Preparation of quinazolinediones and their uses

Abstract

The invention described herein relates to compounds having structure I for treating diseases and disorders in which inhibition or modulation of the ubiquitin ligase COP1 enzyme results in a physiologically beneficial response, particularly for the treatment of non-alcoholic fatty liver disease (NAFLD). These compounds having structure I are capable of increasing the levels of adipose triglyceride lipase (ATGL). Also provided are methods for preparing compounds having structure I. JPEG2025501770000071.jpg37170

Description

[Technical field]

[0001] [ka]

[0002] The present invention relates to the preparation of novel compounds having structure I, in free or acceptable salt form, for the regulation of ubiquitin ligase COP1 through its stabilization as a potential therapeutic target for non-alcoholic fatty liver disease (NAFLD).

[0003] The present invention provides a novel ubiquitin ligase capable of increasing the levels of adipose triglyceride lipase (ATGL) through its regulation via stabilization of the ubiquitin ligase COP1 as a potential therapeutic target for nonalcoholic fatty liver disease (NAFLD). 1 , R 2 , R 3 relates to a small molecule as defined herein. [Background technology]

[0004] Nonalcoholic fatty liver disease (NAFLD) has attracted considerable attention due to the increasing prevalence of this disease spectrum worldwide. NAFLD is a general term that encompasses simple steatosis that progresses to steatohepatitis, fibrosis, cirrhosis, and HCC. Steatosis is a condition that is often reversible, in which lipid droplets, often in the form of triglycerides, accumulate in the liver without significant hepatocellular injury. Steatohepatitis (NASH) is a condition in which hepatocytes are severely damaged and is characterized histologically by the presence of balloon-like hepatocytes, Mallory-Denk bodies, glycogenic nuclei, and other distinctive features. NASH may progress in some cases to the more critical stages of fibrosis and cirrhosis, in which extracellular matrix proteins, especially collagen fibers, accumulate in the liver surrounding hepatocytes, forming scar tissue that irreversibly damages the normal physiological function of the liver. The prevalence of NAFLD is reported to be 20%-30% in Western countries and 5%-18% in Asia. While the incidence of NAFLD is rising at an alarming rate and it is now considered the second most common reason for liver transplantation, no potent treatment is available to reverse the advanced stages of the condition.

[0005] NAFLD is a complex multifactorial disorder involving the interaction of several molecules and their associated signaling pathways. Numerous risk factors are believed to contribute to the development of NAFLD, with type 2 diabetes and metabolic syndrome being considered the most important ones. As is evident, the most prominent feature of NAFLD is the deposition of excess TAG in hepatocytes, and thus, deregulation of enzymes responsible for the control of intracellular lipid turnover and homeostasis may play a key role in NAFLD (Ong et al., Hepatology. 2011, 53, 116-126). The key enzyme associated with the intracellular degradation of TAG is adipose triglyceride lipase (ATGL), also known as patatin-like phospholipase domain-containing protein 2 (PNPLA2). ATGL catalyzes the early, rate-limiting step in the lipolytic cascade of TAG. Indeed, studies have shown that ATGL levels are reduced in patients with NAFLD and that liver injury is exacerbated in mice with liver-specific ATGL depletion (Jha et al., Hepatology, 2014, 59, 858-869).

[0006] The ubiquitin-proteasome system is a key pathway for the regulation of protein turnover in cells. Protein ubiquitination requires the stepwise participation of three enzymes: E1, a ubiquitin-activating enzyme, E2, a ubiquitin-conjugating enzyme, and E3, a ubiquitin ligase. COP1 is one such evolutionarily conserved ubiquitin ligase that plays a central role in a myriad of important cellular pathways, such as insulin secretion from pancreatic β-cells, control of p53 stability, etc.

[0007] Our previous study identified a novel interaction between COP1 and the VP motif of ATGL. This interaction targets ATGL primarily at the lysine 100 residue for proteasomal degradation by K-48-linked polyubiquitination. In NAFLD, enhanced degradation of ATGL by COP1 results in more TAG accumulation in the liver, leading to more severe forms of the disease (Ghosh et al., Diabetes, 2016, 65, 3561-3572). Thus, reducing ubiquitin-mediated degradation of ATGL by inhibiting COP1 could be a potential area for therapeutics. Indeed, in the same study, it was verified that steatosis in mouse liver could be alleviated by adenovirus-mediated depletion of COP1 in mice. In this context, developing small molecules with the potential to target the interaction site between COP1 and ATGL and interfering with its ability to target COP1 and ubiquitinate ATGL would allow ATGL to hydrolyze TAG accumulated in the liver and abrogate the progression of NAFLD. Thus, achieving this increased lipolysis in the liver in the context of NAFLD would provide a powerful therapy to combat the progression of steatosis to steatohepatitis and ultimately limit NAFLD at its onset (Niyogi et al., Biochemical and Biophysical Research Communications, 2019, 512, 806-811).

[0008] Currently, treatment strategies are mainly directed at various targets that mediate hepatocyte dysregulation, inflammation, apoptosis, and oxidative stress. Extrahepatic targets such as the microbiome, gut-liver axis, and organs such as muscle and adipose tissue, which have been implicated in NASH, are also being considered to design therapeutic targets. Certain drugs are in clinical trials in various phases. In particular, elafibranor (PPAR-α / δ ligand), selonsertib (ASK-1 inhibitor), obeticholic acid (FXR agonist), and cenicriviroc (CCR2 / 5 inhibitor) are in phase 3 trials. All of these drugs target fibrosis, which is a fairly advanced stage in NASH. A few drugs, such as Aramchol (SCD-1 inhibitor), IMM-124E (anti-LPS), MGL-3196 (THR-β agonist), NGM282 (FGF19 analogue), PF-05221304 (ACC inhibitor), etc., are in phase 2 clinical trials and are aimed at improving liver fat, thus mainly targeting the steatosis stage. Targeting the fibrosis stage in NASH does not always show benefit, since this stage is often irreversible and much damage has already been done in the liver by the deposition of collagen fibers and the initiation of scar tissue formation. Therefore, if the progression of NASH could be arrested at the reversible steatosis stage by reducing fat deposition, a significantly more effective treatment could be established.

[0009] Thus, taking into account the shortcomings of the prior art reported thus far, there is a need to solve the problem of providing an innovative method of quinazolinedione derivatives for treating diseases and disorders in which inhibition or modulation of the ubiquitin ligase COP1 enzyme leads to a physiologically beneficial response, in particular for the treatment of non-alcoholic fatty liver disease (NAFLD). [Prior art documents] [Non-patent literature]

[0010] [Non-Patent Document 1] Ong et al., Hepatology. 2011, 53, 116-126 [Non-Patent Document 2] Jha et al., Hepatology, 2014, 59, 858-869 [Non-Patent Document 3] Ghosh et al., Diabetes, 2016, 65, 3561-3572 [Non-Patent Document 4] Niyogi et al., Biochemical and Biophysical Research Communications, 2019, 512, pp. 806-811 Summary of the Invention [Problem to be solved by the invention]

[0011] The primary object of the present invention is to provide compounds having structure I.

[0012] Another object of the present invention is to provide a process for the preparation of compounds having structure I.

[0013] Yet another object of the present invention is to evaluate the effectiveness of active compounds using screening methods involving fluorescence microscopy and measuring the levels of ATGL protein.

[0014] Another object of the present invention is to provide a method for testing the specificity of compounds that target the ATGL-COP1 interaction.

[0015] Yet another object of the present invention is to increase hepatic cell levels of ATGL, which can reduce the levels of cellular lipids.

[0016] Another object of the present invention is to reduce the ubiquitination and proteasomal degradation of ATGL.

[0017] Yet another object of the present invention is to identify the specific E1 and E2 enzymes in the ubiquitination process.

[0018] It is yet another object of the present invention to reduce the level of triglycerides in liver cells.

[0019] Yet another object of the present invention is to test the efficacy of compounds in vivo in preclinical models.

[0020] It is yet another object of the present invention to provide compositions comprising compounds of structure I for use in a number of clinical applications, including medicaments and methods for treating conditions such as nonalcoholic fatty liver disease (NAFLD).

[0021] It is yet another object of the present invention to provide compositions and methods using compounds having general structure I that do not have significant cytotoxicity in hepatocytes. [Means for solving the problem]

[0022] An embodiment of the present invention relates to a compound having structure I, or a pharma- ceutically acceptable salt thereof:

[0023] [ka]

[0024] (In the formula, R 1 is independent,

[0025] [ka]

[0026] is selected from the group consisting of R 2 is independent,

[0027] [ka]

[0028] is selected from the group consisting of R3 is independent,

[0029] [ka]

[0030] (selected from the group consisting of:

[0031] In an embodiment of the invention, a compound having structure I is 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (5), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-methyl-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (8), 1-(3-acetylphenyl)-3-(1-isopropyl-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (11), 1-(3-acetylphenyl)-3-(1-cyclohexyl-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (14), 1-(3-acetylphenyl)-3-(1-cyclopentyl-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (17), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(1-methylpiperidin-4-yl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (20), 1-(3-acetylphenyl)-3-(1-(cyclopropylmethyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (23), 1-(3-acetylphenyl)-3-(1-benzyl-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (26), 1-(3-acetylphenyl)-3-(1-(4-cyanobenzyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (29), 1-(3-acetylphenyl)-3-(1-(4-cyano-2-fluorobenzyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (32), 1-(3-acetylphenyl)-3-(1-(4-fluorobenzyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (35), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(4-nitrobenzyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (38), 1-(3-acetylphenyl)-3-(1-(4-bromobenzyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (41), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(4-(trifluoromethyl)benzyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (44), 1-(3-acetylphenyl)-3-(1-(4-methoxybenzyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (47), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(pyridin-3-ylmethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (50), Methyl 3-((6-(3-(3-acetylphenyl)ureido)-3-(2-methoxyethyl)-2,4-dioxo-3,4-dihydroquinazolin-1(2H)-yl)methyl)benzoate (53), Methyl 4-((6-(3-(3-acetylphenyl)ureido)-3-(2-methoxyethyl)-2,4-dioxo-3,4-dihydroquinazolin-1(2H)-yl)methyl)benzoate (56), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(pyrrolidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (59), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (62), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(2-morpholinoethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (65), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(2-(4-methylpiperazin-1-yl)ethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (68), 1-(1-(2-(1H-imidazol-1-yl)ethyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-acetylphenyl)urea (72), tert-Butyl 4-(2-(6-(3-(3-acetylphenyl)ureido)-3-(2-methoxyethyl)-2,4-dioxo-3,4-dihydroquinazolin-1(2H)-yl)ethyl)piperazine-1-carboxylate (75), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperazin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (76), 1-(1-(2-(1H-pyrrol-1-yl)ethyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-acetylphenyl)urea (79), 1-(1-(2-(1H-indol-1-yl)ethyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-acetylphenyl)urea (82), 1-(1-(2-(1H-benzo[d]imidazol-1-yl)ethyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-acetylphenyl)urea (85), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(3-(pyrrolidin-1-yl)propyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (88), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(3-(piperidin-1-yl)propyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (91), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(3-morpholinopropyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (94), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(3-(4-methylpiperazin-1-yl)propyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (97), 1-(3-acetylphenyl)-3-(3-(3-methoxypropyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (102), 1-(3-acetylphenyl)-3-(3-(2-ethoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (107), 1-(3-acetylphenyl)-3-(3-ethyl-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (112), Ethyl 2-(6-(3-(3-acetylphenyl)ureido)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2-dihydroquinazolin-3(4H)-yl)acetate (117), 1-(3-acetylphenyl)-3-(3-(3-methoxyphenyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (122), 1-(3-acetylphenyl)-3-(3-(2-methoxyphenyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (127), 1-(3-acetylphenyl)-3-(3-(2-(dimethylamino)ethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (132), 1-(3-acetylphenyl)-3-(2,4-dioxo-1,3-bis(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (137), 1-(3-acetylphenyl)-3-(3-(2-aminoethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (142), 1-(3-acetylphenyl)-3-(3-(2-(4-methylpiperazin-1-yl)ethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (147), 1-(3-acetylphenyl)-3-(3-(3-morpholinopropyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (152), 1-(3-acetylphenyl)-3-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (157), 1-(3-acetylphenyl)-3-(3-(1-methylpiperidin-4-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (162), 1-(3-acetylphenyl)-3-(2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-3-(pyridin-4-yl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (167), 1-(3-acetylphenyl)-3-(2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-3-(pyridin-3-yl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (172), 1-(3-acetylphenyl)-3-(2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-3-(pyridin-2-yl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (177), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-methoxyphenyl)urea (178), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(2-methoxyphenyl)urea (179), 1-(3-(1-hydroxyethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (180), 1-(3-ethylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (181), Methyl 3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)benzoate (182), 3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)benzoic acid (183), 3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)-N,N-dimethylbenzamide (184), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(pyrrolidine-1-carbonyl)phenyl)urea (185), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(morpholine-4-carbonyl)phenyl)urea (186), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-phenylurea (187), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(methylamino)phenyl)urea (188), N-(3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)phenyl)acetamide (189), N-(3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)phenyl)-N-methylacetamide (190), N-benzyl-N-(3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)phenyl)acetamide (191), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-1-methylurea (192), 1-(3-acetylphenyl)-1-hydroxy-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (193), (Z)-1-(3-(1-(hydroxyimino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (194), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(2,2,2-trifluoroacetyl)phenyl)urea (195), 1-(3-(1-aminoethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (196), 1-(3-(1-(dimethylamino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (197), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(1-(phenylamino)ethyl)phenyl)urea (199), 1-(3-(1-((2-fluorophenyl)amino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (201), 1-(3-(1-((4-fluorophenyl)amino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (203), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(1-((4-methoxyphenyl)amino)ethyl)phenyl)urea (205), 1-(3-(1-((4-cyanophenyl)amino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (207), 1-(3-(1-(cyclohexylamino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (209), 1-(3-(1-(cyclopentylamino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (211), 6-((5-acetyl-1H-benzo[d]imidazol-2-yl)amino)-3-(2-methoxyethyl)-1-(2-(piperidin-1-yl)ethyl)quinazoline-2,4(1H,3H)-dione (216), 6-((2-((3-acetylphenyl)amino)-3,4-dioxocyclobut-1-en-1-yl)amino)-3-(2-methoxyethyl)-1-(2-(piperidin-1-yl)ethyl)quinazoline-2,4(1H,3H)-dione (217), (R)-1-(3-acetylphenyl)-3-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (222), (S)-1-(3-acetylphenyl)-3-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (226), (R)-3-(3-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)-N,N-dimethylbenzamide (228), (S)-3-(3-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)-N,N-dimethylbenzamide (229), (R)-1-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(2,2,2-trifluoroacetyl)phenyl)urea (230), (S)-1-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(2,2,2-trifluoroacetyl)phenyl)urea (231) is selected from the group consisting of:

