Pyrazolo[3,4-d]pyrimidinone compounds for the treatment of chronic heart failure

Abstract

This application relates to a series of novel compounds that are selective inhibitors of phosphodiesterase type 9 ("PDE9") for the treatment of chronic heart failure. More specifically, this application relates to pyrazolo[3,4-d]pyrimidinone compounds for use in the treatment and prevention of chronic heart failure.

Description

[Technical Field]

[0001] Related applications This application claims priority and benefits of U.S. Provisional Application No. 63 / 533,961, filed on 22 August 2023, and U.S. Provisional Application No. 63 / 643,495, filed on 7 May 2024, the contents of which are incorporated herein by reference in their entirety. [Background technology]

[0002] In the United States, approximately 6 million adults suffer from heart failure, and it is the leading cause of hospitalization among those 65 years of age and older. Heart failure accounts for approximately $30.7 billion in annual healthcare costs in the U.S., and 50% of patients die within five years. New treatments are needed to reduce the incidence and mortality rates of heart failure.

[0003] Recent evidence suggests that PDE9A plays a crucial role in the pathophysiology of heart failure, with increased expression in cardiomyocytes of heart failure patients and inhibiting the beneficial natriuretic peptide receptor / cGMP / protein kinase GI (PKGI) signaling pathway.

[0004] The cyclic guanosine monophosphate (cGMP) / protein kinase GI (PKGI) signaling pathway plays a crucial role in the pathophysiology of the cardiovascular system. For example, PKGI activation inhibits cardiac hypertrophy and remodeling. In cardiomyocytes, signaling via the cGMP / PKGI pathway is mediated by four hormones: nitric oxide (NO) and three natriuretic peptides (atrial natriuretic peptide [ANP], β-natriuretic peptide [BNP], and c-natriuretic peptide [CNP]). NO acts via soluble guanylate cyclase (sGC) receptors located in the cytoplasm, CNP acts via guanylate cyclase type B receptors (GC-B), and both ANP and BNP act via guanylate cyclase type A receptors (GC-A) located in the cell membrane. Stimulation of guanylate cyclase converts guanosine triphosphate to cGMP.

[0005] cGMP is a second messenger molecule that mediates signaling bound to NO and natriuretic peptides. cGMP binds to and activates protein kinase G (PKG), causing phosphorylation changes in PKG substrates in the myocardium. cGMP generally has a protective function for the heart, inhibiting harmful remodeling. Degradation or hydrolysis of cGMP in the cardiovascular system can occur under the action of two enzymes, PDE-5 and PDE-9. Inhibiting PDE-5 and / or PDE-9 can inhibit this degradation and increase cGMP levels. PDE9 inhibitors are described, for example, in U.S. Patent No. 7,964,607, which is incorporated herein by reference in its entirety.

[0006] Both reduced-ejection-fraction heart failure (HFrEF) and preservative-ejection heart failure (HFpEF) are major health problems with significant impacts on morbidity, mortality, quality of life, and healthcare costs. They are leading causes of hospitalization worldwide, and despite advances in treatment, the five-year mortality rate (within five years of initial diagnosis) remains at approximately 50%. Novel therapies, such as PDE-9 inhibitors, are needed to reduce morbidity and mortality in heart failure (HF). [Overview of the project]

[0007] In one embodiment, this specification provides a method for treating chronic heart failure in a subject requiring treatment. The method comprises administering to the subject a therapeutically effective amount of a compound or a pharmaceutically acceptable salt thereof, wherein the compound has the structure of formula (I).

[0008] [ka]

[0009] During the ceremony, R is selected from the group consisting of (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, (C3-C8)cycloalkyl, heterocycloalkyl, aryl, and heteroaryl, each of which may optionally be substituted with 1 to 3 substituents, the substituents independently selected from the group consisting of (C1-C4)alkyl, (C1-C4)alkoxy, halogen, and (C1-C4)haloalkyl. R1 is selected from the group consisting of hydrogen, (C1-C4) alkyl, (C2-C4) lucenyl, (C2-C4) alkynyl, (C1-C4) haloalkyl, and cyclopropyl; R2 is selected from the group consisting of a heteroaryl selected from the group consisting of (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, (C1-C6)haloalkyl, pyridinyl, pyridadinyl, pyrimidinyl, and pyrazinyl, and ER5, and the heteroaryl may optionally be substituted with 1 to 3 substituents independently selected from (C1-C4)alkyl and (C1-C4)haloalkyl; R3 is selected from the group consisting of hydrogen, (C1-C4) alkyl, (C2-C4) alkenyl, (C2-C4) alkynyl, (C3-C6) cycloalkyl, and (C1-C4) haloalkyl; E is selected from the group consisting of -CH2-, -CH2CH2-, -CH2CH2CH2-, and -C(O)-; and R5 is selected from the group consisting of (C3-C8) cycloalkyl, heterocycloalkyl, aryl, aryloxy, and heteroaryl, all of which may be substituted with 1 to 3 substituents, and such substituents are independently selected from the group consisting of (C1-C4) alkyl, (C2-C4) alkenyl, (C2-C4) alkynyl, (C1-C4) hydroxyalkyl, (C1-C4) haloalkyl, (C1-C4) alkoxy, (C1-C4) haloalkoxy, (C3-C8) cycloalkyl, halo, cyano, phenyl, morpholinyl, (C1-C4) alkylamino, pyrazolyl, triazolyl, and imidazolyl.

[0010] In another embodiment, this specification provides a method for treating chronic heart failure in a subject requiring treatment. This method comprises administering to the subject a therapeutically effective amount of a compound, which is 6-((3S,4S)-4-methyl-1-(pyrimidine-2-ylmethyl)pyrrolidine-3-yl)-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one ("Compound A") or a pharmaceutically acceptable salt thereof.

[0011] In some embodiments, the method includes administering 5 mg to 50 mg of the compound to a subject. In some embodiments, the method includes administering 5 mg to 25 mg of the compound to a subject. In some embodiments, the method includes administering 50 mg of the compound to a subject. In some embodiments, the method includes administering the compound twice daily. In some embodiments, the method includes administering the compound every 12 hours. In some embodiments, the method includes administering the compound for at least 2 weeks. In some embodiments, the method includes administering the compound for at least 10 weeks. In some embodiments, the method includes administering the compound for the remainder of the subject's life. In some embodiments, the method includes orally administering the compound of formula (I).

[0012] In some embodiments, chronic heart failure is reduced-ejection-fraction chronic heart failure. In some embodiments, chronic heart failure is preserved-ejection-fraction chronic heart failure. In some embodiments, chronic heart failure is moderate-ejection-fraction chronic heart failure. In some embodiments, chronic heart failure is ultra-high-ejection-fraction chronic heart failure.

[0013] In some embodiments, the subjects have a plasma or serum NT-proBNP level of at least 600 pg / ml prior to administration.

[0014] In some embodiments, the method results in a decrease of about 30% in the level of N-terminal pro-brain natriuretic peptide (NT-proBNP) measured in the plasma or serum of a subject. In some embodiments, the method results in a decrease of about 30% in the level of brain natriuretic peptide (BNP) measured in the plasma or serum of a subject. In some embodiments, the method results in an increase of about 30% in the level of cGMP measured in the plasma or serum of a subject. In some embodiments, the method increases the level of cGMP measured in the urine of a subject by about 30%. In some embodiments, the method increases the ratio of plasma cGMP level to plasma NT-proBNP level by about 30%. In some embodiments, the method increases the ratio of plasma cGMP level to plasma BNP level by about 30%. In some embodiments, the method increases the Kansas City Cardiomyopathy Questionnaire-23 (KCCQ-23) score by at least 5 points.

[0015] In another aspect, the present disclosure provides a method of treating chronic heart failure in a subject in need thereof. The method comprises orally administering to the subject 25 mg of 6-((3S,4S)-4-methyl-1-(pyrimidin-2-ylmethyl)pyrrolidin-3-yl)-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidin-4-one (「Compound A」) or a pharmaceutically acceptable salt thereof every 12 hours.

Brief Description of the Drawings

[0016] [Figure 1] Figure 1 is a schematic diagram of Part A of the test described in Example 3. Abbreviations: D = day; q12h = every 12 hours; V = visit. [Figure 2] Figure 2 is a schematic diagram of Part B of the test described in Example 3. Abbreviations: D = day; V = visit. [Figure 3] Figure 3 is a graph showing the change over time in the geometric mean (ng / mL) of urinary cGMP measured at day 0 (baseline), day 1, day 14, and day 28 of administration for placebo and Compound A. [Figure 4] Figure 4 shows two graphs illustrating the mean percentage change from baseline over time for plasma cGMP measured on day 1 and week 4 of administration. [Figure 5] Figure 5 shows two graphs illustrating the mean percentage change (time-adjusted) from baseline in plasma cGMP measured at week 4 of administration for patients treated with Entrest and patients not treated with Entrest. [Figure 6] Figure 6 is a graph showing the change in the geometric mean (95% confidence interval) of NT-proBNP (pg / mL) measured on days 0, 14, 28, and 84 of administration for patients who received placebo and compound A. [Figure 7] Figure 7 is a graph showing the mean change in KCCQ scores (i.e., Clinical Summary Score (CSS), Overall Summary Score (OSS), and Total Summary Score (TSS)) between the placebo group and compound A group at week 12. [Figure 8] Figure 8 shows three graphs illustrating the percentage of patients in the placebo group and compound A group whose KCCQ summary scores (OSS and CSS) increased by more than 5 points, more than 10 points, and more than 20 points at week 12. [Modes for carrying out the invention]

[0017] This specification provides a method for treating chronic heart failure using a selective PDE9 inhibitor, as well as a single-dose dosage form and a pharmaceutical composition.

[0018] compound This application comprises a selective PDE9 inhibitor represented by formula (I) and pharmaceutically acceptable salts thereof.

[0019] [ka]

[0020] During the ceremony, R is selected from the group consisting of (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, (C3-C8)cycloalkyl, heterocycloalkyl, aryl, and heteroaryl, each of which may optionally be substituted with 1 to 3 substituents, the substituents independently selected from the group consisting of (C1-C4)alkyl, (C1-C4)alkoxy, halogen, and (C1-C4)haloalkyl. R1 is selected from the group consisting of hydrogen, (C1-C4) alkyl, (C2-C4) lucenyl, (C2-C4) alkynyl, (C1-C4) haloalkyl, and cyclopropyl; R2 is selected from the group consisting of a heteroaryl selected from the group consisting of (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)alkynyl, (C1-C6)haloalkyl, pyridinyl, pyridadinyl, pyrimidinyl, and pyrazinyl, and ER5, and the heteroaryl may optionally be substituted with 1 to 3 substituents independently selected from (C1-C4)alkyl and (C1-C4)haloalkyl; R3 is selected from the group consisting of hydrogen, (C1-C4) alkyl, (C2-C4) alkenyl, (C2-C4) alkynyl, (C3-C6) cycloalkyl, and (C1-C4) haloalkyl; E is selected from the group consisting of -CH2-, -CH2CH2-, -CH2CH2CH2-, and -C(O)-; and R5 is selected from the group consisting of (C3-C8) cycloalkyl, heterocycloalkyl, aryl, aryloxy, and heteroaryl, all of which may be substituted with 1 to 3 substituents, and such substituents are independently selected from the group consisting of (C1-C4) alkyl, (C2-C4) alkenyl, (C2-C4) alkynyl, (C1-C4) hydroxyalkyl, (C1-C4) haloalkyl, (C1-C4) alkoxy, (C1-C4) haloalkoxy, (C3-C8) cycloalkyl, halo, cyano, phenyl, morpholinyl, (C1-C4) alkylamino, pyrazolyl, triazolyl, and imidazolyl.

[0021] Preferably, R is selected from the group consisting of ethyl, isopropyl, trifluoroethyl, cyclobutyl, cyclopentyl, difluorocyclohexyl, methoxyphenyl, and tetrahydro-2H-pyran-4-yl; R1 is hydrogen or methyl; R2 is methyl, trifluoroethyl, trifluorobutyl, pyrimidinyl, trifluoromethylpyrimidinyl, or ER5; R3 is methyl, ethyl, isopropyl, trifluoromethyl, trifluoroethyl, or cyclopropyl; E is -CH2- or -C(O)-; and R5 is substituted or unsubstituted cyclopentyl, morpholinyl, phenyl, naphthyl, benzyloxy, pyrimidinyl, or pyridinyl Selected from the group consisting of quinolinil, quinoxalinil, pyrazinil, pyrazolyl, benzimidazolyl, sinnolinil, naphthidolinil, pyrido[2,3-b]pyradinil, imidazo[4,5-c]pyradinil, benzothiadiazolyl, tetrahydropyrazolo[1,5-a]pyradinil, dihydrobenzodioxynil, imidazolyl, dihydrobenzofuranil, triazolyl, oxazolyl, isoxazolyl, benzodioxynil, thiazolyl, imidazo[1,2-a]pyradinil, tetrahydrobenzothiazolyl, dihydrobenzoxazinil, tetrahydropyranil, tetrahydropyrazolo[1,5-a]azepinyl, and dihydropyrrolo[1,2-b]pyrazolyl.

[0022] More preferably, R is selected from the group consisting of isopropyl, cyclobutyl, cyclopentyl, and tetrahydro-2H-pyranyl; R1 is hydrogen; R2 is ER5; R3 is methyl or ethyl; E is -CH2-; and R5 is selected from the group consisting of phenyl, pyrimidine-2-yl, pyridine-2-yl, pyrazine-2-yl, and 5-methylpyrazine-2-yl.

[0023] In other preferred embodiments, the compound is 6-[(3,4-trans)-1-benzyl-4-methylpyrrolidine-3-yl]-1-(2-methoxyphenyl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one 6-[(3,4-trans)-1-benzyl-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-benzyl-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-pyrimidine-2-ylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[4-(trifluoromethyl)pyrimidine-2-yl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzoyl-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(pyridine-3-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[3-(trifluoromethyl)benzyl]pyrrolidine-3-yl}1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(quinoline-2-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(quinoline-4-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0024] 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-[[6-(trifluoromethyl)pyridine-3-yl]methyl]pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(quinoxaline-2-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(quinoxaline-6-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(pyrimidine-5-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1,4-dimethylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(2,2,2-trifluoroethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-(3,4-trans)-4-methyl-1-[(2-methylpyridine-3-yl)methyl]pyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(quinoline-8-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(quinoline-3-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(6-methylpyridine-3-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0025] 6-[(3,4-trans)-1-benzyl-4-isopropylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-methylpyrrolidine-3-yl]-1-cyclopentyl-3-methyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3S,4S)-4-methyl-1-(quinoxaline-6-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(2-phenylethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[(6-methoxypyridine-3-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-[(3-methylpyridine-2-yl)methyl]pyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(3-methylpyridine-2-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-ethylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-cyclopropylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-methylpyrrolidine-3-yl]-1-isopropyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one;

[0026] 6-[(3S,4S)-1-benzyl-4-methylpyrrolidine-3-yl]-1-isopropyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-[(3,4-trans)-4-methyl-1-(quinoline-2-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-[(3,4-trans)-4-methyl-1-(quinoxaline-6-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-[(3,4-trans)-4-methyl-1-(quinoline-3-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-(trifluoromethyl)pyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-[(3S,4S)-4-methyl-1-(quinoxaline-6-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-[(3S,4S)-4-methyl-1-(quinoline-3-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3S,4S)-4-methyl-1-[(5-methylpyrazine-2-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-ethylpyrrolidine-3-yl]-1-isopropyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-benzyl-4-ethylpyrrolidine-3-yl]-1-isopropyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one;

[0027] 1-Isopropyl-6-{(3S,4S)-4-methyl-1-[(2-methylpyrimidine-5-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-(1-benzylpyrrolidine-3-yl)-1-cyclopentyl-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-(3S,4S)-1-[(6-methoxypyridine-3-yl)methyl]-4-methylpyrrolidine-3-yl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-4-ethyl-1-(quinoline-3-ylmethyl)pyrrolidine-3-yl]-1-isopropyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-{(3,4-trans)-4-ethyl-1-[(6-methoxypyridine-3-yl)methyl]pyrrolidine-3-yl}-1-isopropyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-4-ethyl-1-(quinoxaline-6-ylmethyl)pyrrolidine-3-yl]-1-isopropyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3S,4S)-1-[(1,3-dimethyl-1H-pyrazole-5-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3S,4S)-4-methyl-1-(4,5,6,7-tetrahydropyrazolo[1,5-a]pyridine-3-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3S,4S)-4-methyl-1-[(1-methyl-1H-benzimidazole-2-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-{(3S,4S)-4-methyl-1-[(5-methylpyrazine-2-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0028] 6-[(3S,4S)-1-(sinnoline-3-ylmethyl)-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(quinoxaline-6-ylmethyl)-4-(trifluoromethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3S,4S)-4-methyl-1-[(2-methylpyrimidine-4-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3S,4S)-1-{[2-(dimethylamino)pyrimidine-4-yl]methyl}-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-cyclopropyl-1-[(5-methylpyrazine-2-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-cyclopropyl-1-(quinoxaline-6-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-cyclopropyl-1-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-methylpyrrolidine-3-yl]-1-ethyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-{(3,4-trans)-4-ethyl-1-[(2-methylpyrimidine-5-yl)methyl]pyrrolidine-3-yl}-1-isopropyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0029] 6-[(3S,4S)-1-benzyl-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-(2,2,2-trifluoroethyl)pyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-methylpyrrolidine-3-yl]-1-(4,4-difluorocyclohexyl)-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-methylpyrrolidine-3-yl]-1-(2,2,2-trifluoroethyl)-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-[(3S,4S)-4-methyl-1-(1,5-naphthyridine-4-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-[(3S,4S)-4-methyl-1-(1,8-naphthyridine-4-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-[(3S,4S)-4-methyl-1-(quinoline-4-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-[(3S,4S)-4-methyl-1-(pyrido[2,3-b]pyrazine-8-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Isopropyl-6-{(3,4-trans)-1-[(6-methoxy-1,5-naphthyridine-4-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3,4-trans)-1-[(8-fluoroquinoline-2-yl)methyl]-4-methylpyrrolidine-3-yl}-1-isopropyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one;

[0030] 1-Isopropyl-6-{(v)-1-[(6-methoxyquinoline-4-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-methylpyrrolidine-3-yl]-1-cyclobutyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-4-methyl-1-[(2-methylpyrimidine-5-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3,4-trans)-4-ethyl-1-[(2-methylpyrimidine-5-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-4-methyl-1-[(5-methylpyrazine-2-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-1-[(6-methoxypyridine-3-yl)methyl]-4-methylpyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-4-methyl-1-(quinoline-3-ylmethyl)pyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-4-methyl-1-[(2-methylpyrimidine-4-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-4-methyl-1-[(6-methylpyridine-3-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-4-methyl-1-[(6-(trifluoromethyl)pyridine-3-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0031] 6-{(3S,4S)-4-methyl-1-[(1-methyl-1H-imidazo[4,5-c]pyridine-2-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-1-[(1,3-dimethyl-1H-pyrazole-5-yl)methyl]-4-methylpyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclobutyl-6-{(3,4-trans)-4-methyl-1-[(2-methylpyrimidine-5-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-(2,1,3-benzothiadiazole-5-ylmethyl)-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-4-methyl-1-(quinoxaline-2-ylmethyl)pyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-4-methyl-1-(quinoline-4-ylmethyl)pyrrolidine-3-yl]-1-tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-4-methyl-1-(pyridine-2-ylmethyl)pyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-benzyl-4-methylpyrrolidine-3-yl]-3-methyl-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-(3-fluorobenzyl)-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-(3,5-difluorobenzyl)-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0032] 6-{(3S,4S)-4-methyl-1-[4-(trifluoromethyl)benzyl]pyrroridine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-benzyl-4-ethylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-benzyl-4-ethylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 3-methyl-6-[(3S,4S)-4-methyl-1-(pyridine-3-ylmethyl)pyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 3-methyl-6-{(3S,4S)-4-methyl-1-[(2-methylpyrimidine-5-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-1-[(6-methoxypyridine-3-yl)methyl]-4-methylpyrrolidine-3-yl}-3-methyl-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-4-methyl-1-[(6-methylpyridine-2-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-(4-fluorobenzyl)-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-(2-fluorobenzyl)-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-4-methyl-1-[2-(trifluoromethyl)benzyl]pyrroridine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0033] 6-[(3S,4S)-1-(2,4-difluorobenzyl)-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-(4-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-benzyl-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-thiopyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-(2-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-(3-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-4-methyl-1-[3-(trifluoromethyl)benzyl]pyrroridine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-1-(26-difluorobenzyl)-4-methylpyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-4-ethyl-1-[(5-methylpyrazine-2-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-{(3S,4S)-4-ethyl-1-[(6-methoxypyridine-3-yl)methyl]pyrrolidine-3-yl}-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-4-ethyl-1-(pyridine-2-ylmethyl)pyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0034] 6-[(3S,4S)-4-ethyl-1-(quinoxaline-2-ylcarbonyl)pyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3S,4S)-4-methyl-1-(pyrimidine-2-ylmethyl)pyrrolidine-3-yl]-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 2-({(3S,4S)-3-ethyl-4-[4-oxo-1-(tetrahydro-2H-pyran-4-yl)-45-dihydro-1H-pyrazolo[3,4-d]pyrimidine-6-yl]pyrrolidine-1-yl}methyl)benzonitrile; 3-({(3S,4S)-3-ethyl-4-[4-oxo-1-(tetrahydro-2H-pyran-4-yl)-45-dihydro-1H-pyrazolo[3,4-d]pyrimidine-6-yl]pyrrolidine-1-yl}methyl)benzonitrile; 4-({(3S,4S)-3-ethyl-4-[4-oxo-1-(tetrahydro-2H-pyran-4-yl)-45-dihydro-1H-pyrazolo[3,4-d]pyrimidine-6-yl]pyrrolidine-1-yl}methyl)benzonitrile; 1-Cyclopentyl-6-(3,4-trans)-4-methyl-1-[3-(1H-pyrazole-1-yl)benzyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(2-methylpyridine-4-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-(2-chloro-6-fluorobenzyl)-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,3-dimethylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[2-(difluoromethoxy)benzyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-]pyrimidine-4-one

[0035] 1-Cyclopentyl-6-{(3,4-trans)-1-[(2-ethoxypyridine-3-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(4,5,6,7-tetrahydropyrazolo[1,5-a]pyridine-3-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,3-dihydro-1,4-benzodioxin-6-ylmethyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[4-(1H-imidazole-1-yl)benzyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,5-dichlorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(4-methoxy-3-methylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,3-dihydro-1-benzofuran-7-ylmethyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,3-difluorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(5-fluoro-2-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2-fluoro-4-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0036] 1-Cyclopentyl-6-[(3,4-trans)-1-(3-fluoro-4-methylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(2-methyl-1,3-thiazole-5-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[(4-isopropyl-1,3-thiazole-2-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[(1,3-dimethyl-1H-pyrazole-5-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,3-difluoro-4-methylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-[[6-(1H-pyrazole-1-yl)pyridine-2-yl]methyl]pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(4-methylbenzyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(2-naphthylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[(2-methoxypyridine-3-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2-ethoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0037] 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[4-(1H-1,2,4-triazole-1-yl)benzyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3-methoxy-4-methylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(1-naphthylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3-fluoro-4-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,5-dimethoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(5-methylisoxazole-3-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2-fluoro-6-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,4-difluorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(4-fluoro-3-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,3-dihydro-1,4-benzodioxin-5-ylmethyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one;

[0038] 6-[(3,4-trans)-1-(2-chloro-4-fluorobenzyl)-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,4-dimethylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3,5-dimethoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3-ethoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-(4-chloro-2-fluorobenzyl)-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 3-{[(3,4-trans)-3-(1-cyclopentyl-4-oxo-4,5-dihydro-1H-pyrazolo[3,4-d]pyrimidine-6-yl)-4-methylpyrrolidine-1-yl]methyl}benzonitrile; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,5-difluorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 2-{[(3,4-trans)-3-(1-cyclopentyl-4-oxo-4,5-dihydro-1H-pyrazolo[3,4-d]pyrimidine-6-yl)-4-methylpyrrolidine-1-yl]methyl}benzonitrile; 6-[(3,4-trans)-1-(3-chloro-4-fluorobenzyl)-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[4-(difluoromethoxy)benzyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0039] 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(3-methylbenzyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3,4-difluorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,5-dimethylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-(3-chloro-2-fluorobenzyl)-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,3-dichlorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(1,3-thiazole-2-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3-fluoro-2-methylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(2-methylpyrimidine-5-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[(2-ethylpyrimidine-5-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(4-isopropylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0040] 1-Cyclopentyl-6-{(3,4-Trans-1-[(1-ethyl-1H-pyrazole-4-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-Dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[(4-methoxypyridine-3-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(isoxazole-5-ylmethyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(4-ethoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-{[6-(1-hydroxy-1-methylethyl)pyridine-3-yl]methyl}-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[(2,2-dimethyl-2,3-dihydro-1-benzofuran-5-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3,4-dimethoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(5-methylpyrazine-2-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(imidazo[1,2-a]pyridine-2-ylmethyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(2-phenyl-1,3-oxazol-4-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0041] 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(2-methylbenzyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2-isopropoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-(sinnoline-3-ylmethyl)-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[3-(difluoromethoxy)benzyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(4-fluoro-3-methylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[4-(1H-pyrazole-1-yl)benzyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[(2,7-dimethylimidazo[1,2-a]pyridine-3-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3,5-dichlorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(4-isopropoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-[[2-(1-hydroxy-1-methylethyl)pyridine-4-yl]methyl]-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one;

[0042] 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(4,5,6,7-tetrahydro-1,3-benzothiazole-2-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(mesitylmethyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,6-dichlorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 4-{[(3,4-trans)-3-(1-cyclopentyl-4-oxo-4,5-dihydro-1H-pyrazolo[3,4-d]pyrimidine-6-yl)-4-methylpyrrolidine-1-yl]methyl}benzonitrile; 1-Cyclopentyl-6-[(3,4-trans)-1-(2-fluoro-5-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,6-dimethylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[(4-methoxy-3,5-dimethylpyridine-2-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3,5-dimethylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3,4-dimethylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(1-methyl-1H-benzimidazole-2-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0043] 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(4-methyl-3,4-dihydro-2H-1,4-benzoxazine-7-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(3-phenylpropyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[2-(trifluoromethyl)benzyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(4,4,4-trifluorobutyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(cyclopentylmethyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,4-dimethoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[4-(morpholine-4-ylmethyl)benzyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-(2,1,3-benzothiadiazole-5-ylmethyl)-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-{(3,4-trans)-1-[2-(benzyloxy)ethyl]-4-methylpyrrolidine-3-yl}-1-cyclopentyl-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one;

[0044] 1-Cyclopentyl-6-[(3,4-trans)-1-(2,6-difluorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2-methoxybenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(3,5,6-trimethylpyrazine-2-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,4-dichlorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-4-methyl-1-(5,6,7,8-tetrahydro-4H-pyrazolo[1,5-a]azepine-3-ylmethyl)pyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3-fluorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2,3-dihydro-1-benzofuran-5-ylmethyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2-methoxy-5-methylbenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(2-fluorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-(2-chlorobenzyl)-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one;

[0045] 1-Cyclopentyl-6-[(3,4-trans)-1-(3,4-dichlorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 6-[(3,4-trans)-1-(2,1,3-benzothiadiazole-4-ylmethyl)-4-methylpyrrolidine-3-yl]-1-cyclopentyl-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(2-propylpyrimidine-5-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-1-[(1-ethyl-1H-pyrazole-5-yl)methyl]-4-methylpyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[2-(trifluoromethoxy)benzyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[4-(trifluoromethyl)benzyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-{(3,4-trans)-4-methyl-1-[(1-methyl-1H-imidazo[4,5-c]pyridine-2-yl)methyl]pyrrolidine-3-yl}-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; 1-Cyclopentyl-6-[(3,4-trans)-1-(3,5-difluorobenzyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrzolo[3,4-d]pyrimidine-4-one; or 1-Cyclopentyl-6-[(3,4-trans)-1-(5,6-dihydro-4H-pyrrolo[1,2-b]pyrazole-3-ylmethyl)-4-methylpyrrolidine-3-yl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one; Or, a pharmaceutically acceptable salt thereof.

[0046] In a preferred embodiment, the compound is 6-((3S,4S)-4-methyl-1-(pyrimidine-2-ylmethyl)pyrrolidine-3-yl)-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one ("Compound A") or a pharmaceutically acceptable salt thereof.

[0047] The compounds of this application have been surprisingly found to exhibit pharmacological activity, including selective inhibition of PDE9, making them suitable for the treatment, prevention, and / or control of diseases that can be regulated or normalized by inhibition of PDE9.

[0048] The compounds and intermediates of this application may be named according to either the IUPAC (International Union of Pure and Applied Chemistry) or CAS (Chemical Abstracts Service, Columbus, Ohio) nomenclature system.

