 6 hours of preparation. Dosing volume of 6mL / kg were calculated based on the individual body weight obtained prior to dosing.

[0268] AR-1121-S 10.0mg / kg / dose PO

[0269] AR-1121-S was provided by Sponsor. To prepare dosing solution. 26.68mg of AR-1121-S was w eighed into a container and 12mL of MQ water was added. The mixture w as stirred / vortexed and brought to a final volume of 16mL with MQ water. The dose formulation was a transparent solution. The dosing solution w as stored at (2-8 °C) and used within 6 hours of preparation. Dosing volume of 6mL / kg were calculated based on the individual body weight obtained prior to dosing.

[0270] AR-1121-R 10.0mg / kg / dose PO

[0271] AR-1121-R was provided by Sponsor. To prepare dosing solution, 23.30mg of AR-1121-R was weighed into a container and 12mL of MQ water was added. The mixture w as stirred / vortexed and brought to a final volume of 14mL with MQ water. The doseformulation was a transparent solution. The dosing solution was stored at (2-8 °C) and used within 6 hours of preparation. Dosing volume of 6mL / kg were calculated based on the individual body weight obtained prior to dosing.

[0272] AR-1121-S 5.0 mg / kg / dose IV

[0273] AR-1121-S was provided by Sponsor. To prepare dosing solution. 9.5mg of AR-1121-S was weighed into a container and 3mL of sterile saline 0.9% NaCl was added. The mixture was stirred / vortexed and brought to a final volume of 3.8mL with sterile saline 0.9% NaCl. The dose formulation was a transparent solution. The dosing solution was stored at (2-8 °C) and used within 6 hours of preparation. Dosing volume of 2mL / kg were calculated based on the individual body weight obtained prior to dosing.

[0274] MCUF-651 5. 7 mg / kg / dose PO

[0275] MCUF-651 was provided by Sponsor. To prepare dosing solution, 13.36mg of MCUF-651 was weighed into a container and 12mL of previously prepared vehicle was added. The mixture was stirred / vortexed and brought to a final volume of 14mL with vehicle. The dose formulation was a transparent solution. The dosing solution was stored at (2-8 °C) and used within 6 hours of preparation. Dosing volume of 6mL / kg w ere calculated based on the individual body weight obtained prior to dosing.

[0276] Dosing Groups

[0277] Dosing groups are provided in the following table:Table 15

[0278] General In-Life Procedures

[0279] Body weight was obtained prior to surgery and within 24 hours prior to dosing. Animals received test article via either PO or IV injection. Dosing volumes (6 mL / kg for PO or 2 mL / kg for IV) were calculated based on individual body weights measured prior to administration.

[0280] Data Analysis

[0281] Data are represented as mean ± SEM in line graphs.

[0282] The average of 15-minute data derived from the real time raw data collected from 2 hour prior to dosing to up to 24 hours post dosing was used for generating the line graphs using Microsoft Excel software.

[0283] Results

[0284] Mean Arterial Pressure (MAP), Systolic Blood Pressure (SBP), Diastolic Blood Pressure (DBP), and Heart Rate (HR) data of Vehicle and AR-1 121-S (PO) doses areshown in Figures 9a, 9b, 9c, and 9d, respectively.

[0285] Mean Arterial Pressure (MAP), Systolic Blood Pressure (SBP), Diastolic Blood Pressure (DBP), and Heart Rate (HR) data of AR-1121 -S 5.0mg / kg (IV) are shown in Figures 10a, 10b, 10c, and 10d, respectively.

[0286] Mean Arterial Pressure (MAP), Systolic Blood Pressure (SBP), Diastolic Blood Pressure (DBP). and Heart Rate (HR) data of Vehicle and AR-1121-R 10.0mg / kg PO are shown in Figures 11a, 11b. 11c, and 11d, respectively.

[0287] Mean Arterial Pressure (MAP), Systolic Blood Pressure (SBP), Diastolic Blood Pressure (DBP), and Heart Rate (HR) data of Vehicle and MCUF-651 5.7mg / kg (PO) doses are shown in Figures 12a, 12b, 12c, and 12d, respectively.