[0032] Yet another embodiment of the present invention is a method for the preparation of a compound having structure I, comprising: (i) 2-Amino-5-nitrobenzoic acid (compound 1) was reacted with 2-methoxyethylamine, methoxypropylamine, 2-ethoxyethylamine, 2M ethylamine in THF, glycine ethyl ester hydrochloride, m-anisidine, o-anisidine, N,N-dimethylethylenediamine, 1-(2-aminoethyl)piperidine, 4-(2-aminoethyl)morpholine, 1-(2-aminoethyl)-4-methylpiperazine, 3-(4-morpholinyl)propylamine ... with an aliphatic or aromatic amine selected from the group consisting of 2, 98, 103, 108, 113, 118, 123, 128, 133, 138, 143, 148, 153, 158, 163, 168, 173, 218, and 223; (ii) cyclizing the compound selected from the group consisting of 2, 98, 103, 108, 113, 118, 123, 128, 133, 138, 143, 148, 153, 158, 163, 168, 173, 218, and 223 obtained in step (i) using a cyclizing agent CDI in DMF as a solvent at 100° C. for 12 to 16 hours to obtain a compound selected from the group consisting of 3, 99, 104, 109, 114, 119, 124, 129, 134, 139, 144, 149, 154, 159, 164, 169, 174, 219, and 224; (iii) reacting compound 3 obtained in step (ii) with methyl iodide and dry DMF at 0° C. for 12 hours to obtain compound 6; (iv) Compounds 3, 99, 104, 109, 114, 119, 124, 129, 134, 139, 144, 149, 154, 159, 164, 169, 174, 219, and 224 obtained in step (ii) are reacted with 2-iodopropane, bromocyclohexane, bromocyclopentane, 4-bromo-1-methylpiperidine, (bromomethyl)cyclopropane, benzyl bromide, 4-(bromomethyl)benzonitrile, 4-(bromomethyl)-3-fluorobenzonitrile, 4-fluorobenzyl bromide, 4-nitrobenzyl bromide, 4-bromobenzyl bromide, 4-methyl ... a suitable alkyl chloride selected from the group consisting of methyl 3-(bromomethyl)pyridine hydrobromide, methyl 3-(bromomethyl)benzoate, methyl 4-(bromomethyl)benzoate, 1-(2-chloroethyl)pyrrolidine, 1-(2-chloroethyl)piperidine, 4-(2-chloroethyl)morpholine, 1-(2-chloroethyl)-4-methylpiperazine, 1-(3-chloropropyl)pyrrolidine, 1-(3-chloropropyl)piperidine, 4-(3-chloropropyl)morpholine, and 1-(3-chloropropyl)-4-methylpiperazine; and 2 CO 3 and dry DMF at 120° C. for 12 hours to obtain a compound selected from the group consisting of 9, 12, 18, 21, 24, 27, 30, 33, 36, 39, 42, 45, 48, 51, 54, 57, 60, 63, 66, 86, 89, 92, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 220, and 225; (v) The compound 3 obtained in step (ii) is reacted with chlorobromoethane and K 2 CO 3 and dry DMF at 120° C. for 12 hours to obtain compound 69; (vi) Reacting compound 69 obtained in step (v) with a suitable amine selected from the group consisting of imidazole, N-Boc piperazine, pyrrole, indole, and benzimidazole, K 2 CO 3and dry DMF at 120° C. for 12 hours to obtain a compound selected from the group consisting of 70, 73, 77, 80, and 83; (vii) The compound selected from the group consisting of 3, 6, 9, 12, 18, 21, 24, 27, 30, 33, 36, 39, 42, 45, 48, 51, 54, 57, 60, 63, 66, 70, 73, 77, 80, 83, 86, 89, 92, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 220, and 225 obtained in steps (ii), (iii), and (iv) is subjected to a palladium-carbon (5% or 10%, wet) treatment at room temperature for 2 to 5 hours under H 2 to obtain an amine compound selected from the group consisting of 4, 7, 10, 13, 19, 22, 25, 28, 31, 34, 37, 40, 43, 46, 49, 52, 55, 58, 61, 64, 67, 71, 74, 78, 81, 84, 87, 90, 93, 96, 101, 106, 111, 116, 121, 126, 131, 136, 141, 146, 151, 156, 161, 166, 171, 176, 221, 226; (viii) Compounds 4, 7, 10, 13, 19, 22, 25, 28, 31, 34, 37, 40, 43, 46, 49, 52, 55, 58, 61, 64, 67, 71, 74, 78, 81, 84, 87, 90, 93, 96, 101, 106, 111, 116, 121, 126, 131, 136, 141, 146, 151, 156, 161, 166, 171, 176, 221, 226 obtained in step (vii) are treated with 4-nitrophenyl chloroformate in the presence of TEA as a base, followed by incubation at room temperature for 3-8 hours. with an amine selected from the group consisting of 3'-aminoacetophenone in dry THF for 20 min to obtain a compound having structure I selected from the group consisting of 5, 8, 11, 14, 20, 23, 26, 29, 32, 35, 38, 41, 44, 47, 50, 53, 56, 59, 62, 65, 68, 72, 75, 79, 82, 85, 88, 91, 94, 97, 102, 107, 112, 117, 122, 127, 132, 137, 142, 147, 152, 157, 162, 167, 172, 177, 222, 227; (ix) Compounds 61, 221, and 226 obtained in step (vii) are treated with 4-nitrophenyl chloroformate in the presence of TEA as a base, followed by the treatment with m-anisidine, o-anisidine, 1-(3-aminophenyl)ethanol, 3-ethylaniline, methyl 3-aminobenzoate, 3-amino-N,N-dimethylbenzamide, (3-aminophenyl)(pyrrolidin-1-yl)methanone, (3-aminophenyl)(morpholino)methanone, aniline, N1-methylbenzene-1,3-diamine, N-(3-aminophenyl)acetate, etc. in dry THF at room temperature for 3 to 8 hours. 178, 179, 180, 181, 182, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 195, 228, 229, 230, 231; (x) Compound 182 obtained in step (ix) was treated with LiOH H in THF (3:2:1) at room temperature for 12 hours. 2 O, methanol and H 2 with O to obtain compound 183 having structure I; (xi) treating compound 62 obtained in step (viii) with hydroxylamine hydrochloride in EtOH at room temperature for 12 hours to obtain compound 194 having structure I; (xii) treating compound 75 obtained in step (viii) with trifluoroacetic acid in DCM at room temperature for 8 hours to obtain compound 76 having structure I; (xiii) Compound 62 obtained in step (viii) is reacted with NH 3 / methanol, followed by NaBH in methanol 4 to obtain compound 196 having structure I; (xiv) treating compound 196 obtained in step (xiii) with HCOOH and formaldehyde in the presence of concentrated HCl at 100° C. for 8 hours to obtain compound 197 having structure I; (xv) treating compound 62 obtained in step (viii) with various aromatic amines (aniline, 2-fluoroaniline, 4-fluoroaniline, 4-methoxyaniline, 4-aminobenzonitrile, cyclohexylamine, cyclopentylamine) in the presence of p-toluenesulfonic acid (PTSA) in EtOH at 100° C. to obtain compounds having structure I selected from the group consisting of 196, 200, 202, 204, 206, 208, 210; (xvi) Compounds 196, 200, 202, 204, 206, 208, and 210 obtained in step (xv) are dissolved in sodium cyanoborohydride (NaCNBH 3 207, 209, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 2 (xvii) treating commercially available compound 212 with 2-methoxyethylamine in HATU and DMF at room temperature for 2 hours to give compound 213; (xviii) treating the compound obtained in step (xvii) with CDI in DMF at 100° C. for 12 hours to obtain compound 214; (xix) The compound 214 obtained in step (xviii) is treated with 1-(2-chloroethyl)piperidine and K at 120° C. for 12 hours. 2 CO 3 and dry DMF to obtain compound 215; (xx) The compound 215 obtained in step (xix) was treated with 1-(2-amino-1H-benzo[d]imidazol-5-yl)ethanone and Pd as a catalyst at 100° C. for 12 hours. 2 (dba) 3 and X-Phos and base K as ligands 2 CO 3 to obtain compound 216; (xxi) reacting compound 61 obtained in step (vii) with 3,4-diethoxy-3-cyclobutene-1,2-dione and p-toluenesulfonic acid (PTSA) in EtOH and 3-aminoacetophenone at 80° C. for 12 hours to obtain compound 217. The present invention provides a method comprising:

[0033] Yet another embodiment of the present application provides compounds having structure I or salts thereof for use in treating diseases and disorders related to the regulation of COP1 enzyme through its stabilization or the regulation of ATGL.

[0034] Another embodiment of the present invention provides a compound having structure I, or a salt thereof, for use in lowering triglyceride levels in liver cells.

[0035] Yet another embodiment of the present invention provides a compound having structure I, or a salt thereof, for use in treating a disease selected from nonalcoholic fatty liver disease (NAFLD) or nonalcoholic steatohepatitis (NASH).

[0036] Another embodiment of the present invention provides a compound having structure I, or a salt thereof, in combination with a pharma- ceutically acceptable excipient.

[0037] Yet another embodiment of the present invention provides a method for the modulation of the COP1 enzyme through its stabilization with compounds having structure I.

[0038] Another aspect of the present invention provides a method of increasing the level of ATGL with a compound having structure I.

[0039] The present invention has other advantages and features which will become more readily apparent from the following detailed description of the invention and the appended claims, when taken in conjunction with the accompanying drawings. Effect of the Invention

[0040] The compounds of the present invention having structure I have several advantages. Compounds having structure I can regulate COP1 ubiquitin ligase enzyme through stabilization in hepatocytes. Compounds having structure I can reduce the level of triglycerides in hepatocytes. Therefore, they can be used in clinical applications for the treatment of conditions including non-alcoholic fatty liver disease (NAFLD). Compounds having structure I have good ADME properties in vitro (kinetic solubility, Log D and metabolic stability). [Brief description of the drawings]

[0041] [Figure 1] 1A-G illustrate the results of Western blot analysis in HepG2 cells following treatment with compounds 5, 11, 62, 94, 8, 59 and 91, according to an embodiment of the present disclosure. [Diagram 2] FIG. 1 illustrates ATGL protein status in mouse primary hepatocytes after compound treatment in accordance with an embodiment of the present disclosure. [Diagram 3] 3A-C illustrate the results of an immunoprecipitation assay to examine the ubiquitination status of ATGL after treatment with compounds, according to an embodiment of the present disclosure. [Figure 4] FIG. 1 illustrates images of HepG2 cells upon compound treatment using confocal microscopy, in accordance with an implementation of the present disclosure. [Diagram 5] FIG. 1 illustrates in one panel the results of Western blot analysis in HepG2 cells after treatment with compounds 5, 8, 11, 59, 62, 65, 91, 94 at a dose of 5 μM according to the practice of the present disclosure. [Figure 6] FIG. 1 illustrates the status of induction of ATGL and stabilization of COP1 in mouse liver and adipose tissue upon treatment of compound by oral administration at a dose of 80 mg / Kg for 9 days according to the practice of the present disclosure. [Figure 7]7A-E illustrate histology data (H&E) from randomly distributed C57BL / 6 mice fed a HFD for 12 weeks and orally gavaged with vehicle (red) and compound 62 (blue) for the last 4 weeks. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0042] While the invention has been disclosed with reference to particular embodiments, those skilled in the art will recognize that various modifications may be made and equivalents may be substituted without departing from the scope of the invention. In addition, many modifications may be made to adapt a particular situation or material to the teachings of the invention without departing from its scope.

[0043] Throughout the specification and claims, unless the context clearly dictates otherwise, the following terms have the meanings expressly associated therewith herein.

[0044] The meanings of "a," "an," and "the" include plural references. The meaning of "in" includes "in" and "on." When reference is made to the drawings, like numerals refer to like parts throughout the views. Additionally, unless otherwise stated or inconsistent with the disclosure herein, references to the singular include references to the plural.

[0045] definition For convenience, before further description of the present disclosure, certain terms and examples employed herein will now be described in detail. These definitions should be read and understood in light of the remainder of the present disclosure by those skilled in the art. Although the terms used herein have meanings that are recognized and known to those skilled in the art, for convenience and completeness, certain terms and their meanings are described below.

[0046] The articles "a", "an" and "the" are used to refer to one or to more than one (ie, to at least one) of the grammatical object of the article.

[0047] The terms "comprise" and "comprising" are used in an inclusive, open-ended sense in which additional elements may be included. It is not intended to be interpreted as "consisting only of." Throughout this specification, unless the context requires otherwise, the word "comprise" and variations such as "comprises" and "comprising" are understood to imply the inclusion of a stated element or step or group of elements or steps, but not the exclusion of any other element or step or group of elements or steps.

[0048] Complex multifactorial disorders such as NAFLD lead to the deposition of excessive TAG in hepatocytes, and thus deregulation of enzymes responsible for controlling intracellular lipid turnover and homeostasis may play a key role in NAFLD. Adipose triglyceride lipase (ATGL), also known as patatin-like phospholipase domain-containing protein 2 (PNPLA2), is a key enzyme involved in the intracellular degradation of TAG and catalyzes the initial rate-limiting step in the TAG lipolytic cascade.

[0049] In accordance with the above objectives, the present invention relates to compounds having the general structure I or a salt thereof:

[0050] [ka]

[0051] (In the formula, R 1 is independent,

[0052] [ka]

[0053] is selected from the group consisting of R 2 is independent,

[0054] [ka]

[0055] is selected from the group consisting of R 3 is independent,

[0056] [ka]

[0057] (selected from the group consisting of: All compounds having structure I disclosed in this invention are shown in Table 1.