[0049] Compounds of formula (I) may contain a chiral or asymmetric center and therefore may exist in different stereoisomeric forms. Those skilled in the art will understand that, unless otherwise specified, all stereoisomers (e.g., enantiomers and diastereoisomers, and racemic mixtures thereof) of the novel compounds and intermediates described, illustrated and / or discussed herein are within the scope of the claims of this application. Furthermore, unless otherwise specified, this application includes all geometric and positional isomers. (3S,4S) enantiomers of the corepyrrolidinyl structure are preferred.

[0050] Diastereomer mixtures can be separated into individual diastereomers based on their physicochemical differences by methods well known to those skilled in the art, such as chromatography and / or fractionation crystallization. Enantiomers can be separated by reacting the enantiomer mixture with a suitable optically active compound (e.g., an alcohol) to convert it into a diastereomer mixture, separating the diastereomers, and converting the individual diastereomers back into their corresponding pure enantiomers (e.g., by hydrolysis). Additional methods include the separation of racemic mixtures using chiral salts and chiral chromatography.

[0051] Those skilled in the art will further recognize that the compound of formula (I) may exist in crystalline form as a hydrate with water molecules incorporated into its crystal structure, and as a solvate with solvent molecules incorporated. All of these hydrate and solvate forms are considered part of this application.

[0052] Those skilled in the art will understand that certain compounds of formula (I) may exist as tauthelomeric isomers, that is, an equilibrium exists between two isomers that are in rapid equilibrium with each other. A common example of tautomerism is keto-enol tautomerism, which is as follows:

[0053] [ka]

[0054] The extent to which one tautomer exists relative to the other depends on various factors, including the substitution pattern and the type of solvent. Other examples that conform to this application will be known to those skilled in the art. Unless otherwise specified, all tautomer forms of formula (I) are included in the scope of this application.

[0055] This application also includes prodrugs of the compounds of this application. A "prodrug" refers to a drug precursor that releases a drug in vivo through chemical or physiological processes (e.g., upon reaching physiological pH or by enzymatic activity). A prodrug may itself have biological activity or may be converted into a biologically active compound during residence in the body (e.g., by metabolism or hydrolysis). Considerations regarding the preparation and use of prodrugs are described in "Prodrugs as Novel Delivery Systems" by Higuchi & Stella (A.C.S. Symposium Series, Volume 14) and "Bioreversible Carriers in Drug Design" edited by Edward B. Roche (American Pharmaceutical Association and Pergamon Press, 1987). All prodrugs of the various compounds of formula (I) are included within the scope of this application.

[0056] This application also encompasses isotopically labeled compounds of formula (I) that are identical to the compounds described herein, except that one or more atoms are replaced with atoms having an atomic weight or mass number different from the atomic weight or mass number normally found in nature. Examples of isotopes that can be incorporated into the compounds of formula (I) include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, fluorine, and chlorine, such as H, 3 H, 13 C, 14 C, 15 N, 18 O, 17 O, 31 P, 32 P, 35 S, 18 F and 36 Cl and the like. Compounds of formula (I) containing the aforementioned isotopes and / or other isotopes of other atoms, as well as their pharmaceutically acceptable salts, are included within the scope of this application.

[0057] Certain isotopically labeled compounds of formula (I), such as 3 H and 14Products incorporating radioactive isotopes such as 13C are useful for tissue distribution assays of drugs and / or substrates. Tritiation, i.e. 3 H and 14 Isotopes of 1C are particularly preferred due to their ease of preparation and detectability. Furthermore, deuterium, i.e. 2 Substitution with heavier isotopes such as H may yield certain therapeutic benefits due to improved metabolic stability (e.g., extended in vivo half-life) and may therefore be preferable in some situations. The isotope-labeled compounds of formula (I) and their pharmaceutically acceptable salts can generally be prepared by performing procedures similar to those disclosed in the following scheme and / or examples, and by substituting the non-isotopically labeled reagent with a readily available isotope-labeled reagent.

[0058] definition The specific terms used herein are generally defined as follows:

[0059] In this specification, the number of carbon atoms in various hydrocarbon-containing subgroups may be expressed by prefixes indicating the minimum and maximum number of carbon atoms in the subgroup. For example, (C1-C6)alkyl refers to an alkyl group containing 1 to 6 carbon atoms.

[0060] The term "alkoxy" refers to a linear or branched monovalent saturated aliphatic hydrocarbon radical bonded to an oxygen atom in the core structure. Examples of alkoxy groups include methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, tert-butoxy, and pentoxy.

[0061] The term "alkyl" refers to a saturated, monovalent, linear or branched aliphatic hydrocarbon radical. Examples of alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, sec-butyl, pentyl, isopentyl, neopentyl, hexyl, and isohexyl.

[0062] The term "alkenyl" refers to a partially unsaturated, linear or branched aliphatic hydrocarbon radical having one or more double bonds. Examples of alkenyl groups include ethenyl (also known as "vinyl"), allyl, 1-propenyl, isopropenyl, n-butenyl, and n-pentenyl. The term "alkenyl" includes radicals having "cis" and "trans" orientations, or "Z" and "E" orientations.

[0063] The term "alkynyl" refers to a partially unsaturated, linear or branched aliphatic hydrocarbon radical having one or more double bonds. Examples of alkynyl groups include 1-propynyl, 2-propynyl (also known as "propargyl"), 1-butynyl, 2-butynyl, and 1-pentynyl.

[0064] The term "aryl" refers to monocyclic or polycyclic aromatic ring systems, such as anthracenyl, benzyl, fluorenyl, indenyl, naphthyl, phenantrenyl, and phenyl. The term "aryl" is also intended to include partially hydrogenated derivatives of such ring systems, such as 1,2,3,4-tetrahydronaphthyl.

[0065] The term "aryloxy" refers to an aryl radical, such as benzyloxy, which is bonded to an oxygen atom attached to a skeletal structure.

[0066] The term "cycloalkyl" refers to saturated monocyclic or bicyclic cycloalkyl groups. Examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.

[0067] The term "halogen" or "halo" refers to the atoms and radicals of chlorine, bromine, fluorine, and iodine.

[0068] The term "haloalkyl" refers to an alkyl or cycloalkyl substituent in which at least one hydrogen radical is substituted with a halogen group. If multiple hydrogens are substituted with halogens, the halogens may be the same or different. Examples of haloalkyl radicals include trifluoromethyl, 2,2,2-trifluoroethyl, 4,4,4-trifluorobutyl, 4,4-difluorocyclohexyl, chloromethyl, dichloromethyl, trichloromethyl, and 1-bromoethyl.

[0069] The term "haloalkoxy" refers to an alkoxy radical in which at least one hydrogen atom radical is substituted with a halogen atom. If multiple hydrogens are substituted with halogens, the halogens may be the same or different. Examples of haloalkoxy radicals include difluoromethoxy, trifluoromethoxy, 2,2,2-trifluoroethoxy, chloromethoxy, and bromomethoxy.

[0070] As used herein, the term "heteroaryl" includes unsaturated heterocyclic systems containing one or more heteroatoms, such as nitrogen, oxygen, and sulfur. If a heteroaryl group contains one or more heteroatoms, these heteroatoms may be identical or distinct. Heteroaryl radicals may be bonded via carbon atoms or heteroatoms. The term "heteroaryl" also includes partially hydrogenated derivatives of such cyclic systems. Examples of heteroaryl groups include furanyl (also known as "furyl"), imidazolinyl, imidazolyl (also known as "1,3-diazolyl"), indolyl, oxadiazolyl, oxazinyl, oxazolyl, isoxazolyl, pyranyl, pyrazinyl (also known as "1,4-diadinyl"), pyrazolyl (also known as "1,2-diazolyl"), pyrazolinyl, pyrazyl, pyridadinyl (also known as "1,2-diadinyl"), pyridyl (also known as "pyridinyl"), pyrimidinyl (also known as "1,3-diadinyl" and "pyrimidyl"), pyrrolyl, thiadiadinyl, thiadiazolyl, thiatriazolyl, thiazolyl, isothiazolyl, thienyl, thiofuranyl (also known as "thiophenyl"), thiopyranyl, triazinyl, triazolyl, and similar groups.

[0071] The term "heteroaryl" includes radicals in which two or three rings are fused, with at least one of the rings containing a heteroatom as a ring atom, including (a) a group in which a heterocycloalkyl ring is fused with an aryl ring or a heteroaryl ring, or (b) a radical in which a cycloalkyl ring is fused with a heteroaryl ring. Examples of heteroaryl compounds formed by the condensation of two rings include benzodioxynyl, dihydrobenzodioxynyl, benzofuranyl, dihydrobenzofuranyl, isobenzofuranyl, benzimidazolyl, benzothiadiazolyl, tetrahydrobenzothiadiazolyl, benzothiazolyl, benzothienyl (also known as "benzothiophenyl", "thionaphthenyl", and "benzothiofuranyl"), benzoxazinyl, dihydrobenzoxazinyl, benzoxazolyl, chromanil, isochromanil, chromenyl, sinnolinyl (also known as "1,2-benzodiadinyl"), imidazopyridinyl (e.g., imidazo[1,2-a]pyridinyl or imidazo[4,5-c]pyridinyl), indazolyl, indolinyl, isoindolinyl, indolyl, isoindolyl, naphthyridinyl, oxathiopyrrolyl, pteridinyl, phthalazinyl, purinyl (also known as "imidazo[4, [5-d]pyrimidinyl), pyranopyrrolyl, pyrazoloazepinyl, tetrahydropyrazoroazepinyl (e.g., tetrahydropyrazoro[1,5-a]azepinyl), pyrazolopyridinyl, tetrahydropyrazoropyridinyl (e.g., tetrahydropyrazoro[1,5-a]pyridinyl), pyrazolopyridinyl (e.g., pyrazolo[3,4-d]pyridinyl), pyridopyridinyl (e.g., pyrido[2,3-b]pyridinyl), pyridopyridinyl Examples include nyl, pyrrolopyrazolyl, dihydropyrrolopyrazolyl (e.g., dihydropyrrolo[1,2-b]pyrazolyl), quinazolinyl (also known as "1,3-benzodiadinyl"), quinolinyl (also known as "1-benzazinyl"), isoquinoline group (also known as "2-benzazinyl"), quinolidinyl group, quinolyl group, isoquinolyl group, quinoxalinyl group, dithianaphthalenyl group, and thienofuranyl group (e.g., thieno[3,2-b]furanyl group).

[0072] Examples of heteroaryls having a 3-position fused ring include acridinyl, diazaantryl, triazaphenanthrene, carbazolyl, carbolinyl, furosinolinyl, perimidinyl, phenanthridine, phenanthrolinyl, phenazinyl, phenothiazinyl, phenoxazinyl, phenoxazinyl, thianthrenyl, and xanthenyl.

[0073] The term "heterocycloalkyl" refers to a saturated monocyclic or polycyclic cycloalkyl group in which at least one carbon atom is substituted with a heteroatom such as nitrogen, oxygen, or sulfur. If the heterocycle contains one or more heteroatoms, these heteroatoms may be the same or different. Heterocycloalkyl radicals may be bonded via carbon atoms or heteroatoms. Examples of heterocycloalkyl groups include azetidinyl, dioxacyclohexyl, 1,3-dioxolanyl, imidazolidinyl, morpholinyl, piperadinyl, piperidinyl, pyrazolidinyl, pyrrolidinyl, tetrahydrofuranyl, tetrahydropyranyl, tetrahydrothiopyranyl, and thiazanyl.

[0074] A cyclic group may be bonded to other groups in multiple ways. Unless a specific bond configuration is specified, all possible configurations are intended. For example, the term "pyridyl" includes 2-, 3-, or 4-pyridyl (2-, 3-, or 4-pyridinyl).

[0075] The term "oxo" refers to a carbonyl group formed by the bonding of a carbon atom and an oxygen atom.

[0076] The term "salt" refers to both organic and inorganic salts of the compound of formula (I). Such salts can be prepared in situ during the final isolation and purification of the compound, or by reacting the compound, prodrug, or stereoisomer of formula (I), a suitable organic or inorganic acid, or a base individually, and then isolating the salts thus formed. Typical anionic salts include hydrobromide, hydrochloride, hydroiodide, sulfate, bisulfate, nitrate, acetate, trifluoroacetate, oxalate, besylate, palmitate, pamoate, malonate, stearate, laurate, malate, borate, benzoate, lactate, phosphate, hexafluorophosphate, benzenesulfonate, tosylate, formate, citrate, maleate, fumarate, succinate, tartrate, naphthylate, mesylate, glucoheptonate, lactobionate, and laurylsulfonate. Typical cationic salts include sodium, potassium, calcium, and magnesium salts. For an overview, see, for example, Berge, et al., J. Pharm. Sci., 66, 1-19 (1977).

[0077] Salts of the compound of formula (I) can be readily prepared by mixing a solution of the compound of formula (I) with a desired acid or base (as appropriate). The salt precipitates from the solution and is recovered by filtration or by evaporation of the solvent.

[0078] The term "radical" refers to a group of atoms that behave as a single reactant in a chemical reaction. For example, an organic radical is a group of atoms that gives a compound containing it characteristic properties, or a group of atoms that remain unchanged throughout a series of reactions or transformations.

[0079] The symbol "-" represents a covalent bond.

[0080] The term "reaction-inert solvent" or "inert solvent" refers to a solvent or mixture of solvents that does not interact with the starting material, reagent, intermediate, or product in a manner that would adversely affect their desired properties.

[0081] As used herein, the terms “to treat,” “to treat,” “treated,” or “treatment” include preventative (e.g., prophylactic), palliative, or curative uses or outcomes.

[0082] Pharmaceutical composition This application also includes a pharmaceutical composition comprising a predetermined amount of a compound represented by formula (I) or a pharmaceutically acceptable salt thereof, and optionally a pharmaceutically acceptable medium, carrier, or diluent. In some embodiments, the pharmaceutical composition essentially comprises a compound represented by formula (I) or a pharmaceutically acceptable salt thereof. In a preferred embodiment, the pharmaceutical composition is in an amount effective to inhibit the enzyme PDE9 in a mammal. In another preferred embodiment, the mammal is a human.

[0083] Methods for preparing various pharmaceutical compositions containing predetermined amounts of active ingredients are known to those skilled in the art or are evident in light of the disclosures herein. See, for example, "Remington's Pharmaceutical Sciences" (Mack Publishing Co., Easton, Pa., 19th edition (1995)).

[0084] Suitable medicinal carriers, media, and diluents for such formulations include inert solid diluents or fillers, sterile aqueous solutions, and various organic solvents. Formulations formed by combining the compounds of this application with pharmaceutically acceptable carriers, media, or diluents can be readily administered in various dosage forms such as tablets, powders, lozenges, syrups, and injectable solutions.

[0085] Solid dosage forms for oral administration include capsules, tablets, powders, and granules. In such solid dosage forms, the active ingredient is at least one inert conventional pharmaceutical excipient (or carrier) such as sodium citrate, calcium carbonate, or dicalcium phosphate, or (a) fillers or bulking agents such as starch, lactose, sucrose, mannitol, and silicic acid; (b) binders such as carboxymethylcellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose, and acacia; (c) humectants such as glycerol; (d) disintegrants, such as agar, calcium carbonate, potato starch, or (e) Tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (f) Dissolution retarders, e.g., paraffin; (g) Absorption enhancers, e.g., quaternary ammonium compounds; (g) Wetting agents, e.g., cetyl alcohol and glycerol monostearate; (h) Adsorbents, e.g., kaolin and bentonite; and / or (i) Lubricants, e.g., talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, or mixtures thereof, may be mixed with the product. In the case of capsules and tablets, the dosage form may further contain a buffer.

[0086] Solid dosage forms can be prepared as sustained-release and pulsed-release formulations by including additional additives that function as release rate modifiers, in addition to the additives described above for immediate-release formulations, and by coating and / or incorporating these into the formulation body. Examples of release rate modifiers include hydroxypropyl methylcellulose, methylcellulose, sodium carboxymethylcellulose, ethylcellulose, cellulose acetate, polyethylene oxide, xanthan gum, ammonium methacrylate copolymer, hydrogenated castor oil, carnauba wax, paraffin wax, cellulose acetate phthalate, hydroxypropyl methylcellulose phthalate, methacrylic acid copolymer, and mixtures thereof. Sustained-release and pulsed-release dosage forms may contain one or a combination thereof of excipients that adjust the release rate.

[0087] The pharmaceutical compositions of this application may further include rapidly dispersible or rapidly dissolving formulations (FDDFs). The terms “dispersion” or “dissolution” used herein to describe FDDFs depend on the solubility of the active pharmaceutical ingredient (API) used. That is, if the API is insoluble, a rapidly dispersible formulation can be prepared, and if the API is soluble, a rapidly dissolving formulation can be prepared.

[0088] Similar types of solid compositions can also be used as fillers for soft or hard gelatin capsules, using excipients such as lactose or milk sugar and high molecular weight polyethylene glycol.

[0089] Solid dosage forms such as tablets, sugar-coated tablets, capsules, and granules can be prepared using enteric coatings, other coatings well known to those skilled in the art, and shells. These may contain opacifying agents and may have compositions that allow for delayed, sustained, or controlled release of the active ingredient. Examples of usable encapsulation compositions include polymers and waxes. Furthermore, the active ingredient may be microencapsulated using one or more of the above-mentioned additives, if necessary.

[0090] Liquid formulations for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs. In addition to the active ingredient, liquid formulations may contain water or other solvents, solubilizers, emulsifiers, and other inert diluents commonly used in this field. Examples include ethanol, isopropanol, ethyl carbonate, benzyl benzoate, propylene glycol, 1,3-butylene glycol, oils and fats (especially cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, and sesame oil), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycol, and fatty acid esters of sorbitan, or mixtures of these substances.

[0091] In addition to the active ingredient, the pharmaceutical composition may contain suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar, tragacanth, or mixtures thereof. It may also contain sweeteners, flavorings, and fragrances.

[0092] The pharmaceutical composition of this application may further contain excipients such as preservatives, wetting agents, emulsifiers, and dispersants. Microbial contamination of the composition can be prevented using various antibacterial and antifungal agents such as parabens, chlorobutanol, phenol, and sorbic acid. It may also be desirable to include isotonic agents, such as sugars and sodium chloride. To delay the absorption of the injectable pharmaceutical composition, absorption retarders such as aluminum monostearate or gelatin can be used.

[0093] For parenteral administration, solutions using sesame oil or peanut oil, propylene glycol aqueous solution, or sterile aqueous solution may be used. Such aqueous solutions should be appropriately buffered as needed, and the liquid diluent should first be isotonicized with sufficient saline or glucose. These aqueous solutions are particularly suitable for intravenous, intramuscular, subcutaneous, and intraperitoneal administration. In this regard, the sterile aqueous media used are readily available by standard techniques well known to those skilled in the art.

[0094] For intranasal or inhalation administration, the compound of formula (I) is conveniently administered in the form of a solution or suspension from a pump-type spray container squeezed or pumped by the patient, or in the form of an aerosol spray from a pressurized container or nebulizer using a suitable propellant (e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide, or other suitable gas). In the case of pressurized aerosols, the dose unit can be determined by providing a valve for dispensing a fixed amount. The pressurized container or nebulizer may contain a solution or suspension of the compound of this application. Capsules and cartridges (e.g., made of gelatin) for use in inhalers or inhalation devices may be prepared to contain the compound of this application or a powder mixture of the compound and a suitable powder base such as lactose or starch.

[0095] The pharmaceutical compositions of this application may also be formulated for therapeutic use in veterinary applications. In this case, the compounds of this application, or their veterinarily acceptable salts, or their veterinarily acceptable solvates or prodrugs, are administered as appropriately acceptable formulations in accordance with normal veterinary practice, and a veterinarian determines the most appropriate administration plan and route for a particular animal.

[0096] Generally, compounds of formula (I) and pharmaceutically acceptable salts thereof can be prepared according to the exemplary synthetic routes disclosed in the chemical formula and examples disclosed in U.S. Patent No. 7,964,607, which is incorporated herein by whole reference, as well as by other conventional preparation procedures that are well known to those skilled in the art or are obvious in light of this disclosure.

[0097] Treatment method The present application also includes a method for treating chronic heart failure in a subject requiring treatment, the method comprising, to the subject, a therapeutically effective amount of a pharmaceutical composition containing (a) a compound of formula (I) or a pharmaceutically acceptable salt thereof, or (b) a compound of formula (I) or a pharmaceutically acceptable salt thereof, in a pharmaceutically acceptable medium, carrier or diluent, either alone or in combination with the second agent described above.

[0098] This application also includes using (a) a compound represented by formula (I) or a pharmaceutically acceptable salt thereof, or (b) a pharmaceutical composition comprising a compound represented by formula (I) or a pharmaceutically acceptable salt thereof, for the manufacture of a pharmaceutical for the treatment of chronic heart failure in subjects requiring such treatment.

[0099] This application also includes (a) a compound of formula (I) or a pharmaceutically acceptable salt thereof; or (b) a pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof, for use in the treatment of chronic heart failure in subjects requiring such treatment.

[0100] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt thereof, is 6-((3S,4S)-4-methyl-1-(pyrimidine-2-ylmethyl)pyrrolidine-3-yl)-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one ("Compound A") or a pharmaceutically acceptable salt thereof.

[0101] Dosage and method of administration The appropriate administration regimen, dosage, and dosing interval of this compound will vary, in particular, depending on the use of the compound of formula (I) in this application, the type of pharmaceutical composition used, the characteristics of the patient, and the type and severity of the disease being treated. Generally, the effective dose of the compound of formula (I) or a pharmaceutically acceptable salt thereof is in the range of about 5 mg to about 100 mg per day. In some embodiments, the compound of formula (I) is administered in doses of 0.1 mg to 0.5 mg, 0.5 mg to 1 mg, 1 mg to 5 mg, 5 mg to 90 mg, 5 mg to 80 mg, 5 mg to 70 mg, 5 mg to 60 mg, 5 mg to 50 mg, 5 mg to 45 mg, 5 mg to 40 mg, 5 mg to 35 mg, 5 mg to 30 mg, 5 mg to 25 mg, 5 g to 20 mg, 5 mg to 15 mg, 5 mg to 10 mg, 10 mg to 50 mg, 5 g to 100 mg, 100 mg to 150 mg, 150 mg to 200 mg, 200 mg to 250 mg, or 250 mg to 300 mg. In some embodiments, the compound of formula (I) is administered in doses of 1 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, 90 mg, 100 mg, 150 mg, 200 mg, 250 mg, or 300 mg.

[0102] The drug may be administered as a single dose (e.g., once daily) or multiple times. In some embodiments, the compound of formula (I) may be administered every 4 hours, every 8 hours, every 12 hours, every 18 hours, or every 24 hours, once daily, every other day, every 3 days, or once weekly. In some embodiments, 25 mg of the compound of formula (I) may be administered every 12 hours. In some embodiments, 50 mg of the compound of formula (I) may be administered per day. In a preferred embodiment, the compound of formula (I) may be administered twice daily. In another preferred embodiment, the compound of formula (I) may be administered every 12 hours. In some embodiments, the drug may be administered for 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 4 months, 6 months, 8 months, 10 months, 1 year, or longer. The drug may be continued until the patient's symptoms improve. In some embodiments, the compound of formula (I) is administered over the remainder of the patient's life. Administration may be continuous or intermittent.

[0103] The general dosage range may require some variation depending on the age and weight of the subject being treated, the intended route of administration, and the specific compound being administered. Determining the dosage range and optimal dose for a specific mammalian subject is possible for a person skilled in the art who is familiar with the contents of this disclosure.

[0104] Compounds of formula (I) include oral, buccal, sublingual, ocular, topical (e.g., transdermal), parenteral (e.g., intravenous, intramuscular, or subcutaneous), rectal, intraventricular, vaginal, intraperitoneal, intrabladder, topical (e.g., powder, ointment, or eye drops), intranasal and / or inhalation formulations, or “flush” formulations, i.e., methods for dissolving the drug in the mouth without the need for water. As those skilled in the art will understand, an appropriate dosing plan, the amount of each dose, and the dosing interval of the compound will vary depending on the compound of formula (I) or its prodrug used, the type of formulation used, the characteristics of the person being treated, and / or the severity of the disease being treated. In a preferred embodiment, the compound of formula (I) is administered orally.

[0105] This application also includes a method for inhibiting PDE9 in mammals, comprising administering to mammals requiring PDE9 inhibition a formulation containing (a) a compound of formula (I) or a pharmaceutically acceptable salt thereof, or (b) a compound of formula (I) or a pharmaceutically acceptable salt thereof, in a pharmaceutically acceptable medium, carrier, or diluent, either alone or in combination with the second agent described above.

[0106] Target patient group The therapeutic methods described herein are useful for treating cardiac diseases, particularly those related to abnormal PDE9 / cGMP signaling. In a preferred embodiment, the patient treated according to the methods described herein suffers from heart failure.

[0107] In some embodiments, the subjects treated according to the methods described herein have chronic heart failure. In some embodiments, the patient has chronic heart failure with reduced ejection fraction (EF). In some embodiments, the patient has chronic heart failure with preserved EF. In some embodiments, the patient has chronic heart failure with moderate EF (between 40% and 60%, or between 40% and 50%). In some embodiments, the patient has chronic heart failure with superior EF (greater than 60%). In some embodiments, the patient has left ventricular hypertrophy. In some embodiments, the patient has congestive heart failure.

[0108] In some embodiments, the subjects have an ejection fraction (EF) of less than 40% as determined by echocardiography. In some embodiments, the subjects have an EF of 40% to 60% as determined by echocardiography. In some embodiments, the subjects have an EF of more than 60% as determined by echocardiography. In some embodiments, the subjects have an EF of less than 60% as measured by echocardiography. In some embodiments, the subjects have an EF of more than 40% as measured by echocardiography and exhibit left atrial enlargement. Treatment of chronic heart failure may lead to improvement in EF.

[0109] In some embodiments, the target plasma NT-proBNP concentration is at least 600 pg / ml. In some embodiments, the target plasma NT-proBNP concentration is at least 300 pg / ml. In some embodiments, the subject has atrial fibrillation or atrial flutter and an NT-proBNP concentration of 1000 pg / mL or higher. In some embodiments, the subject has atrial fibrillation or atrial flutter and an NT-proBNP concentration of 500 pg / mL or higher. Treatment of chronic heart failure may improve the patient's NT-proBNP concentration (e.g., plasma NT-proBNP concentration or serum NT-proBNP concentration).

[0110] In this specification, the terms “subject” and “patient” are used interchangeably. In some embodiments, the patient is a human being.

[0111] Determination of effectiveness

[0112] The term “therapeutic dose” as used with respect to compounds of formula (I) refers to an amount of the compound of formula (I) sufficient to produce one or more desired therapeutic effects. Such effects include, for example, improvement of symptoms or clinical markers. Any suitable method known to those skilled in the art or described herein may be used to measure the effectiveness of the treatments described herein.

[0113] In some embodiments, the methods described herein result in a decrease in N-terminal pro-B natriuretic peptide (NT-pro-BNP) levels and / or B natriuretic peptide (BNP) levels in the plasma of the subject. NT-pro-BNP and BNP are circulating biomarkers detectable in the blood. Both are derived from the same propeptide produced by the heart in response to cardiomyocyte stretching. BNP induces diuresis, vasodilation, and decreased secretion of renin and aldosterone (see Hall, J Card Fail 2005 Jun;11(5 Suppl):S81-3). Elevated levels of NT-pro-BNP and / or BNP are biomarkers of poor prognosis in several types of heart disease (Salah et al., Heart 2019; 105:1182-1189). The normal range for NT-pro-BNP is 0–125 pg / mL in patients under 74 years of age and 0–450 pg / mL in patients 74 years of age and older. In patients under 50 years of age, values ​​exceeding 450 pg / mL, and in patients 50 years of age or older, values ​​exceeding 900 pg / mL, suggest impaired cardiac function. BNP levels below 100 pg / mL are considered normal, while values ​​above 100 pg / mL indicate impaired cardiac function.

[0114] In some embodiments, the treatments described herein reduce NT-proBNP levels measured in the plasma of the subject by about 10–20%, about 20–30%, about 30–40%, or about 40–50%. In some embodiments, the treatments described herein reduce NT-proBNP levels measured in the plasma of the subject by about 10%, about 20%, about 30%, about 40%, about 50%, or more than 50%.

[0115] In some embodiments, the treatments described herein reduce BNP levels measured in the plasma of the subject by about 10–20%, about 20–30%, about 30–40%, or about 40–50%. In some embodiments, the treatments described herein reduce BNP levels measured in the plasma of the subject by about 10%, about 20%, about 30%, about 40%, about 50%, or more than 50%.

[0116] In some embodiments, the therapies described herein result in an increase in cGMP levels in the plasma or urine of the subject. 3',5'-cyclic guanosine monophosphate (cGMP) is a second messenger involved in various signaling pathways, produced by guanylate cyclase. cGMP is broken down by phosphodiesterases 5 and 9 (PDE5 and PDE9) in the myocardium. Treatment with compounds of formula (I) that inhibit PDE9 is thought to increase cGMP levels.