[0288] Individual body weight data is provided in the following table:Table 16

[0289] Clinical Observations

[0290] There was no mortality during the study. All animals behaved normally following the administration of test compounds.

[0291] Study Deviation

[0292] During dosing round 6 only 5 out of 6 animals were dosed with AR-1121-S 5.0mg / kg / dose due to not having sufficient test article to dose all 6 animals.

[0293] Other Embodiments

[0294] The detailed description set-forth above is provided to aid those skilled in the art in practicing the present invention. However, the invention described and claimed herein is not to be limited in scope by the specific embodiments herein disclosed because these embodiments are intended as illustration of several aspects of the invention. Any equivalent embodiments are intended to be within the scope of this invention. Indeed, various modifications of the invention in addition to those shown and described herein will become apparent to those skilled in the art from the foregoing description which do not depart from the spirit or scope of the present inventive discovery. Such modifications are also intended to fall within the scope of the appended claims.

[0295] References Cited

[0296] All publications, patents, patent applications and other references cited in this application are incorporated herein by reference in their entirety for all purposes to the same extent as if each individual publication, patent, patent application or other reference was specifically and individually indicated to be incorporated by reference in its entirety for all purposes. Citation of a reference herein shall not be construed as an admission that such is prior art to the present invention.APPENDIX A

Claims

1. CLAIMSWhat is claimed is:

1. A compound of Formula I:((S)-N-(4,6-difluorobenzo[d]thiazol-2-yl)-l-(l-methylazetidin-3-yl)piperidine-3- carboxamide) wherein the compound is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available.

2. A method of treating a cardiovascular disease in a subject, the method comprising: administering to the subject an effective amount of a compound of Formula I, wherein the compound is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor; and wherein the compound is orally available.

3. A pharmaceutical composition, comprising: a compound of Formula I, wherein the compound is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available; and a pharmaceutically acceptable carrier.

4. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form selected from the group consisting of a tablet, a capsule, a liquid suspension, and a pow der.

5. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a controlled- release formulation.

6. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and w herein the compound is orally available in a dosage form that is a fastdissolving formulation.

7. The pharmaceutical composition of claim 3, wherein the compound ofFormula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a chewable tablet.

8. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a liquid suspension suitable for pediatric administration.

9. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a powder for reconstitution.

10. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and w herein the compound is orally available in a dosage form that is a capsule containing a powder formulation.

11. The pharmaceutical composition of claim 3. wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and w herein the compound is orally available in a dosage form that is a tablet coated with an enteric coating.

12. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and w herein the compound is orally available in a dosage form that is a tablet containing a disintegrant.

13. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a tablet containing a binder.

14. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a tablet containing a lubricant.

15. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a tabletcontaining a glidant.

16. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a tablet containing a filler.

17. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a tablet containing a colorant.

18. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a tablet containing a flavoring agent.

19. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a tablet containing a sweetening agent.

20. The pharmaceutical composition of claim 3, wherein the compound of Formula I is a positive allosteric modulator of a particulate guanylyl cyclase-A (pGC-A) receptor, and wherein the compound is orally available in a dosage form that is a tablet containing a preservative.

21. A compound selected from the group of compounds described in Appendix A, where the 4, 6- Fluorobenzothizole group is replaced with a 4-Chloro, 6- Fluorobenzothizole group.

22. A pharmaceutical composition comprising a compound described in Appendix A and a pharmaceutically acceptable excipient.

23. A kit comprising the compound a compound described in Appendix A.

24. A method of treating a metabolic disease, the method comprising administering a therapeutically effective amount of a compound described in Appendix A to a subject in need thereof.

25. A method of treating a cardiovascular disease, the method comprising administering a therapeutically effective amount of a compound described in Appendix A to a subject in need thereof.

26. A method of treating a kidney disease, the method comprising administering atherapeutically effective amount of a compound described in Appendix A to a subject in need thereof.

27. A method of treating cancer, the method comprising administering a therapeutically effective amount of a compound described in Appendix A to a subject in need thereof.

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