[0058] [ka]

[0059] [Table 1A]

[0060] [Table 1B]

[0061] [Table 1C]

[0062] [Table 1D]

[0063] [Table 1E]

[0064] [Table 1F]

[0065] [Table 1G]

[0066] [Table 1H]

[0067] [Table 1I]

[0068] [Table 1J]

[0069] [Table 1K]

[0070] [Table 1L]

[0071] [Table 1M]

[0072] [Table 1N]

[0073] [Table 1O]

[0074] In an embodiment of the invention, there is provided a method for the preparation of a compound having structure I, comprising: (i) 2-Amino-5-nitrobenzoic acid (compound 1) was reacted with 2-methoxyethylamine, methoxypropylamine, 2-ethoxyethylamine, 2M ethylamine in THF, glycine ethyl ester hydrochloride, m-anisidine, o-anisidine, N,N-dimethylethylenediamine, 1-(2-aminoethyl)piperidine, 4-(2-aminoethyl)morpholine, 1-(2-aminoethyl)-4-methylpiperazine, 3-(4-morpholinyl)propane, 1-(2-aminoethyl)-4-methylpiperazine ... with an aliphatic or aromatic amine selected from the group consisting of 2, 98, 103, 108, 113, 118, 123, 128, 133, 138, 143, 148, 153, 158, 163, 168, 173, 218, 223; (ii) cyclizing the compound selected from the group consisting of 2, 98, 103, 108, 113, 118, 123, 128, 133, 138, 143, 148, 153, 158, 163, 168, 173, 218, 223 obtained in step (i) using a cyclizing agent CDI in DMF as a solvent at 100° C. for 12-16 hours to obtain a compound selected from the group consisting of 3, 99, 104, 109, 114, 119, 124, 129, 134, 139, 144, 149, 154, 159, 164, 169, 174, 219, 224; (iii) reacting compound 3 obtained in step (ii) with methyl iodide and dry DMF at 0° C. for 12 hours to obtain a compound selected from the group consisting of 6; (iv) Compound 3, 99, 104, 109, 114, 119, 124, 129, 134, 139, 144, 149, 154, 159, 164, 169, 174, 219, 224 obtained in step (ii) is reacted with 2-iodopropane, bromocyclohexane, bromocyclopentane, 4-bromo-1-methylpiperidine, (bromomethyl)cyclopropane, benzyl bromide, 4-(bromomethyl)benzonitrile, 4-(bromomethyl)-3-fluorobenzonitrile, 4-fluorobenzyl bromide, 4-nitrobenzyl bromide, 4-bromobenzyl bromide, 4-methanol a suitable alkyl chloride selected from the group consisting of methyl 3-(bromomethyl)pyridine hydrobromide, methyl 3-(bromomethyl)benzoate, methyl 4-(bromomethyl)benzoate, 1-(2-chloroethyl)pyrrolidine, 1-(2-chloroethyl)piperidine, 4-(2-chloroethyl)morpholine, 1-(2-chloroethyl)-4-methylpiperazine, 1-(3-chloropropyl)pyrrolidine, 1-(3-chloropropyl)piperidine, 4-(3-chloropropyl)morpholine, and 1-(3-chloropropyl)-4-methylpiperazine; and K 2 CO 3 and dry DMF at 120° C. for 12 hours to obtain a compound selected from the group consisting of 9, 12, 18, 21, 24, 27, 30, 33, 36, 39, 42, 45, 48, 51, 54, 57, 60, 63, 66, 86, 89, 92, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 220, and 225; (v) The compound 3 obtained in step (ii) is reacted with chlorobromoethane and K 2 CO 3 and dry DMF at 120° C. for 12 hours to obtain a compound selected from the group consisting of 69; (vi) Reacting compound 69 obtained in step (v) with a suitable amine selected from the group consisting of imidazole, N-Boc piperazine, pyrrole, indole, and benzimidazole, K 2 CO 3and dry DMF at 120° C. for 12 hours to obtain a compound selected from the group consisting of 70, 73, 77, 80, and 83; (vii) The compound selected from the group consisting of 3, 6, 9, 12, 18, 21, 24, 27, 30, 33, 36, 39, 42, 45, 48, 51, 54, 57, 60, 63, 66, 70, 73, 77, 80, 83, 86, 89, 92, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 220, and 225 obtained in steps (ii), (iii), and (iv) is subjected to a palladium-carbon (5% or 10%, wet) treatment at room temperature for 2 to 5 hours under H 2 to obtain an amine compound selected from the group consisting of 4, 7, 10, 13, 19, 22, 25, 28, 31, 34, 37, 40, 43, 46, 49, 52, 55, 58, 61, 64, 67, 71, 74, 78, 81, 84, 87, 90, 93, 96, 101, 106, 111, 116, 121, 126, 131, 136, 141, 146, 151, 156, 161, 166, 171, 176, 221, 226; (viii) Compounds 4, 7, 10, 13, 19, 22, 25, 28, 31, 34, 37, 40, 43, 46, 49, 52, 55, 58, 61, 64, 67, 71, 74, 78, 81, 84, 87, 90, 93, 96, 101, 106, 111, 116, 121, 126, 131, 136, 141, 146, 151, 156, 161, 166, 171, 176, 221, 226 obtained in step (vii) are treated with 4-nitrophenyl chloroformate in the presence of TEA as a base, followed by incubation at room temperature for 3-8 hours. with an amine selected from the group consisting of 3'-aminoacetophenone in dry THF for 20 min to obtain a compound having structure I selected from the group consisting of 5, 8, 11, 14, 20, 23, 26, 29, 32, 35, 38, 41, 44, 47, 50, 53, 56, 59, 62, 65, 68, 72, 75, 79, 82, 85, 88, 91, 94, 97, 102, 107, 112, 117, 122, 127, 132, 137, 142, 147, 152, 157, 162, 167, 172, 177, 222, 227; (ix) Compounds 61, 221, and 226 obtained in step (vii) are treated with 4-nitrophenyl chloroformate in the presence of TEA as a base, followed by the treatment with m-anisidine, o-anisidine, 1-(3-aminophenyl)ethanol, 3-ethylaniline, methyl 3-aminobenzoate, 3-amino-N,N-dimethylbenzamide, (3-aminophenyl)(pyrrolidin-1-yl)methanone, (3-aminophenyl)(morpholino)methanone, aniline, N1-methylbenzene-1,3-diamine, N-(3-aminophenyl)acetate, etc. in dry THF at room temperature for 3 to 8 hours. 178, 179, 180, 181, 182, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 195, 228, 229, 230, 231; (x) Compound 182 obtained in step (ix) was treated with LiOH H in THF (3:2:1) at room temperature for 12 hours. 2 O, methanol and H 2 with O to obtain a compound having structure I, selected from the group consisting of 183; (xi) treating compound 62 obtained in step (viii) with hydroxylamine hydrochloride in EtOH at room temperature for 12 hours to obtain a compound having structure I, selected from the group consisting of 194; (xii) treating compound 75 obtained in step (viii) with trifluoroacetic acid in DCM at room temperature for 8 hours to obtain a compound having structure I, selected from the group consisting of 76; (xiii) Compound 62 obtained in step (viii) is reacted with NH 3 / methanol, followed by NaBH in methanol 4to obtain a compound having structure I, selected from the group consisting of 196; (xiv) treating compound 196 obtained in step (xiii) with HCOOH and formaldehyde in the presence of concentrated HCl at 100° C. for 8 hours to obtain a compound having structure I, selected from the group consisting of 197; (xv) treating compound 62 obtained in step (viii) with various aromatic amines (aniline, 2-fluoroaniline, 4-fluoroaniline, 4-methoxyaniline, 4-aminobenzonitrile, cyclohexylamine, cyclopentylamine) in the presence of p-toluenesulfonic acid (PTSA) in EtOH at 100° C. to obtain compounds having structure I selected from the group consisting of 196, 200, 202, 204, 206, 208, 210; (xvi) Compounds 196, 200, 202, 204, 206, 208, and 210 obtained in step (xv) are dissolved in sodium cyanoborohydride (NaCNBH 3 207, 209, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 2 (xvii) treating commercially available compound 212 with 2-methoxyethylamine in HATU and DMF at room temperature for 2 hours to obtain a compound having structure I, selected from the group consisting of 213; (xviii) treating the compound obtained in step (xvii) with CDI in DMF at 100° C. for 12 hours to obtain a compound having structure I, selected from the group consisting of 214; (xix) The compound 214 obtained in step (xviii) is treated with 1-(2-chloroethyl)piperidine and K at 120° C. for 12 hours. 2 CO 3 and dry DMF to obtain a compound selected from the group consisting of 215; (xx) The compound 215 obtained in step (xix) was reacted with 1-(2-amino-1H-benzo[d]imidazol-5-yl)ethenone and Pd as a catalyst at 100° C. for 12 hours. 2 (dba)3 and X-Phos and base K as ligands 2 CO 3 to obtain a compound selected from the group consisting of 216; (xxi) reacting compound 61 obtained in step (vii) with 3,4-diethoxy-3-cyclobutene-1,2-dione and p-toluenesulfonic acid (PTSA) in EtOH and 3-aminoacetophenone at 80° C. for 12 hours to obtain a compound selected from the group consisting of 217. A method is provided that includes:

[0075] [ka]

[0076] Table 2 shows the structures of the reactants and products obtained by reaction via a chloroformate intermediate.

[0077] [Table 2A]

[0078] [Table 2B]

[0079] [Table 2C]

[0080] [Table 2D]

[0081] [Table 2E]

[0082] [Table 2F]

[0083]

Table 2G

[0084]

Table 2H

[0085]

Table 2I

[0086]

Table 2J

[0087]

Table 2K

[0088]

Table 2L

[0089]

Table 2M

[0090]

Table 2N

[0091]

Table 3

[0092] In an embodiment of the present invention, there is provided a compound having structure I for use in treating diseases and disorders related to the regulation of COP1 enzyme through its stabilization or the regulation of ATGL.

[0093] In another embodiment of the present invention, there is provided a compound having structure I for use in lowering triglyceride levels in liver cells.

[0094] In yet another embodiment of the present invention, there is provided a compound having structure I for use in treating a disease selected from non-alcoholic fatty liver disease (NAFLD) or non-alcoholic steatohepatitis (NASH).

[0095] In yet another embodiment of the present invention, there is provided a composition comprising a compound having structure I in combination with a pharma- ceutically acceptable excipient.

[0096] Another embodiment of the present invention provides a method for the modulation of the COP1 enzyme through its stabilization by compounds having structure I.

[0097] Another embodiment of the present invention provides a method of increasing the level of ATGL with a compound having structure I.

[0098] Although the subject matter has been described with reference to specific embodiments, this description is not meant to be construed in a limiting sense. Various modifications of the disclosed embodiments, as well as alternative embodiments of the subject matter, will become apparent to those skilled in the art upon reference to the description of the subject matter. It is therefore contemplated that such modifications can be made without departing from the spirit or scope of the subject matter as defined. EXAMPLES

[0099] The following examples, including preferred embodiments, serve to illustrate the practice of this invention, it being understood that the particulars given are by way of example and for purposes of illustrative discussion of preferred embodiments of the invention.

[0100] Temperatures are given in degrees Celsius. The structure of final products, intermediates and starting materials is confirmed by standard analytical methods, spectroscopic characterization, e.g. MS, NMR. The abbreviations used are those conventional in the art.

[0101] All starting materials, reagents, catalysts, building blocks, acids, bases, dehydrating agents and solvents utilized to synthesize the compounds of the present invention are either commercially available or can be prepared by organic synthesis methods known in the art.

[0102] Example 1 General Procedure A: Amide Formation Reaction 5-Nitroanthranilic acid (1 mmol) was taken in DMF (1-2 mL), HATU (1-1.2 equiv.) was added and the reaction mixture was stirred at room temperature for 15 min-1 h. To the reaction mixture, a suitable substituted aliphatic or aromatic amine (1-1.5 equiv.) was added dropwise followed by TEA (2.5-3 equiv.) and the contents of the reaction mixture were stirred for another 45 min. The reaction was monitored by checking TLC. Upon completion, the reaction mixture was washed thoroughly with ice-cold water to remove DMF and extracted with EtOAc. Column chromatography was performed to obtain the pure product.

[0103] Example 2 General Procedure B: Cyclization using CDI The amide compound (1 mmol) prepared by general procedure A provided in Example 1 was taken in DMF (1-2 mL) and added with CDI (1-1.5 equiv.) and heated at 110° C. for 8-12 h. The reaction was monitored by checking TLC. Upon completion, the reaction mixture was washed with water followed by extraction with EtOAc. Column chromatography was performed to obtain the pure product.

[0104] Example 3 General Procedure C: Substitution Reactions The compound (quinazolinedione) prepared by general procedure B provided in Example 2 was taken up in DMF (1-2 mL) and 2 CO 3(1-1.5 equiv.) and an appropriate alkyl chloride (aromatic or aliphatic) were added and heated at 120° C. for 12 h. The reaction was monitored by checking TLC. Upon completion, the reaction mixture was washed with water followed by extraction with EtOAc or chloroform. Column chromatography was performed to obtain the pure product.

[0105] Example 4 General Procedure D: Reduction Compounds (1 mmol) prepared by general procedures B and C provided in Examples 2 and 3 were dissolved in methanol (2-5 mL) and a small amount of 10% wet Pd-C was added. The reaction mixture was purged with nitrogen and H 2 The mixture was degassed for 2-5 hours by passing gas through it to obtain the fully reduced compound. The reaction was monitored thoroughly by checking TLC. Upon completion of the reaction, the Pd-C was filtered through a bed of celite and the methanol was evaporated in vacuum to obtain the desired compound. Column chromatography was performed to obtain the pure product.