[0117] In some embodiments, the treatments described herein increase cGMP levels by about 10–20%, about 20–30%, about 30–40%, or about 40–50%. In some embodiments, the treatments described herein increase cGMP levels by about 10%, about 20%, about 30%, about 40%, about 50%, or more than 50%. In some embodiments, cGMP levels are measured in the urine of the subject. In some embodiments, cGMP levels are measured in the blood of the subject. In some embodiments, cGMP levels are measured in the plasma of the subject.

[0118] In some embodiments, the therapies described herein modulate the ratio of cGMP to NT-proBNP and / or the ratio of cGMP to BNP. A decrease in the cGMP-to-BNP ratio has been reported as a poor prognostic factor in patients with heart failure (Lourenco et al., Eur J Heart Fail. 2009 Feb. 11(2): 185-190).

[0119] In some embodiments, the therapies described herein increase the cGMP to NT-proBNP ratio by about 10–20%, about 20–30%, about 30–40%, or about 40–50%. In some embodiments, the therapies described herein increase the cGMP to NT-proBNP ratio by about 10%, about 20%, about 30%, about 40%, about 50%, or more than 50%. In some embodiments, cGMP levels are measured in the subject's blood. In some embodiments, cGMP levels are measured in the subject's plasma. In some embodiments, cGMP levels are measured in the subject's urine. In some embodiments, NT-proBNP levels are measured in the subject's plasma.

[0120] In some embodiments, the treatments described herein increase the cGMP-to-BNP ratio by about 10–20%, about 20–30%, about 30–40%, or about 40–50%. In some embodiments, the treatments described herein increase the cGMP / BNP ratio by about 10%, about 20%, about 30%, about 40%, about 50%, or more than 50%. In some embodiments, the cGMP concentration is measured in the subject's blood. In some embodiments, the cGMP concentration is measured in the subject's plasma. In some embodiments, the BNP level in the subject's blood is measured. In some embodiments, the BNP level in the subject's plasma is measured. In some embodiments, the cGMP level in the subject's urine is measured.

[0121] In some embodiments, the therapies described herein result in an increase in the patient's Kansas City Cardiomyopathy Questionnaire (KCCQ) score. The KCCQ is an FDA-approved clinical outcome assessment tool and is recommended as a performance indicator to quantify the quality of care in heart failure (see Spertus et al., JACC 2020, 76(20)). It can measure the impact of heart failure on a patient's life and is strongly correlated with clinical events over time. Several versions of the KCCQ exist, including a 12-item version (KCCQ-12) and a 23-item version (KCCQ-23). ​​The KCCQ score ranges from 1 to 100, representing health status as follows: 0-24: Very Poor to Poor; 25-49: Poor to Fair; 50-74: Fair to Good; 75-100: Good to Very Good.

[0122] In some embodiments, the treatments described herein result in an improvement of 5–10 points, 10–15 points, 15–20 points, 20–25 points, 25–30 points, 30–35 points, 35–40 points, 40–45 points, 45–50 points, or 50 points or more on the KCCQ. In some embodiments, the treatments described herein result in an improvement of at least 5 points, at least 10 points, at least 15 points, at least 20 points, at least 25 points, at least 30 points, at least 35 points, at least 40 points, at least 45 points, or at least 50 points on the KCCQ.

[0123] The effectiveness of the treatments described herein can be measured at any appropriate point in time after administration. In some embodiments, effectiveness is determined 12 weeks after the first dose. In some embodiments, effectiveness is determined 6 months after the first dose. In some embodiments, effectiveness is determined 12 months after the first dose.

[0124] In some embodiments, the treatments described herein result in an improvement in the mortality rate of the subjects. In some embodiments, the methods described herein reduce the mortality rate by approximately 10–20%, 20–30%, 30–40%, or 40–50% compared to subjects of the same age diagnosed with chronic heart failure who have not been treated according to the methods described herein. In some embodiments, mortality is the mortality rate due to heart failure. In some embodiments, mortality is the cardiovascular mortality rate. In some embodiments, mortality is the all-cause mortality rate.

[0125] In some embodiments, the treatments described herein result in a reduction in hospitalizations due to heart failure in patients. In some embodiments, the method reduces hospitalizations due to heart failure by 10–20%, approximately 20–30%, approximately 30–40%, or approximately 40–50% compared to subjects of the same age diagnosed with chronic heart failure who have not been treated according to the method described herein.

[0126] Combination therapy This application includes the combination of a PDE9 inhibitor compound represented by formula (I) with one or more additional pharmaceutically active ingredients. When the active ingredients are used in combination, they can be administered sequentially or simultaneously, in separate dosage forms or combined into a single dosage form. Accordingly, this application also includes pharmaceutical compositions comprising the following amounts: (a) a first agent comprising the compound of formula (I) or a pharmaceutically acceptable salt thereof; (b) a second pharmacologically active substance; and (c) a pharmaceutically acceptable carrier, medium, or diluent.

[0127] Depending on the disease, disorder, or condition being treated, various pharmacologically active substances may be selected for use in combination with the compound of formula (I). Pharmacologically active substances that may be used in combination with the compositions of this application include, but are not limited to, the following:

[0128] (vi) Beta-adrenergic receptor blockers (beta-blockers), such as carteolol, esmolol (BREVIBLOC), labetalol (NORMODYNE, TRANDATE), oxprenolol (LARACOR, TRASACOR), pindolol (VISKEN), propranolol (INDERAL), sotalol (BETAPACE, SOTALEX, SOTACOR), timolol (BLOCADREN, TIMOPTIC), acebutolol (SECTRAL, PRENT), nadolol (CORGARD), metoprolol tartrate (LOPRESSOR), metoprolol succinate (TOPROL-XL), atenolol (TENORMIN), butoxamine, and SR 59230A (Sanofi).

[0129] (x) Calcium blockers, e.g., nilvadipine (ESCOR, NIVADIL), amlodipine (NORVASC, ISTIN, AMLODIN), felodipine (PLENDIL), nicardipine (CARDENE), nifedipine (ADALAT, PROCARDIA), MEM 1003 and its parent compounds nimodipine (NIMOTOP), nisoldipine (SULAR), nitrendipine, lacidipine (LACIPIL, MOTENS), lercanidipine (ZANIDIP), diltiazem (CARDIZEM), verapamil (CALAN, VERELAN), and enecazine (also known as NS-7).

[0130] (xi) Catechol O-methyltransferase (COMT) inhibitors, such as tolcapone (TASMAR), entacapone (COMTAN), and tropolone.

[0131] (xii) Central nervous system stimulants, such as caffeine, fenmetrazine, fendimetrazine, pemoline, fencamfamine (glucoenergan, reactvan), phenethylline (captagon), piperadol (meretran), deanol (also known as dimethylaminoethanol), methylphenidate (daytrana), methylphenidate hydrochloride (ritalin), dexmethylphenidate (focalin), amphetamine (alone or in combination with other central nervous system stimulants, such as adderall (amphetamine aspartate, amphetamine sulfate, dextroamphetamine saccharate, and dextroamphetamine sulfate)), dextroamphetamine sulfate (dexedrine, dextrostat), methamphetamine (desoxyn), lisdexamfetamine (vyvanse), and benzfetamine (didrex).

[0132] (xiii) Corticosteroids, such as prednisone (STERAPRED, DELTASONE), prednisolone (PRELONE), prednisolone acetate (OMNIPRED, PRED MILD, PRED FORTE), sodium prednisolone phosphate (ORAPRED ODT), methylprednisolone (MEDROL); methylprednisolone acetate (DEPO-MEDROL), and sodium methylprednisolone succinate (A-METHAPRED, SOLU-MEDROL).

[0133] (xiv) Dopamine receptor agonists, e.g., apomorphine (APOKYN), bromocriptine (PARLODEL), cabergoline (DOSTINEX), dihydroexidine, dihydroergocriptine, phenoldopam (CORLOPAM), lizlid (DOPERGIN), pergolide (PERMAX), pyribezil (TRIVASTAL, TRASTAL), pramipexole (MIRAPEX), quinpyrrole, ropinirole (REQUIP), and rotigotine (NEUPRO).

[0134] (xv) Dopamine receptor antagonists, e.g., tetrabenazine (NITOMAN, XENAZINE), 7-hydroxyamoxapine, droperidol (INAPSINE, DRIDOL, DROPLETAN), domperidone (MOTILIUM), L-741742, L-745870, lacloprid, SCH-23390, ecopipam, SKF-83566, and metoclopramide (REGLAN).

[0135] (xvi) Dopamine reuptake inhibitors, such as nomifensin maleate (MERITAL), banoxerine (also known as GBR-12909) and its decanoate ester DBL-583, and amineptin.

[0136] (xvii) Gamma-aminobutyric acid (GABA) receptor agonists, such as baclofen (LIORESAL, KEMSTRO), pentobarbital (NEMBUTAL), progavid (GABRENE), and clomethiazole.

[0137] (xviii) Immunomodulators, e.g., glatiramer acetate (also known as copolymer-1; COPAXONE), MBP-8298 (synthetic myelin basic protein peptide), dimethyl fumarate, fingolimod (also known as FTY720), linomide, lakinimod (also known as ABR-215062 and SAIK-MS), ABT-874 (human anti-IL-12 antibody), rituximab (RITUXAN), alemtuzumab (CAMPATH), daclizumab (ZENAPAX), and natalizumab (TSABRI).

[0138] (xix) Immunosuppressants, such as methotrexate (TREXALL, RHEUMATREX), mitoxantrone (NOVANTRONE), mycophenolate mofetil (CELLCEPT), sodium mycophenolate (MYFORTIC), azathioprine (AZASAN, IMURAN), mercaptopurine (PURI-NETHOL), cyclophosphamide (NEOSAR, CYTOXAN), chlorambucil (LEUKERAN), cladribine (LEUSTATIN, MYLINAX), alpha-fetoprotein, etanercept (ENBREL), and 4-benzyloxy-5-((5-undecyl-2H-pyrrole-2-ylidene)methyl)-2,2'-bi-1H-pyrrole (also known as PNU-156804).

[0139] (xx) Interferon, for example, interferon beta-1a (AVONEX, REBIF) and interferon beta-1b (BETASERON, BETAFERON).

[0140] (xxi) Levodopa (or its methyl or ethyl ester) alone, or in combination with a dopa decarboxylase inhibitor (e.g., carbidopa (SINEMET, CARBILEV, ARCOPA, V1512), benserazide (MADOPAR), α-methyldopa, monofluoromethyldopa, difluoromethyldopa, brocresin, or m-hydroxybenzylhydrazine).

[0141] (xxii) N-methyl-D-aspartate (NMDA) receptor antagonists, e.g., memantine (NAMENDA, AXURA, EBIXA), amantadine (SYMMETREL), acamprosate (CAMPRAL), besonprodil (also known as PD-196,860 or CI-1041), ketamine (KETALAR), dersemin (also known as NPS 1506), dexanabinol (also known as HU-211), dextromethorphan, dextrophan, traxoprodil (also known as CP-101,606), himantan, idantadol (also known as V-3381), lancisemin (also known as AR-R 15896), levorphanol (DROMORAN), methadone (DOLOPHINE), neramexane (MRZ Perzinfotel (also known as 2 / 579), phencyclidine, thianeptine (STABLON), dizosilpine (also known as MK-801), ibogaine, boacangine, tiletamine, riluzole (RILUTEK), aptiganel (CERESTAT), gabestinel, and remasimide.

[0142] (xxiii) Monoamine oxidase (MAO) inhibitors, e.g., selegiline (EMSAM), selegiline hydrochloride (I-deprenyl, ELDEPRYL, ZELAPAR), dimethyl selegiline, brophalomine, phenelzine (NARDIL), tranylcypromine (PARNATE), moclobemide (AURORIX, MANERIX), befloxatone, safinamide (also known as PNU-151774E), isocarboxazide (MARP) LAN), niaramid (NIAMID), rasagerine (AZILECT), iproniazid (MARSILID, IPROZID, IPRONID), iproclozid, troxatone (HUMORYL, PERENUM), bifemeran, deoxypeganin, harmine (also known as telepatin or banasterine), harmanine, linezolid (ZYVOX, ZYVOXID), and pargiline (EUDATIN, SUPIRDYL).

[0143] (xxiv) Muscarinic receptor (especially M1 subtype) agonists, e.g., betanethyl chloride (DUVOID, URECHOLINE), pilocarpine (SALAGEN), NGX267, arecoline, L-687306, L-689660, flutrethonium iodide (FURAMON, FURANOL), flutrethonium benzenesulfonate, flutrethonium p-toluenesulfonate, McN-A-343, oxotremoline, and carbachol (CARBASTAT, MIOSTAT, CARBOPTIC).

[0144] (xxv) Nicotinic receptor agonists, e.g., epibatidine, ABT-089, ABT-594, AZD-0328, R4996 (also known as MEM-63908), TC-5619, and EVP-6124.

[0145] (xxvi) Neuroprotective agents, e.g., 2,3,4,9-tetrahydro-1H-carbazole-3-onoxime, AL-108, ACD3480 (also known as TC-1734), bis(4-β-D-glucopyranosyloxybenzyl)-2-β-D-glucopyranosyl-2-isobutyl tartrate (also known as dactyloline B or DHB), zariproden (XAPRILA), dimevorin hydrochloride (DIMEBON), Disfenton (NXY-059, CEROVIVE), alundic acid (ONO-2506, PROGLIA, CEREACT), citicoline (also known as cytidine 5'-choline diphosphate), edaravone (RA DICUT), AEOL-10150, AGY-94806 (also known as SA-450, Msc-1), granulocyte colony-stimulating factor (AX-200), BAY-387271 (also known as KN-387271), DP-b99, HF-0220 (17-β-hydroxyepiandrosterone), HF-0420 (also known as oligotropin), pyridoxal 5'-phosphate (also known as MC-1), microplasmin, S-18986, piclozotan (also known as SUN-N4057), NP031112, L-seryl-L-methionyl-L-alanyl-L-lysyl-L-glutamyl-glycyl-L-valine, and SUN-N8075.

[0146] (xxvii) Norepinephrine (norepinephrine) reuptake inhibitors, such as atomoxetine (STRATTERA), doxepin (APONAL, ADAPIN, SINEQUAN), nortriptyline (AVENTYL, PAMELOR, NORTRILEN), amoxapine (ASENDIN, DEMOLOX, MOXIDIL), reboxetine (EDRONAX, VESTRA), biloxazine (VIVALAN), maprotiline (DEPRILEPT, LUDIOMIL, PSYMION), bupropion (WELLBUTRIN), and radaxafine.

[0147] (xxviii) Other PDE9 inhibitors, e.g., BAY 73-6691, and those disclosed in U.S. Patents 2003 / 0195205, 2004 / 0220186, 2006 / 0111372, and 2006 / 0106035.

[0148] (xxix) Other phosphodiesterase (PDE) inhibitors, e.g., (a) PDE1 inhibitors (e.g., vinpocetine (CAVINTON, CERACTIN, INTELECTOL) and those disclosed in U.S. Patent No. 6,235,742), (b) PDE2 inhibitors (e.g., erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), BAY 60-7550 and those described in U.S. Patent No. 6,174,884), (c) PDE4 inhibitors (e.g., rolipram, Ro 20-1724, ibdilast (KETAS), picramiraste (also known as RP73401), CDP840, siromiriraste (ARIFLO), roflumiriraste, tofimilast, ogremiriraste (also known as GRC) (3886), tetomirast (also known as OPC-6535), lilimiafast, theophylline (UNIPHYL, THEOLAIR), allophylline (also known as LAS-31025), doxophilin, RPR-122818, or mesembrine), and (d) PDE5 inhibitors (e.g., sildenafil (VIAGRA, REVATIO), tadalafil (CIALIS), val Denafil (LEVITRA, VIVANZA), Udenafil, Avanafil, Dipyridamole (PERSANTINE), E-4010, E-4021, E-8010, Zaprinast, PF489791, UK-357903, DA-8159, and those disclosed in international patent applications WO05 / 049616, WO06 / 120552, and WO07 / 122466.

[0149] (xxx) Quinolines, for example, quinine (including its hydrochloride, dihydrochloride, sulfate, bisulfate and gluconate), chloroquine, hydroxychloroquine (PLAQUENIL), mefloquine (LARIAM), and amodiaquine (CAMOQUIN, FLAVOQUINE).

[0150] (xxxi)β-secretase inhibitors, e.g., WY-25105, (+)-fencerine tartrate (POSIPHEN), LSN-2434074 (also known as LY-2434074), PNU-33312, KMI-574, SCH-745966, Ac-rER(N 2-Acetyl-D-arginyl-L-arginine), roxistatin (also known as E64d), and CA074Me.

[0151] (xxxii)y-secretase inhibitors, e.g., LY-411,575, LY-685,458, ELAN-G, ELAN-Z, 4-chloro-N-[2-ethyl-1(S)-(hydroxymethyl)butyl]benzenesulfonamide.

[0152] (xxxiii Serotonin (5-hydroxytryptamine) 1A (5-HT 1A ) Receptor antagonists, such as spiperone, levopindolol, BMY 7378, NAD-299, S(-)-UH-301, NAN 190, WAY 100635, and recozotan (also known as SRA-333).

[0153] (xxxiv) Serotonin (5-hydroxytryptamine) 6 (5-HT6) receptor antagonists, e.g., mianserin (TORVOL, BOLVIDON, NORVAL), methiotepine (also known as metitepine), ritanserin, ALX-1161, ALX-1175, MS-245, LY-483518 (also known as SGS518), MS-245, Ro 04-6790, RO 43-68544, Ro 63-0563, RO 65-7199, Ro 65-7674, SB-399885, SB-214111, SB-258510, SB-271046, SB-357134, SB-699929, SB-271046, SB-742457 and PRX-07034.

[0154] (xxxv) Serotonin (5-HT) reuptake inhibitors, such as alaprocrat, citalopram (CELEXA, CIPRAMIL), escitalopram (LEXAPRO, CIPRALEX), clomipramine (ANAFRANIL), duloxetine (CYMBALTA), femoxetine (MALEXIL), fenfluramine (MALEXIL), norfenfluramine, fluoxetine (PROZAC), fluvoxamine (LUVOX), indalpine, milnacipran (IXEL), paroxetine (PAXIL, SEROXAT), sertraline (ZOLOFT, LUSTRAL), trazodone (DESYREL, MOLIPAXIN), venlafaxine (EFFEXOR), dimerizine (NORMUD, ZELMID), bicifadine, desvenlafaxine (PRISTIQ), prasofensin, and tesofensin.

[0155] (xxxvi) Nutritional factors, such as nerve growth factor (NGF), basic fibroblast growth factor (bFGF), neurotrophin-3 (NT-3), brain-derived neurotrophic factor (BDNF), and glial-derived neurotrophic factor (GDNF), as well as propentophilin, idebenone, and AIT-082 (NEOTROFIN); and similar substances. [Examples]

[0156] The following examples are for illustrative purposes only and do not limit the specification, including the claims, in any way. The compound of formula (I) is also referred to as "Cpd(I)" for short.

[0157] Example 1: Nonclinical evaluation of selective PDE9A inhibitors In recent years, scientific evidence has emerged indicating that PDE9A plays a crucial role in the pathophysiology of heart failure and that its expression is elevated in cardiomyocytes of heart failure patients. Compounds of formula (I) (e.g., compound A) are small molecule inhibitors of PDE9A. The optimal therapeutic agent is one that exhibits high affinity for PDE9A and selectively inhibits PDE9A in a way that sustainably enhances the beneficial effects of cGMP in cardiomyocytes without harmful off-target effects.

[0158] In vitro pharmacology Compounds of formula (I) (e.g., compound A) are inhibitors of cGMP-specific PDE9A enzymes with more than 100-fold selectivity compared to enzymes derived from other phosphodiesterase gene families. They are competitive inhibitors of human recombinant PDE9A with an IC50 of 12 nM (Kleiman et al., J Pharmacol Exp Ther. 2012;341:396-409).

[0159] In vivo pharmacology Compounds of formula (I) (e.g., compound A) were tested in a mouse model of pressure-overload-induced heart failure with aortic stenosis (TAC). Compounds of formula (I) (e.g., compound A) significantly reduced left ventricular mass and improved left ventricular function after aortic stenosis.

[0160] Secondary pharmacodynamics The activity of the compound represented by formula (I) (e.g., compound A) against a wide range of receptors, enzymes, and ion channels was evaluated at a single concentration of 10 μM (3955 ng / mL). With the exception of the melatonin 2 receptor (Ki=3.8 μM) and phosphodiesterase 6 (IC50=6.9 μM), inhibition of binding or enzyme activity of less than 50% was observed for all targets. These values ​​significantly exceed the Cmax of the compound of formula (I) (e.g., compound A) at the highest dose (320 ng / mL) tested in the Phase 2 trial, indicating that this compound is unlikely to exhibit significant cross-reactivity with targets other than PDE9A at clinically meaningful exposure levels.

[0161] Safety pharmacology A series of safety pharmacology studies were conducted on the compound of formula (I) (e.g., compound A). Oral administration was chosen for these studies, as this is the planned route of clinical administration (note that studies using anesthetized guinea pigs were conducted via intravenous injection). Cardiovascular evaluation was performed using beagle dogs equipped with telemetry devices in a crossover study design.

[0162] When the potential effects on the lungs and central nervous system (CNS) of the compound of formula (I) (e.g., compound A) were evaluated in male rats at doses up to 300 mg / kg, only mild effects on the CNS (decreased body temperature and exercise activity) were observed at the high dose of 300 mg / kg, but no biologically significant effects on respiratory parameters (tidal volume, respiratory rate, or minute ventilation) were observed.

[0163] When the cardiovascular effects of the compound of formula (I) (e.g., compound A) were evaluated, it was found that in the hERG assay, a statistically significant reduction in the amplitude of the current in the hERG assay occurred at 10, 30, and 100 μM (corresponding to 3,955, 11,864, and 39,546 ng / mL, respectively). As a result, the calculated maximum inhibitory concentration (IC50) was 55.1 μM (21,790 ng / mL). This corresponds to a margin of approximately 73 times compared to the estimated plasma concentration after a clinical dose of 25 mg twice daily (q12h) administered every 12 hours. In an in vivo cardiovascular screening study using anesthetized guinea pigs, the compound of formula (I) (e.g., compound A) was shown to potentially cause mild cardiovascular effects (e.g., decreased blood pressure and increased heart rate) at plasma concentrations of 2,333 ng / mL or higher (7 times the expected maximum plasma concentration (Cmax) of 322.7 ng / mL in humans at steady state).

[0164] In GLP in vivo cardiovascular evaluations, male beagle dogs were administered 3, 10, or 30 mg / kg of the compound of formula (I) (e.g., compound A). At 3 mg / kg, no biologically significant changes were observed in blood pressure, left ventricular pressure, or electrocardiogram. However, between 1 and 5.5 hours post-administration, significant drug-related increases were observed in heart rate (+23 and +24 beats / min), myocardial contractility (left ventricular maxima +dP / dt, 490 and 626 mmHg / sec), and corrected QT values ​​(QTc; +9 and +13 milliseconds). The shortening of the PR interval (-9 to -10 milliseconds) was considered to be attributable to the increase in heart rate. No significant cardiovascular effects were observed during the subsequent observation period of this study, suggesting that the effects observed at the initial stage are reversible.

[0165] Pharmacokinetics and drug metabolism in animals Analysis method Tandem mass spectrometry (LC-MS / MS) was used to measure drug concentrations in plasma and serum samples during pharmacokinetic and toxicological studies.

[0166] Absorption - Single-dose pharmacokinetics After intravenous administration of 1 mg / kg to male rats, the compound of formula (I) (e.g., compound A) showed moderate plasma clearance (CL) of 37 mL / min / kg (hepatic blood flow rate 70 mL / min / kg). The steady-state volume of distribution (Vss) was 1.9 L / kg, resulting in a short terminal half-life (t1 / 2) of 1.5 hours (Table 1). After a single oral administration (1 mg / kg) to rats and dogs, the oral bioavailability of the compound of formula (I) (e.g., compound A) was high, at 73% and 91% in rats and dogs, respectively (Table 1).

[0167] [Table 1]

[0168] AUCinf = Area under the concentration-time curve from time 0 to infinity; CL = Total drug clearance from plasma; max = maximum observed concentration; h = time; IV = intravenous administration; M = male; F = female; N = number of animals; PO = oral administration; t1 / 2 = terminal phase half-life; Tmax = time until Cmax first appears; Vss = steady-state volume of distribution; F = absolute bioavailability.

[0169] Absorption-repeated dose pharmacokinetics (toxicological kinetics) The binding rate of compounds of formula (I) (e.g., compound A) to plasma proteins is low regardless of species (the release rate is approximately 70-90%). Therefore, the safety margin can be calculated based on the total plasma concentration.

[0170] Pharmacokinetic data were obtained from primary (definitive) toxicity studies conducted in rats and dogs over periods of up to 6 months and 39 weeks, respectively. These studies evaluated sex differences in serum exposure to the compound of formula (I) (e.g., compound A), dose-exposure relationships, and time-dependent changes in systemic exposure.

[0171] The serum pharmacokinetics of the compound of formula (I) (e.g., compound A) were measured in male and female rats after oral administration of 10, 50, and 350 mg / kg / day once daily for 3 months. Since no difference in exposure levels was observed between male and female animals, the mean data from both sexes are summarized in Table 2. Exposure levels on day 1 and day 85 were similar, and therefore, no drug accumulation was observed throughout the administration period.

[0172] Furthermore, the compound of formula (I) (e.g., compound A) was orally administered to male and female rats at doses of 30, 100, and 350 mg / kg / day for 6 months, from postnatal day 28 to day 209 (PND), and the serum pharmacokinetics were measured. No significant sex differences were observed in exposure levels. The mean systemic exposure, assessed by AUC0-24, increased almost proportionally with increasing dose at PND 208 (Table 2).

[0173] The serum pharmacokinetics of the compound of formula (I) (e.g., compound A) were measured after oral administration of 1, 4, and 12 mg / kg / day once daily to male and female dogs for 3 months. No significant sex differences in exposure were observed between male and female animals, exposure levels on day 1 and day 88 were similar, and no significant drug accumulation was observed. Systemic exposure, assessed by Cmax and area under the concentration-time curve (AUC), increased with increasing dose. The mean pharmacokinetic parameters of the compound of formula (I) (e.g., compound A) are summarized in Table 2.

[0174] Furthermore, the serum pharmacokinetics were measured after orally administering the compound of formula (I) (e.g., compound A) to male and female dogs at doses of 1, 4, and 12 mg / kg / day for 39 weeks.

[0175] Qualitative analysis of the data revealed that at doses of 4 and 12 mg / kg / day, systemic exposure (assessed by AUC0-24) was higher in males than in females, although exposure levels varied. As the dose increased, the mean systemic exposure at days 1 and 270 increased. Based on mean AUC values, little to no accumulation was observed between days 1 and 270 (Table 2).

[0176] [Table 2]

[0177] a. Based on molecular weight 395.5. b. Day 208 after birth (rats were administered daily from day 28 to day 208 after birth; total 181 doses). c. Non-continuous sampling, n=3 / sex / time of measurement. AUC0-24 = area under the concentration-time curve from 0 to 24 hours; Cmax = maximum observed concentration; h = time; M = male; F = female; N = number of animals; PO = oral.

[0178] Distribution - Plasma protein binding and distribution into the bloodstream The degree of binding of the compound of formula (I) (e.g., compound A) to plasma proteins was measured by equilibrium dialysis at concentrations of 0.1 and 1.0 μg / mL in mouse, rat, guinea pig, dog, and human plasma. This compound showed low binding to plasma proteins in all species evaluated. The mean free fractions were 0.782, 0.800, 0.684, 0.851, and 0.930 in mouse, rat, guinea pig, dog, and human plasma, respectively. Protein binding did not show concentration dependence in the range of 0.1–1.0 μg / mL.

[0179] Initial studies have shown that the blood distribution of compounds of formula (I) (e.g., compound A) is similar across the species studied, with blood / plasma ratios ranging from 1.01 to 1.14.

[0180] Distribution-tissue permeability The compound of formula (I) (e.g., compound A) is a substrate of the P-glycoprotein efflux transporter. In mice and rats, the mean brain / plasma ratios were 0.88 and 0.59, respectively, indicating that this compound can cross the blood-brain barrier. In dogs, the brain osmosis, as assessed by the cerebrospinal fluid / free plasma ratio, was 0.93, indicating that the compound also penetrates the brain in dogs.

[0181] Metabolism in vitro Preliminary experiments using experimental animals and human-derived in vitro systems have shown that the compound of formula (I) (e.g., compound A) undergoes cytochrome P450-mediated metabolism, being reduced to a dihydric alcohol (M3) via N-dealkylation (M1) and ring-opening of the tetrahydropyran ring. The main isoforms involved in the metabolism are CYP3A4 / 5, with CYP2D6 and CYP2C8 also contributing to a small extent.

[0182] In vivo metabolism Evaluation of circulating metabolites in humans after a single oral administration of 150 mg revealed the presence of M1 and M3, as well as a third metabolite (M2) produced by hydroxylation of the pyrimidine ring. A comparison of exposure to these three metabolites in humans, rats, and dogs showed that while the concentrations of M1 and M3 were high in animals, the concentration of metabolite M2 was low.

[0183] Measurements of M2 in plasma samples from rats administered 300 mg / kg / day orally daily and from humans administered 35 mg twice daily (BID) showed that the average concentration in humans was 74.3 ng / mL, compared to 26.9 ng / mL in rats. Preliminary measurements of this metabolite in the urine of humans and rats showed that rats excreted relatively more M2 than humans (after adjusting for body weight).

[0184] discharge In preliminary elimination studies in rats and dogs after oral administration of 1 mg / kg of the compound of formula (I) (e.g., compound A), 4.5% and 8.5% of the dose were excreted unchanged in the urine of rats and dogs, respectively.

[0185] Pharmacokinetic interactions DDI risk was assessed using a steady-state Cmax of 323 ng / mL (total 817 nM, unbound 760 nM) after oral administration of 25 mg twice daily to humans. This steady-state Cmax was calculated using the observed day 1 Cmax of 248.2 ng / mL, assuming an accumulation ratio of 1.3 to derive the steady-state Cmax.

[0186] Potential for drug-drug interactions via enzymes The potential of a compound of formula (I) (e.g., compound A) to inhibit human cytochrome P450 (CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, or CYP3A) was investigated in a preliminary study using human liver microsomes. Since the compound (30 μM) did not show inhibition of 49% or more for any of the activities, it was not possible to calculate the IC50 value.

[0187] After a 30-minute preliminary incubation with NADPH, no evidence was found that the compound of formula (I) (e.g., compound A) (30 μM) inhibited microsomal metabolism via CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, or CYP3A4 (midazolam or testosterone) in a time-dependent manner.

[0188] Potential drug transporter inhibition The potential of compounds of formula (I) (e.g., compound A) to inhibit various transporters was evaluated using stable-expressing mammalian cell lines. The transporters under test were stably expressed in cultured cell lines, and the ability of cells to transport typical molecules specific to the expressed transporters was evaluated in the presence and absence of the compound.

[0189] The inhibitory ability of the efflux transporter P-gp was evaluated in MDCKII-MDR1 cells by comparing the efflux ratio of digoxin (10 μM) with and without the addition of the compound at a maximum concentration of 300 μM. Compound (I) inhibited P-gp at an IC50 value of 56.9 μM. This indicates that the compound of formula (I) (e.g., compound A) may inhibit P-gp. However, it is unlikely that this compound will inhibit P-gp at clinically relevant concentrations.

[0190] The potential of compounds of formula (I) (e.g., compound A) to inhibit the human hepatic uptake transporters OATP1B1 and OATP1B3 expressed in human embryonic kidney (HEK) 293 cells was evaluated at concentrations up to 300 μM. The compound did not inhibit the transport of rozvastatin (5 μM) via OATP1B1 and OATP1B3, even at the highest concentration tested. This indicates that the compound is unlikely to inhibit OATP1B1 or OATP1B3 at clinically relevant concentrations.

[0191] The inhibitory ability of the compound of formula (I) (e.g., compound A) against OAT1, OAT3, MATE1, MATE2K, and OCT2, which are human kidney transporters expressed in HEK293 cells, was evaluated using [3H]-para-aminohippurate (2 μM), [3H]-estrone-3-sulfate (0.2 μM), and [14C]-metformin (10 μM) as probe substrates. This compound did not inhibit the transport via OAT1 up to the highest concentration tested (150 μM). Concentration-dependent inhibition of transport via MATE1 and OCT2 was observed, and inhibition of 43.7% and 46.1% was recognized, respectively, at the highest concentration tested (150 μM). Concentration-dependent inhibition of transport via OAT3 and MATE2K by the compound of formula (I) (e.g., compound A) was observed, and the estimated IC50 values were 29.5 μM and 5.83 μM, respectively. This indicates that this compound has a low possibility of inhibiting OAT1, MATE1, OCT2, or OAT3 at clinically relevant concentrations, but the compound of formula (I) (e.g., compound A) may inhibit MATE2K at clinically relevant concentrations.

[0192] The pharmacokinetics-pharmacodynamics To evaluate the exposure-response relationship of the combined administration of the compound of formula (I) (e.g., compound A) and hydroxyurea on neutrophil adhesion and neutrophil / platelet aggregation in animals, a TNFα-treated mouse model was used as a preclinical pharmacological model. Based on these pharmacodynamic endpoints, a statistically significant pharmacodynamic effect was recognized in the concentration range of 76 - 157 nM (free), corresponding to the minimum concentration (Cmin) at 1 - 10 mg / kg. Although the pharmacodynamic evaluation items in heart failure are different, these data suggest a correlation between the target binding of PDE9A and the pharmacodynamic endpoints. Therefore, assuming that the efficacy of this compound is equivalent between mice and humans and considering the unbound fraction in humans, a dose that maintains Cmin at approximately 32 - 67 ng / mL (total amount) is expected to show pharmacological activity in humans.

[0193] Toxicity Single-dose toxicity The single-dose toxicity of the compound of formula (I) (e.g., Compound A) was investigated by oral gavage at doses of 750, 1000, or 1500 mg / kg in male rats and 10, 30, 100, 150, 200, or 300 mg / kg in male dogs. In rats, single doses of 1000 mg / kg or higher were not tolerated, and neurological signs such as reduced activity, tremors, partial closure of the eyes, and a hunched posture were observed prior to death at doses of 1000 mg / kg or higher.

[0194] In the repeated-dose study in dogs, single doses of 150 mg / kg or higher were not tolerated based on severe clinical signs including convulsions. At the maximum tolerated dose (MTD) of 100 mg / kg in the dog study, reduced activity and vomiting were observed.

[0195] Repeated-dose toxicity For the single agent of the compound of formula (I) (e.g., Compound A), exploratory and definitive repeated-dose toxicity studies were conducted in rats and dogs for up to 6 months and 39 weeks, respectively.

[0196] In the studies in rats and dogs for up to 6 months and 39 weeks, respectively, oral administration of this compound was well tolerated at doses up to 100 mg / kg per day in rats and 12 mg / kg per day in dogs.

[0197] In a short-term study, male rats were administered a dose of 500 mg / kg / day of compound (I) (e.g., compound A) for 4 days. Good tolerance was observed, with only mild decreases in sodium and chloride levels. After 7 days of administration, death was observed in both male and female rats at doses of 500, 750, and 1000 mg / kg / day. Prior to death, signs of tremor, convulsions, or general weakness were observed. Female rats were more sensitive than male rats to the effects of this compound on the central nervous system, and all female rats at doses of 750 mg / kg / day or higher were subsequently euthanized. Some of these animals did not show any prodromal clinical signs on day 1. Other neurological signs included hyperreactivity and tremor in the 1000 mg / kg / day dose group, and decreased activity, partial eye closure, and salivation in the 500 mg / kg / day and higher dose groups. Transient hunching or recumbency was observed at all doses. Microscopically, lesions (wall degeneration) were observed in the cardiovascular system of males at doses of 500 mg / kg / day or higher, and lesions (myocardial degeneration and inflammation, with occasional hemorrhage) were observed in the myocardium of both males and females at doses of 750 mg / kg / day or higher. The Cmax and AUC24 at a dose of 500 mg / kg / day were 21,400 ng / mL and 309,000 ng·h / mL, respectively.

[0198] In a dose-finding study using young rats (28-42 days old), administration of 350 mg / kg / day reduced the average weight gain in both males and females after the start of administration, resulting in a decrease in average weight only in females throughout the study period. In a 6-month rat study, rats (15 rats of each sex and dose group) were administered 30, 100, and 350 mg / kg / day orally once daily from day 28 to day 209 after birth. Between PND 115 and 208, the 350 mg / kg / day dose group showed deaths related to the test substance (5 males and 1 female) and adverse clinical findings (convulsions, decreased activity, prone posture, and pallor and / or coldness). Histological findings in the 5 male rats that died were considered to be general, mild, and incidental background lesions, and the cause of death was not clear microscopically. The cause of death in the female was attributed to chronic progressive nephropathy with tubular dilation, and macroscopically, multilocular cysts were observed. No deaths related to the test substance were observed in the major 1-month and 3-month studies.

[0199] When the compound of formula (I) (e.g., compound A) was administered to rats daily for one month at doses of 10, 50, and 300 mg / kg / day, good tolerance was observed, and the increased salivary secretion observed with the 300 mg / kg / day dose was not considered an adverse event. In a three-month study, decreased activity and eye squinting were observed in males at oral doses of 10, 50, and 350 mg / kg / day, and in females at 50 and 350 mg / kg / day. In males in the 350 mg / kg / day dose group, mean body weight and mean body weight gain related to the test substance decreased. The effect of the 350 mg / kg / day dose on body weight was judged to be harmful due to the magnitude of the change. After six months of administration, a decrease in mean body weight gain related to the test substance was observed with the 350 mg / kg / day dose, and as a result, body weight at 209 days postnativity decreased only in males (0.90 times that of the control group). Furthermore, in the male and female groups administered 350 mg / kg / day, delays related to the test substance were observed in achieving penile foreskin separation and vaginal patency, respectively. Since no other effects suggestive of endocrine disruption were observed in this study, these delays were not considered harmful.

[0200] In rat studies at 1, 3, and 6 months, effects on hematological and clinical chemical parameters related to the test substance were observed, but these were not considered harmful due to the mild to slight severity of the findings. In the 1-month study, female rats administered 300 mg / kg / day showed increases in reticulocyte count, red blood cell distribution width (RDW), white blood cell count, and lymphocyte count. At 3 months, no effects were observed on reticulocyte or red blood cell (RBC) parameters, but the mean total white blood cell count and lymphocyte count increased slightly to moderately in the 50 and 350 mg / kg / day dose groups, and the number of large unstained cells (LUCs) increased slightly in female rats administered 350 mg / kg / day. In the 6-month rat study, no changes in hematological parameters related to the test substance were observed.

[0201] In the 1-month and 3-month rat primary studies, the high-dose groups showed changes in clinical chemistry parameters including decreased blood glucose, total protein, and albumin, increased blood glucose, elevated blood urea nitrogen (BUN, males), and elevated inorganic phosphorus (P). Other test substance-related effects on clinical chemistry parameters in the 1-month study included decreased chloride and increased cholesterol, alanine aminotransferase (ALT), and calcium. In the 3-month study, other test substance-related changes included a slight increase in alkaline phosphatase (ALP) in female rats receiving 350 mg / kg / day, and a slight decrease in the albumin / globulin (A / G) ratio in female rats receiving 10, 50, and 350 mg / kg / day. One female rat receiving 350 mg / kg / day showed a slight decrease in sodium and chloride levels. In urinalysis, test substance-related changes included slight increases in urine volume and pH, and a decrease in specific gravity in males receiving 50 mg / kg / day, and in both males and females receiving 350 mg / kg / day. After 6 months of administration to rats, test substance-related changes in clinical chemistry parameters of PND210 were observed in the 350 mg / kg / day group. These included a slight increase in alkaline phosphatase in females (1.45 times that of the control group) and an increase in phosphorus (1.20 times that of males and 1.16 times that of females). These findings were not considered harmful due to the small magnitude of the differences. No test substance-related changes were observed in urinalysis parameters.

[0202] In a one-month rat study, liver changes were observed in female rats. An increase in absolute liver weight (18%) was observed in the 300 mg / kg / day dose group, but no microscopic findings associated with this change were observed. In male rats at the same dose, there was a slight increase in the number of hyaline droplets in the renal tubular epithelium, but this finding was specific to male rats and is not a predictor for humans (Hard et al, 1993). No kidney changes were observed in a three-month rat study. At a dose of 350 mg / kg / day, an increase in cytoplasmic vacuolation related to the test substance was observed in the adrenal cortex of male rats, and similar findings, though much milder, were observed in female rats. Since no clinicopathological findings corresponding to this change were observed, this may be an exaggerated physiological response. In male rats, there was a slight increase in the severity of chronic progressive nephropathy, which is considered a naturally occurring finding in rats. In a one-month trial, cardiomyopathy was observed in four male and one female rats administered 300 mg / kg / day, but this finding was also observed in four male rats in the solvent control group, and was the most severe. No cardiomyopathy was observed in a three-month trial. Cardiomyopathy is a common spontaneous finding in rats, and the incidence and severity of this finding in the control and treatment groups were considered to be within expected background levels and not related to the test substance (Lewis, 1992). In a six-month trial, in the 350 mg / kg / day group, the absolute and relative weights of the adrenal glands, heart, liver, and ovaries were higher than in the control group (1.14 to 1.56 times higher than in the control group) in relation to the test substance, and in male rats in the 100 mg / kg / day group, the adrenal gland weight was higher than in the control group (1.18 to 1.22 times higher than in the control group). No correlation was found between increased organ weight and microscopic or clinicopathological findings, and, with the exception of liver weight, no such findings were observed during autopsy at the time of retrieval. For these reasons, increased organ weight was not considered harmful. No microscopic findings related to the test substance were observed during the initial autopsy or autopsies during the retrieval period.

[0203] In a 6-month rat study, the NOAEL (No Observed Adverse Emission Limit) was set at 100 mg / kg / day, based on mortality and adverse events at a dose of 350 mg / kg / day. The mean Cmax and AUC0-24 associated with a 100 mg / kg / day dose were 12,200 ng / mL and 141,000 ng·h / mL, respectively.

[0204] In a 10-day study in dogs, neurological and gastrointestinal effects associated with the study drug were observed, and seizures were seen in one dog receiving 100 mg / kg / day. Prior to the onset of seizures, head shaking, vertical movement, and head flexion were observed. Other signs included backward gait, gait instability, and intermittent tremors. Although the dose was reduced to 75 mg / kg / day, most of the same clinical signs were still observed. Vomiting was observed after doses of 30 mg / kg / day or higher, and post-administration salivation was observed after doses of 75 mg / kg / day or higher. At the maximum tolerated dose (MTD) of 30 mg / kg / day, the total Cmax was 20,600 ng / mL and the total AUC0-24 was 71,400 ng·h / mL.

[0205] In a major one-month study in dogs, oral administration of the compound of formula (I) (e.g., compound A) showed good tolerability at doses of 3 and 10 mg / kg / day for one month, as no findings related to the test substance were observed. However, doses of 25 and 50 mg / kg / day were untolerated due to neurological effects (seizures).

[0206] Initially, a high dose of 50 mg / kg / day was selected, but this dose was not tolerated and was limited to a single dose. One female in the 50 mg / kg / day group experienced a seizure lasting 26 minutes on day 1 without warning and was euthanized on day 2. This animal required two doses of diazepam to respond. In the remaining animals in the 50 mg / kg / day dose group, vomiting, tremors, excessive salivation, and / or decreased activity were observed after administration, and administration was discontinued. After a 3-day recovery period, administration to these animals was resumed at 25 mg / kg / day, replacing one female. One male in the 25 mg / kg / day dose group experienced a seizure and tremor lasting 20 minutes without warning on day 17 and was euthanized. This animal responded after two doses of diazepam. One male and one female in the 25 mg / kg / day dose group showed decreased activity. At doses of 10 mg / kg / day or higher, the incidence of vomiting and watery / unformed stools increased in a dose-dependent manner, and increased saliva secretion was also observed. No effects were observed on other laboratory parameters.

[0207] In major 3-month and 39-week studies in dogs (study numbers 6348-574 and 8297047), the compound of formula (I) (e.g., compound A) was administered daily at doses of 1, 4, and 12 mg / kg / day and was well tolerated. No deaths, adverse findings, or changes in body weight, food intake, ophthalmic findings, electrocardiogram, heart rate, blood pressure, hematology, coagulation, urinalysis, macroscopic or microscopic parameters related to the test substance were observed. An increased incidence of abnormal stools (watery, mucous, unformed) related to the test substance was observed at doses of 4 and / or 12 mg / kg / day, but this was not considered harmful.

[0208] Genotoxicity A series of genotoxicity studies were performed on the compound of formula (I) (e.g., compound A), consisting of a microbial reverse mutagenesis test, an in vitro cytogenetic test (in human lymphocytes), and an in vitro rat micronucleus test. All in vitro tests were performed with or without exogenous metabolic activation, up to concentrations limited by cytotoxicity or insolubility.

[0209] The compound of formula (I) (e.g., Compound A) did not show mutagenicity in bacteria regardless of the presence or absence of metabolic activation (Table 3). However, the compound of formula (I) (e.g., Compound A) induced structural chromosomal aberrations in human lymphocytes in vitro in the presence of metabolic activation, and this finding was reproducible. The compound of formula (I) (e.g., Compound A) did not induce chromosomal damage in the bone marrow in a 1-month pivotal study using rats at doses that produced Cmax and AUC0-24 values significantly exceeding the predicted clinical exposure levels.

[0210] [Table 3]

[0211] a. Conducted as part of a 1-month pivotal study using rats. The mean AUC24 and Cmax values at a dose of 300 mg / kg / day on day 25 were 220,000 ng·h / mL and 17,800 ng / mL, respectively.

[0212] Reproductive and Developmental Toxicity To determine the dosing levels for the major reproductive and developmental toxicity studies, non-major preliminary oral EFD studies using pregnant animals were conducted. These non-major oral dose-finding studies in rats and rabbits are not described in this report but are included in Table 4.

[0213] [Table 4-1]

[0214] <� [Table 4-2]

[0215] [Table 4-3]

[0216] [Table 4-4]

[0217] ALP = Alkaline phosphatase; ALT = Alanine aminotransferase; AUC24 = Area under the concentration-time curve from 0 to 24 hours; BID = Twice a day; BLQ = Below the limit of quantification; BP = Blood pressure; bpm = Beats per minute; BUN = Blood urea nitrogen; Cmax = Calculated maximum value; F = Female; HCT = Hematocrit; HGB = Hemoglobin; HR = Heart rate; LUC = Large unstained cell; LV = Left ventricle; MAPD50 = Monophase activity at 50% repolarization Potential action duration; MAPD90 = duration of monophase action potential at 90% repolarization (sinus = at basal heart rate; pace = during ventricular pacing with a cycle length of 200 milliseconds; MCHC = mean corpuscular hemoglobin concentration; msec = milliseconds; M = male; NA = not applicable; NC = not calculated; ND = not measured; NOAEL = no adverse effect level; P = phosphorus; RBC = red blood cells; q12h = every 12 hours; QTc = corrected QTc interval; TK = toxic pharmacokinetics; WBC = white blood cells). a. AUC24 and Cmax values ​​represent the mean serum concentration (for the entire compound in formula (I), e.g., compound A). In repeated-dose studies, reported values ​​were measured near the end of the study. In single-dose studies, AUC and Cmax values ​​were measured on day 1. If the study included both sexes, the combined values ​​for both sexes were used. b. The exposure margin was calculated based on the total AUC24 of 3042 ng·h / mL and total Cmax of 322.7 ng / mL in humans when a clinical dose of 25 mg was administered twice daily (every 12 hours). Total plasma or serum concentrations were used for all calculations. c. Toxicity pharmacokinetic parameters were not evaluated in this study. The values ​​shown are those measured on day 1 in male rats during a 1-month study using male rats at doses of 50 and 300 mg / kg / day. d. The total dose includes an intravenous loading dose of 0.1 mL / kg / min over 5 minutes, followed by a maintenance infusion dose of 11.67 μL / kg / min over 10 minutes. e. Pharmacokinetic parameters at 3 mg / kg were not evaluated in this study. The values ​​represent male dogs in the 3 mg / kg / day dose group on day 1 of a 1-month study. f. Because sample substitution was suspected, the AUC24 was recalculated using unverified software (Pharsight WinNonLin version 5.2). This value is the recalculated value. The original mean AUC24 was 35,000 ng·h / mL. g. This is not the true Cmax value. It is the mean total plasma concentration 6 hours after administration (the equivalent safety margin is indicated in the Cmax exposure column). One subject (male) died 20 minutes after administration, on day 1. i. Four and a half hours after administration, the seizures recurred despite the administration of diazepam, so the animal was euthanized on day 1. j. In the primary and pharmacodynamic studies, deaths were observed in 2 out of 18 animals in the 500 mg / kg / day dose group, and in 11 out of 18 animals in the 750 and 1000 mg / kg / day dose groups. Female animals that survived in the 750 or 1000 mg / kg / day dose groups were euthanized on day 2 due to adverse symptoms. k. Female dogs were administered 100 mg / kg / day on days 1 and 2, and then euthanized on day 2. For male dogs, the dose was reduced from 100 mg / kg / day to 75 mg / kg / day on day 3. Values ​​on day 10 of remaining male animals after receiving 1.75 mg / kg / day for 8 days. This value represents the concentration 0.26 hours after administration on the second day following an administration of 100 mg / kg / day in female dogs that underwent unplanned euthanasia. n. For each NOAEL, AUC and Cmax are listed for each gender. o. Value on day 1. Five females died between gestational days (GD) 8 and 17. Three of these deaths were determined to be related to the test substance, and two were determined to be due to administration errors. q. All females died or were selectively euthanized by day 8 of gestation (GD). No significant macroscopic findings were observed, and all females were determined to be pregnant. r. This is not true Cmax. Serum samples were collected 2 hours after administration on day 8 of pregnancy. The AUC value is AUC2.

[0218] Embryo-fetal development after oral administration in rats and rabbits In the main EFD study involving pregnant female rats (22 rats / dose), doses of 0, 50, 150, or 350 mg / kg / day were administered from day 6 to day 17 of gestation (GD). Three deaths related to the test substance were observed in the 350 mg / kg / day group. In the 350 mg / kg / day group, two female rats in the embryo / fetal development stage and one female rat in the pharmacokinetic stage were found dead on day 8 of GD. Prior to death, these female rats showed weight loss and decreased food intake, and one female rat had a red substance around her mouth and nose, but no macroscopic findings were observed. Two other deaths were attributed to the administration procedure and were not considered to be related to the compound administration. Adverse effects on body weight and food intake were observed in the 350 mg / kg / day dose group. Throughout the entire administration period, the mean maternal weight gain (0.70 times that of the control group) and mean body weight (0.96 times that of the control group) were significantly lower. These differences were consistent with the decrease in average maternal food intake observed throughout the administration period. The average uterine weight during pregnancy in this group was significantly lower than that of the control group (0.86 times that of the control group). No adverse effects on body weight or food intake were observed in rats administered 50 mg / kg / day or 150 mg / kg / day.

[0219] When maternal toxicity was observed at a dose of 350 mg / kg / day, developmental toxicity occurred, and the mean fetal weight decreased to 0.86 times that of the control group, partly due to a decrease in maternal weight. No effect on fetal weight was observed at doses of 50 mg / kg / day and 150 mg / kg / day, and no effects related to the test substance were observed on fetal survival rate or morphology at any dose. The NOAEL for maternal toxicity was set at 150 mg / kg / day, based on mortality, weight loss, and decreased food intake at 350 mg / kg / day. The NOEL for embryo / fetal development was set at 150 mg / kg / day, based on the decrease in fetal weight at a dose of 350 mg / kg / day. The 150 mg / kg / day dose corresponded to an AUC24 of 183,000 ng·h / mL and a Cmax of 15,400 ng / mL at gestational day 17.

[0220] In the main EFD study involving pregnant rabbits (20 rabbits / dose), no effects related to the test substance were observed in maternal body weight, weight gain, net body weight and weight gain, uterine weight during pregnancy, feed intake, or gross examination at any of the administered doses. No effects related to the test substance were observed in fetal body weight, fetal survival rate, or fetal morphology. The NOAEL for maternal toxicity and embryo / fetal development in this study was 50 mg / kg / day (maximum study dose), which corresponds to a Cmax of 9220 ng / mL and an AUC24 of 53,400 ng·h / mL, respectively.

[0221] Preliminary dose-ranging studies and 6-month rat studies were conducted using young rats.

[0222] immunotoxicity Repeated-dose studies in preclinical settings revealed no evidence of immunosuppressive or inflammatory adverse effects, or adverse effects on lymphoid tissue. These studies, combined with hematological, organ weight, and macroscopic and histopathological evaluations, provided compelling evidence that the compound of formula (I) (e.g., compound A) is not associated with any immunotoxicity concerns.

[0223] Relationship between findings and pharmacokinetics Exposure (defined by Cmax and AUC24) to the compound of formula (I) (e.g., compound A) in rats and dogs increased with increasing dose within the tested dose range. In repeated-dose studies with once-daily administration, no differences were observed between exposure parameters on day 1 and those on subsequent days. In the primary studies in rats and dogs, conducted over periods of up to 6 months and 39 weeks, respectively, no sex differences in exposure were observed at any dose. The serum / plasma concentration thresholds of the compound related to the primary responses, and the calculated exposure margins for these primary responses, can be found in Table 4.

[0224] Target organ toxicity Based on non-clinical studies using compounds of formula (I) (e.g., compound A), the central nervous system and cardiovascular system were identified as potential targets in humans. Other findings included effects on the digestive system in dogs and minor changes in clinical chemistry parameters in rats, but these were not considered harmful.

[0225] Following administration of the compound of formula (I) (e.g., compound A), effects on the central nervous system were observed in single-dose safety pharmacology studies and single-dose and repeated-dose studies in rats and dogs. Adverse neurological signs in the repeated-dose studies were similar to those in the single-dose studies. In neurological safety pharmacology studies, a single high-dose administration of 300 mg / kg to rats resulted in decreased body temperature and horizontal and vertical motor activity.

[0226] In rats, a single dose (1000 mg / kg or more) of the compound of formula (I) (e.g., compound A) resulted in findings such as tremors, decreased activity, partial clotting of the eyes, hunched posture, salivation, recumbency, and coldness to the touch prior to death. In repeated dose studies, tremors, convulsions, or generalized weakness were observed prior to death at doses of 500 mg / kg / day or more. In other animals, generalized weakness, including decreased activity, partial clotting of the eyes, and hunched posture, was also observed.

[0227] In dose-escalation single-dose and repeated-dose studies using the compound of formula (I) (e.g., compound A), adverse central nervous system effects were observed in dogs. In the dose-escalation single-dose study, seizures were observed at 150 mg / kg and 300 mg / kg, and euthanasia was required at 300 mg / kg. Other neurological signs included head shaking, limb rigidity / extension, and inability to stand, preceding seizures at 300 mg / kg; head shaking / tremor, head flexion, and backward gait observed at 200 mg / kg; decreased activity at 100 mg / kg; and generalized tremor at 30 mg / kg. In a 10-day study in dogs, seizures, head shaking / tremor, gait instability, tremor, and excessive salivation were observed as central nervous system side effects associated with administration of 100 mg / kg / day, and the dose was reduced to 75 mg / kg / day after 2 days of administration. In a one-month pivotal study, unplanned euthanasia due to seizures occurred on day 2 with a dose of 50 mg / kg / day, and on day 17 with a reduced dose of 25 mg / kg / day. Other neurological findings at the 50 mg / kg / day dose included tremors, decreased activity, and excessive salivation. In a three-month study using dogs, no effects on the central nervous system were observed at doses up to 12 mg / kg / day.

[0228] Because dogs are a more susceptible species in non-clinical studies, neurological effects in single-dose escalation and repeated-dose studies occurred at serum concentrations more than 51 times the human total mean Cmax of 322.7 ng / mL. The relevant AUC24 values ​​in dogs were more than 28 times the predicted total AUC24 (3042 ng·h / mL) in humans at a clinical dose of 25 mg BID (every 12 hours). These neurological effects, particularly seizures, generally occurred immediately after administration of the compound of formula (I) (e.g., compound A) and were associated with Cmax.

[0229] In single-dose safety pharmacology studies in guinea pigs and dogs, and in repeated-dose studies in rats and dogs, administration of the compound of formula (I) (e.g., compound A) resulted in cardiovascular effects.

[0230] In anesthetized guinea pigs, when the blood concentration of the compound of formula (I) (e.g., compound A) was more than seven times (≥2333 ng / mL) the predicted total mean Cmax of 322.7 ng / mL in humans when a clinical dose of 25 mg was administered every 12 hours, a decrease in blood pressure and a shortening of the monophasic action potential duration were observed, and at 12,588 ng / mL, an increase in heart rate was observed.

[0231] In a 7-day exploratory study of the compound of formula (I) (e.g., compound A) in rats, wall degeneration in the major coronary arteries was observed at doses of 500 mg / kg / day or higher, and ventricular myocardial degeneration accompanied by bleeding and / or inflammation was observed at doses of 750 mg / kg / day or higher. The etiology of the cardiac lesions could not be determined, but local hemodynamic insufficiency is considered a contributing factor consistent with the vasodilatory effect of the treatment. This is expected to be associated with a temporary or sustained decrease in coronary blood flow (Greaves, 2007; Histopathology of preclinical toxicology studies. 3rd ed. New York; Elsevier; 2007:299). At doses of 500 mg / kg or higher, the total AUC24 was 309,000 ng·h / mL, which is approximately 122 times the total AUC24 (3042 ng·h / mL) in humans at a clinical dose of 25 mg BID (every 12 hours).