[0106] Example 5 General Procedure E: Urea Derivative Formation via Chloroformate Intermediate The compound (1 mmol) prepared by general procedure C provided in Example 4 was dissolved in dry THF (5-10 mL). 4-Nitrophenyl chloroformate (1-1.5 eq.) was added in small portions and the reaction mixture was stirred for 15 min-3 h until the amine was consumed. The reaction was monitored by checking TLC. Further, the appropriate amine (1-1.5 eq.) was added to the reaction mixture followed by TEA (2-4 eq.) and the reaction mixture was stirred for another 2-8 h. Upon completion of the reaction, the reaction mass was evaporated in vacuum to remove THF and saturated NaHCO 3 The mixture was washed with ethyl acetate and extracted with EtOAc, and the pure product was obtained by column chromatography. Preparation of Compounds 5, 8, and 11 (Scheme 1, Scheme 2, Scheme 3)

[0107] [ka]

[0108] Example 6 Synthesis of 2-amino-N-(2-methoxyethyl)-5-nitrobenzamide (2) (DSR-01-27): The following was made according to general procedure A using 2-amino-5-nitrobenzoic acid 1 (4 g, 21.97 mmol), DMF (12 mL), HATU (9.1 g, 24.17 mmol), 2-methoxyethylamine (2.1 mL, 24.17 mmol) and TEA (7.6 mL, 54.93 mmol). After evaporation, the crude mass was diluted with chloroform and petroleum ether was added to give a precipitate. The precipitate was washed with petroleum ether to give compound 2 (4.2 g, 80%) as a yellow solid. 1 H NMR (400 MHz, d 6 -DMSO) δ (ppm) 8.71 (br.s, -NH), 8.46 (d, J= 2.8 Hz, 1H), 7.96 (dd, J= 9.4 Hz, 2.4 Hz, 1H), 7.71 (br.s, 2H), 6.75 (d, J= 9.2 Hz, 1H), 3.43- 3.40 (m, 2H), 3.37- 3.33 (m, 2H), 3.23 (s, 3H).ESI-HRMS m / z 240.0995 (M+H + ). Melting point: 168°C.

[0109] Example 7 Synthesis of 2-hydroxy-3-(2-methoxyethyl)-6-nitroquinazolin-4(3H)-one (3) (TSG-03-19): Compound 2 was prepared by general procedure B (0.3 g, 1.25 mmol), CDI (0.304 g, 1.88 mmol) was dissolved in DMF and stirred at 140° C. for 8 h. After completion of the reaction, the reaction mass was washed with water, extracted with ethyl acetate and purified by column chromatography (silica gel, mesh size 100-200) eluting with (50% ethyl acetate-petroleum ether) to give compound 3 (0.286 g, 86%) as an off-white solid, as shown in Scheme 1. 1H NMR(400MHz, DMSO-d6): δ= 12.08(s, 1H), 8.64-8.60(m, 1H), 8.49-8.45(m, 1H), 7.32(d, J= 12.0Hz, 1H), 4.09(t, J= 8.0Hz, 2H), 3.54(t, J= 8.0Hz, 2H), 3.25(s, 3H).ESI-HRMS m / z478.2463 (M+H + ).

[0110] Example 8 Synthesis of 6-amino-2-hydroxy-3-(2-methoxyethyl)quinazolin-4(3H)-one (4) (TSG-03-20): The following compound was made by general procedure D using 3 (0.220 g, 0.83 mmol), methanol (5 mL), and a small amount of 10% wet Pd-C to give compound 4 (0.121 g, 62%) as a light brown solid as shown in Scheme 1. 1 H NMR (400MHz, DMSO-d6): δ= 11.03(s, 1H), 7.11-7.08(m, 1H), 6.94-6.90(m, 1H), 5.18(s, 2H), 4.06(t, J= 8.4Hz, 2H), 3.50(t, J= 8.0Hz, 2H), 3.23(s, 3H). ESI-HRMS m / z 236.1034 (M+H + ).

[0111] Example 9 Synthesis of 1-(3-acetylphenyl)-3-(2-hydroxy-3-(2-methoxyethyl)-4-oxo-3,4-dihydroquinazolin-6-yl)urea (5) (TSG-03-23): The following compound was made by general procedure E using 4 (0.080 g, 0.34 mmol), dry THF (3 mL), 4-nitrophenyl chloroformate (0.082 g, 0.40 mmol), 3′-aminoacetophenone (0.054 g, 0.40 mmol), TEA (0.094 mL, 0.68 mmol) to give compound 5 shown in Scheme 1 (0.056 g, 45%) as a light brown solid. 1 H NMR (400MHz, DMSO-d6), δ= 11.29(s, 1H), 8.85(d, J= 10.8Hz, 1H), 8.07(d, J= 2.4Hz, 1H), 8.04-8.02(m, 1H), 7.64(dd, J= 9.2Hz, 2.8Hz, 2H), 7.55-7.53(m, 1H), 7.41-7.37(m, 1H), 7.09(d, J= 8.8Hz, 1H), 4.05(t, J= 6.0Hz, 2H), 3.49(t, J= 6.0Hz, 2H), 3.20(s, 3H), 2.52(s, 3H).ESI-HRMS m / z397.1515 (M+H + ).

[0112] Example 10 Synthesis of 3-(2-methoxyethyl)-1-methyl-6-nitroquinazoline-2,4(1H,3H)-dione (6) (TSG-03-103): Compound 3 (0.5 g, 2.00 mmol) was dissolved in dry DMF (5 mL) and then NaH (0.057 g, 2.4 mmol) was added at 0 °C with N 2 The atmosphere was maintained while adding the mixture, and the reaction mixture was stirred at room temperature for 1 h. Then CH 3 I (0.36 ml, 2.4 mmol) was added dropwise at 0° C. and the reaction mixture was stirred at room temperature overnight. Upon completion of the reaction, the reaction mass was worked up with EtOAc and water and then purified by column chromatography (silica gel, mesh size 100-200) eluting with (50% EtOAc / petroleum ether) to obtain compound 6 (0.442 g, 84%) as shown in scheme 2 as an off-white solid.

[0113] Example 11 Synthesis of 6-amino-3-(2-methoxyethyl)-1-methylquinazoline-2,4(1H,3H)-dione (7) (TSG-03-104): Compound 6 (0.4 g, 1.5 mmol) was dissolved in methanol (8 mL) and a small amount of 10% wet Pd-C was added. The reaction mixture was purged with nitrogen and H 2The mixture was degassed by passing gas through it for 2 hours to obtain the fully reduced compound. The reaction was monitored closely by checking TLC. Upon completion of the reaction, the Pd-C was filtered through a bed of Celite and the methanol was evaporated in vacuum to obtain compound 7 (0.25 g, 72%) as shown in scheme 2 as a light brown solid.

[0114] Example 12 Synthesis of 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-methyl-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (8) (TSG-03-105): Compound 7 (0.1 g, 0.4 mmol) was dissolved in dry THF (3 mL), 4-nitrophenyl chloroformate (0.097 g, 0.48 mmol) was added in small portions and the reaction mixture was stirred for 2 h until the amine was consumed. The reaction was monitored by checking TLC. 3'-aminoacetophenone (0.065 g, 0.48 mmol) was added followed by TEA (0.12 mL, 0.8 mmol) and the reaction mixture was stirred for another 3 h. Upon completion of the reaction, the reaction mass was evaporated in vacuum to remove THF and saturated NaHCO 3 The solution was washed with and extracted with EtOAc to give a yellow crude mass, which was then purified by column chromatography (silica gel, mesh size 100-200) eluting with (80% EtOAc / petroleum ether) to give compound 8 (0.085 g, 52%) as shown in Scheme 2 as an off-white solid. 1 H NMR (400 MHz, d 6-DMSO) δ (ppm) 8.92(s, 1H), 8.90(s, 1H), 8.17(d, 2.4Hz, 1H), 8.04-8.02(m, 1H), 7.77(dd, J= 9.2Hz, 2.8Hz, 1H), 7.66-7.63(m, 1H), 7.56-7.53(m,1H), 7.42-7.34(m, 2H), 4.10(t, J= 6.0Hz, 2H), 3.49(t, J= 6.0Hz, 2H), 3.45(s, 3H), 3.20(s, 3H), 2.52(s, 3H). ESI-HRMS m / z 411.1659 (M+H + ).

[0115] (Example 13) Synthesis of 1-isopropyl-3-(2-methoxyethyl)-6-nitroquinazoline-2,4(1H,3H)-dione (9) (TSG-03-106): Compound 1 (0.3g, 1.13mmol), K 2 CO 3 (0.314 g, 2.26 mmol), 2-iodopropane (0.288 g, 1.69 mmol) were taken in dry DMF in a pressure tube and stirred overnight at 120° C. Upon completion of the reaction, the reaction mass was worked up with EtOAc and water and then purified by column chromatography (silica gel, mesh size 100-200) eluting with (50% EtOAc / petroleum ether) to obtain compound 8 (0.225 g, 65%) as an off-white solid as shown in Scheme 3. 1 H NMR (400 MHz, CDCl3) δ (ppm) 9.06(d, J= 2.8Hz, 1H), 8.42(dd, J= 9.2Hz, 2.58Hz, 1H), 7.46(d, J= 9.2Hz, 1H), 5.17-4.99 (m, 1H), 4.30(t, J= 6.0Hz, 2H), 3.67(t, J= 6.0Hz, 2H), 3.34(s, 3H), 1.62(d, J= 6.8Hz, 6H). + ).

[0116] Example 14 Synthesis of 6-amino-1-isopropyl-3-(2-methoxyethyl)quinazoline-2,4(1H,3H)-dione (10) (TSG-03-112): Compound 9 (0.180 g, 0.58 mmol) was dissolved in methanol (5 mL) and a small amount of 10% wet Pd-C was added. The reaction mixture was quenched with nitrogen and H 2 The mixture was degassed by passing gas through it for 2 hours to obtain the fully reduced compound. The reaction was monitored closely by checking TLC. Upon completion of the reaction, the Pd-C was filtered through a bed of Celite and the methanol was evaporated in vacuum to obtain compound 10 (0.12 g, 75%) as shown in scheme 3 as a light brown solid.

[0117] 1 H NMR (400 MHz, CDCl 3 ) δ (ppm) 7.49(d, J= 2.8Hz, 1H), 7.17(d, J= 8.8Hz, 1H), 6.97(dd, J= 8.8Hz, 2.8Hz, 1H), 5.05-4.92 (m, 1H), 4.28(t, J= 6.0Hz, 2H), 3.74(s, 2H), 3.64(t, J= 6.0HZ, 2H), 3.35(s, 3H), 1.56(d, 6H). ESI-HRMS m / z 278.1503 (M+H + ).

[0118] Example 15 Synthesis of 1-(3-acetylphenyl)-3-(1-isopropyl-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (11) (TSG-03-114): Compound 10 (0.08 g, 0.39 mmol) was dissolved in dry THF (3 mL), 4-nitrophenyl chloroformate (0.095 g, 0.48 mmol) was added in small portions and the reaction mixture was stirred for 2 h until the amine was consumed. The reaction was monitored by checking TLC. 3'-aminoacetophenone (0.065 g, 0.48 mmol) was added followed by TEA (0.12 mL, 0.8 mmol) and the reaction mixture was stirred for another 3 h. Upon completion of the reaction, the reaction mass was evaporated in vacuum to remove THF and saturated NaHCO 3 The solution was washed with ethyl acetate and extracted with EtOAc to give a yellow crude mass, which was then purified by column chromatography (silica gel, mesh size 100-200) eluting with (70% EtOAc / petroleum ether) to give compound 11 (0.057 g, 35%) as shown in Scheme 3 as an off-white solid. 1 H NMR (400 MHz, CDCl 3 ) δ (ppm) 8.36(s, 1H), 8.26(dd, J= 9.2Hz, 2.8Hz, 1H), 8.17(s, 1H), 7.92(d, J= 2.8Hz, 1H), 7.89-7.87(m, 1H), 7.86-7.83(m, 1H), 7.61-7.58(m, 1H), 7.40-7.38(m, 1H), 7.33(d, J= 9.2Hz, 1H), 5.08-4.93(m, 1H), 4.32(t, J= 5.6Hz, 2H), 3.71(t, J= 5.6Hz, 2H), 3.29(s, 3H), 2.60(s, 3H), 1.56(d, J= 6.8Hz, 6H). ESI-HRMS m / z 439.1971 (M+H + ). Preparation of Compounds 14, 17, 20, 23, 26, 29, 32, 35, 38, 41, 44, 47, 50, 53, and 56 (Scheme 4 to Scheme 18)

[0119] [ka]

[0120] [ka]

[0121] [ka]

[0122] (Example 16) Preparation of compounds 59 and 62 (Scheme 19, Scheme 20)

[0123] [ka]

[0124] Synthesis of 3-(2-methoxyethyl)-6-nitro-1-(2(pyrrolidin-1-yl)ethyl)quinazoline-2,4(1H,3H)-dione (57) (TSG-03-130): Compound 3 (0.4 g, 1.50 mmol) was dissolved in dry DMF (5 mL), then sodium tert-butoxide (0.29 g, 3 mmol) was added and the reaction mass was stirred at room temperature for 1 h. Then a mixture of 1-(2-chloroethyl)pyrrolidine hydrochloride (0.403 g, 2.25 mmol) in dry TEA (1 mL) was added and the reaction mass was stirred at 100° C. overnight. Upon completion of the reaction, the reaction mass was worked up with EtOAc and water and then purified by column chromatography (silica gel, mesh size 100-200) eluting with (3% MeOH / CHCl 3 ) to afford compound 57 (0.409 g, 75%) as shown in Scheme 19 as an off-white solid. 1 H NMR (400 MHz, CDCl 3) δ (ppm) 9.04(d, J= 2.8Hz, 1H), 8.46(dd, J= 9.2Hz, 2.8Hz, 1H), 7.46(d, J= 9.2Hz, 1H), 4.36-4.30(m, 4H), 3.67(t, J= 5.6Hz, 2H), 3.33(s, 3H), 2.84-2.80(m, 2H), 2.70-2.67(m, 4H), 1.83-1.80(m, 4H). ESI-HRMS m / z 363.1665 (M+H + ).

[0125] (Example 17) Synthesis of 6-amino-3-(2-methoxyethyl)-1-(2-(6-phenylin-1-yl)ethyl)quinazoline-2,4(1H,3H)-dione (58) (TSG-03-132): Compound 57 (0.2 g, 0.47 mmol) was dissolved in methanol (5 mL) and a small amount of 10% wet Pd-C was added. The reaction mixture was purged with nitrogen and H 2 The mixture was degassed by passing gas through it for 2 hours to obtain the fully reduced compound. The reaction was closely monitored by checking TLC. Upon completion of the reaction, the Pd-C was filtered through a bed of Celite and the methanol was evaporated in vacuum to obtain compound 58 (0.110 g, 60%) as shown in Scheme 19 as a light brown solid. 1 H NMR (400 MHz, CDCl 3 ) δ (ppm) 7.62(d, J= 8.8Hz, 1H), 7.41(d, J= 2.4Hz, 1H), 7.09(dd, J= 8.8Hz, 2.8Hz, 1H), 4.61(t, J= 8.0Hz, 2H), 4.25(t, J= 6.0Hz, 2H), 3.62(t, J= 6.0Hz, 2H), 3.31(s, 3H), 3.29-3.27(m,2H), 2.17-2.11(m, 4H). ESI-HRMS m / z 333.1929 (M+H + ).