[0232] In dogs, cardiovascular changes were similar across all studies and consistent with the known pharmacological effects of the compound of formula (I) (e.g., compound A). In safety pharmacology studies, cardiovascular evaluations showed significant findings, including increases in heart rate (HR), myocardial contractility (LV maxima + dP / dt), and QTc, when the compound of formula (I) (e.g., compound A) was administered at doses of 10 mg / kg or higher. In a 39-week GLP study in dogs, no effect of this compound on cardiovascular parameters was observed, and the total Cmax at doses that caused cardiovascular effects provided an exposure margin of more than 9 times the mean total Cmax in humans of 322.7 ng / mL.

[0233] In a primary study in dogs, dose-dependent gastrointestinal disorders, including vomiting and changes in stool, were observed at doses of 4 mg / kg / day or higher in association with the administration of the compound of formula (I) (e.g., compound A). No changes related to body weight or food intake were observed, and no histopathological findings were found. Based on the Cmax and AUC24 at the NOAEL dose of 12 mg / kg / day, the safety margins for exposure at the predicted clinical dose of 25 mg twice daily were 14-fold and 11-fold, respectively.

[0234] In 1, 3, and 6-month studies using rats, effects on hematological and clinical chemical parameters related to the test substance were observed, but these were not considered harmful due to the mild to slight severity of the findings. In the 1-month study, female rats administered 300 mg / kg / day showed increases in reticulocyte count, RDW, white blood cell count, and lymphocyte count. At 3 months, no effects on reticulocytes or red blood cell parameters were observed, but the mean total white blood cell count and lymphocyte count increased slightly to moderately in the 50 and 350 mg / kg / day groups, and female rats administered 350 mg / kg / day showed a slight increase in lymphocyte count. In the 6-month rat study, findings were limited to increases in alkaline phosphatase and phosphorus levels in the high-dose (350 mg / kg / day) group, and no harmful effects were observed.

[0235] No hematological findings were observed in the main studies using 3-month and 39-week-old dogs.

[0236] Example 2: Effects of the compound of formula (I) (e.g., compound A) on humans Data from eight Phase 1 trials, one Phase 1b trial, and one Phase 2 trial are available for the compound of formula (I) (e.g., compound A). The Phase 1 trials were designed to evaluate the safety, tolerability, pharmacokinetics (PK), and pharmacodynamics of single and repeated doses of the compound in healthy volunteers (18–85 years of age). The Phase 1b trial includes data on the safety, pharmacokinetics, and pharmacodynamics of the compound of formula (I) (e.g., compound A) administered with or without concomitant use of hydroxyurea in stable patients with sickle cell disease.

[0237] The primary objective of Study 1 was to evaluate the safety, tolerability, pharmacokinetics (PK), and pharmacokinetics of a compound of formula (I) (e.g., compound A) after single-dose and dose-escalation (oral) administration. The primary objective of Study 2 was to evaluate the safety, tolerability, and pharmacokinetics of repeated administration of a compound of formula (I) (e.g., compound A) in healthy elderly subjects. This was an investigator-blinded, subject-blinded, sponsor-open, placebo-controlled study. At the end of Study 2, a dose of 5 mg was administered every 12 hours (q12h) for 6 days, after which participants moved on to Study 3. The primary objective of Study 3 was to evaluate the safety, tolerability, and pharmacokinetics (PK) of repeated administration of a compound of formula (I) (e.g., compound A) in healthy elderly subjects. In the first three cohorts, the compound of formula (I) (e.g., compound A) was administered for 7 days using a dose-escalation method. Cohorts 1 (5 mg q12h) and 2 (15 mg q12h) each consisted of 8 subjects (randomization ratio of 3:1 between the drug group and placebo group), while Cohort 3 (35 mg every 12 hours) was administered to two consecutive groups of 4 subjects each (randomization ratio of 3:1 between the drug group and placebo group). Cohort 4 (35 mg every 12 hours) consisted of 8 subjects (randomization ratio of 3:1 between the drug group and placebo group) and was administered for 14 days. A total of 32 subjects participated in this study, of whom 24 received treatment with a compound of formula (I) (e.g., compound A).

[0238] Study 4 involved a total of 17 participants, 15 of whom received treatment with the compound of formula (I) (e.g., compound A). The participants consisted of 15 men and 2 women, all of whom were Japanese. In Cohort 1, a total of 9 participants were assigned to the study treatment. Six participants received two doses of the compound of formula (I) (e.g., compound A) at doses of 3 mg, 10 mg, or 25 mg, while all 9 participants received one dose of placebo. In Cohort 2, a total of 8 participants were assigned to the study treatment. Six of them received 25 mg of the compound every 12 hours, and two received placebo every 12 hours.

[0239] The potential pharmacokinetic interactions between the compound of formula (I) (e.g., compound A) and donepezil, and its safety and tolerability when administered in the background of donepezil therapy, were evaluated in two small phase 1 trials (one in healthy volunteers (Trial 5) and the other in patients with Alzheimer's disease (AD) (Trial 6)). In Trial 6, the safety and pharmacokinetics of administration of 25 mg every 12 hours for 7 days were evaluated in these subjects. Five subjects received placebo, and 10 subjects received the compound of formula (I) (e.g., compound A). In Trial 5, the safety and tolerability of combination therapy of the compound with donepezil at steady state, the effect of steady-state donepezil on the steady-state pharmacokinetics of the compound of formula (I) (e.g., compound A), and the effect of steady-state the compound of formula (I) (e.g., compound A) on the steady-state pharmacokinetics of donepezil were evaluated. In Study 7, the effect of food on the pharmacokinetics of a compound of formula (I) (e.g., compound A) was evaluated in 10 healthy adult subjects. Subjects received one 35 mg dose each of freshly dispensed (EP) tablets on an empty stomach and after a meal, with a one-week washout period in between.

[0240] In Phase 2 Study 8, the efficacy and safety of administering 25 mg of compound (I) (e.g., compound A) every 12 hours for 12 weeks were evaluated compared to placebo (1:1 randomized). The primary objective of this protocol was to evaluate the efficacy of compound (I) (e.g., compound A) compared to placebo on performance-based cognitive function assessment measures (Alzheimer's Disease Rating Scale - Cognitive Function [ADAS-cog]) in patients with mild to moderate Alzheimer's disease (AD). Secondary objectives of this study were to evaluate the effects of compound (I) (e.g., compound A) on other clinically relevant indicators, including behavior and overall change assessment by clinicians; to evaluate safety and tolerability compared to placebo; and to evaluate the pharmacokinetics (PK) of compound (I) (e.g., compound A).

[0241] Study 9 aimed to evaluate the safety and tolerability of the compound of formula (I) (e.g., compound A) in adult subjects with stable sickle cell disease (SCD) (including subjects receiving and not receiving hydroxyurea as background therapy), and to obtain exploratory biomarker data. Two doses (25 mg and 5 mg, both administered every 12 hours) were evaluated over 28 days.

[0242] A detailed QT / QTc study (Study 10) was conducted. Participants were randomly assigned to receive a single oral dose of either 25 mg of compound (e.g., compound A), 100 mg of compound (e.g., compound A) (or more than therapeutic dose), placebo, or 400 mg of moxifloxacin (positive control). At therapeutic and ultra-therapeutic doses, compound (e.g., compound A) was not associated with QTc interval prolongation meeting the threshold of clinical concern based on ICH E14 criteria, and therefore met the criteria for a negative QT / QTc test. Compound (e.g., compound A) was generally well-tolerated in this population.

[0243] Based on these Phase I and Phase II data, the compound of formula (I) (e.g., compound A) demonstrated good tolerability. The most frequently reported adverse events in relation to treatment were headache, diarrhea, and nausea, which were generally mild in severity.

[0244] Pharmacokinetics and Metabolism in Humans Single dose escalation study In the single dose escalation study (Study 1), the mean C max The values ​​ranged from 8.4 to 1198 ng / mL, and the area under the plasma concentration-time curve (AUC(0-last)) from 0 to the final measurement time ranged from 54.7 to 10,340 ng·h / mL. The compound of formula (I) (e.g., compound A) was rapidly absorbed, and the plasma concentration reached its maximum value (T) 0.75 to 1.25 hours after administration. max ) reached.

[0245] The oral volume of distribution (Vz / F) of the compound of formula (I) (e.g., compound A) ranged from 128.7 to 713.2 L, suggesting broad distribution to tissues. In the target cohort, concentrations in cerebrospinal fluid (CSF) were measured. The CSF / plasma ratio, based on the area under the concentration-time curve (AUC(0-8)) from 0 to 8 hours, was 0.61. The mean cGMP concentration in CSF increased by 246% from baseline, showing a maximum effect at 6 hours. Pharmacokinetic parameters for Study 1 are summarized in Table 5. The compound of formula (I) (e.g., compound A) exhibited a biphasic distribution profile, and the estimated terminal half-life (t1 / 2) in cohorts where the biphasic profile was well characterized (30 mg, 75 mg, and 150 mg) was approximately 19 to 31 hours. In other cohorts, the biphasic characteristics of this compound were not fully evaluated due to subquantitative samples or insufficient sampling time points. Overall, exposure levels (Cmax and AUC) increased proportionally with dose.

[0246] [Table 5]

[0247] Geometric mean (%CV) of AUCinf, AUClast, Cmax, CL / F, and Vz / F; arithmetic mean (SD) of t1 / 2; median (range) of Tmax; AUCinf = area under the plasma concentration-time curve from 0:00 to infinity; AUClast = area under the plasma concentration-time curve from 0:00 to the last measurement; CL / F = oral clearance; Cmax = maximum plasma concentration; CV = coefficient of variation; N = number of subjects; n = number of subjects contributing to the mean values ​​of AUCinf, t1 / 2, CL / F, and Vz / F; SD = standard deviation; Tmax = time when Cmax was observed; t1 / 2 = terminal elimination half-life; Vz / F = terminal distribution volume.

[0248] Repeated administration of the compound (I) every 12 hours (q12h) for 7 or 14 days resulted in a dose-proportional increase in exposure to the compound (I) (e.g., compound A). Absorption of the compound (I) (e.g., compound A) was rapid, with a median Tmax of 0.50–1.00 hours after administration. The observed accumulation ratio of the compound (I) (e.g., compound A) was approximately 1.2–1.5. Steady state appeared to be achieved on day 3. Approximately 17–27% of the oral dose was excreted unchanged in the urine, suggesting that renal excretion is not the primary elimination route for the compound (I) (e.g., compound A). Pharmacokinetic parameters are summarized in Table 6.

[0249] [Table 6]

[0250] Studies on the effects of diet In Study 7, the effect of food administration on the pharmacokinetics of the compound of formula (I) (e.g., compound A) was investigated. When an EP tablet formulation of the compound of formula (I) (e.g., compound A) was administered orally as a single dose, followed by administration with a high-fat meal, the mean Cmax decreased by approximately 20%. No significant changes were observed in AUC.

[0251] subgroups of the population Single-dose studies (Study 1) and repeated-dose studies (Studies 2 and 3) were conducted in healthy elderly subjects. The effect of age on plasma exposure to the compound of formula (I) (e.g., compound A) was minimal. In Study 4, the pharmacokinetics of the compound were investigated in healthy adult and healthy elderly Japanese subjects. Pharmacokinetic parameters were similar to those observed in the initial human study (Study 1) and repeated-dose studies (Study 3). Exposure increased proportionally with dose, reaching a steady state within 3 days after administration. The dose-normalized Cmax value in Japanese subjects was slightly higher than previously observed.

[0252] Drug-drug interactions Studies investigating potential drug interactions between the compound of formula (I) (e.g., compound A) and donepezil were conducted in healthy subjects (Study 5) and patients with mild to moderate Alzheimer's disease (Study 6). In neither study was any effect of the compound of formula (I) (e.g., compound A) on the pharmacokinetics of donepezil, nor was any effect of donepezil on the pharmacokinetics of the compound of formula (I) (e.g., compound A). A study investigating potential drug interactions with itraconazole is planned.

[0253] Metabolism in humans Preliminary analysis of human plasma samples revealed the presence of three circulating metabolites of compound (I) (e.g., compound A) in both single-dose studies (Study 1) and repeated-dose studies (Study 3).

[0254] QT extension (TQT) trial Study 10 was a single-dose study in healthy subjects to evaluate the effect of a compound of formula (I) (e.g., compound A) on the QT interval. At therapeutic and exceeding therapeutic doses, the compound was not associated with QTc interval prolongation meeting the threshold of clinical concern based on ICH E14 criteria; therefore, this study met the negative criteria for a TQT study. The exceeding therapeutic dose in this study was set to provide a margin of approximately four times the expected mean maximum exposure at steady state when 25 mg was administered every 12 hours. The appropriateness of the study's sensitivity was confirmed through the effect of moxifloxacin, used as a positive control, on the QTc interval.

[0255] Safety and effectiveness The Phase I trial program was designed to evaluate the safety, tolerability, pharmacokinetics (PK), and pharmacodynamics (PD) of the compound of formula (I) (e.g., compound A) in single and repeated doses in healthy young adults (18–55 years) and healthy older adults (65–85 years). One of the Phase II trials (Study 8) was designed to evaluate the efficacy, safety, tolerability, and PK of 12 weeks of treatment with the compound of formula (I) (e.g., compound A) in patients with mild to moderate Alzheimer's disease (AD). Furthermore, a Phase Ib trial was designed to evaluate the safety, pharmacokinetics (PK), and pharmacodynamics (PD) of the compound of formula (I) (e.g., compound A) in patients with stable sickle cell disease, and a thorough QT / QTc trial was conducted, showing that the compound of formula (I) (e.g., compound A) was not associated with clinically significant QTc prolongation at therapeutic and exceeding doses.

[0256] Test 1 Study 1 was a physician-blinded, subject-disclosed, sponsor-disclosed study involving escalating oral doses of compound (I) (e.g., compound A) in healthy young and elderly subjects. In both the young and elderly cohorts, the study was designed as a three-period randomized crossover trial with placebo substitution, where each subject received two consecutive dose levels of compound (I) (e.g., compound A) (single doses of 1–150 mg) followed by placebo, with a one-week interval between doses. In addition, another cohort consisting of healthy young subjects received a single dose of 40 mg or placebo to evaluate the effects of compound (I) (e.g., compound A) on cerebrospinal fluid (CSF) cGMP regulation and central nervous system (CNS) penetration. A total of 27 subjects (23 healthy young adult volunteers and 4 healthy elderly volunteers) received a single oral dose of the compound of formula (I) (e.g., compound A) in the range of 1 to 150 mg. Two subjects in the CSF cohort received a single dose of placebo only. The number of subjects treated at each dose level is shown in Table 7.

[0257] [Table 7]

[0258] No deaths, serious adverse events (SAEs), or discontinuation of administration due to adverse events (AEs) were reported. Except for the CSF cohort, no serious AEs were observed. This included two patients who reported headache (one after 40 mg and one after placebo) and one patient who reported neck pain (after 40 mg). Eight adverse events (AEs) were reported from 18 young patients not included in the CSF cohort, all of which were mild. These included one AE after 3 mg of compound (I) (e.g., compound A), two after 30 mg, four after 150 mg, and one after placebo. No adverse events were reported after 1 mg, 10 mg, or 75 mg doses. The most frequent adverse events were nausea (3 cases: all in the 150 mg dose group) and abdominal distension (2 cases: 1 in the placebo group and 1 in the 3 mg dose group). Both of these adverse events were determined by the principal investigator to be related to the investigational drug.

[0259] In the four elderly subjects, a total of 10 adverse events were reported: 3 after 30 mg administration, 4 after 75 mg administration, 1 after 120 mg administration, and 2 after placebo administration. The most frequent adverse events were limb pain (3 cases: 1 each in the 30 mg and 75 mg groups, and 1 in the placebo group) and arthropod bites (1 case with 2 events: after 30 mg and 75 mg administration), all of which were determined by the principal investigator to be unrelated to the investigational drug. All adverse events in the elderly cohort were mild, with the exception of arthropod bites, which were rated as moderate, and dry skin, which was rated as moderate in one subject after 120 mg administration.

[0260] In the seven young participants comprising the CSF cohort, a total of 26 adverse events were reported: 12 in four participants after 40 mg administration and 14 in two participants after placebo administration. All 12 adverse events reported after 40 mg administration were judged to be related to spinal catheter insertion. Of the 14 adverse events reported after placebo administration in the CSF cohort, only two (ear discomfort and flushing) were judged to be unrelated to spinal catheter insertion. No adverse events were judged to be related to the investigational drug. No clinically significant changes from baseline were observed in clinical laboratory values, vital signs, electrocardiogram (ECG) findings, or cardiac telemetry monitoring.

[0261] Test 2 Study 2 was a repeated-dose, dose-escalation, investigator-blinded, sponsor-open, placebo-controlled study involving healthy elderly subjects and a compound of formula (I) (e.g., compound A). Ten subjects received either a 5 mg dose or placebo every 12 hours (q12h) for 6 days, after which this study was terminated and the participants switched to Study 3 (described below).

[0262] There were no deaths among the subjects during the study period. One subject in the group administered with compound (I) (e.g., compound A) experienced a serious adverse event (SAE) of atrial fibrillation during the study period, but this was determined to be unrelated to the administration of the study drug.

[0263] No subjects discontinued the study or had their dosage reduced due to adverse events (AEs). Of the 10 subjects, 9 (2 in the placebo group and 7 out of 8 in the group treated with compound (I) (e.g., compound A)) experienced one or more adverse events during the study. Adverse events observed in multiple subjects are shown in Table 8.

[0264] [Table 8]

[0265] Treatment-related adverse events included diarrhea, frequent urination, nausea, and abdominal pain, all of which were mild. The majority of adverse events not considered treatment-related by the principal investigator occurred more than 10 days after the start of the study. Among those who reported adverse events, several presented with fever (n=6) and cough (n=6), some of whom also experienced additional adverse events such as nausea, fatigue, sore throat, runny nose, chest congestion, or pneumonia. The principal investigator believed that the series of symptoms experienced by these subjects were caused by a viral illness. No clinically significant abnormalities in laboratory values, vital signs, or electrocardiogram findings were observed during the study period.

[0266] Test 4 The primary objective of Study 4 was to evaluate the safety and tolerability of compound (I) (e.g., compound A) after single and repeated oral administration in healthy young and elderly Japanese subjects, and to clarify the pharmacokinetics (PK) of compound (I) (e.g., compound A) after administration. This study was a three-group crossover study with a three-stage dose-escalation single dose and a one-period repeated-dose study. It was a randomized, investigator-blinded, sponsor-open, placebo-controlled study. A total of 17 subjects participated in Study 4, of whom 15 received treatment with compound (I) (e.g., compound A). All subjects completed the study, and there were no dropouts.

[0267] No deaths, serious adverse events (SAEs), dropouts due to adverse events (AEs), or temporary discontinuation or reduction of treatment due to AEs were reported. A summary of the number of subjects who experienced AEs is shown in Table 9. This table shows both the number of AEs due to all causes and the number of AEs evaluated as treatment-related.

[0268] In Cohort 1, only one subject experienced abnormal laboratory values ​​that met the criteria for potential clinical concern during the study period. This subject received 25 mg of compound (I) (e.g., compound A). In Cohort 2, three subjects who received 25 mg of compound (I) (e.g., compound A) every 12 hours, and one subject who received placebo, all showed abnormal laboratory values ​​that met the criteria for potential clinical concern during the study. In Cohort 2, one subject who received 25 mg of compound (I) (e.g., compound A) every 12 hours experienced a laboratory adverse event: hematuria. This event was mild and was determined by the principal investigator to be related to the investigational drug. This event did not meet the criteria for clinical concern. One subject showed elevated aspartate aminotransferase (AST) and ALT levels, which did not meet the criteria for potential clinical concern, but were temporally related to the administration of the investigational drug. Throughout the study period, changes from baseline in systolic and diastolic blood pressure (BP), pulse rate, and electrocardiogram (ECG) values ​​in both the supine and standing positions were not considered clinically significant.

[0269] [Table 9]

[0270] This includes all data collected since the first dose of the investigational drug. Except for the number of adverse events (AEs), each row counts only once per treatment. AE = Adverse event; q12h = Every 12 hours; SAE = Serious adverse event.

[0271] Test 7 Study 7 was an open-label, randomized, two-period, two-treatment, two-order (AB and BA), crossover, single-dose study involving 10 healthy adult males and females aged 18 to 55 years. This study was designed to clarify the effect of food on the pharmacokinetics (PK) of a compound of formula (I) (e.g., compound A) administered as an immediately-dispensed (EP) tablet formulation. This was the first clinical trial using the EP tablet formulation, which was planned for use in subsequent clinical trials. The 35 mg dose was selected because it was within the range of single doses that were well-tolerated in healthy subjects. The expected exposure was below the individual maximum plasma concentration (Cmax) exposure limit of 600 ng / mL (approximately 1 / 7 of the mean Cmax observed from the NOAEL dose in a one-month canine toxicity study) and was expected to significantly increase cGMP in cerebrospinal fluid (CSF). PK sampling times were selected based on the observed PK profile of the oral solution.

[0272] Table 10 shows an overview of treatment-related adverse events (TEAEs) that occurred during treatment, including both treatment-related adverse events of all causes and treatment-related adverse events.

[0273] [Table 10]

[0274] AE = Adverse event; EP = Dispensing; SAE = Serious adverse event.

[0275] Table 11 shows an overview of the incidence of TEAEs, categorized by systemic organ classification (SOC) and recommended terminology (PT).

[0276] [Table 11]

[0277] MedDRA(v12.0) = Medical Terminology Glossary for Pharmaceutical Regulatory Activities (Version 12.0); EP = Dispensing; n = Number of Subjects; SOC = Systematic Organ Classification; PT = Recommended Terms.

[0278] No clinically significant changes in vital signs or electrocardiograms were observed. In one fasted subject, orthostatic systolic blood pressure increased by up to 30 mmHg from baseline, and in another fasted subject, orthostatic systolic blood pressure decreased by up to 30 mmHg from baseline. In one fasted subject, the QTcF interval increased by up to 30-60 milliseconds from baseline. No subjects showed corrected or uncorrected QT values ​​of up to 500 milliseconds or more.

[0279] In conclusion, a single dose (35 mg) of an EP tablet formulation of the compound of formula (I) (e.g., compound A) was judged to be safe and well-tolerated in healthy subjects after meals and on an empty stomach.

[0280] Test 6 This study was a double-blind, sponsor-open, randomized, placebo-controlled, parallel-group, repeated-dose study of the compound of formula (I) (e.g., compound A) in patients with mild to moderate Alzheimer's disease (AD) who were stable on donepezil treatment. The study evaluated the safety and pharmacokinetics (PK) of the compound of formula (I) (e.g., compound A) administered orally at a dose of 25 mg every 12 hours for 7 days.

[0281] In this study, no deaths, serious adverse events (SAEs), dropouts due to adverse events, or temporary discontinuation or reduction of administration due to adverse events were reported. Ten subjects who received the compound of formula (I) (e.g., compound A) every 12 hours reported 15 adverse events (13 of which were treatment-related), while five subjects who received placebo every 12 hours and donepezil once daily reported no adverse events. Of the 15 reported adverse events, seven were mild and eight were moderate. A summary of the number of subjects experiencing all-cause and treatment-related adverse events is shown in Table 12.

[0282] [Table 12]

[0283] This includes all data collected since the first dose of the investigational drug. Except for the number of adverse events (AEs), each entry in each row is counted only once per treatment. AE = Adverse event; q12h = Every 12 hours; QD = Once a day; SAE = Serious adverse event.

[0284] None of the electrocardiogram or vital sign abnormalities reported in this study met the criteria for reporting adverse events (AEs). None of the reported laboratory value abnormalities met the criteria for reporting adverse events.

[0285] Table 13 shows an overview of TEAE incidence rates by SOC and PT.

[0286] [Table 13]

[0287] Includes all data collected after the first dose of the investigational drug. AE = Adverse Event; MedDRA = Glossary of Drug Regulatory Activity Terms; PT = Recommended Terms; q12h = Every 12 Hours; QD = Once Daily; SOC = Systemic Organ Classification.

[0288] The results of this study showed that multiple doses of compound (I) (e.g., compound A) administered over q12h, combined with donepezil administered over QD, were safe and well-tolerated in generally healthy men and women with mild to moderate Alzheimer's disease (AD), and no deaths, serious adverse events (SAEs), or discontinuation due to adverse events were reported. Over 7 days, the mean blood concentrations of donepezil were similar between subjects who received compound (I) (e.g., compound A) and those who received the corresponding placebo.

[0289] Test 3 Study 3 was a multi-stage dose-escalation (up to 14 days), investigator-blinded, sponsor-disclosed, placebo-controlled, randomized trial involving a compound of formula (I) (e.g., compound A) in healthy elderly subjects. A total of four consecutive subject cohorts received treatment.

[0290] All subjects completed the study. Throughout the study period, changes from baseline in systolic and diastolic blood pressure, pulse rate, and electrocardiogram values ​​in the supine and standing positions were minimal and not clinically significant. Reported TEAEs are shown in Table 14. In one subject who received 15 mg every 12 hours, there was one case of headache that was rated as moderate but unrelated to the study drug; all other AEs were mild. The most frequent AEs were diarrhea and headache, which were generally transient, lasting from a few minutes to a few hours. No deaths, SAEs, or study interruptions were observed.

[0291] Table 14 shows an overview of the number of subjects who experienced adverse events (all causes and treatment-related).

[0292] [Table 14]

[0293] This includes all data collected since the first dose of the investigational drug. Except for the number of adverse events (AEs), each entry in each row is counted only once per treatment. AE = Adverse event; q12h = Every 12 hours; SAE = Serious adverse event.

[0294] Table 15 shows an overview of the incidence rates of TEAEs by SOC and PT in subjects with two or more individuals overall.

[0295] [Table 15]

[0296] In a specific treatment group, if the same subject experienced two or more events within the same priority term event category, only the most severe event was included. All data collected after the first dose of the investigational drug are included. MedDRA = Drug Regulatory Activity Glossary; PT = Priority Term; q12h = Every 12 Hours; SOC = Systemic Organ Classification.

[0297] The most frequent adverse events considered treatment-related were diarrhea and headache. No clinically significant cardiovascular or neurological adverse events were reported.

[0298] All adverse events in this study were considered mild, with the exception of one case of headache, which was of moderate severity and deemed unrelated to the study drug.

[0299] Test 5 This study was a double-blind, sponsor-open, randomized, placebo-controlled, parallel-group, 3-period, repeated-dose study of the compound of formula (I) (e.g., compound A) in healthy subjects receiving concomitant donepezil. The study evaluated the effect of steady-state donepezil on the steady-state pharmacokinetics (PK) of the compound of formula (I) (e.g., compound A), and the effect of the steady-state compound on the steady-state PK of donepezil. In healthy subjects receiving concomitant donepezil, multiple doses of 25 mg of the compound of formula (I) (e.g., compound A) were found to be safe and well-tolerated.

[0300] Five subjects discontinued the study due to adverse events (AEs). Four experienced nausea, headache, and vomiting while receiving 10 mg of donepezil, and one experienced abnormal behavior while receiving 10 mg of donepezil in combination with 25 mg of compound (I) (e.g., compound A). One serious adverse event (acute psychosis) was reported and considered a serious adverse event during the study (reported by one subject after drug washout and evaluated as unrelated to the study drug or donepezil). Prior to the occurrence of the serious adverse event (SAE) of acute psychosis, the subject had reported abnormal behavior, which began with 10 mg of donepezil and continued while receiving 10 mg of donepezil in combination with compound (I) (e.g., compound A). All five adverse events leading to discontinuation were, in the opinion of the principal investigator, attributable to donepezil and all resolved by the end of the reporting period.

[0301] All adverse events reported with the combination of donepezil 10 mg and placebo were isolated occurrences, with the exception of nausea (3 cases) and abdominal discomfort, vomiting, drowsiness, and lethargy (2 cases each). All adverse events reported with the combination of donepezil 10 mg and a compound of formula (I) (e.g., compound A) were isolated occurrences, with the exception of dizziness (4 cases), abnormal dreams (3 cases), and diarrhea and headache (2 cases each).

[0302] Changes from baseline in systolic and diastolic blood pressure, pulse rate, and electrocardiogram values ​​in both supine and standing positions during the study period were not considered clinically significant and did not meet the criteria for reporting adverse events.

[0303] Table 16 summarizes the number of subjects who experienced adverse events (all-cause and treatment-related). The majority of reported adverse events were judged to be of mild severity. Table 17 summarizes the incidence of TEAEs by SOC and PT.

[0304] [Table 16]

[0305] This includes all data collected after the first dose of the investigational drug. Except for adverse events (AEs), each subject is counted only once per treatment in each row. Serious adverse events (SAEs) are based on the investigator's assessment. AE = Adverse event; SAE = Serious adverse event.

[0306] [Table 17-1]

[0307] [Table 17-2]

[0308] In a specific treatment group, if the same subject experiences the same priority term event category more than once, only the most severe occurrence will be counted. All data collected since the first dose of the investigational drug is included. Within each treatment row, a subject is counted only once per treatment. MedDRA = Drug Regulatory Activity Glossary; n = Number of subjects

[0309] Test 8 This study is a multicenter, randomized, double-blind, placebo-controlled, parallel-group Phase II trial evaluating the efficacy and safety of 12 weeks of treatment with the compound of formula (I) (e.g., compound A) compared to placebo. The primary objective of this study was to evaluate the efficacy of the compound of formula (I) (e.g., compound A) compared to placebo on performance-based cognitive function assessment measures in patients with mild to moderate Alzheimer's disease (AD). Secondary objectives of this study were to evaluate the effects of the compound of formula (I) (e.g., compound A) on other clinically relevant indicators, including behavioral and clinician-assessed overall change; to evaluate the safety and tolerability of the compound of formula (I) (e.g., compound A) compared to placebo in patients with mild to moderate AD; and to evaluate the pharmacokinetics (PK) of the compound of formula (I) (e.g., compound A).

[0310] The results of this study did not show that formula (I) (e.g., compound A) was more effective than placebo in cognitive function as measured by the Alzheimer's Disease Assessment Scale-Cog (ADAS-cog), behavior as measured by the Neuropsychiatric Symptom Assessment Scale (NPI), and clinician-measured overall change as measured by the Clinical Global Improvement-I.

[0311] Table 18 shows a summary of TEAEs, all-cause adverse events, and treatment-related adverse events by treatment group. The incidence of TEAEs was slightly higher in the group treated with the compound of formula (I) (e.g., compound A) (all-cause: 63.7%; treatment-related: 34.1%) than in the placebo group (all-cause: 58.0%; treatment-related: 27.0%).

[0312] [Table 18]

[0313] a. After the trial ended and the database was locked, an additional serious adverse event (SAE) was reported to the sponsor. In one subject who received the compound of formula (I) (e.g., compound A), an SAE occurred 87 days after the last dose of the investigational drug, but this event was not considered treatment-related and is not included in this table. b. Subject 10611002 (placebo group) discontinued treatment on day 44 due to an adverse event (AE) and died on day 98. This subject is included in this table as a case of treatment discontinuation due to an AE. Except for the number of adverse events (AEs), subjects were counted only once per treatment in each row. Serious adverse events (SAEs) are based on the assessment of the principal investigator. AE = Adverse event, SAE = Serious adverse event.

[0314] The incidence rates of treatment-related events (≥2% of the population in any treatment group) for TEAEs, all causes, and MedDRA (version 13.0) SOC and PT are shown by treatment in Table 19. In this report, a 2% cutoff value is used to provide a more comprehensive summary of TEAEs.

[0315] When evaluated by SOC, all-cause adverse events belonging to the gastrointestinal disorder class were observed at a higher rate in the group treated with compound (I) (e.g., compound A) (19.8%) than in the placebo group (5.0%). These gastrointestinal disorders included abdominal pain, diarrhea, dry mouth, dyspepsia, gastroesophageal reflux disease, intestinal obstruction, nausea, and upper gastrointestinal bleeding. No other major differences were observed in the incidence of adverse events by SOC among the treatment groups. Adverse events occurring in more than 5% of subjects in either treatment group were diarrhea, nausea, nasopharyngitis, and headache. The majority of TEAEs were judged to be of mild severity by the principal investigator. The incidence of abnormal clinical laboratory values, abnormal vital signs, and abnormal electrocardiograms was similar between the compound (I) (e.g., compound A) group and the placebo group. In the group administered the compound of formula (I) (e.g., compound A), one case each of decreased blood pressure and increased blood pressure was reported, and these were determined to be adverse events by the principal investigator. These adverse events were not reported to be associated with dizziness, vertigo, or syncope.

[0316] In a patient administered the compound of formula (I) (e.g., compound A), one electrocardiogram abnormality was reported as an adverse event. This was a right bundle branch block, which was considered clinically insignificant.

[0317] The majority of TEAEs were judged to be of mild severity by the principal investigator. Serious adverse events that occurred in the group administered with compound (I) (e.g., compound A) included one case of agitation, one case of bowel obstruction, one case of abdominal pain, one case of cerebral hematoma, one case of fall, and one case of hip fracture. Serious adverse events that occurred in the placebo group included one case of cardiac arrest, one case of respiratory arrest, one case of traumatic brain injury, one case of chronic liver failure, one case of cirrhosis, and one case of postoperative wound infection.

[0318] [Table 19]

[0319] In each row, subjects were counted only once per treatment. Coding according to MedDRA (v13.0) was applied. N = number of subjects evaluated for adverse events; n = number of subjects who exhibited adverse events.

[0320] Seven subjects permanently discontinued treatment and the study due to adverse events (AEs): six in the treatment group receiving compound (I) (e.g., compound A) and one in the placebo group. Of the six subjects who received compound (I) (e.g., compound A), three permanently discontinued treatment and the study due to gastrointestinal adverse events (dyspepsia, abdominal pain, and diarrhea) considered to be treatment-related (Table 19).

[0321] One patient in the placebo group discontinued treatment on day 44 due to an adverse event (AE) and died on day 98. This patient is listed as "Death" in Table 22 and as treatment discontinuation due to AE in Table 18.

[0322] [Table 20]

[0323] a. Age at screening. b. Number of days since the start of investigational drug administration. Investigational drug administration start date = Day 1. c. Coding according to the Medical Terminology for Regulatory Activities (MedDRA, v13.0) was applied. d. Serious adverse events (SAEs) based on the assessment of the principal investigator. This table includes subjects who discontinued treatment and the trial due to adverse events (AEs). Subject 10611002 (placebo group) discontinued treatment due to an adverse event (AE) on day 44 and died on day 98. This subject is included in Table 22 as "Death" and in Table 18 as treatment discontinuation due to AE. AE = Adverse event; SAE = Serious adverse event.

[0324] In the group administered with the compound of formula (I) (e.g., compound A), a total of 4 subjects experienced SAEs (no deaths), 5 subjects experienced serious AEs, and 6 subjects discontinued treatment due to one or more AEs. After the end of the study and database locking, additional SAEs not included in Table 18 were reported to the sponsor.

[0325] In one patient administered the compound of formula (I) (e.g., compound A), a serious adverse event (SAE) of gastric cancer was observed 87 days after the last dose of the investigational drug. This event was not considered treatment-related.

[0326] In the placebo group, four subjects experienced serious adverse events (SAEs) (including two deaths), three subjects experienced severe adverse events (AEs), and two subjects discontinued treatment due to one or more AEs. All SAEs are listed in Table 21 for each subject and treatment group. Only one SAE of syncope in the placebo group was considered treatment-related by the principal investigator.

[0327] [Table 21]

[0328] a. Coding according to MedDRA v13.1 was applied. b. Causal relationships were based on the investigator's assessment. SAE = Serious Adverse Event; MedDRA = Medical Terminology for Regulatory Activities.

[0329] Two participants in the placebo group died, while there were no deaths in the group treated with the compound of formula (I) (e.g., compound A). A summary of the two participants who died during the study is shown in Table 22.

[0330] [Table 22]

[0331] a. Age at the onset of symptoms. b. Date of death was calculated as date of death - date of initiation of first effective treatment + 1. c. Coding from the Regulatory Medical Terminology (MedDRA, v13.1) was applied. Subject 10611002 (placebo group) discontinued treatment due to an adverse event on day 44 and died on day 98. This subject is listed as "Death" in this table and as "Discontinuation due to adverse event" in Table 18.

[0332] Test 9 This study was a phase 1b, randomized, double-blind (sponsor-open), placebo-controlled trial to evaluate the safety, tolerability, pharmacokinetics, and pharmacodynamics of administering a compound of formula (I) (e.g., compound A) in combination with and without hydroxyurea in patients with sickle cell disease.

[0333] In total, 30 subjects (12 males and 18 females) were randomly assigned to this study (7 subjects in the treatment group receiving 5 mg of compound (I) (e.g., compound A) twice daily, 16 subjects in the group receiving 25 mg of compound (I) (e.g., compound A) twice daily, and 7 subjects in the placebo group). Of the 30 subjects, 29 were assigned to the study treatment and received the drug, but one subject in the group receiving 25 mg of compound (I) (e.g., compound A) twice daily discontinued the study after randomization but before the start of the study treatment. Of the 29 subjects who received the investigational drug, 28 completed the study, and one subject in the group receiving 25 mg of compound (I) (e.g., compound A) twice daily discontinued the study due to a serious adverse event (SAE).

[0334] All 29 subjects who received either the compound of formula (I) (e.g., compound A) or a placebo were included in the safety analysis. Overall, 21 TEAEs were observed in 7 subjects in the group receiving 5 mg of the compound of formula (I) (e.g., compound A) twice daily, 56 TEAEs occurred in the group receiving 25 mg of the compound of formula (I) (e.g., compound A) twice daily, and 22 TEAEs occurred in the placebo group. The majority of events were mild (15 out of 21 in the group receiving 5 mg of the compound of formula (I) (e.g., compound A) twice daily, 43 out of 56 in the group receiving 25 mg of the compound of formula (I) (e.g., compound A) twice daily, and 20 out of 22 in the placebo group). Serious adverse events (SAEs) were reported in 3 subjects. One patient in the treatment group receiving 25 mg of compound (I) twice daily discontinued the study due to a serious adverse event (SAE). No dose reductions or temporary discontinuations due to adverse events (AEs) were observed in any patient. No deaths were reported. Of the 56 TEAEs in the group receiving 25 mg of compound (I) twice daily, 18 were considered treatment-related by the principal investigator, and of the 22 TEAEs in the placebo group, 5 were considered treatment-related. No TEAEs were considered treatment-related in the group receiving 5 mg of compound (I) twice daily.

[0335] Serious adverse events (SAEs) that occurred during treatment are summarized in Table 23 (All causal relationships). Serious adverse events (SAEs) were reported in three subjects: two subjects in the group administered 5 mg of compound (I) twice daily, and one subject in the group administered 25 mg of compound (I) twice daily. None of these SAEs were determined by the principal investigator to be related to the investigational drug.

[0336] [Table 23]

[0337] Abbreviations: AE = Adverse Event, BID = Twice a Day, MedDRA = Medical Terminology for Regulatory Activities, SOC = Organ Classification. MedDRA version 19.0 coding dictionary applied.

[0338] Non-serious adverse events (AEs) that occurred during treatment and were observed in 5% or more of the subjects are summarized in Table 24 (All causal relationships) and Table 25 (Treatment-related). The systemic organ classification (SOC) with the highest number of subjects experiencing all-cause adverse events during treatment (TEAEs) was nervous system disorders (57.1% in the group administered 5 mg of compound (I) twice daily, 60.0% in the group administered 25 mg of compound (I) twice daily, and 28.6% in the placebo group), and systemic disorders and injection site abnormalities (42.9% in the group administered 5 mg of compound (I) twice daily, 46.7% in the group administered 25 mg of compound (I) twice daily, and 57.1% in the placebo group). The most frequently reported TEAEs were headache (57.1% in the group administered 5 mg of compound (I) twice daily (e.g., compound A), 33.3% in the group administered 25 mg of compound (I) twice daily (e.g., compound A), and 28.6% in the placebo group) and fatigue (28.6% in the group administered 5 mg of compound (I) twice daily (e.g., compound A), 40.0% in the group administered 25 mg of compound (I) twice daily (e.g., compound A), and 28.6% in the placebo group).

[0339] [Table 24-1]

[0340] [Table 24-2]

[0341] Abbreviations: AE = Adverse Event, BID = Twice a Day, MedDRA = Medical Terminology for Regulatory Activities, SOC = Systemic Organ Classification. MedDRA version 19.0 coding dictionary is used.

[0342] [Table 25]

[0343] Abbreviations: AE = Adverse Event, BID = Twice a day, MedDRA = Medical Dictionary for Regulatory Activities, SOC = Systemic Organ Classification. MedDRA version 19.0 coding dictionary is used.

[0344] Abnormal laboratory values ​​were observed in all 29 subjects assigned to the study treatment group. One subject with a history of elevated baseline total bilirubin and AST levels was enrolled in the group receiving compound (I) (e.g., compound A) at a dose of 5 mg BID. On day 28 of the study, the AST level rose to more than three times the upper limit of normal (ULN), and this increase was reported as an adverse event (AE), but the principal investigator determined that it was not related to the study drug.

[0345] In this study, no clinically significant findings were observed in vital signs or electrocardiogram data.

[0346] Test 10 This study was a Phase I, single-dose, randomized, 4-treatment, 4-period crossover, double-blind (open-label for the positive control) sponsor-initiated open-label trial using placebo and a positive control. The plan was to enroll approximately 44 healthy adults in total to ensure about 40 participants complete the study. Participants who dropped out were not replaced.

[0347] Participants were randomly assigned to one of four administration sequences and received a single oral dose of 25 mg of compound (e.g., compound A), 100 mg of compound (e.g., compound A), placebo, and 400 mg of moxifloxacin in a randomized order (Williams Square).

[0348] The primary objective of this study was to evaluate the effect of a single oral dose of the compound of formula (I) (e.g., compound A) on QTc compared to placebo. Table 26 summarizes the statistical analysis comparing 25 mg of the compound of formula (I) (e.g., compound A) and 100 mg of the compound of formula (I) (e.g., compound A) with placebo for baseline-adjusted QTcF at each post-dosing time point. At all time points, 100 mg of the compound of formula (I) (e.g., compound A) had an average effect on QTcF of up to approximately 5 milliseconds compared to placebo. However, the upper limit of the two-sided 90% (equivalent to one-sided 95%) confidence interval at all post-dosing time points was less than 10 milliseconds, with a maximum value of 7.14 milliseconds, thus meeting the negative criteria for QT / QTc trials based on ICH E14 guidance.

[0349] [Table 26]

[0350] Baseline was defined as the mean of three measurements taken at -1 hour, -0.5 hours, and 0 hours prior to administration during each period. The mean of repeated measures was used in the calculation. Baseline was used as a covariate in the model. In the repeated measures model, sequence, period, time, treatment, and the interaction between time and treatment were treated as fixed effects, while subjects within the sequence were treated as random effects. Abbreviations: QTcF = corrected QT interval using the Friedelic formula; vs = comparison.

[0351] A categorical summary of the absolute value of the QTcF interval and the maximum increase from baseline is shown in Table 27. In two subjects (4.8%) administered 100 mg of compound (I) (e.g., compound A) and one subject (2.4%) administered moxifloxacin, an absolute maximum QTcF interval of 450 msec to less than 480 msec was observed. No subjects had a QTcF interval of 480 msec or greater.

[0352] In this study, no subjects experienced a change in QTcF interval from baseline exceeding 30 milliseconds.

[0353] [Table 27]

[0354] Baseline was defined as the average of three measurements taken before administration (-1, -0.5, 0 hours) in each period. Pre-administration on day 1 of each period. Abbreviations: N = number of subjects; n = number of subjects with measurements; QTcF = corrected QT interval using the Friedelic formula.

[0355] Table 28 summarizes the number of subjects who experienced all-cause and treatment-related TEAEs. With the exception of one TEAE, all TEAEs were mild in severity. One subject reported moderate nausea, an AE, during administration of 25 mg of compound (I) (e.g., compound A), resulting in discontinuation of treatment. No deaths, serious adverse events (AEs), or serious adverse events (SAEs) were reported. Two subjects permanently dropped from the study due to TEAEs (mild rhabdomyolysis and moderate nausea). Both of these TEAEs were reported during administration of 25 mg of compound (I) (e.g., compound A), but the principal investigator determined that none were related to the study drug. No temporary discontinuation of treatment due to adverse events was reported.

[0356] [Table 28]

[0357] Data includes data up to 28 days after the final dose of the investigational drug. Except for adverse events (AEs), each subject was counted only once per treatment in each row. Serious adverse events (SAEs) are based on the investigator's assessment. Severity is counted based on the maximum severity or grade of the event. The MedDRA (version 19.0) coding dictionary was applied. Abbreviations: AE = Adverse event; MedDRA = Medical terminology for drug regulatory activities; SAE = Serious adverse event.

[0358] The incidence rates of all-cause and treatment-related TEAEs (by systemic organ class and preferred term) are summarized in Table 29.

[0359] The most frequently reported TEAEs were nausea, headache, and orthostatic dizziness. These were reported more frequently after treatment with moxifloxacin 400 mg (1, 2, and 1 case, respectively) or placebo of the compound of formula (e.g., compound A) (1, 1, and 0 cases, respectively) compared to treatment with moxifloxacin 400 mg (1, 2, and 1 case, respectively) or the compound of formula (e.g., compound A) (1, 1, and 0 cases, respectively).

[0360] With the exception of one case of TEAE, all TEAEs were of mild severity. One subject reported moderate nausea three days after administration of 25 mg of compound (I) (e.g., compound A), and as a result, administration was discontinued.

[0361] [Table 29]

[0362] Includes data up to 28 days after the final dose of the investigational drug. In each row, the subject was counted only once per treatment. The MedDRA (version 19.0) coding dictionary was applied. Abbreviations: AE = Adverse Event; MedDRA = Medical Terminology for Regulatory Activities; SAE = Serious Adverse Event.

[0363] Two subjects discontinued the study due to TEAEs. Both occurred during administration of 25 mg of compound (I) (e.g., compound A), but the principal investigator determined that neither was related to the investigational drug. One subject discontinued the study after receiving 25 mg of compound (I) (e.g., compound A) in period 1 due to mild rhabdomyolysis caused by strenuous exercise. This AE was first observed on day 7 of period 1 (6 days after administration) and resolved after 9 days. Another subject discontinued the study after receiving 100 mg of compound (I) (e.g., compound A) in period 1 and 25 mg of compound (I) (e.g., compound A) in period 2 due to moderate nausea for other or unknown reasons. This adverse event was first observed on day 4 of period 2 (3 days after administration) and resolved after 6 days.

[0364] Overall, abnormal laboratory values ​​were observed in 28 subjects, but no observable trends were found between treatment groups for any of the laboratory parameters. Three subjects showed clinically significant changes in liver function values, but none were considered to be related to the investigational drug by the principal investigator.

[0365] The investigators determined that none of the blood pressure changes were clinically significant. Based on data from these Phase I and Phase II trials involving healthy volunteers, patients with mild to moderate Alzheimer's disease (AD), and patients with stable sickle cell disease, the compound of formula (I) (e.g., compound A) was well-tolerated. The most frequent adverse events (AEs) were headache, diarrhea, and nausea, which were generally mild in severity. Overall, no safety findings were observed that would hinder further development of this compound.

[0366] Example 3: Phase 2 randomized, double-blind, placebo-controlled clinical trial to evaluate the efficacy of a compound of formula (I) (e.g., compound A) in patients with chronic heart failure. As described in Example 2, the compound of formula (I) (e.g., compound A) has shown generally good tolerability in all clinical trials to date. To date, no serious safety concerns have been identified with the compound of formula (I) (e.g., compound A). No seizures or other serious central nervous system (CNS) adverse events were observed in healthy volunteers or patients. No QTc prolongation was observed at therapeutic doses or doses above therapeutic dose. No other cardiovascular adverse events were observed.

[0367] This study is a Phase II, two-part, seamless, randomized, double-blind, placebo-controlled, parallel-group clinical trial. Part A is a dose-exploratory study in patients with CHFrEF. Part B is a proof-of-concept study in patients with CHFrEF or CHFpEF.

[0368] Test design In Part A, eligible subjects with CHFrEF are randomly assigned in a 2:1 ratio to receive either the compound of formula (I) (e.g., compound A) or a placebo. As shown in Figure 1, subjects begin Period 1 with either 10 mg of the compound of formula (I) (e.g., compound A) orally twice daily (BID) or the corresponding placebo. Subjects continue Period 1 for two weeks. On day 15, subjects enter Period 2, receiving either 25 mg of the compound of formula (I) (e.g., compound A) orally twice daily, or the corresponding placebo.

[0369] Participants will continue Phase 2 for 2 weeks. On day 29, participants will enter Phase 3, continuing with oral administration of compound (I) (e.g., compound A) 25 mg twice daily, or the corresponding placebo. Participants will continue Phase 3 for 8 weeks, completing a total of 12 weeks of administration of compound (I) (e.g., compound A) or the corresponding placebo. Safety and tolerability will be continuously evaluated. A follow-up visit will be conducted at week 16. In Part A, the efficacy of compound (I) (e.g., compound A) will be evaluated in the entire population of participants with CHFrEF, and in two subgroups defined by whether or not they used sacubitril / valsartan (Entresto) (yes or no), depending on the number of participants in each strata. Randomization will be stratified based on whether or not Entresto was used (yes or no).

[0370] The study configuration for Part B is shown in Figure 2. Part B will begin enrollment after the data review of Part A. The starting dose for Part B will be determined based on all available data from Part A, including safety, tolerability, target binding, and pharmacokinetics (PK). Part A subjects are not eligible to participate in Part B. The Part B population includes subjects with CHFrEF and CHFpEF (systolic function < 60%) below normal. Subjects will be randomly assigned in a 1:1 ratio to receive either the compound of formula (I) (e.g., compound A) or the corresponding placebo for 12 weeks. Subject follow-up will be conducted in a decentralized manner, using a combination of in-person and remote visits to monitor tolerability and safety. Subjects will be followed up for 28 days after the end of the treatment period to confirm safety. In Part B, the efficacy of the compound of formula (I) (e.g., compound A) will be evaluated in the entire population of patients with heart failure (EF < 60%) and in the following eight pre-designated subgroups: CHFrEF, CHFpEF, Entresto group, Entresto-free group, Entresto-combined CHFrEF group, Entresto-free CHFrEF group, Entresto-combined CHFpEF group, and Entresto-free CHFpEF group. Randomization will be stratified based on Entresto use (use vs. not use) and EF (≤40% vs. >40%).

[0371] In both Part A and Part B of this study, a participant is considered to have completed the study if they complete their visit at week 16. The end date of the study is defined as the last visit date of the last participant participating in the study.

[0372] Dosage selection in this study

[0373] In Part A, subjects randomly assigned to receive the compound of formula (I) (e.g., compound A) will receive the compound or the corresponding placebo orally twice daily for two weeks, followed by ten weeks of oral administration of the compound or the corresponding placebo. In Part B, subjects randomly assigned to receive the compound of formula (I) (e.g., compound A) will receive treatment at one dose level of the compound for twelve weeks. The selection of the dose level will be based on all available data obtained from Part A, including safety, tolerability, target binding, and pharmacokinetics (PK).

[0374] This clinical trial will evaluate the dose of a compound of formula (I) (e.g., compound A) whose safety has been previously established in humans. The pharmacokinetic profile, safety, and tolerability of the compound of formula (I) (e.g., compound A) have been well-documented in humans over 14 days with administration of up to 35 mg every 12 hours and a single dose of 150 mg, and no specific safety concerns have been identified. The planned maximum dose in this clinical trial is 25 mg twice daily, which is expected to result in an average Cmax of approximately 344 ng / mL.

[0375] The minimum percentage of enzyme inhibition required for a compound of formula (I) (e.g., compound A) to be effective in heart failure (HF) is not definitively known, but it is assumed that exposure to PDE-9 above the maximum median inhibitory concentration (IC50) is necessary. Dosage regimens of 10 mg twice daily and 25 mg twice daily of a compound of formula (I) (e.g., compound A) are expected to achieve exposure levels above the IC50 of PDE-9, while remaining well below the 10-fold margin of no adverse effect level (NOAEL), allowing for sufficient consideration of the effect of PDE-9 inhibition above IC50.

[0376] Objective (Part A) major • To evaluate the safety and tolerability of the compound of formula (I) (e.g., compound A) in patients with heart failure with an ejection fraction (EF) of 40% or less (CHFrEF). In patients with CHFrEF, plasma cGMP levels at week 4 will be compared between a compound of formula (I) (e.g., compound A) and a placebo.

[0377] secondary • Evaluate the pharmacokinetics (PK) of the compound of formula (I) (e.g., compound A) in patients with CHFrEF.

[0378] exploratory In patients with CHFrEF, the effects of each compound (I) (e.g., compound A) on N-terminal pro-B natriuretic peptide (NT-pro-BNP) compared to placebo will be evaluated at the dose and time point of each compound (I) (e.g., compound A). • In patients with CHFrEF, evaluate the effect of different dose levels of the compound of formula (I) (e.g., compound A) on BNP compared to placebo. • In patients with CHFrE, evaluate the effects of different dose levels of the compound of formula (I) (e.g., compound A) compared to placebo on plasma and urinary cGMP. • In patients with CHFrEF, evaluate the effect of different doses of the compound of formula (I) (e.g., compound A) on the cGMP / NT-proBNP ratio. • In patients with CHFrEF, evaluate the effect of different doses of the compound of formula (I) (e.g., compound A) on the cGMP / BNP ratio compared to placebo. • In patients with CHFrEF, the placebo-compared effect of compound (I) (e.g., compound A) on the Kansas City Cardiomyopathy Questionnaire-23 (KCCQ-23) score will be evaluated at week 12. • In patients with heart failure (CHF), the effect of compound (I) (e.g., compound A) compared to placebo will be evaluated on the proportion of patients whose KCCQ-23 clinical summary score (KCCQ-23-CS) improved by 5 points or more from baseline at 12 weeks.

[0379] Objective (Part B) major • In CHF patients, determine at week 12 whether the compound of formula (I) (e.g., compound A) reduces NT-proBNP compared to placebo.

[0380] secondary • In patients with heart failure (CHF), evaluate the effect of the compound of formula (I) (e.g., compound A) on BNP compared to placebo at 12 weeks. • In patients with heart failure (CHF), evaluate the effect of the compound of formula (I) (e.g., compound A) on plasma cGMP compared to placebo at 12 weeks. • In patients with heart failure (CHF), evaluate the effect of the compound of formula (I) (e.g., compound A) compared to placebo at 12 weeks on the plasma cGMP / NT-proBNP ratio. • In patients with heart failure (CHF), the comparative effect of the compound of formula (I) (e.g., compound A) with placebo on the plasma cGMP / BNP ratio at week 12 will be evaluated. • In patients with heart failure (CHF), the comparative effect of a compound of formula (I) (e.g., compound A) against placebo on KCCQ-23-CS at week 12 will be evaluated. • In patients with heart failure (CHF), the effect of compound (I) (e.g., compound A) compared to placebo will be evaluated in terms of the proportion of subjects with a KCCQ-23-CS improvement of 5 points or more from baseline at 12 weeks. • To clarify the relationship between the plasma pharmacokinetic (PK) exposure of a compound of formula (I) (e.g., compound A) and changes in NT-proBNP. To evaluate the safety and tolerability of a compound of formula (I) (e.g., compound A) in patients with heart failure (CHF).

[0381] exploratory • In patients with heart failure (CHF) and all eight subgroups, the effect of compound (I) (e.g., compound A) compared to placebo on NT-proBNP will be evaluated in terms of mean values ​​after baseline, values ​​at each time point, and changes over time. • In CHF patients and all eight subgroups, the effect of compound (I) (e.g., compound A) on BNP compared to placebo will be evaluated in terms of mean values ​​after baseline, values ​​at each time point, and changes over time. • In patients with heart failure (CHF) and all eight subgroups, evaluate the effect of compound (I) (e.g., compound A) compared to placebo on plasma cGMP in mean, time-point, and temporal changes from baseline. • In patients with heart failure (CHF) and all eight subgroups, evaluate the effect of compound (I) (e.g., compound A) compared to placebo on the plasma cGMP to NT-pro BNP ratio, in mean values ​​from baseline, at each time point, and over time. • In patients with heart failure (CHF) and all eight subgroups, the effect of compound (I) (e.g., compound A), compared to placebo, on the plasma cGMP / BNP ratio as a mean value after baseline at each time point and over time will be evaluated. In patients with heart failure (CHF) and all eight subgroups, the effects of compound (I) (e.g., compound A), compared to placebo, on the KCCQ-23-CS, KCCQ-23 overall summary score (KCCQ-23-OS), and eight domains, as mean values ​​after baseline at 12 weeks, will be evaluated. • In all eight target subgroups, evaluate the relationship between the plasma concentration of the compound of formula (I) (e.g., compound A) and the change in plasma cGMP.

[0382] Evaluation items Main effectiveness Part A: Changes in plasma cGMP from baseline (-1 day) at week 4. Part B: Changes in NT-Pro BNP from baseline (Day 1) at week 12.

[0383] Major safety features (Part A only): ·AE ·Physical findings • Vital signs • 12-lead electrocardiogram • Clinical tests to confirm safety (biochemical tests, urinalysis, blood tests)

[0384] Secondary benefits (Part B only) • Change in BNP from baseline (day 1) at week 12. • Change in plasma cGMP from baseline (day 1) at week 12. • Change in plasma cGMP to NT-proBNP ratio from baseline (day 1) at week 12. • Change in plasma cGMP / BNP ratio from baseline (day 1) at week 12. • Changes in KCCQ-23-CS from baseline (day 1) at week 12. • The percentage of subjects whose KCCQ-23-CS score improved by 5 points or more from baseline at week 12.