[0126] (Example 18) Synthesis of 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(pyrrolidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (59) (TSG-03-134): Compound 58 (0.08 g, 0.23 mmol) was dissolved in dry THF (3 mL), 4-nitrophenyl chloroformate (0.069 g, 0.34 mmol) was added in small portions and the reaction mixture was stirred for 2 h until the amine was consumed. The reaction was monitored by checking TLC. 3'-aminoacetophenone (0.046 g, 0.34 mmol) was added followed by TEA (0.06 mL, 0.46 mmol) and the reaction mixture was stirred for another 3 h. Upon completion of the reaction, the reaction mass was evaporated in vacuum to remove THF and saturated NaHCO 3 The solution was washed with ethyl acetate and extracted with EtOAc to give a yellow crude mass, which was then subjected to column chromatography (silica gel, mesh size 100-200) while eluting with (10% MeOH / CHCl 3 ) Purification to give compound 59 (0.049 g, 42%) as an off-white solid. 1 H NMR (400 MHz, CDCl 3 ) δ (ppm) 8.74(s,1H), 8.50(s, 1H), 8.02-7.99(m, 1H), 7.87(d, J= 2.4Hz, 1H), 7.80-7.77(m, 1H), 7.66-7.64(m, 1H), 7.54-7.51(m, 1H), 7.33-7.29(m, 1H), 7.12-7.09(m, 1H), 4.33(t, J= 6.4Hz, 2H), 4.21(t, J= 5.2Hz, 2H), 3.61(t, J= 5.2Hz, 2H), 3.28(s, 3H), 3.16-3.13(m, 2H), 3.04-2.98(m, 4H), 2.55(s, 3H), 1.94-1.89(m, 4H). ESI-HRMS m / z 449.2403 (M+H + ).

[0127] (Example 19) Synthesis of 3-(2-methoxyethyl)-6-nitro-1-(2-(piperidin-1-yl)ethyl)quinazoline-2,4(1H,3H)-dione (60) (TSG-03-107): Compound 3 (0.3g, 1.13mmol), K 2 CO 3 (0.312 g, 2.26 mmol), 1-(2-chloroethyl)piperidine hydrochloride (0.312 g, 1.69 mmol) were taken in dry DMF in a pressure tube and stirred overnight at 120° C. On completion of the reaction, the reaction mass was worked up with EtOAc and water and then purified by column chromatography (silica gel, mesh size 100-200) eluting with (80% EtOAc / petroleum ether) to obtain compound 60 (0.289 g, 68%) as an off-white solid. 1 H NMR (400 MHz, CDCl 3 ) δ (ppm) 9.03(d, J= 2.8Hz, 1H), 8.47(dd, J= 9.2Hz, 2.8Hz, 1H), 7.53(d, J= 9.2Hz, 1H), 4.35(t, J= 7.2Hz, 2H), 4.31(t, J= 5.6Hz, 2H), 3.67(t, J= 5.6Hz, 2H), 3.33(s, 3H), 2.69(t, J= 7.6Hz, 2H), 2.59-2.57(m, 4H), 1.64-1.58(m, 4H), 1.48-1.43(m, 2H). ESI-HRMS m / z 377.1837 (M+H + ).

[0128] (Example 20) Synthesis of 6-amino-3-(2-methoxyethyl)-1-(2-(piperidin-1-yl)ethyl)quinazoline-2,4(1H,3H)-dione (61) (TSG-03-116): Compound 60 (0.2 g, 0.51 mmol) was dissolved in methanol (5 mL) and a small amount of 10% wet Pd-C was added. The reaction mixture was irradiated with nitrogen and H 2The mixture was degassed by passing gas through it for 2 hours to obtain the fully reduced compound. The reaction was closely monitored by checking TLC. Upon completion of the reaction, the Pd-C was filtered through a bed of Celite and the methanol was evaporated in vacuum to obtain compound 61 (0.118 g, 64%) as a light brown solid. 1 H NMR (400 MHz, DMSO-d6) δ (ppm) 7.18(d, J= 2.8Hz, 2H), 6.98(dd, J= 8.8Hz, 2.8Hz, 1H), 5.25(br.s 2H), 4.18-4.09(m, 2H), 4.06(t, J= 6Hz, ESI-HRMS m / z 347.2078 (M+H + ).

[0129] Example 21 Synthesis of 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (62) (TSG-03-117): Compound 61 (0.08 g, 0.23 mmol) was dissolved in dry THF (3 mL), 4-nitrophenyl chloroformate (0.071 g, 0.34 mmol) was added in small portions and the reaction mixture was stirred for 2 h until the amine was consumed. The reaction was monitored by checking TLC. 3'-aminoacetophenone (0.048 g, 0.34 mmol) was added followed by TEA (0.07 mL, 0.46 mmol) and the reaction mixture was stirred for another 3 h. Upon completion of the reaction, the reaction mass was evaporated in vacuum to remove THF and saturated NaHCO 3 The solution was washed with ethyl acetate and extracted with EtOAc to give a yellow crude mass, which was then subjected to column chromatography (silica gel, mesh size 100-200) while eluting with (10% MeOH / CHCl 3) Purification to give compound 62 (0.057 g, 48%) as an off-white solid. 1 H NMR (400 MHz, DMSO-d6) δ (ppm) 9.11-9.05(m, 2H), 8.18(d, J= 2.4Hz, 1H), 8.04-8.03(m , 1H), 7.76(dd, J= 8.8Hz, 2.4Hz, 1H), 7.66-7.63(m, 1H), 7.55(d, J= 7.6Hz, 1H), 7.41-7.38(m, 2H), 4.24-4.19(m, 2H), 4.10(t, J= 6.0Hz, 2H), 3.50(t, J= 6.0Hz, 2H), 3.51-3.48(m, 6H), 3.20(s, 3H), 2.52(s, 3H), 1.54-1.42(m, 4H), 1.39-1.29(m, 2H). ESI-HRMS m / z 508.2555 (M+H + ). Preparation of Compounds 65, 68, 72, 75, 76, 79, 82, 85, and 88 (Scheme 21 to Scheme 29)

[0130] [ka]

[0131] [ka]

[0132] Example 22 Synthesis of 3-(2-methoxyethyl)-1-(2-morpholinoethyl)-6-nitroquinazoline-2,4(1H,3H)-dione (63) (TSG-03-129): Compound 3 (0.4 g, 1.50 mmol) was dissolved in dry DMF (5 mL), then sodium tert-butoxide (0.29 g, 3 mmol) was added and the reaction mass was stirred at room temperature for 1 h. Then a mixture of 4-(2-chloroethyl)morpholine hydrochloride (0.421 g, 2.25 mmol) in dry TEA (1 mL) was added and the reaction mass was stirred at 100° C. overnight. Upon completion of the reaction, the reaction mass was worked up with EtOAc and water and then purified by column chromatography (silica gel, mesh size 100-200) eluting with (3% MeOH / CHCl 3 ) Purification to give compound 63 (0.225 g, 65%) as an off-white solid. 1 H NMR (400 MHz, CDCl 3 ) δ (ppm) 9.06(d, J= 2.8Hz, 1H), 8.47(dd, J= 9.2Hz, 2.4Hz, 1H), 7.35(d, J= 9.2Hz, 1H), 4.34-4.29(m, 4H), 3.69-3.65(m, 6H), 3.34(s, 3H), 2.66(t, J= 6.8Hz, 2H), 2.55-2.53(m, 4H). ESI-HRMS m / z 379.1611 (M+H + ).

[0133] (Example 23) Synthesis of 6-amino-3-(2-methoxyethyl)-1-(2-morpholinoethyl)quinazoline-2,4(1H,3H)-dione (64) (TSG-03-131): Compound 63 (0.180 g, 0.47 mmol) was dissolved in methanol (5 mL) and a small amount of 10% wet Pd-C was added. The reaction mixture was quenched with nitrogen and H 2 The mixture was degassed by passing gas through it for 2 hours to obtain the fully reduced compound. The reaction was closely monitored by checking TLC. Upon completion of the reaction, the Pd-C was filtered through a bed of Celite and the methanol was evaporated in vacuum to obtain compound 64 (0.107 g, 65%) as a light brown solid. 1 H NMR (400 MHz, CDCl 3) δ (ppm) 7.45(d, J= 2.4Hz, 1H), 7.05-6.95(m, 2H), 4.29(t, J= 6.0Hz, 2H), 4.20(t, J= 7.2Hz, 2H), 3.77(s, 2H), 3.69-3.64(m, 6H), 3.34(s, 3H), 2.62(t, J= 7.2Hz, 2H), 2.56-2.54(m, 4H). ESI-HRMS m / z 349.1877 (M+H + ).

[0134] (Example 24) Synthesis of 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(2-morpholinoethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (65) (TSG-03-133): Compound 64 (0.08 g, 0.23 mmol) was dissolved in dry THF (3 mL), 4-nitrophenyl chloroformate (0.069 g, 0.34 mmol) was added in small portions and the reaction mixture was stirred for 2 h until the amine was consumed. The reaction was monitored by checking TLC. 3'-aminoacetophenone (0.046 g, 0.34 mmol) was added followed by TEA (0.06 mL, 0.46 mmol) and the reaction mixture was stirred for another 3 h. Upon completion of the reaction, the reaction mass was evaporated in vacuum to remove THF and saturated NaHCO 3 Washing with solution and extraction with EtOAc gave a yellow crude mass, which was then purified by column chromatography (silica gel, mesh size 100-200) eluting with (70% EtOAc / petroleum ether) to give compound 65 (0.052 g, 45%) as an off-white solid. 11H NMR (400 MHz, CDCl3) δ (ppm) 8.32(s, 1H), 8.24(dd, J = 9.2Hz, 2.4Hz, 1H), 8.14(s, 1H), 7.89(d, J = 2.8Hz, 1H), 7.87 - 7.84(m, 2H), 7.61 - 7.58(m, 1H), 7.41 - 7.36(m, 1H), 7.18(d, J = 9.2Hz, 1H), 4.32(t, J = 5.6Hz, 2H), 4.22(t, J = 6.8Hz, 2H), 3.72 - 3.66(m, 6H), 3.30(s, 3H), 2.65 - 2.62(m, 2H), 2.61(s, 3H), 2.57 - 2.53(m, 4H). ESI - HRMS m / z 510.2355 (M + H + ). Preparation of Compounds 91, 94, 97, 102, 107, 112, 117, 122, 127, 132, 137, 142, 147, 152, 157, 162, 167, 172, 177 - 193 (Schemes 30 - 59)

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Chem.

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Chem.

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Chem.

[0141] (Example 25) Synthesis of 3-(2-methoxyethyl)-6-nitro-1-(3-(piperidin-1-yl)propyl)quinazoline-2,4(1H,3H)-dione (89) (TSG-03-156): Compound 3 (0.3g, 1.13mmol), K 2 CO 3 (0.312 g, 2.26 mmol), 1-(3-chloropropyl)piperidine hydrochloride (0.269 g, 1.69 mmol) were taken in dry DMF in a pressure tube and stirred overnight at 120° C. On completion of the reaction, the reaction mass was worked up with EtOAc and water and then purified by column chromatography (silica gel, mesh size 100-200) eluting with (80% EtOAc / petroleum ether) to obtain compound 89 (0.258 g, 62%) as an off-white solid.

[0142] 1 H NMR (400 MHz, CDCl3) δ (ppm) 9.05(d, J= 2.4Hz, 1H), 8.44(dd, J= 9.2Hz, 3.2Hz, 1H), 7.61(d, J= 9.2Hz, 1H), 4.32(t, J= 6.0Hz, 2H), 4.22(t, J= 7.2Hz, 2H), 3.67(t, J= 6.0Hz, 2H), 3.34(s, 3H), 2.42-2.32 (m, 6H), 1.93-1.86(m, 2H), 1.60-1.53 ​​(m, 4H), 1.48-1.41(m, 2H).

[0143] (Example 26) Synthesis of 6-amino-3-(2-methoxyethyl)-1-(3-(piperidin-1-yl)propyl)quinazoline-2,4(1H,3H)-dione (90) (TSG-03-181): Compound 89 (0.2 g, 0.51 mmol) was dissolved in methanol (5 mL) and a small amount of 10% wet Pd-C was added. The reaction mixture was purged with nitrogen and H 2The mixture was degassed by passing gas through it for 2 hours to obtain the fully reduced compound. The reaction was closely monitored by checking TLC. Upon completion of the reaction, the Pd-C was filtered through a bed of Celite and the methanol was evaporated in vacuum to obtain compound 90 (0.122 g, 65%) as a light brown solid. 1 H NMR (400 MHz, CDCl3) δ (ppm) 7.45(d, J= 2.8Hz, 1H), 7.15(d, J= 8.8Hz, 1H), 6.97(dd, J= 8.8Hz, 2.8Hz, 1H), 4.28(t, J= 6.0Hz, 2H), 4.09(t, J= 7.2Hz, 2H), 3.64(t, J= 6.0Hz, 2H), 3.33(s, 3H), 2.46-2.35(m, 4H), 1.46-1.39(m, 2H). ESI-HRMS m / z 361.2246(M+H + ).

[0144] Example 27 Synthesis of 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(3-(piperidin-1-yl)propyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (91) (TSG-03-183): Compound 90 (0.08 g, 0.22 mmol) was dissolved in dry THF (3 mL), 4-nitrophenyl chloroformate (0.072 g, 0.34 mmol) was added in small portions and the reaction mixture was stirred for 2 h until the amine was consumed. The reaction was monitored by checking TLC. 3'-aminoacetophenone (0.048 g, 0.34 mmol) was added followed by TEA (0.07 mL, 0.46 mmol) and the reaction mixture was stirred for another 3 h. Upon completion of the reaction, the reaction mass was evaporated in vacuum to remove THF and saturated NaHCO 3 The solution was washed with ethyl acetate and extracted with EtOAc to give a yellow crude mass, which was then subjected to column chromatography (silica gel, mesh size 100-200) while eluting with (10% MeOH / CHCl 3 ) Purification to give compound 91 (0.052 g, 46%) as an off-white solid.