[0385] Exploratory effectiveness Part A: • Changes in NT-Pro BNP from baseline (day 1) to week 2, week 2 to week 4, and week 2 to week 12. Changes in BNP from baseline (day 1) to week 2, week 2 to week 4, and week 2 to week 12. • Changes in cGMP in plasma, spot urine, and 6-hour urine from baseline (-1 day) to week 2, week 2 to week 4, and week 2 to week 12 (the "week 2 to week 12" period does not apply to 6-hour urine cGMP). • Changes in the ratio of cGMP (plasma, spot urine, and 6-hour urine) to NT-proBNP from baseline (-1 day) to week 2, week 2 to week 4, and week 2 to week 12 (the "weeks 2 to 12" period does not apply to 6-hour urine cGMP). • Changes in the ratio of cGMP (plasma, spot urine, and 6-hour urine) to BNP from baseline (-1 day) to week 2, week 2 to week 4, and week 2 to week 12 (the "weeks 2 to 12" period does not apply to cGMP from 6-hour urine). • Changes from baseline in KCCQ-23-CS, KCCQ-23-OS, and eight domains (physical limitations, symptom stability, symptom frequency, symptom burden, total symptom score, quality of life, self-efficacy, and social limitations) at week 12. • The percentage of subjects whose KCCQ-23-CS score improved by 5 points or more from baseline at week 12.

[0386] Part B: • Mean NT-proBNP levels after baseline (at weeks 2, 6, and 12) • Mean BNP levels after baseline (weeks 2, 6, and 12) • Mean plasma cGMP levels after baseline (weeks 2, 6, and 12) • Mean plasma cGMP / NT-proBNP ratio after baseline (weeks 2, 6, and 12) • Mean plasma cGMP / BNP ratio after baseline (weeks 2, 6, and 12) • Mean values ​​of KCCQ-23-OS and the eight domains at 12 weeks after baseline.

[0387] Secondary safety (Part B only) ·AE • Physical examination • Vital signs • 12-lead electrocardiogram • Clinical laboratory tests (biochemistry, urinalysis, hematology)

[0388] [Table 30]

[0389] Abbreviations: ALP = alkaline phosphatase; ALT = alanine aminotransferase; AST = aspartate aminotransferase; aPTT = activated partial thromboplastin time; BUN = blood urea nitrogen; CBC = complete blood count; CO2 = carbon dioxide; GGT = gamma-glutamyl transpeptidase; INR = international normalized ratio; MCH = mean corpuscular hemoglobin; MCHC = mean corpuscular hemoglobin concentration; MCV = mean corpuscular volume; PT = prothrombin time; RBC = red blood cell; WBC = white blood cell. * The estimated glomerular filtration rate (eGFR), calculated using the simplified mediocryodietary diet (MDRD) formula, is based on creatinine levels.

[0390] Pharmacokinetics Part A: The PK endpoint is the following PK parameter of a compound of formula (I) (e.g., compound A): Cmax ·Cmax arrival time (tmax) ·AUC0-last • Area under the plasma concentration-time curve (AUCtau) during the dosing interval. • Apparent full-body clearance (CL / F, excluding day 1) • Accumulation rate for AUCtau[Racc(AUCtau)] and Cmax[Racc(Cmax)]

[0391] Part B: • Measurement of the plasma concentration of a compound of formula (I) (e.g., compound A) and its correlation with changes in NTproBNP. • Correlation between the plasma concentration of a compound of formula (I) (e.g., compound A) and the change in plasma cGMP.

[0392] estimator Tables 31 and 32 describe the construction of estimators for the primary objectives of Part A and the primary and main secondary objectives of Part B, respectively.

[0393] [Table 31]

[0394] Abbreviations: AE = Adverse event; cGMP = Cyclic guanosine monophosphate; CHFrEF = Chronic heart failure with reduced ejection fraction; EF = Ejection fraction; mITT = Modified Intention to Treat; SAE = Serious adverse event

[0395] [Table 32-1]

[0396] [Table 32-2]

[0397] [Table 32-3]

[0398] Abbreviations: BNP = type β natriuretic peptide; cGMP = cyclic guanosine monophosphate; CHF = chronic heart failure; CI = confidence interval; EF = ejection fraction; KCCQ-23-CS = Kansas City Cardiomyopathy Questionnaire-23 Clinical Summary Score; LS = least squares method; mITT = modified intention-based treatment (mITT); MMRM = repeated measures mixed model; NT-proBNP = N-terminal pro-type B natriuretic peptide

[0399] Selection criteria Unless otherwise specified, the subject must meet all of the following criteria at the time of the screening visit: 1. The candidate must understand and comply with all examination procedures, understand the risks associated with the examination, and be able to provide written informed consent. 2. Adults aged 18 or older at the time of screening (regardless of gender). 3. The patient's medical history must include findings supporting a diagnosis of clinical heart failure (HF) syndrome that have persisted for at least six months prior to screening, and must fall within the New York Heart Association (NYHA) functional class II–III. Heart failure (HF) syndrome is defined by the presence of one or more of the following findings: • Dyspnea, paroxysmal nocturnal dyspnea; • Decreased exercise tolerance, prolonged recovery time after exercise; ·fatigue; • Peripheral edema (lower leg, ankle); or • Hospitalization or emergency outpatient visit within the past 6 months due to heart failure requiring intravenous diuretic therapy.

[0400] 4. In the case of Part A: • Efficacy (EF) of 40% or less as determined by echocardiography during screening. • NT-proBNP level of 600 pg / ml or higher at screening. For patients with atrial fibrillation or atrial flutter at screening, the NT-proBNP level of 1000 pg / ml or higher at screening is required.

[0401] For Part B: • Targets with an EF of 40% or less: ○ Efficacy (EF) must be 40% or less on the echocardiogram performed during screening. ○ NT-proBNP levels must be 600 pg / ml or higher at the time of screening. For patients with atrial fibrillation or atrial flutter at the time of screening, NT-proBNP levels must be 1000 pg / mL or higher at the time of screening. • Eligible for EF over 40%: 〇Screening echocardiography must show an ejection fraction (EF) of >40% and left atrial enlargement. ○ NT-proBNP levels must be 300 pg / ml or higher at the time of screening. For patients with atrial fibrillation or atrial flutter at the time of screening, NT-proBNP levels must be 500 pg / mL or higher at the time of screening.

[0402] 5. The patient has been receiving individually optimized heart failure treatment at a stable dose based on standard clinical guidelines for at least four weeks prior to screening, and no additional guideline-based heart failure treatment has been administered within three months of screening. Treatment for heart failure should include beta-blockers, renin-angiotensin-aldosterone system (RAAS) inhibitors (angiotensin-converting enzyme (ACE) inhibitors, angiotensin II receptor blockers (ARBs)), or combination therapy of ARBs and neprilysin inhibitors (sacubitril / valsartan [Entresto®]), mineralocorticoid receptor antagonists (MRAs), sodium-glucose-coupled transporter 2 (SGLT-2) inhibitors, diuretics, or other standard therapies based on clinical guidelines and the patient's tolerance.

[0403] 6. Women of childbearing potential may be eligible for enrollment if they are not pregnant, not breastfeeding, do not plan to become pregnant during the trial period, and are using appropriate contraception as defined in selection criterion #7. A woman is considered of childbearing potential unless she meets at least one of the following two criteria: a. Having undergone a hysterectomy, bilateral salpingectomy, or bilateral oophorectomy before signing the Informed Consent Form (ICF) and having gone through at least one menstrual cycle; or b. Being postmenopausal: For women aged 55 or older, more than one year must have passed since their last menstrual period, or for women under 55, more than one year must have passed since their last menstrual period and their follicle-stimulating hormone (FSH) levels must be within the postmenopausal range.

[0404] 7. Women of potential pregnancy must use double contraception, including a highly effective method, from screening until one month after the last dose. Approved double contraception includes the use of one of the following methods in addition to an intrauterine device or hormonal contraceptive: (1) condoms, or (2) diaphragms or cervical caps. Women maintaining a monogamous relationship with a male partner who has on record undergone a vasectomy are also eligible for approved contraception.

[0405] 8. Men with a female partner who may become pregnant must agree to use highly effective contraception from the screening visit until three months after the last dose of the investigational drug. Highly effective contraception includes recorded vas deferens ligation, abstinence from sexual intercourse, double barrier methods (e.g., the man uses a condom and the woman uses a diaphragm or cervical cap), or a combination of barrier methods and hormonal contraception (e.g., the man uses a condom and the woman uses hormonal contraception or an intrauterine device). Due to the risk of drug secretion into semen, men whose partners are currently pregnant (including men who have undergone vas deferens ligation) must use barrier methods throughout the study and for three months after the last dose of the investigational drug. Men whose partners are confirmed to be postmenopausal are not required to use contraception.

[0406] Exclusion criteria Unless otherwise specified, subjects who meet any of the following criteria at the time of their screening visit will be excluded from this study. 1. It is recorded that the EF was 60% or higher within 6 months of screening. 2. Recent exacerbation of heart failure (HF) requiring hospitalization or administration of intravenous diuretics within 60 days of screening. 3. Symptomatic hypotension or poorly controlled hypertension (multiple measurements of systolic blood pressure > 180 mmHg or diastolic blood pressure > 110 mmHg) are observed at the time of screening or baseline visit. 4. Individuals with a history of epileptic seizures or those considered to be at high risk of epileptic seizures. 5. Electrocardiogram abnormalities that, in the judgment of the principal investigator, may pose a risk to the safety of the subjects or affect the validity of the trial results. 6. Participants who will receive scheduled interventions (such as percutaneous coronary intervention or implantation of medical devices) during their participation in this study. 7. Individuals who have undergone acute coronary syndrome, stroke, transient ischemic attack, cardiac, carotid artery, or other major cardiovascular surgery, or carotid angioplasty within 60 days of screening. 8. Patients clinically suspected of having heart failure secondary to infiltrative cardiomyopathy (e.g., amyloid, sarcoidosis), hypertrophic cardiomyopathy (obstructive or non-obstructive), or severe valvular heart disease, active myocarditis, active pericarditis, or clinically significant congenital heart disease. 9. I have previously received an allogeneic heart transplant, or I am scheduled to receive one. 10. If a mechanical circulatory support device is currently in use or is scheduled to be used.

[0407] 11. If you are receiving long-term treatment with PDE-5 inhibitors (such as Viagra, Cialis, Revatio, Adcirca, etc.). 12. At the time of screening, the estimated glomerular filtration rate (eGFR) measured using the Modification of Diet in Renal Disease was 30 mL / min / 1.73m². 2 If it is less than. 13. Liver dysfunction as defined by alanine aminotransferase / aspartate aminotransferase levels being 3 times or more above the upper limit of normal (ULN) and / or total bilirubin levels being 2 times or more above the ULN at the time of screening. 14. Concomitant use of drugs known as potent inhibitors or inducers of cytochrome P450 3A4 (CYP3A4). 15. Within 30 days of randomization, or within 5 half-lives of the previous administration of the investigational drug (whichever is longer), the participant must have participated in another clinical trial or received the other investigational drug. 16. The principal investigator determines that the subject has a history of alcohol or drug abuse that would impair their ability to comply with the protocol requirements. 17. There is evidence of a clinically significant cardiovascular, endocrine, gastrointestinal, hematological, hepatic, immunological, neurological, oncological, respiratory, psychiatric, or renal disorder, or, in the judgment of the principal investigator, evidence of any other condition that is not explained by a diagnosis of heart failure and which threatens the safety of the subject or affects the validity of the study results. 18. Subjects with a history of malignant tumors. However, subjects who have been disease-free for more than 5 years prior to screening, or subjects who have only basal cell carcinoma or squamous cell carcinoma and whose treatment has been successful are excluded. 19. Pregnant or breastfeeding women. Here, pregnancy refers to a woman's condition from conception to the end of pregnancy, and is defined as a positive result confirmed by a human chorionic gonadotropin (hCG) test. 20. A woman of childbearing potential as defined in selection criterion 6, except if she is using contraception as defined in selection criterion 7.

[0408] Cancellation Criteria Screening Ineligible Screening ineligibility refers to subjects who consented to participate in a clinical trial but were not subsequently enrolled in the trial for failing to meet the inclusion / exclusion criteria or for other reasons. To ensure transparent reporting on screening ineligible subjects in order to meet the publication requirements of the Consolidated Standards of Reporting Trials and to respond to inquiries from regulatory authorities, a minimum amount of screening ineligibility information is required. This minimum information includes demographics, details of screening ineligibility, eligibility criteria, and serious adverse events (SAEs).

[0409] Re-screening

[0410] For subjects who do not meet the criteria for participation in this study, a rescreening may be performed once, provided that the cause of initial screening ineligibility is not found in subsequent screenings. Subjects who undergo rescreening will be assigned a new subject number.

[0411] Temporary or permanent discontinuation of investigational drug In rare cases, it may be necessary for a participant to temporarily or permanently discontinue the administration of the investigational drug. If a participant requires a temporary discontinuation of the investigational drug, the study team must consult with the clinical trial monitor and the sponsor. This study will make every effort to prevent data loss and to ensure complete follow-up of all participants throughout the study period. Principal investigators will be trained to minimize complete dropouts whenever possible. Participants who permanently discontinue the investigational drug do not necessarily have to drop out of the study. These participants will be encouraged to continue their participation in the study and will be asked to attend the remaining study visits as specified in the protocol.

[0412] Cancellation / Withdrawal from the target exam Participants may withdraw from the examination at any time at their own discretion, and may also be withdrawn at any time at the examiner's discretion for safety, behavioral, or administrative reasons. However, every effort should be made to retain participants in the examination. The reasons why a participant did not complete the examination will be recorded.

[0413] If a participant withdraws their consent to future information disclosure, the sponsor may retain and continue to use the data collected before the withdrawal of consent. The participant may also request the destruction of any collected but untested specimens. The principal investigator must record this in the facility's clinical trial record and notify the sponsor or its designated representative.

[0414] Participants withdrawing from the study will be required to complete the final study procedure (visit 9 for Part A, visit 8 for Part B) as outlined in the evaluation schedule (Tables 33 and 34). All ongoing (S)AEs at the time of withdrawal will be followed up for 28 days or until resolved or stabilized.

[0415] If a subject repeatedly misses scheduled medical appointments and the testing facility is unable to contact them, they will be considered untraceable.

[0416] Candidates who drop out of the exam will not be replaced.

[0417] Cancellation of the exam The sponsor reserves the right, at its sole discretion, to close the clinical trial site or discontinue the clinical trial at any time for any reason. The clinical trial site will be closed upon completion of the clinical trial. The clinical trial site will be deemed closed when all necessary documentation and trial materials have been collected and the site closure visit has been conducted. Reasons for discontinuing the clinical trial include, but are not limited to, the following: • Unknown adverse events (i.e., those whose nature, severity, and / or duration have not been previously reported in similar clinical trials of the investigational drug) • Increased frequency and / or severity and / or duration of known, anticipated, or previously reported adverse events (AEs) (this may also apply to adverse events defined as baseline signs and symptoms at the time of enrollment). • Medical or ethical reasons that would affect the continuation of the trial. • Recruitment of the target group is difficult. • Discontinuation of drug development.

[0418] The principal investigator may initiate the closure of a clinical trial site at any time, provided there are reasonable grounds and sufficient notice has been given well in advance of the planned termination date. Reasons for early closure of a clinical trial site by the sponsor or principal investigator include, but are not limited to, the following: If the principal investigator fails to comply with the clinical trial protocol, the requirements of the institutional review board / ethics board or local health authorities, the sponsor's procedures, or the Good Clinical Practice (GCP) guidelines. • If the recruitment of participants by the principal investigator is insufficient.

[0419] Experimental treatment Treatments administered

[0420] The investigational drug in this study is a compound of formula (I) (e.g., compound A), and is provided in the form of orally administered tablets in two tablet strengths (5 mg and 15 mg). The control drug in this study consists of two matching placebo tablets (one placebo tablet that matches the appearance of the 5 mg active ingredient tablet, and one placebo tablet that matches the appearance of the 15 mg active ingredient tablet). The matching placebo tablets are identical in color, size, and shape to the active ingredient tablets.

[0421] In Part A of this study, subjects will receive either 10 mg (two 5 mg tablets) of the compound of formula (I) (e.g., compound A) orally twice daily, or a control placebo for two weeks during Period 1. During Periods 2 and 3, subjects will receive either 25 mg (two 5 mg tablets and one 15 mg tablet) of the compound of formula (I) (e.g., compound A) orally twice daily, or a control placebo for ten weeks. In Part A, on the 5th and 7th visits, subjects will be instructed not to take the investigational drug before blood and urine samples are taken. Subjects will not take the investigational drug on the 9th visit. They will be instructed to take the investigational drug until the day before the 9th visit.

[0422] In Part B of this study, subjects will receive one dose level of the compound of formula (I) (e.g., compound A) (selected based on all available data, including safety, tolerability, target binding, and pharmacokinetics (PK) from Part A) or the corresponding placebo for 12 weeks. In Part B, on the 4th and 6th visits, subjects will be instructed not to take the study drug before pre-administration blood sampling. Subjects will not take the study drug on the 8th visit. Subjects will be instructed to take the study drug until the day before the 8th visit.

[0423] In both Part A and Part B of this study, participants will be instructed to take the investigational drug orally twice daily (BID) at the same time each day, whenever possible.

[0424] Preparation, storage, handling and management The investigational drug will be supplied by the sponsor or its designated party. The packaging of 10 mg of compound (I) (e.g., compound A) and the corresponding placebo will be clearly distinguished from the packaging of 25 mg of compound (I) (e.g., compound A) and the corresponding placebo. However, at each dose level, the compound (I) (e.g., compound A) and the corresponding placebo will be labeled in a double-blind manner.

[0425] The principal investigator or a person designated by the principal investigator must verify that all investigational drugs received were kept under appropriate temperature conditions during transport, and if any deficiencies are found, they must be reported and corrected before use of the investigational drug.

[0426] All investigational drugs must be stored at room temperature in a locked, secure, environmentally controlled, and (manually or automatically) monitored location within the facility, and access must be restricted to the principal investigator and approved facility staff.

[0427] Only subjects enrolled in this study will receive the investigational drug. During the study visit in Part A or B of this study (as summarized in the evaluation schedule (Tables 33 and 34)), the facility pharmacist will be blinded to the treatment allocation and will dispense a sufficient quantity of the compound of formula (I) (e.g., compound A) or its corresponding placebo tablet for outpatient self-administration. Subjects will be instructed to take the investigational drug according to the provided instructions.

[0428] During trial visits in Parts A and B of this study (as summarized in the evaluation schedule (Tables 33 and 34)), participants will be asked to return all used and unused drug containers. The returned investigational drug must not be re-dispensed to participants.

[0429] Treatment allocation method All participants will be centrally randomized using an interactive web response system (IWRS). Prior to the start of the study, each site will be provided with IWRS login information and operating instructions. On day 1 of Part A of the study, eligible participants will be randomly assigned in a 2:1 ratio to either the compound of formula (I) (e.g., compound A) or placebo using the IWRS. On day 1 of Part B of the study, eligible participants will be randomly assigned in a 1:1 ratio to either the compound of formula (I) (e.g., compound A) or placebo using the IWRS. Randomization will be stratified in Part A by the use of Entresto (used vs. not used), and in Part B by the use of Entresto (used vs. not used) and EF (≤40% vs. >40%). In Part B, capping measures will be applied to ensure appropriate sample sizes in each subgroup to evaluate the study objectives. The principal investigator or their designator will access the IWRS to obtain the randomization number assigned to the subject after confirming that the subject meets all inclusion criteria and does not fall under any exclusion criteria.

[0430] No dose changes other than those specified in the protocol's dose escalation are permitted.

[0431] Procedures for blinding and deblinding This study is a double-blind study. On day 1 of Part A, eligible subjects will be randomly assigned in a 2:1 ratio to either the compound of formula (I) (e.g., compound A) or placebo. On day 1 of Part B, eligible subjects will be randomly assigned in a 1:1 ratio to either the compound of formula (I) (e.g., compound A) or placebo. Throughout the study period, both the principal investigator and the subjects will remain blinded to the investigational drug assigned to each subject. The following control measures will be taken to maintain the double-blind status of this study: For each dosage, tablets of the compound of formula (I) (e.g., compound A) and the corresponding placebo tablets are supplied in the same packaging and are identical in color, odor, taste, and appearance. The principal investigator and other staff involved in this study will remain blinded to the treatment allocation codes. • Pharmacists at facilities responsible for dispensing investigational drugs will be blinded to the treatment allocation process.

[0432] If it becomes necessary to decode the subject during the trial, the date, time, and reason will be recorded in the subject's original data. In such circumstances, the subject will be excluded from the trial, administration of compound (I) (e.g., compound A) or the corresponding placebo will be discontinued, and treatment termination procedures will be carried out after documenting any adverse events (AEs) related to the reason for deblinding.

[0433] The IWRS includes a programmed procedure for deblinding. Blinding in a trial may be deblinded if the principal investigator determines that knowing the subject's assignment of the investigational drug is in the subject's best interest. Where possible, the sponsor must be notified before deblinding. However, this does not apply in medical emergencies where knowledge of a specific blinded investigational drug would affect the immediate management of the subject's condition (e.g., an antidote is available) and identification of the investigational drug is necessary. In this case, the sponsor must be notified within 24 hours of deblinding. The date and reason for deblinding must be recorded in the source documents and electronic case report forms (eCRF), where applicable.

[0434] If a quality assurance audit is conducted, auditors may access open-label investigational drug administration records (if any) at the facility to verify that randomization and dispensing were carried out correctly.

[0435] treatment compliance The investigational drug will be administered during visits for Parts A and B of this study. Details are summarized in the evaluation schedule (Tables 33 and 34). Participants will be instructed to take the investigational drug according to the provided instructions.

[0436] During visits in Parts A and B of this study (as summarized in the evaluation schedule (Tables 33 and 34)), participants will be asked to return all used and unused medication containers. Medication management and participant compliance will be evaluated by maintaining appropriate medication dispensing and return records.

[0437] This study is a Phase 2, two-part, seamless, randomized, double-blind, placebo-controlled, parallel-group clinical trial. Part A is a dose-finding study in patients with CHFrEF. Part B is a proof-of-concept study in patients with CHFrEF or CHFpEF.

[0438] [Table 33-1]

[0439] [Table 33-2]

[0440] Abbreviations: AE = Adverse event; aPTT: Activated partial thromboplastin time; BNP = type b natriuretic peptide; cGMP = Cyclic guanosine monophosphate; ECG = Electrocardiogram; ET = End of study; FSH = Follicle-stimulating hormone; hr = Time; INR = International normalized ratio; KCCQ-23 = Kansas City Cardiomyopathy Questionnaire-23; NT-proBNP = N-terminal pro-type B natriuretic peptide; PK = Pharmacokinetics; PT = Prothrombin time a. Unless otherwise specified, all evaluations are performed before administration. b. To complete eligibility assessment within the screening period, all screening tests should ideally be performed 15 days (-15 days) before the start of the trial. In rare cases, if the reporting of test results is delayed due to unforeseen circumstances (e.g., shipping delays, loss of specimens, unresolved inquiries), screening results up to 21 days prior may be accepted. c. Limited to women who may be pregnant. d. FSH measurement will only be performed on women under 55 years of age who have not had a menstrual period for 12 months or more at the time of screening. e. Vital signs, including heart rate, blood pressure (systolic and diastolic), respiratory rate, and body temperature, will be measured at the times specified in the table above, and, if necessary, during unscheduled visits. Vital signs will be recorded while the subject is seated and has rested for at least 5 minutes. All measurements will be taken individually. If abnormalities are found in vital signs, further evaluation will be performed to the extent deemed necessary by the principal investigator. If rescreening is performed, all vital sign parameters will be remeasured as described above. f. The physical examination includes assessment of the heart, lungs, limbs, weight, and height. Height will be measured only at the time of screening. Weight will be measured at the time of screening and at the ninth visit. All other assessments will be performed at all applicable times and, if necessary, at all unscheduled visits. g. A resting 12-lead electrocardiogram (ECG) is recorded at the times specified in the table above and at all applicable unscheduled visits. The resting 12-lead ECG is recorded after the subject has been resting in a supine position for at least 5 minutes. h. Blood samples are collected 12 to 24 hours after the final dose of the investigational drug. i. Clinical safety tests include biochemical tests, urinalysis, and hematological tests. See Table 30 for specific test items. Clinical safety tests are performed at the times specified in the table above and at all applicable unscheduled visits. All clinical safety tests are analyzed in the central laboratory. j. On visits 5 and 7, subjects will be instructed not to take the investigational drug before blood and urine samples are taken. Subjects will not take the investigational drug on visit 9. They will be instructed to take the investigational drug until the day before visit 9. k. For the measurement points at 1 hour, 2 hours, 3 hours, and 6 hours, a time frame of ±15 minutes applies. l.KCCQ-23 is evaluated electronically. m. A facility pharmacist, with treatment allocation blinded, dispenses the investigational drug for self-administration by the patient as an outpatient. The patient is instructed to take the investigational drug according to the provided instructions. n. Participants must return all used and unused medication containers. o. If a subject refuses remote sample collection, or if national / institutional policy does not permit blood collection outside the facility by a third party, the subject may instead be permitted to receive treatment at the facility.

[0441] [Table 34]

[0442] Abbreviations: AE = Adverse event; aPTT = Activated partial thromboplastin time; BNP = Type B natriuretic peptide; cGMP = Cyclic guanosine monophosphate; ECG = Electrocardiogram; EF = Ejection fraction; ET = End of study; FSH = Follicle-stimulating hormone; INR = International normalized ratio; KCCQ-23 = Kansas City Cardiomyopathy Questionnaire-23; NT-proBNP = N-terminal pro-Type B natriuretic peptide; PK = Pharmacokinetics; PT = Prothrombin time a. Unless otherwise specified, all evaluations are performed before administration. b. To complete eligibility assessment within the screening period, all screening tests should ideally be performed 14 days (-14 days) before the start of the trial. In rare cases, if the reporting of test results is delayed due to unforeseen circumstances (e.g., shipping delays, loss of specimens, unresolved inquiries), screening results up to 21 days before the start of the trial will be accepted. c. Limited to women who may be pregnant. d. Vital signs, including heart rate, blood pressure (systolic and diastolic), respiratory rate, and body temperature, will be measured at the times specified in the table above, and, if necessary, during unscheduled visits. Vital signs will be measured while the subject is seated and after at least 5 minutes of rest. All measurements will be performed individually. If abnormalities are found in vital signs, further evaluation will be performed to the extent deemed necessary by the principal investigator. If rescreening is performed, all vital sign parameters will be remeasured as described above. e.FSH is measured only in women under 55 years of age who have not had a menstrual period for 12 months or more at the time of screening. f. The physical examination includes assessment of the heart, lungs, limbs, weight, and height. Height will be measured only at the time of screening. Weight will be measured at the time of screening and at the eighth visit. All other assessments will be performed at all applicable times and, if necessary, at unscheduled visits. g. A resting 12-lead electrocardiogram (ECG) is recorded at the times specified in the table above and, if necessary, during unscheduled visits. The resting 12-lead ECG is recorded after the subject has been resting in a supine position for at least 5 minutes. h. For patients with an EF of 40% or less: EF must be 40% or less as measured by echocardiography at the time of screening. For patients with an EF greater than 40%: EF must be greater than 40% as measured by echocardiography at the time of screening, and left atrial enlargement must also be observed. i.KCCQ-23 is evaluated electronically. j. Clinical tests include biochemical tests, urinalysis, and hematological tests. See Table 30 for specific tests. Clinical tests are performed at the times specified in the table above and, if necessary, during all unscheduled visits. All clinical tests are analyzed in the central laboratory. k. On the 4th and 6th visits, subjects will be instructed not to take the investigational drug before pre-administration blood sampling. On the 8th visit, subjects will not take the investigational drug. Subjects will be instructed to take the investigational drug until the day before the 8th visit. l. A facility pharmacist, with treatment allocation blinded, dispenses the investigational drug to enable self-administration by the patient as an outpatient. The patient is instructed to take the investigational drug according to the provided instructions. m. Participants must return all used and unused medication containers. n. Depending on the wishes of the individual and the facility, the individual may voluntarily visit the facility.

[0443] Combination therapy Any medications or vaccines (including over-the-counter drugs, prescription drugs, vitamins and / or herbal supplements) taken by the subject since the date of ICF signing must be recorded along with the following information: ·Reason for use • Administration dates including the start and end dates of administration • Dosage information including dosage and frequency

[0444] If you have any questions regarding combination therapy or previous treatments, you should contact your medical monitor.

[0445] Except for the prohibited medications listed below, any medication may be administered concomitantly if necessary for the patient's well-being, for the treatment of comorbidities or to address potential adverse events (AEs). At each visit to the hospital for this study, the study staff will confirm the patient's use of concomitant medications and record this information in the eCRF.

[0446] Prohibited combination therapies The following individuals must not take any medications from the time they sign their ICF: PDE-5 inhibitors (e.g., Viagra, Cialis, Revatio, Adcirca) Known potent inhibitors or inducers of CYP3A4

[0447] Test evaluation and procedures The trial procedures and their timing are summarized in the evaluation schedule (Tables 33 and 34). Adherence to the trial design requirements, including those specified in the evaluation schedule (Tables 33 and 34), is essential and mandatory for conducting the trial. All screening evaluations must be completed and reviewed to ensure that all candidate subjects meet all eligibility criteria. The principal investigator shall maintain a screening log to record details of all subjects screened and to verify eligibility, or, where applicable, to record reasons for screening ineligibility.

[0448] All clinical tests in this study will be performed in the central laboratory. The maximum amount of blood collected from each subject during the study period will comply with local regulations. Re-collection of samples or unscheduled sample collection may occur for safety reasons or due to technical problems with the samples.

[0449] Effectiveness evaluation NT-ProBNP is evaluated at the points specified in the evaluation schedule (Tables 33 and 34).

[0450] The KCCQ-23-CS, KCCQ-23-OS, and the KCCQ-23 score, which includes eight domains (physical limitations, symptom stability, symptom frequency, symptom burden, total symptom score, quality of life, self-efficacy, and social limitations), are assessed electronically at the points specified in the assessment schedule (Tables 33 and 34).

[0451] BNP is evaluated at the points specified in the evaluation schedule (Tables 33 and 34).

[0452] Plasma cGMP is evaluated at the time points specified in the evaluation schedule (Tables 33 and 34).

[0453] In Part A of this study only, urinary cGMP (6-hour urine and spot urine) will be measured at the points specified in the "Evaluation Schedule" (Tables 33 and 34).

[0454] Safety and tolerability evaluation The definition, severity, and causal relationship of adverse events (AEs) are detailed below. Adverse events (AEs) are elicited from the subject (or, where appropriate, their caregiver, representative, or legal guardian) by the staff of the clinical trial site using non-leading questions such as, "How are you feeling today?" or "Have you had any health concerns since your last visit?". Subjects are encouraged to voluntarily report any adverse events that occur at any point during the trial period. Adverse events are recorded in the eCRF.

[0455] If a female participant becomes pregnant during the study period, administration of the investigational drug must be immediately discontinued. Any pregnancy events during the study period will be reported and monitored until the end of the study.

[0456] Blood and urine samples will be collected at the times specified in the evaluation schedule (Tables 33 and 34), and, if necessary, during unscheduled visits, and safety tests including hematological, biochemical, coagulation, and urinalysis will be performed. If clinically significant abnormal laboratory values ​​are observed at the discretion of the principal investigator, they should be reported as adverse events.

[0457] Vital signs, including heart rate, blood pressure (systolic and diastolic), respiratory rate, and body temperature, will be measured at the times specified in the "Evaluation Schedule" (Tables 33 and 34), and at all unscheduled visits as needed. Vital signs will be recorded while the subject is seated and has rested for at least 5 minutes. All measurements will be taken individually. If abnormalities are found in vital signs, further evaluation will be performed to the extent deemed necessary by the principal investigator. If rescreening is performed, all vital sign parameters will be remeasured as described above.

[0458] Physical examination includes assessment of the heart, lungs, limbs, weight, and height. Height is measured only at the time of screening. Weight is measured at the time of screening and on day 85 (the 9th visit in Part A and the 8th visit in Part B). All other assessments are performed at the times specified in the "Assessment Schedule" (Tables 33 and 34), and at any unscheduled visits as needed. Any clinically significant changes observed from the initial physical findings at screening to subsequent visits are considered adverse events (AEs).

[0459] A resting 12-lead electrocardiogram is recorded after the subject has rested in a supine position for at least 5 minutes, at the points specified in the "evaluation schedule" (Tables 33 and 34), and at all unscheduled visits as needed.

[0460] An adverse event (AE) is an undesirable medical event that occurs in a human being in connection with the use of a drug, regardless of whether or not it is related to the drug. This includes: • Clinically significant worsening of the existing condition • Recurrence of pre-existing disease • Adverse events resulting from an overdose of the sponsor's investigational drug (whether accidental or intentional) (i.e., administration exceeding the dose prescribed by a healthcare professional for clinical reasons). • Adverse events resulting from misuse of investigational drugs (i.e., use for reasons other than clinical necessity) • Adverse events related to discontinuation of the investigational drug

[0461] The severity of adverse events is assessed using the following categories: • Mild: Usually transient and may require only minimal treatment or intervention. The event generally does not interfere with normal daily life. • Moderate: Usually mitigated by additional specific therapeutic interventions. The event interferes with normal daily life and causes discomfort, but does not pose a risk of serious or permanent harm to the individual. • Severe: Interferes with normal daily life, significantly impacts clinical condition, or may require intensive therapeutic intervention.

[0462] Any adverse and unintended reaction to the investigational drug (IMP; i.e., when there is a reasonable possibility of a causal relationship between the IMP and the AE), regardless of the dose, should be considered a drug adverse event.

[0463] For commercially available medications, any adverse and unintended drug reaction occurring at doses normally used in humans for the prevention, diagnosis, treatment, or regulation of physiological functions is considered an adverse drug event.

[0464] An unexpected adverse drug event is defined as an adverse event whose nature or severity does not match the product information.

[0465] A serious adverse event (SAE) is an adverse event that occurs at any dose and meets one or more of the following criteria: • Those that result in death • It is life-threatening (see below) • Those requiring hospitalization or extension of an existing hospitalization period (see below) • Causes persistent or significant impairment or functional loss (see below) • Causes congenital abnormalities or birth defects. • Those that cause serious medical events (see below)

[0466] A life-threatening adverse event refers to any adverse event that, in itself, poses an immediate risk of death to the subject. A life-threatening event does not include events that, if they had occurred in a more severe form, could have caused death, but did not actually pose an immediate risk of death at the time they occurred.

[0467] Hospitalization or extension of hospitalization is a criterion for considering an adverse event to be serious.

[0468] Disability is defined as a condition in which an individual's ability to perform normal daily living functions is significantly impaired (i.e., when an adverse event results in a significant, persistent, or permanent change, impairment, injury, or impairment to the physical function / structure, physical activity, or quality of life in question).

[0469] Pharmacokinetic analysis Blood samples for concentration analysis of the compound of formula (I) (e.g., compound A) are collected by venipuncture at the time points specified in the evaluation schedule (Tables 33 and 34). The procedures for collection, processing, and shipping of PK blood samples are detailed in the laboratory manual. PK samples are evaluated in the central laboratory.

[0470] The plasma concentration of the compound of formula (I) (e.g., compound A) shall be measured using a validated analytical procedure. Details of the analytical method shall be described in the biological analysis report.

[0471] Sample size and data analysis Determination of sample size Part A includes approximately 60 CHFrEF patients. The sample size was not determined by power calculation, but rather by ensuring a reasonable number of subjects to evaluate the study's objectives.

[0472] Part B will include approximately 600 subjects in total, consisting of approximately 300 CHFrFF patients and approximately 300 CHFpEF patients. Subjects will be randomly assigned in a 1:1 ratio to either the compound of formula (I) (e.g., compound A) group or the control placebo group. Randomization will be stratified based on Entresto use (yes / no) and EF (≤40% / ≥40%). By assigning 300 subjects to each treatment group, regardless of EF or Entresto use, a t-test will have 99% power when used across the entire subject population to detect a 25% reduction in NT-proBNP (i.e., a reduction of 0.288 units in log-transformed BNP).

[0473] This sample size also ensures sufficient power to detect similar decreases in NT-proBNP at a one-sided significance level of 0.025 in the analysis subgroups for CHFrEF and CHFpEF (150 patients in each treatment group, power = 87%). In the analysis subgroups for patients using Entresto (135 patients in each treatment group, power = 84%) and patients not using Entresto (165 patients in each treatment group, power = 90%), there is sufficient power to detect similar decreases in NTproBNP at a one-sided significance level of 0.025.

[0474] There is considerable uncertainty regarding the use of Entrest, particularly in patients with CHFpEF. Due to this uncertainty, enrollment in each stratum will be closed once the planned number of subjects in that stratum has been randomized. For example, once 180 patients with CHFrEF receiving Entrest are randomized, this study will be closed for patients with CHFrEF receiving Entrest, but enrollment will continue for other subjects whose subgroups have not been closed. By implementing upper limit measures, sufficient sample size will be ensured in each subgroup to evaluate the study objectives.

[0475] Target population for analysis This study includes the following populations for analysis: • Intended Treatment (ITT): The ITT population consists of all randomized subjects. Subjects are analyzed based on the randomized treatment, regardless of the treatment actually received, treatment adherence, or treatment duration. The ITT population is used for sensitivity analysis. • Modified ITT (mITT): The mITT population consists of all randomized subjects who received at least one dose of the compound of formula (I) (e.g., compound A) or placebo. Subjects are analyzed according to the randomized treatment, regardless of the actual treatment received, treatment adherence, or duration of treatment. The mITT population is used as the primary efficacy analysis population. • Safety: The safety analysis population consists of all subjects who received at least one dose of the compound of formula (I) (e.g., compound A) or placebo. Subjects are analyzed based on the first treatment they received. The safety analysis population is used as the primary safety analysis population. • Pharmacokinetics: The PK analysis population consists of all subjects who have received at least one dose of the compound of formula (I) (e.g., compound A) and who do not exhibit any protocol deviations that are considered to affect the PK outcome. • Protocol Adherence (PP): The PP population consists of all mITT subjects with no significant protocol deviations. Subjects are analyzed based on their randomized treatment group. The PP population is used for sensitivity analysis.

[0476] Unless otherwise specified, data is summarized using descriptive statistics for continuous endpoints and frequency and proportion for categorical endpoints. The proportion for each category is calculated based on the number of subjects with no missing data (i.e., the total is 100%).

[0477] Unless otherwise specified, baseline is defined as the last complete observation prior to the first dose. In Part B, capping measures are applied to ensure an appropriate sample size for each subgroup to assess the study objectives.

[0478] Efficacy analysis Primary efficacy endpoints Part A: The primary endpoint is the change in plasma cGMP from baseline (-1 day) at week 4. This is analyzed using an analysis of covariance (ANCOVA) model that includes categorical terms for treatment (2 levels) and entrainment (2 levels), as well as continuous terms for EF and baseline plasma cGMP, to compare the compound of formula (I) (e.g., compound A) with placebo in the mITT population. The compound of formula (I) (e.g., compound A) consists of a two-week treatment of 10 mg every 12 hours for two weeks, followed by 25 mg every 12 hours for two weeks, but is analyzed as a single treatment group. The least squares mean and 95% confidence interval (CI) of the change from baseline in each treatment group, as well as the least squares mean treatment difference and its 95% CI, are calculated from the ANCOVA model.

[0479] Part B: The primary endpoint is the change from baseline in NT-proBNP at week 12. NT-proBNP is measured at five time points: screening (day -14 to day -1), day 1 (pre-administration), week 2 (day 15 ± 2 days, pre-administration), week 6 (day 43 ± 3 days, pre-administration), and week 12 (day 85 ± 5 days). Baseline serum NT-proBNP is defined as the measurement on day 1. The change from baseline in serum NT-proBNP over time is analyzed in the mITT population using a mixed repeated measures model (MMRM). This model includes treatment, visit, Entresto use, treatment-visit interaction, treatment-Entresto interaction, visit-Entresto interaction, and baseline EF as covariates, and uses random effects on subjects, intercepts, and slope. The covariance structure is unstructured. Missing data will not be imputed. The least squares mean and associated two-sided 95% confidence intervals for each treatment group, estimated mean differences between treatment groups, and corresponding two-sided 95% confidence intervals obtained from MMRM are presented for all visit times. Treatment superiority is confirmed if the one-sided p-value for the treatment difference at week 12 is less than 0.025. Similar analyses are repeated for each of the eight subgroups within the mITT population.

[0480] Sensitivity analysis: • Using MMRM in the primary analysis of the above primary endpoints, an analysis will be conducted for the PP population. In the ITT population, we will perform the analysis using MMRM, which was used in the primary analysis of the aforementioned primary endpoint, but we will consider different model assumptions using different covariance structures (i.e., AR(1), CS). Using the observed baseline and week 12 NT-pro BNP values, an analysis will be performed using a nonparametric ANCOVA model. In the mITT population, the primary analysis of the above primary endpoints will be performed using MMRM, but missing serum NT-proBNP values ​​will be substituted using multiple imputation and the Ruben rule. • In the PP population, the primary analysis of the above primary endpoints will be performed using MMRM, but missing serum NT-proBNP values ​​will be imputed using multiple imputation and the Ruben rule. The statistical analysis plan (SAP) will contain details of these analyses.

[0481] Secondary efficacy endpoints (item B only) The change in BNP from baseline (day 1) is analyzed using MMRM in the same manner as the primary efficacy endpoint. The difference between treatment groups in the change from baseline at week 12 is presented.

[0482] The change in plasma cGMP from baseline (day 1) is analyzed using MMRM in the same manner as the primary efficacy endpoint. The difference between treatment groups in the change from baseline at week 12 is presented.

[0483] The changes from baseline (day 1) in the ratio of plasma cGMP to NT-proBNP and the ratio of cGMP to BNP will be analyzed using MMRM, similar to the primary efficacy endpoint. The differences between treatment groups in the changes from baseline at week 12 will be presented.

[0484] The KCCQ-23 includes a clinical summary score, a comprehensive summary score, and eight domains: physical limitations, symptom stability, symptom frequency, symptom burden, total symptom score, quality of life, self-efficacy, and social limitations.

[0485] In patients with heart failure (CHF), the proportion of patients whose KCCQ-23-CS score increased by 5 points or more from baseline to week 12 was calculated for each treatment group. The treatment effect at week 12 was estimated by logistic regression analysis using Entresto use, baseline KCCQ-23-CS score, and baseline ejection fraction (EF) as covariates. The odds ratios and their 95% confidence intervals between treatment groups obtained from the logistic regression analysis are presented.

[0486] Furthermore, descriptive statistics on the change in KCCQ-23-CS scores from baseline to week 12 are presented. Additionally, the mean difference in the change from baseline at week 12 is estimated between treatment groups using the ANCOVA model. This analysis is repeated in eight subgroups of interest.

[0487] Safety analysis In the safety dataset, Part A summarizes all safety data for each treatment group and administration period (period 1, period 2, period 3, combined period 2 and period 3, and overall), while Part B summarizes all safety data for each treatment group.

[0488] For Part A, the primary safety analysis reports exposure-adjusted incidence rates (per target year). This is defined as the number of subjects who experienced adverse events (AEs) or serious adverse events (SAEs) corresponding to a specific systemic organ class or priority term during the relevant period, divided by the total time exposed to risk. Total risk time is the sum of exposure time (in years) for each period. Adjusted incidence rates are presented separately for each systemic organ class and priority term for Period 1, and separately for Periods 2 and 3 combined. The Hodges-Lehmann method is used to calculate point estimates and interval estimates of treatment differences within the subjects.

[0489] For both Part A and Part B, the overall incidence of new adverse events (EAEs) (i.e., all events that occurred, worsened, or became serious after the initial administration of the investigational drug) is presented by organ system and priority term, based on the Medical Dictionary of Regulatory Activities (MDRC). Similar descriptions are provided for serious EAEs, severe EAEs, treatment-related EAEs, and EAEs that led to discontinuation of the investigational drug. Adverse events resulting in death are also summarized.

[0490] For both Part A and Part B, descriptive statistics will be performed on the observed values ​​at each visit, as well as on the changes from baseline and relative changes. Box plots will be created for a pre-selected set of vital signs.

[0491] For both Part A and Part B, summarize the results of the quantitative routine 12-lead electrocardiogram (ECG) for each visit, including observed values, changes from baseline, and relative changes. Create box plots for a selection of ECGs. Include interpretations and quantitative values ​​of the 12-lead ECG data.

[0492] For both Part A and Part B, descriptive statistics will be performed on the observed values, changes, and relative changes from baseline at each visit. The number and percentage of subjects with values ​​outside the reference range (outside the clinically significant range for each parameter) will also be presented. Box plots will be created for some pre-selected test parameters.

[0493] Pharmacokinetic analysis In Part A, the PK parameter is calculated using a non-compartmental method with the plasma concentration of the compound of formula (I) (e.g., compound A). In Part B, the PK parameter is not calculated due to limited PK sampling. All plasma concentrations of the compound of formula (I) (e.g., compound A) are summarized for each study part, dose, visit date, and measurement time point. In Part A, the PK parameter is summarized for each dose and visit date.

[0494] Example 4: Phase II, randomized, double-blind, placebo-controlled trial to evaluate the safety and pharmacodynamic effects of a compound of formula (I) (e.g., compound A) in patients with chronic heart failure. The natriuretic peptide (NP) signaling pathway plays a vital role in cardiovascular pathophysiology. Upon stimulation, the second messenger cyclic guanosine monophosphate (cGMP) amplifies NP signaling. The importance of this signaling pathway in heart failure (HF) has been demonstrated by multiple clinical HF trials of sacubitril / valsartan, which showed clinical efficacy. NP signaling is reduced by hydrolysis of cGMP by PDE9. Myocardial biopsies from heart failure patients have shown elevated PDE9 expression in both HFrEF and HFpEF, confirming that PDE9 reduces cGMP signaling generated by NP receptor activation in heart failure. Therefore, PDE9 inhibition may enhance cGMP signaling via PDE9 inhibition and is an important new target in clinical trials for heart failure. Increased cGMP levels after administration of PDE9 inhibitors in heart failure patients are a novel biomarker indicating potential clinical efficacy.

[0495] This study evaluates the safety and tolerability of PDE9 inhibition in 60 patients with reduced ejection fraction heart failure (HFrEF), as well as the placebo-compared effect of a compound of formula (I) (e.g., compound A) on plasma and urinary cGMP at week 4.

[0496] The study included adults aged 18 years or older with a history of heart failure of at least 6 months, classified as NYHA class II or III, and having one or more of the following: dyspnea, paroxysmal nocturnal dyspnea (PND), edema, fatigue, exertional intolerance, or hospitalization or emergency department visit due to heart failure (requiring intravenous diuretic administration), and having a left ventricular ejection fraction (EF) of less than 40% and an NT-proBNP level of 600 pg / ml or higher, or 1000 pg / ml or higher if atrial fibrillation or atrial flutter was present, and having received guideline-based heart failure treatment at a stable dose for at least 4 weeks prior to screening. Eligible subjects were randomly assigned in a 2:1 ratio to receive either the compound of formula (I) (e.g., compound A) or placebo. Participants began oral administration of the compound of formula (I) (e.g., compound A) or the corresponding placebo for two weeks, followed by increased doses for another 10 weeks, completing a total of 12 weeks of administration of the compound of formula (I) (e.g., compound A) or the corresponding placebo.

[0497] Safety was assessed by adverse events, physical examination, vital signs, electrocardiogram, and safety-related clinical laboratory tests. The primary pharmacodynamic endpoint was the change from baseline in plasma cGMP at week 4 between treatment groups. Secondary endpoints included evaluating the pharmacokinetics of the compound of formula (I) (e.g., compound A) in subjects with HFrEF.

[0498] Example 5: Phase II, randomized, double-blind, placebo-controlled clinical trial to evaluate the efficacy of a compound of formula (I) (e.g., compound A) in patients with chronic heart failure - Results of Part A Adverse events that occurred during treatment Compound A showed good tolerability in a population of HFrEF patients undergoing GDMT.

[0499] In this Phase 2 Part A trial involving 60 patients with HFrEF, compound A demonstrated good tolerability (Table 35).

[0500] [Table 35]

[0501] Effects of PDE9 inhibition by compound A on plasma and urinary cGMP As shown in Figure 3, PDE9 inhibition by compound A resulted in a consistent and significant increase in urinary cGMP compared to placebo (Tables 36 and 37).

[0502] [Table 36]

[0503] [Table 37]

[0504] Significant increases in the median percentage change from baseline in plasma cGMP were observed on days 1, 15, and 29 after administration of compound A. The increase in plasma cGMP occurred early after administration of compound A and was consistently observed throughout the study period (Table 38). The increases in cGMP at all measurement time points were statistically significant compared to placebo (Table 39).

[0505] The mean percentage change from baseline in plasma cGMP was measured on days 1, 15, and 24 after administration of compound A (Table 40). As shown in Figure 4, administration of compound A increased cGMP at all measurement points and maintained high levels throughout the first and second phases of the administration period (Table 41).

[0506] [Table 38]

[0507] [Table 39]

[0508] [Table 40]

[0509] [Table 41]

[0510] Effects of Compound A and Entrest on cGMP The median percentage change in plasma cGMP measured on days 1, 15, and 29 after administration of compound A was similar between patients with and without a history of Entrest treatment (Table 42). Since three-quarters of patients were taking Entrest during the study period (73% in the compound A group and 75% in the placebo group), the increase in plasma cGMP with compound A was additive to the effect of Entrest. As shown in Figure 5, the time-adjusted mean percentage change in plasma cGMP from baseline at week 4 was consistently higher with compound A compared to placebo in both patients with and without a history of Entrest treatment, further demonstrating the additive effect of compound A on Entrest.

[0511] [Table 42]

[0512] The baseline was set as day -1. Days 1, 15, and 29 are compared to the same point in time as day -1.

[0513] NT-Pro BNP Levels Figure 6 shows that no difference in NT-proBNP levels was observed in the group analysis of this small trial of 60 patients (Table 43). However, retrospective exploratory analysis suggests a potential therapeutic effect on NT-proBNP in patients with more severe heart failure at baseline.

[0514] [Table 43]

[0515] KCCQ score Figure 7 shows that the mean change from baseline in multiple KCCQ domains improved in compound A compared to placebo, with some domains approaching statistical significance (Table 44). Analysis of NT-proBNP changes in subgroups based on baseline KCCQ values ​​showed a significant treatment-related decrease in NT-proBNP in patients with lower baseline KCCQ values ​​(more severe heart failure) (Table 45). As shown in Figure 8, administration of compound A increased the proportion of patients showing a larger category change compared to placebo in patients whose KCCQ summary score increased by 5, 10, and 20 points.

[0516] [Table 44]

[0517] [Table 45]

[0518] Pharmacokinetics of Compound A Plasma concentrations of compound A were measured on day 1 after a single dose of 10 mg twice daily (BID), at week 2 after a single dose of 25 mg twice daily (BID), and at week 4 after a two-week course of 25 mg twice daily (BID). Plasma concentrations of compound A showed dose-proportional exposure between the 10 mg twice daily and 25 mg twice daily doses, with mild accumulation observed with the 25 mg twice daily dose during the two-week administration period (Table 46). These findings were consistent with compound A exposure levels in other clinical trials.

[0519] [Table 46]

[0520] 1 Day 1, single dose; 2 Week 2, single dose; 3 Week 4, after 2 weeks of administration; N / A (Not applicable)

Claims

1. A method for treating chronic heart failure in a subject requiring treatment, the method comprising administering to the subject a therapeutically effective amount of a compound or a pharmaceutically acceptable salt thereof, wherein the compound is of formula (I): 【Chemistry 1】 [During the ceremony, R is selected from the group consisting of (C １ -C ６ )alkyl, (C ２ -C ６ )alkenyl, (C ２ -C ６ )alkynyl, (C ３ -C ８ )cycloalkyl, heterocycloalkyl, aryl, and heteroaryl, each of which may optionally be substituted with 1 to 3 substituents, and the substituents are independently (C １ -C ４ )alkyl, (C １ -C ４ )alkoxy, halogen, and (C １ -C ４ )haloalkyl, selected from the group consisting of, R １ is hydrogen, (C １ -C ４ ) alkyl, (C ２ -C ４ ) Lukenil, (C ２ -C ４ ) Alkinnil, (C １ -C ４ ) Selected from the group consisting of haloalkyl and cyclopropyl; R ２ is, (C １ -C ６ ) alkyl, (C ２ -C ６ ) Alkenil, (C ２ -C ６ ) Alkinnil, (C １ -C ６ ) Heteroaryl selected from the group consisting of haloalkyl, pyridinyl, pyridadinyl, pyrimidinyl, and pyrazinyl, and ER ５ A heteroaryl is selected from the group consisting of (C １ -C ４ ) alkyl and (C １ -C ４ ) It may be substituted with 1 to 3 substituents independently selected from the haloalkyl group; R ３ is hydrogen, (C １ -C ４ ) alkyl, (C ２ -C ４ ) Alkenil, (C ２ -C ４ ) Alkinnil, (C ３ -C ６ ) cycloalkyl and (C １ -C ４ ) Selected from the group consisting of haloalkyl; E is -CH ２ -ien-CH ２ CH ２ -ien-CH ２ CH ２ CH ２ Selected from the group consisting of -, and -C(O)-; and R ５ is, (C ３ -C ８ ) Selected from the group consisting of cycloalkyl, heterocycloalkyl, aryl, aryloxy, and heteroaryl, each of which may be substituted with 1 to 3 substituents, and such substituents are independently (C １ -C ４ ) alkyl, (C ２ -C ４ ) Alkenil, (C ２ -C ４ ) Alkinnil, (C １ -C ４ ) Hydroxyalkyl, (C １ -C ４ ) Haloalkyl, (C １ -C ４ ) Alkoxy, (C １ -C ４ ) Haloalkoxy, (C ３ -C ８ ) Cycloalkyl, halo, cyano, phenyl, morpholinyl, (C １ -C ４ (Independently selected from the group consisting of alkylamino, pyrazolyl, triazolyl, and imidazolyl) The above method having the structure.

2. A method for treating chronic heart failure in a subject requiring treatment, the method comprising administering a therapeutically effective amount of a compound to the subject, wherein the compound is 6-((3S,4S)-4-methyl-1-(pyrimidine-2-ylmethyl)pyrrolidine-3-yl)-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one or a pharmaceutically acceptable salt thereof.

3. The method according to claim 1 or 2, comprising administering 5 mg to 50 mg of the compound to a subject.

4. The method according to claim 1 or 2, comprising administering 5 mg to 25 mg of the compound to a subject.

5. The method according to claim 1 or 2, comprising administering 25 mg of the compound to a subject.

6. The method according to claim 1 or 2, comprising administering 50 mg of the compound to a subject.

7. The method according to any one of claims 1 to 6, comprising administering the compound twice a day.

8. The method according to any one of claims 1 to 7, comprising administering the compound every 12 hours.

9. The method according to any one of claims 1 to 8, comprising administering the compound for at least two weeks.

10. The method according to any one of claims 1 to 9, comprising administering the compound for at least 10 weeks.

11. The method according to any one of claims 1 to 9, comprising administering the compound over the remainder of the subject's life.

12. The method according to any one of claims 1 to 11, comprising orally administering a compound of formula (I).

13. The method according to any one of claims 1 to 12, wherein the chronic heart failure is chronic heart failure accompanied by a reduced ejection fraction.

14. The method according to any one of claims 1 to 12, wherein the chronic heart failure is chronic heart failure with preserved ejection fraction.

15. A method according to any one of claims 1 to 12, wherein the chronic heart failure is chronic heart failure with a moderate ejection fraction.

16. The method according to any one of claims 1 to 12, wherein the chronic heart failure is chronic heart failure with an extremely high ejection fraction.

17. The method according to any one of claims 1 to 16, wherein the target plasma or serum NT-proBNP level is at least 600 pg / ml before administration.

18. The method according to any one of claims 1 to 17, wherein the method reduces the N-terminal pro B-type natriuretic peptide (NT-pro BNP) level by approximately 30% when measured in the plasma or serum of the subject.

19. The method according to any one of claims 1 to 18, wherein the method reduces the level of type b natriuretic peptide (BNP) by approximately 30% when measured in the plasma or serum of the subject.

20. The method according to any one of claims 1 to 18, wherein the method increases the cGMP level by approximately 30% when measured in the target plasma or serum.

21. The method according to any one of claims 1 to 20, wherein the method increases the cGMP level by approximately 30% when measured in the urine of the subject.

22. The method according to any one of claims 1 to 21, wherein the method increases the ratio of plasma cGMP to plasma NT-proBNP levels by about 30%.

23. The method according to any one of claims 1 to 22, wherein the method increases the ratio of plasma cGMP to plasma BNP levels by about 30%.

24. The method according to any one of claims 1 to 23, wherein the method increases the Kansas City Cardiomyopathy Questionnaire-23 (KCCQ-23) score by at least 5 points.

25. A method for treating chronic heart failure in a subject requiring treatment, the method comprising orally administering to the subject 25 mg of 6-((3S,4S)-4-methyl-1-(pyrimidine-2-ylmethyl)pyrrolidine-3-yl)-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one or a pharmaceutically acceptable salt thereof every 12 hours.

26. Use of 6-((3S,4S)-4-methyl-1-(pyrimidine-2-ylmethyl)pyrrolidine-3-yl)-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one or a pharmaceutically acceptable salt thereof in the manufacture of a pharmaceutical product for the treatment of chronic heart failure in a patient requiring treatment.

27. 6-((3S,4S)-4-methyl-1-(pyrimidine-2-ylmethyl)pyrrolidine-3-yl)-1-(tetrahydro-2H-pyran-4-yl)-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one or a pharmaceutically acceptable salt thereof for use in the treatment of chronic heart failure in patients requiring treatment.

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