[0145] 1 H NMR (400 MHz, CDCl3) δ (ppm) 9.08(s,1H), 9.04(s, 1H), 8.22(d, J= 1.6Hz, 1H), 8.09-8.07(m, 1H), 7.82-7.89(m, 1H), 7.70-7.68(m, 1H), 7.59(d, J= 5.2Hz, 1H), 7.51(d, J= 6.0Hz, 1H), 7.46-7.42(m, 1H), 4.14(t, J= 4.0Hz, 2H), 4.11(t, J= 4.4Hz, 2H), 3.53(t, J= 4.0Hz, 2H), 3.24(s, 3H), 2.56(s, 3H), 2.44-2.24(m, 6H), 1.81-1.75(m, 2H), 1.49-1.43(m, 4H), 1.40-1.33(m, 2H). ESI-HRMS m / z 522.2722 (M+H + ).

[0146] (Example 28) Synthesis of 3-(2-methoxyethyl)-1-(3-morpholinopropyl)-6-nitroquinazoline-2,4(1H,3H)-dione (92) (TSG-03-155): Compound 3 (0.4 g, 1.50 mmol) was dissolved in dry DMF (5 mL), then sodium tert-butoxide (0.29 g, 3 mmol) was added and the reaction mass was stirred at room temperature for 1 h. Then a mixture of 4-(3-chloropropyl)morpholine (0.296 g, 1.8 mmol) in dry TEA (1 mL) was added and the reaction mass was stirred at 100° C. overnight. Upon completion of the reaction, the reaction mass was worked up with EtOAc and water and then purified by column chromatography (silica gel, mesh size 100-200) while eluting with (3% MeOH / CHCl 3 ) Purification to give compound 92 (0.366 g, 62%) as an off-white solid.

[0147] 1H NMR (400 MHz, CDCl3) δ (ppm) 9.05(d, J= 2.4Hz, 1H), 8.44(dd, J= 9.2Hz, 2.8Hz, 1H), 7.49(d, J= 9.2Hz, 1H), 4.32(t, J= 5.6Hz, 2H), 4.26(t, J= 7.2Hz, 2H), 3.70-3.66(m, 6H), 3.34(s, 3H), 2.46-2.41(m, 6H), 1.92-1.87(m, 2H).

[0148] (Example 29) Synthesis of 6-amino-3-(2-methoxyethyl)-1-(3-morpholinopropyl)quinazoline-2,4(1H,3H)-dione (93) (TSG-03-158): Compound 92 (0.2 g, 0.51 mmol) was dissolved in methanol (5 mL) and a small amount of 10% wet Pd-C was added. The reaction mixture was purged with nitrogen and H 2 The mixture was degassed by passing gas through it for 2 hours to obtain the fully reduced compound. The reaction was closely monitored by checking TLC. Upon completion of the reaction, the Pd-C was filtered through a bed of Celite and the methanol was evaporated in vacuum to obtain compound 93 (0.118 g, 64%) as a light brown solid.

[0149] 1 H NMR (400 MHz, CDCl3) δ (ppm) 7.39(d, J= 2.8Hz, 1H), 7.07(d, J=8.8Hz, 1H), 6.98-6.95(m, 1H), 4.23(t, J= 6.0Hz, 2H), 4.06(t, J= 7.2Hz, 2H), 3.65-3.63(m, 4H), 3.59(t, J= 6.0Hz, 2H), 3.29(s, 3H), 2.42-2.37(m, 6H), 1.87-1.79(m, 2H).

[0150] (Example 30) Synthesis of 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(3-morpholinopropyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (94) (TSG-03-159): Compound 93 (0.08 g, 0.23 mmol) was dissolved in dry THF (3 mL), 4-nitrophenyl chloroformate (0.071 g, 0.34 mmol) was added in small portions and the reaction mixture was stirred for 2 h until the amine was consumed. The reaction was monitored by checking TLC. 3'-aminoacetophenone (0.048 g, 0.34 mmol) was added followed by TEA (0.07 mL, 0.46 mmol) and the reaction mixture was stirred for another 3 h. Upon completion of the reaction, the reaction mass was evaporated in vacuum to remove THF and saturated NaHCO 3 The solution was washed with ethyl acetate and extracted with EtOAc to give a yellow crude mass, which was then subjected to column chromatography (silica gel, mesh size 100-200) while eluting with (10% MeOH / CHCl 3 ) Purification to give compound 94 (0.061 g, 53%) as an off-white solid. 1 H NMR (400 MHz, DMSO-d6) δ (ppm) 8.94(s, 1H), 8.91(s, 1H), 8.17(d, J= 2.8Hz, 1H), 8.04-8.02(m, 1H), 7.75(dd, J= 8.8Hz, 2.4Hz, 1H), 7.67-7.63(m, 1H), 7.56-7.53(m, 1H), 7.45(d, J= 9.2Hz, 1H), 7.42-7.37(m, 1H), 4.13-4.05(m, 4H), 3.52-3.45(m, 6H), 3.20(s, 3H), 2.52(s, 3H), 2.37-2.32 (m, 2H), 2.30-2.22(m, 4H), 1.78-1.69(m, 2H). Preparation of compounds 194, 195, 197, 199, 201, 203, 205, 207, 209, 211, 216, 217, 222, 227, 228-231 (Scheme 60-Scheme 81)

[0151] [ka]

[0152] [ka]

[0153] [ka]

[0154] (Example 31) Synthesis of (E)-1-(3-(1-(hydroxyimino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (194) (TSG-03-186): Compound 62 (0.1 g, 0.19 mmol) was dissolved in ethanol (3 mL), then hydroxylamine hydrochloride (0.028 g, 0.38 mmol) was added and the reaction mixture was refluxed for 4 h. After completion of the reaction, ethanol was evaporated and the reaction mass was worked up with EtOAc and water, then purified by column chromatography (silica gel, mesh size 100-200) with elution (3% MeOH / CHCl 3 ) to afford compound 194 (0.058 g, 59%) as an off-white solid.

[0155] 1H NMR (400 MHz, DMSO-d6) δ (ppm) 11.17(s, 1H), 8.20(d, J= 2.8Hz, 1H), 7.80-7.78(m, 1H), 7.75(d, J= 2.4Hz, 1H), 7.59(d, J= 9.2Hz, 1H), 7.47-7.43(m, 1H), 7.27-7.22(m, 1H), 7.20-7.16(m, 1H), 4.45(t, J= 6.8Hz, 2H), 4.09(t, J= 6.4Hz, 2H), 3.50(t, J= 6.4Hz, 2H), 3.40-3.37(m, 2H), 3.35-3.32(6H), 3.21(s, 3H), 2.09(s, 3H), 1.94-1.84(m, 4H).

[0156] Biology アッセイ To select compounds capable of inhibiting ATGL ubiquitination by COP1 targeting the VP motif, confocal microscopy was performed with the provided molecules. If the compound was effective in inhibiting the interaction, there would be a reduction in the number of lipid droplets in the cells after treatment. This was because the increase in ATGL levels hydrolyzed the TAG accumulated in oleic acid-transduced HepG2 cells, resulting in the reduction mentioned above. With this rationale in mind, HepG2 cells were induced to accumulate lipid droplets after treatment with 250 μM oleic acid and 10 μM of specific compounds were added. The possibility of the compounds resulting in a reduction in lipid droplet number was then examined by comparison with oleic acid-transduced cells, the number of lipid droplets of approximately 20 cells from each treatment was counted, and the average number of lipid droplets in both cells was calculated. The selected compounds were then subjected to dose-dependent treatment, and those that maintained the ability to reduce lipid droplets at low doses were then selected for Western blot analysis. Compounds that could reduce the number of lipid droplets in cells were expected to increase the levels of ATGL, as they appeared to prevent COP1 from ubiquitinating ATGL. Because ubiquitination is a post-transcriptional modification, this increase was only seen at the protein level, and gene expression would remain unchanged. For this reason, Western blots were performed to examine the levels of ATGL in cells with the selected molecules.

[0157] Example 32 Western blot HepG2 cells were treated with compounds 5, 8, 11, 59, 62, 91, 94 (50nM, 100nM, 200nM, 500nM, 1μM and 5μM for dose-dependent assay) for 24 hours. After removing the medium from the cells, the wells were washed twice with 1×PBS to remove any remaining medium. The cells were then lysed in a lysis buffer containing 50mM Tris-HCl (pH 7.4), 100mM NaCl, 1mM EDTA, 1mM EGTA, 1% Triton X-100 and a protease inhibitor cocktail (Millipore, Billerica, MA, USA). Following centrifugation at 20,000g for 20 minutes, the protein solution was extracted from the cells. Proteins were assessed using the Bradford assay. Bradford reagent (BioRad) was diluted 1:4 with double distilled water. 2 μl of protein sample was added to 100 μl of reagent and absorbance was measured at 595 nm. 30 μg of protein was diluted in lysis buffer. 1× loading buffer diluted from a 5× stock containing 250 mM Tris-HCl (pH 6.8), 10% SDS, 50% glycerol, 0.1% bromophenol blue and 10% β-mercaptoethanol was added. Protein samples were then heated to 95° C. for 10 min, cooled and centrifuged at 12,000 g for 2 min before loading.

[0158] For Western blot, proteins were resolved in 10% SDS PAGE (discontinuous buffer system). Gels were run at 80V for approximately 2 hours using 1x running buffer containing SDS, Tris base and glycine. Transfer was performed using a PVDF membrane with a pore size of 0.45 μm (Millipore). 1x transfer buffer containing Tris base, glycine and 20% methanol was used for wet transfer. Transfer was performed at 90V for 3 hours. Following transfer, the PVDF membrane containing the proteins was washed with 1x PBS and 1x PBST containing 1% Tween 20 (Sigma Aldrich). The membrane was then incubated in 5% skim milk powder at room temperature for 1 hour to block non-specific sites. After multiple washes with 1x PBST to wash away any remaining blocking buffer, the required primary antibody (COP1 [Bethyl Laboratories], ATGL [Cell Signalling Technology] or Actin [Cell Signalling Technology]) prepared in 1x PBST, 1% bovine serum albumin and 0.04% sodium azide was added to the membrane and incubated overnight at 4°C. The next day, the membrane was again washed multiple times with 1x PBST to remove any unbound primary antibody. The membrane was then incubated with goat anti-rabbit secondary antibody (Genei) for 1 hour at room temperature and again washed multiple times with 1x PBST. The membrane was then developed using Clarity™ ECL Western Blotting Substrate (BioRad) and viewed on a ChemiDoc (BioRad).

[0159] Figure 1 (A-G) illustrates the results of Western blot analysis in HepG2 cells after treatment with compounds 5, 8, 11, 59, 62, 91 and 94. The figures also depict the dose-dependent Western blot analysis of the following compounds 5, 8, 11, 59, 62, 91 and 94. These compounds showed elevated ATGL levels regardless of the dose of treatment. The increased intensity of the ATGL and COP1 bands relative to the control indicates that the respective protein levels increased upon compound treatment. Actin was used as a loading control.

[0160] (Example 33) Effects of compounds in primary mouse hepatocytes To further enhance the efficacy of compound 62, primary hepatocytes were isolated from mice and treated with these compounds in a dose-dependent manner for 24 hours. Primary hepatocytes and adipose tissue explants were isolated from mice and subjected to compound treatment at doses of 10nM, 20nM, 50nM, 100nM and 500nM for 24 hours. To examine ATGL levels, Western blot was performed with lysates after cell harvest. Antibodies for ATGL and COP1 were used. Actin served as a loading control purpose.

[0161] Culture of primary mouse hepatocytes Hepatocytes - Fed black male mice (C57bl / 6) aged 2 to 4 months were sacrificed using chloroform (SRL) and cleaned with 70% ethanol. Under aseptic conditions, the ventral side of the mouse was incised up to the liver, and the portal vein (PV) and inferior vena cava (IVC) were fully exposed. Blood was removed from the heart to prevent backflow into the liver during perfusion. A butterfly cannula was inserted into the PV, and the liver was perfused with HBSS (Hank's Balanced Salt Solution; 5 mM KCl, 0.4 mM KH). 2 PO 4 , 4 mM NaHCO 3 , 140 mM NaCl, 0.3 mM Na 2 HPO 4 , 6 mM glucose, HEPES, 0.5 mM MgCl 2 6H 2HO, 0.4 mM MgSO4 7H 2 HO, 0.5 mM EDTA; 1 mM CaCl 2 20 ml of buffer (containing no EDTA) was passed through the liver (perfusion) at a constant flow rate of 3 ml / min maintained by a Masterflex digital peristaltic pump (Cole-Parmer). Immediately after the buffer started passing through the liver, the IVC was cut, allowing blood and perfusate from the liver to flow out through the IVC. Immediately after this treatment, the liver became decolorized and pale in color.

[0162] After passage of HBSS, HBSS (1 mM CaCl 2 25 ml of collagenase (Roche) solution (1 mg / ml) in HBSS (containing 1 mM CaCl) was passed through the liver at a constant flow rate of 2 ml / min. After this digestion, the flow was stopped, the cannula was removed, and the pale, mushy liver lobes were gently excised from the body. The gallbladder was removed from the isolated liver. The digested liver tissue pieces were then resuspended in HBSS (containing 1 mM CaCl 2 The cells were minced on a 10 cm culture plate in 100 μl of PBS (containing 100 μl of PBS). The resulting suspension was then passed through a 100 μl cell strainer (SPL) such that the hepatocytes passed through to the filtrate, while the cell clumps and undigested tissue were retained. The filtrate was centrifuged at 50 g for 2 min at 4° C. The supernatant was discarded and the cell pellet was carefully resuspended in DMEM. The resulting suspension was centrifuged at 50 g for 2 min at 4° C. The supernatant was discarded and the cell pellet was carefully resuspended in the required volume of DMEM for plating.

[0163] Hepatocytes were plated as per the experimental protocol and incubated at 37°C with 5% CO 2 After plating, the cells were washed once with HBSS and DMEM for 6-7 hours, and the attached hepatocytes were maintained and subjected to necessary treatment.

[0164] Figure 2 illustrates the ATGL protein status in mouse primary hepatocytes after compound treatment.The level of ATGL was found to increase in a dose-dependent manner.This provides more important and direct evidence of the effectiveness of compound.Compound 62 increases the level of ATGL in primary mouse hepatocytes, as evidenced by the increase in the intensity of the corresponding band in Western blot analysis compared to control.

[0165] (Example 34) ATGL ubiquitination status upon compound treatment assessed by immunoprecipitation assay COP1 is an E3 ubiquitin ligase, and ATGL is one of its targets, which is ubiquitinated and eventually degraded via the proteasome-mediated pathway. Therefore, it was indeed expected that molecules that inhibit COP1 by targeting the VP motif of ATGL would result in a reduction in the ubiquitination level of ATGL. The compounds of the present invention showed a corresponding increase in ATGL protein level with a reduction in lipid droplet number, while gene expression remained unchanged. However, it was of utmost importance to examine the changes in the ubiquitination level of ATGL that occurred upon treatment with the compounds.

[0166] For this purpose, immunoprecipitation assays were performed in which HepG2 cells overexpressing myc-ATGL were transfected with HA-ubiquitin and treated with 5 μM of compounds 5, 62, and 94. MG-132, a proteasome inhibitor, was added 4 h before cells were harvested. Immune complexes were pulled down with anti-myc antibodies and immunoblotted with anti-HA antibodies. The smears obtained in the blots reflect the ubiquitination status of ATGL.

[0167] FIG. 3 (A-C) illustrates the results of immunoprecipitation assays to examine the ubiquitination status of ATGL and COP1 after treatment with compounds 62, 94, and 5. For compounds 5 and 62, a clear reduction in the ubiquitination smear was observed upon treatment compared to the control. However, this reduction was not seen in cells treated with compound 94, indicating that compounds 5 and 62 may be more potent in inhibiting COP1 by blocking ATGL ubiquitination to some extent compared to compound 94. Compounds 62 and 5 were effective in reducing ATGL ubiquitination by COP1 in HepG2 cells, as evidenced by the reduced intensity of the polyubiquitin smear, whereas compound 94 did not exert such an effect.

[0168] (Example 35) Confocal microscopy HepG2 cells were plated on confocal dishes (SPL, Genetix Biotech Asia Pvt. Ltd.). Cells were allowed to attach and divide for 16 hours. Compound 62 was dissolved in DMSO and added to the cells at concentrations of 100 nM and 500 nM for dose-dependent assay. 250 μM oleic acid was used for transfection. BSA (Sigma Aldrich) was used as a negative control. After 24 hours of treatment, the medium was decanted from the cells and washed with 1×PBS solution to remove any debris. 200 μl of staining solution containing 200 ng / ml BODIPY (Invitrogen) and 25 μg / ml HOECKST342 (Invitrogen) was added to the cells and incubated at 37° C. in the dark for 30 minutes. Cells were washed three times with 1×PBS to remove excess stain. Cells were visualized using FLUOVIEW FV10i (Olympus).

[0169] Figure 4 illustrates the screening image of compound 62 in HepG2 cells using confocal microscopy. The green foci in the cells represent lipid droplets. Therefore, an increase or decrease in the number of green foci indicated the corresponding status of lipid droplets in the cells. The introduction of oleate increased lipid droplets, but treatment with compound 62 caused a decrease in the number of lipid droplets when oleate was introduced.

[0170] (Example 36) Western blot HepG2 cells were treated with compounds 5, 8, 11, 59, 62, 65, 91, 94 at 5 μM for 24 hours. After removing the medium from the cells, the wells were washed twice with 1×PBS to remove any remaining medium. The cells were then lysed in a lysis buffer containing 50 mM Tris-HCl (pH 7.4), 100 mM NaCl, 1 mM EDTA, 1 mM EGTA, 1% Triton X-100 and a protease inhibitor cocktail (Millipore, Billerica, MA, USA). Following centrifugation at 20,000 g for 20 min, the protein solution was extracted from the cells. Protein was assessed using the Bradford assay. Bradford reagent (BioRad) was diluted in a 1:4 ratio with double distilled water. 2 μl of protein sample was added to 100 μl of reagent and the absorbance was measured at 595 nm. 30 μg of protein was diluted in the lysis buffer. 1x loading buffer diluted from a 5x stock containing 250mM Tris-HCl (pH 6.8), 10% SDS, 50% glycerol, 0.1% bromophenol blue and 10% β-mercaptoethanol was added, and the protein samples were then heated to 95°C for 10 min, cooled and centrifuged at 12,000g for 2 min before loading.

[0171] For Western blot, proteins were resolved in 10% SDS PAGE (discontinuous buffer system). Gels were run at 80V for approximately 2 hours using 1x running buffer containing SDS, Tris base and glycine. Transfer was performed using a PVDF membrane with a pore size of 0.45 μm (Millipore). 1x transfer buffer containing Tris base, glycine and 20% methanol was used for wet transfer. Transfer was performed at 90V for 3 hours. Following transfer, the PVDF membrane containing the proteins was washed with 1x PBS and 1x PBST containing 1% Tween 20 (Sigma Aldrich). The membrane was then incubated in 5% skim milk powder at room temperature for 1 hour to block non-specific sites. After multiple washes with 1x PBST to wash away any remaining blocking buffer, the required primary antibody (COP1 [Bethyl Laboratories], ATGL [Cell Signalling Technology] or Actin [Cell Signalling Technology]) prepared in 1x PBST, 1% bovine serum albumin and 0.04% sodium azide was added to the membrane and incubated overnight at 4°C. The next day, the membrane was again washed multiple times with 1x PBST to remove any unbound primary antibody. The membrane was then incubated with goat anti-rabbit secondary antibody (Genei) for 1 hour at room temperature and again washed multiple times with 1x PBST. The membrane was then developed using Clarity™ ECL Western Blotting Substrate (BioRad) and viewed on a ChemiDoc (BioRad).

[0172] Figure 5 shows a comparison of the induction of ATGL by treatment with compounds 5, 8, 11, 59, 62, 65, 91, 94 at 5 μM. All compounds have the ability to induce ATGL and stabilize COP1 by modulating its activity.

[0173] (Example 37) In vivo study of compounds to investigate the expression levels of ATGL and COP1 in the liver Healthy male C57BL / 6 mice aged 6-8 weeks (average body weight 28g) were recruited for the study. They were then divided into two groups with 3 mice per group (control, compound 62). Mice were orally administered 80mg / kg of compound 62. The compound was dissolved in 10% EtOH, 40% PEG, 20% PG, 30% NaCl solution (0.9%). The control group received only the solvent in which the compound was dissolved. Mice were sacrificed on the 9th day after one feeding, and the excised liver tissue and adipocyte portions were homogenized in lysis buffer containing a protease inhibitor cocktail. The homogenate was centrifuged at 20,000g for 30 minutes, followed by collection of the supernatant containing the protein lysate. The lysate was then diluted accordingly and protein evaluation was performed by Bradford assay. This was followed by Western blot to examine the levels of ATGL and COP1. Actin was used as a loading control.

[0174] Figure 6 illustrates the results of an in vivo study measuring ATGL and COP1 levels in mice. Western blots from liver and adipose tissue lysates revealed that compound 62 was effective in increasing ATGL and COP1 levels.

[0175] (Example 38) In vivo study of HFD-induced fatty liver disease model using compounds to investigate the expression levels of ATGL and COP1 in the liver A preclinical treatment model was developed by feeding C57BL / 6 mice with a 60% kcal high-fat diet (HFD) for 12 weeks and orally feeding them with compound 62 at a dose of 80 mg / kg body weight for the final 4 weeks.

[0176] Figure 7 illustrates that a substantial depletion of hepatic lipid droplets (Figure 7A) and amelioration of liver fibrogenesis (Figure 7B) in mice was observed in total liver weight (Figure 7B), body weight (Figure 7C), serum cholesterol levels (Figure 7D) and liver triglyceride content (Figure 7E).

[0177] (Example 39) Determination of EC50

[0178] [Table 4]

[0179] (Example 40) Fold change of ATGL (induced by compound treatment) Densitometry analysis was performed using ImageJ to calculate the fold change of ATGL relative to control. Values ​​were normalized to actin.

[0180] [Table 5A]

[0181] [Table 5B]

[0182] (Example 41) Lipophilicity Assay Assay procedure 1.56g NaH 2 PO 4 2H 2 O was dissolved in 0.5 L of water in a 1 L beaker. The volume was made up to 1 L after adjusting the pH to 7.4 using NaOH solution. Equal volumes of sodium phosphate buffer (10 mM, pH 7.4) and n-octanol were added to the separatory funnel and mixed thoroughly by shaking and up and down the funnel several times. The two layers were allowed to separate overnight and then dispensed into two different glass bottles. A 10 mM stock solution was prepared in 100% DMSO and stored at 4°C. 495 μL of the organic phase (1-octanol) was added to each well of a 2 mL deep well plate, followed by 495 μL of buffer and 10 μL of the test substance. The plate was incubated at room temperature for 3 hours on a plate shaker at 500 rpm. After incubation, the samples were equilibrated for 20 minutes and then centrifuged at 4000 rpm for 30 minutes to allow complete phase separation and analyzed by LC-UV. Log D = Log(area of ​​octanol / area of ​​buffer solution)

[0183] [Table 6A]

[0184] [Table 6B]

[0185] (Example 42) Plasma stability assay Assay procedure 1 mM test compound stocks were prepared from an initial 10 mM stock solution of compound by diluting 10 μL of 10 mM stock with 90 μL of DMSO. 10 μL of 1 mM stock was then diluted with 90 μL of DMSO to obtain a concentration of 100 μM. Frozen plasma was thawed at room temperature and centrifuged at 1400 rpm for 15 minutes at 4° C. Approximately 90% of the clear supernatant fraction was transferred to a separate tube and used for the assay. A final working stock of 1 μM was prepared by diluting 3 μL of 100 μM with 297 μL of plasma. Plasma containing test compound was incubated in a shaker at 500 rpm at 37° C. for 120 minutes. 50 μL aliquots of sample were precipitated with 150 μL of acetonitrile containing internal standard for 0, 15, 30, 60 and 120 minutes and centrifuged at 4000 rpm for 20 minutes at 4° C. 120 μL of the supernatant was diluted with 120 μL of water and analyzed by LC-MS / MS.

[0186] [Table 7]

[0187] (Example 43) In vitro assessment of metabolic stability using human liver microsomes (human liver microsome stability HLM) Assay procedure A 1 mM stock solution of test compound was prepared in DMSO and diluted with acetonitrile:water (1:1) to give a working concentration of 100 μM. 2 HPO 4 (1.398g) and KH 2 PO 4 (0.27 g) to give a potassium phosphate buffer solution with a final pH of 7.4. A microsome suspension of 3.333 mg / mL was prepared by diluting 499.95 μL of 20 mg / mL microsome stock to 2500.05 μL with buffer. 532.5 μL of 16 mM NADPH stock was added to 2467.5 μL potassium phosphate buffer to give a working stock of 2.84 mM. 75 μL of 3.333 mg / mL liver microsome working stock and 85 μL of buffer were added to 2.5 μL of test compound (100 μM). The above mixture was preincubated at 37° C. for 15 minutes. After preincubation, 32.5 μL of the mixture was added to 17.5 μL of buffer, which was then incubated at 37° C. for 60 minutes [60 minutes without cofactor (NADPH)]. 16.25 μL of pre-incubated mixture and 8.75 μL of cofactor were added to 150 μL of acetonitrile containing internal standard [0 min sample]. 62 μL of cofactor was added to the remaining pre-incubated mixture [incubation mixture]. 25 μL of incubation mixture was precipitated with 150 μL of acetonitrile containing internal standard without cofactor for 0, 5, 15, 30, 60 min and 60 min, vortexed and centrifuged at 4000 rpm, 4° C. for 20 min. 120 μL of supernatant was diluted with 120 μL of water and analyzed by LC-MS / MS [sample preparation].

[0188] [Table 8A]

[0189] Example 4 In vitro assessment of kinetic solubility The PBS sachet was dissolved in 0.9 L of Milli Q water and the pH was adjusted to 7.4. The final volume was brought up to 1 L with water and stored at ambient temperature 21-25 °C. Stock solutions (50 mM) of each compound (5, 11, 59, 62, 65, 94) were prepared in DMSO and stored at 4-8 °C. 4 μL from the 50 mM stock solution was added to a deep-well plate containing 396 μL of phosphate buffer, pH 7.4, mixed and incubated at room temperature at 300 rpm with constant mixing for 24 h. The plate was well sealed during the incubation period. After incubation, the samples were centrifuged at 4000 rpm for 20 min. The supernatant was analyzed by HPLC-UV. The DMSO content in the samples was 1.0%. The final concentration of the compounds in the deep-well plate was 500 μM.

[0190] [Table 9]

[0191] Although the subject matter has been described in considerable detail with reference to specific examples and implementations thereof, other implementations are possible.

Claims

1. A compound having structure I or a salt thereof: 【Chemistry 1】 (In the formula, R 1 is independent, 【Chemistry 2】 is selected from the group consisting of R 2 is independent, 【Chemistry 3】 is selected from the group consisting of R 3 is independent, 【Chemistry 4】 (selected from the group consisting of:

2. 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (5), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-methyl-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (8), 1-(3-acetylphenyl)-3-(1-isopropyl-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (11), 1-(3-acetylphenyl)-3-(1-cyclohexyl-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (14), 1-(3-acetylphenyl)-3-(1-cyclopentyl-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (17), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(1-methylpiperidin-4-yl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (20), 1-(3-acetylphenyl)-3-(1-(cyclopropylmethyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (23), 1-(3-acetylphenyl)-3-(1-benzyl-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (26), 1-(3-acetylphenyl)-3-(1-(4-cyanobenzyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (29), 1-(3-acetylphenyl)-3-(1-(4-cyano-2-fluorobenzyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (32), 1-(3-acetylphenyl)-3-(1-(4-fluorobenzyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (35), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(4-nitrobenzyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (38), 1-(3-acetylphenyl)-3-(1-(4-bromobenzyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (41), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(4-(trifluoromethyl)benzyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (44), 1-(3-acetylphenyl)-3-(1-(4-methoxybenzyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (47), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(pyridin-3-ylmethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (50), Methyl 3-((6-(3-(3-acetylphenyl)ureido)-3-(2-methoxyethyl)-2,4-dioxo-3,4-dihydroquinazolin-1(2H)-yl)methyl)benzoate (53), Methyl 4-((6-(3-(3-acetylphenyl)ureido)-3-(2-methoxyethyl)-2,4-dioxo-3,4-dihydroquinazolin-1(2H)-yl)methyl)benzoate (56), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(pyrrolidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (59), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (62), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(2-morpholinoethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (65), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(2-(4-methylpiperazin-1-yl)ethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (68), 1-(1-(2-(1H-imidazol-1-yl)ethyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-acetylphenyl)urea (72), tert-Butyl 4-(2-(6-(3-(3-acetylphenyl)ureido)-3-(2-methoxyethyl)-2,4-dioxo-3,4-dihydroquinazolin-1(2H)-yl)ethyl)piperazine-1-carboxylate (75), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperazin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (76), 1-(1-(2-(1H-pyrrol-1-yl)ethyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-acetylphenyl)urea (79), 1-(1-(2-(1H-indol-1-yl)ethyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-acetylphenyl)urea (82), 1-(1-(2-(1H-benzo[d]imidazol-1-yl)ethyl)-3-(2-methoxyethyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-acetylphenyl)urea (85), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(3-(pyrrolidin-1-yl)propyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (88), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(3-(piperidin-1-yl)propyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (91), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(3-morpholinopropyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (94), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-1-(3-(4-methylpiperazin-1-yl)propyl)-2,4-dioxo-1,2,3,4-tetrahydroquinazolin-6-yl)urea (97), 1-(3-acetylphenyl)-3-(3-(3-methoxypropyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (102), 1-(3-acetylphenyl)-3-(3-(2-ethoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (107), 1-(3-acetylphenyl)-3-(3-ethyl-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (112), Ethyl 2-(6-(3-(3-acetylphenyl)ureido)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2-dihydroquinazolin-3(4H)-yl)acetate (117), 1-(3-acetylphenyl)-3-(3-(3-methoxyphenyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (122), 1-(3-acetylphenyl)-3-(3-(2-methoxyphenyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (127), 1-(3-acetylphenyl)-3-(3-(2-(dimethylamino)ethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (132), 1-(3-acetylphenyl)-3-(2,4-dioxo-1,3-bis(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (137), 1-(3-acetylphenyl)-3-(3-(2-aminoethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (142), 1-(3-acetylphenyl)-3-(3-(2-(4-methylpiperazin-1-yl)ethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (147), 1-(3-acetylphenyl)-3-(3-(3-morpholinopropyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (152), 1-(3-acetylphenyl)-3-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (157), 1-(3-acetylphenyl)-3-(3-(1-methylpiperidin-4-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (162), 1-(3-acetylphenyl)-3-(2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-3-(pyridin-4-yl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (167), 1-(3-acetylphenyl)-3-(2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-3-(pyridin-3-yl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (172), 1-(3-acetylphenyl)-3-(2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-3-(pyridin-2-yl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (177), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-methoxyphenyl)urea (178), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(2-methoxyphenyl)urea (179), 1-(3-(1-hydroxyethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (180), 1-(3-ethylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (181), Methyl 3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)benzoate (182), 3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)benzoic acid (183), 3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)-N,N-dimethylbenzamide (184), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(pyrrolidine-1-carbonyl)phenyl)urea (185), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(morpholine-4-carbonyl)phenyl)urea (186), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-phenylurea (187), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(methylamino)phenyl)urea (188), N-(3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)phenyl)acetamide (189), N-(3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)phenyl)-N-methylacetamide (190), N-benzyl-N-(3-(3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)phenyl)acetamide (191), 1-(3-acetylphenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-1-methylurea (192), 1-(3-acetylphenyl)-1-hydroxy-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (193), (Z)-1-(3-(1-(hydroxyimino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (194), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(2,2,2-trifluoroacetyl)phenyl)urea (195), 1-(3-(1-aminoethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (196), 1-(3-(1-(dimethylamino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (197), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(1-(phenylamino)ethyl)phenyl)urea (199), 1-(3-(1-((2-fluorophenyl)amino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (201), 1-(3-(1-((4-fluorophenyl)amino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (203), 1-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(1-((4-methoxyphenyl)amino)ethyl)phenyl)urea (205), 1-(3-(1-((4-cyanophenyl)amino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (207), 1-(3-(1-(cyclohexylamino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (209), 1-(3-(1-(cyclopentylamino)ethyl)phenyl)-3-(3-(2-methoxyethyl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (211), 6-((5-acetyl-1H-benzo[d]imidazol-2-yl)amino)-3-(2-methoxyethyl)-1-(2-(piperidin-1-yl)ethyl)quinazoline-2,4(1H,3H)-dione (216), 6-((2-((3-acetylphenyl)amino)-3,4-dioxocyclobut-1-en-1-yl)amino)-3-(2-methoxyethyl)-1-(2-(piperidin-1-yl)ethyl)quinazoline-2,4(1H,3H)-dione (217), (R)-1-(3-acetylphenyl)-3-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (222), (S)-1-(3-acetylphenyl)-3-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)urea (226), (R)-3-(3-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)-N,N-dimethylbenzamide (228), (S)-3-(3-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)ureido)-N,N-dimethylbenzamide (229), (R)-1-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(2,2,2-trifluoroacetyl)phenyl)urea (230), (S)-1-(3-(1-methoxypropan-2-yl)-2,4-dioxo-1-(2-(piperidin-1-yl)ethyl)-1,2,3,4-tetrahydroquinazolin-6-yl)-3-(3-(2,2,2-trifluoroacetyl)phenyl)urea (231) 2. The compound having structure I of claim 1, selected from the group consisting of:

3. A process for the preparation of a compound having structure I according to claim 1, comprising: (i) 2-Amino-5-nitrobenzoic acid (compound 1) was reacted with 2-methoxyethylamine, methoxypropylamine, 2-ethoxyethylamine, 2M ethylamine in THF, glycine ethyl ester hydrochloride, m-anisidine, o-anisidine, N,N-dimethylethylenediamine, 1-(2-aminoethyl)piperidine, 4-(2-aminoethyl)morpholine, 1-(2-aminoethyl)-4-methylpiperazine, 3-(4-morpholinyl)propane, 1-(2-aminoethyl)-4-methylpiperazine ... with an aliphatic or aromatic amine selected from the group consisting of 2, 98, 103, 108, 113, 118, 123, 128, 133, 138, 143, 148, 153, 158, 163, 168, 173, 218, 223; (ii) cyclizing the compound selected from the group consisting of 2, 98, 103, 108, 113, 118, 123, 128, 133, 138, 143, 148, 153, 158, 163, 168, 173, 218, 223 obtained in step (i) using a cyclizing agent CDI in DMF as a solvent at 100° C. for 12-16 hours to obtain a compound selected from the group consisting of 3, 99, 104, 109, 114, 119, 124, 129, 134, 139, 144, 149, 154, 159, 164, 169, 174, 219, 224; (iii) reacting compound 3 obtained in step (ii) with methyl iodide and dry DMF at 0° C. for 12 hours to obtain compound 6; (iv) Compound 3, 99, 104, 109, 114, 119, 124, 129, 134, 139, 144, 149, 154, 159, 164, 169, 174, 219, 224 obtained in step (ii) is reacted with 2-iodopropane, bromocyclohexane, bromocyclopentane, 4-bromo-1-methylpiperidine, (bromomethyl)cyclopropane, benzyl bromide, 4-(bromomethyl)benzonitrile, 4-(bromomethyl)-3-fluorobenzonitrile, 4-fluorobenzyl bromide, 4-nitrobenzyl bromide, 4-bromobenzyl bromide, 4-methanol a suitable alkyl chloride selected from the group consisting of methyl 3-(bromomethyl)pyridine hydrobromide, methyl 3-(bromomethyl)benzoate, methyl 4-(bromomethyl)benzoate, 1-(2-chloroethyl)pyrrolidine, 1-(2-chloroethyl)piperidine, 4-(2-chloroethyl)morpholine, 1-(2-chloroethyl)-4-methylpiperazine, 1-(3-chloropropyl)pyrrolidine, 1-(3-chloropropyl)piperidine, 4-(3-chloropropyl)morpholine, and 1-(3-chloropropyl)-4-methylpiperazine; and K 2 CO 3 and dry DMF at 120° C. for 12 hours to obtain a compound selected from the group consisting of 9, 12, 18, 21, 24, 27, 30, 33, 36, 39, 42, 45, 48, 51, 54, 57, 60, 63, 66, 86, 89, 92, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 220, and 225; (v) The compound 3 obtained in step (ii) is reacted with chlorobromoethane and K 2 CO 3 and dry DMF at 120° C. for 12 hours to obtain compound 69; (vi) Reacting compound 69 obtained in step (v) with a suitable amine selected from the group consisting of imidazole, N-Boc piperazine, pyrrole, indole, and benzimidazole, K 2 CO 3 and dry DMF at 120° C. for 12 hours to obtain a compound selected from the group consisting of 70, 73, 77, 80, and 83; (vii) The compound selected from the group consisting of 3, 6, 9, 12, 18, 21, 24, 27, 30, 33, 36, 39, 42, 45, 48, 51, 54, 57, 60, 63, 66, 70, 73, 77, 80, 83, 86, 89, 92, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 220, and 225 obtained in steps (ii), (iii), and (iv) is subjected to a palladium-carbon (5% or 10%, wet) treatment at room temperature for 2 to 5 hours under H 2 to obtain an amine compound selected from the group consisting of 4, 7, 10, 13, 19, 22, 25, 28, 31, 34, 37, 40, 43, 46, 49, 52, 55, 58, 61, 64, 67, 71, 74, 78, 81, 84, 87, 90, 93, 96, 101, 106, 111, 116, 121, 126, 131, 136, 141, 146, 151, 156, 161, 166, 171, 176, 221, 226; (viii) Compounds 4, 7, 10, 13, 19, 22, 25, 28, 31, 34, 37, 40, 43, 46, 49, 52, 55, 58, 61, 64, 67, 71, 74, 78, 81, 84, 87, 90, 93, 96, 101, 106, 111, 116, 121, 126, 131, 136, 141, 146, 151, 156, 161, 166, 171, 176, 221, 226 obtained in step (vii) are treated with 4-nitrophenyl chloroformate in the presence of TEA as a base, followed by incubation at room temperature for 3-8 hours. with an amine selected from the group consisting of 3'-aminoacetophenone in dry THF for 20 min to obtain a compound having structure I selected from the group consisting of 5, 8, 11, 14, 20, 23, 26, 29, 32, 35, 38, 41, 44, 47, 50, 53, 56, 59, 62, 65, 68, 72, 75, 79, 82, 85, 88, 91, 94, 97, 102, 107, 112, 117, 122, 127, 132, 137, 142, 147, 152, 157, 162, 167, 172, 177, 222, 227; (ix) Compounds 61, 221, 226 obtained in step (vii) are treated with 4-nitrophenyl chloroformate in the presence of TEA as a base, followed by the treatment with m-anisidine, o-anisidine, 1-(3-aminophenyl)ethanol, 3-ethylaniline, methyl 3-aminobenzoate, 3-amino-N,N-dimethylbenzamide, (3-aminophenyl)(pyrrolidin-1-yl)methanone, (3-aminophenyl)(morpholino)methanone, aniline, N1-methylbenzene-1,3-diamine, N-(3-aminophenyl)acetate, etc. in dry THF at room temperature for 3-8 hours. 178, 179, 180, 181, 182, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 195, 228, 229, 230, 231; (x) Compound 182 obtained in step (ix) was treated with LiOH·H in THF in a ratio of (3:2:1) at room temperature for 12 hours. 2 O, methanol and H 2 with O to obtain compound 183 having structure I; (xi) treating compound 62 obtained in step (viii) with hydroxylamine hydrochloride in EtOH at room temperature for 12 hours to obtain compound 194 having structure I; (xii) treating compound 75 obtained in step (viii) with trifluoroacetic acid in DCM at room temperature for 8 hours to obtain compound 76 having structure I; (xiii) Compound 62 obtained in step (viii) is reacted with NH 3 / methanol, followed by NaBH in methanol 4 to obtain compound 196 having structure I; (xiv) treating compound 196 obtained in step (xiii) with HCOOH and formaldehyde in the presence of concentrated HCl at 100° C. for 8 hours to obtain compound 197 having structure I; (xv) treating compound 62 obtained in step (viii) with various aromatic amines (aniline, 2-fluoroaniline, 4-fluoroaniline, 4-methoxyaniline, 4-aminobenzonitrile, cyclohexylamine, cyclopentylamine) in the presence of p-toluenesulfonic acid (PTSA) in EtOH at 100° C. to obtain compounds having structure I selected from the group consisting of 196, 200, 202, 204, 206, 208, 210; (xvi) Compounds 196, 200, 202, 204, 206, 208, and 210 obtained in step (xv) are dissolved in sodium cyanoborohydride (NaCNBH 3 207, 209, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 2 (xvii) treating commercially available compound 212 with 2-methoxyethylamine in HATU and DMF at room temperature for 2 hours to obtain compound 213 having structure I; (xviii) treating the compound obtained in step (xvii) with CDI in DMF at 100° C. for 12 hours to obtain compound 214 having structure I; (xix) The compound 214 obtained in step (xviii) is treated with 1-(2-chloroethyl)piperidine and K at 120° C. for 12 hours. 2 CO 3 and dry DMF to obtain compound 215; (xx) The compound 215 obtained in step (xix) was reacted with 1-(2-amino-1H-benzo[d]imidazol-5-yl)ethenone and Pd as a catalyst at 100° C. for 12 hours. 2 (dba) 3 and X-Phos and base K as ligands 2 CO 3 to obtain compound 216; (xxi) reacting compound 61 obtained in step (vii) with 3,4-diethoxy-3-cyclobutene-1,2-dione and p-toluenesulfonic acid (PTSA) in EtOH and 3-aminoacetophenone at 80° C. for 12 hours to obtain compound 217. The method includes:

4. 2. The compound of claim 1 for use in treating diseases and disorders associated with the regulation of COP1 enzyme through its stabilization or the regulation of ATGL.

5. 13. A compound according to claim 1 for use in reducing the level of triglycerides in liver cells.

6. 10. The compound of claim 1 for use in the treatment of a disease selected from non-alcoholic fatty liver disease (NAFLD) or non-alcoholic steatohepatitis (NASH).

7. 13. A composition comprising a compound having structure I of claim 1 together with a pharma- ceutically acceptable excipient.

8. 2. A method for modulating the COP1 enzyme through stabilization thereof with a compound having structure I of claim 1.

9. A method of increasing the level of ATGL by a compound having structure I of claim 1.

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