Triazolo compounds for treatment of bile acid diarrhea and autoimmune enteropathy

Abstract

The present invention relates to a compound of Formula (I), in particular, the compound of Formula (II) or an isotopic variant, a pharmaceutically acceptable salt or solvate thereof for use in a method of treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof.

Description

[0001] I75301WO BOEHMERT & BOEHMERT

[0002] Triazolo compounds for treatment of bile acid diarrhea and autoimmune enteropathy

[0003] SUMMARY OF THE INVENTION

[0004] The present invention relates to a compound of Formula (I), in particular, a compound of Formula (II) or an isotopic variant, a pharmaceutically acceptable salt or solvate thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof.

[0005] BACKGROUND

[0006] Bile acid diarrhea (BAD) is caused by dysregulated recycling of bile acids (BAs) within the enterohepatic cycle. Either excessive biosynthesis / secretion of bile acids by the liver / gallbladder or impaired re-absorption of bile acids in the terminal ileum leads to the delivery of abnormally high levels of unabsorbed BAs to the lower Gl tract / colon. This stimulates water and electrolyte secretion, increases gastrointestinal motility, damages mucosa and increases intestinal permeability, next to inducing alteration in the stool microbiome composition, which is associated with a reduced conversion of primary to secondary BAs. Consequently, patients suffer from severe chronic diarrhea, frequent and urgent bowel movements, fecal incontinence, as well as abdominal bloating or swelling [Gastroenterol. Hepatol. (NY) 2023;19:520; Clin. Gastroenterol. Hepatol. 2020;18:24; Int. J.

[0007] Mol. Sci. 2024;25:1544],

[0008] A number of conditions or diseases lead to an abnormal amount of bile acid entering the large intestines. Such conditions include ileal resection, ileal inflammation (e.g. in Crohn's disease, a type of inflammatory bowel disease), idiopathic bile acid malabsorption or overproduction, cholecystectomy (gallbladder removal), vagotomy, small intestinal bacterial overgrowth, radiation enteropathy, celiac disease and chronic pancreatitis.

[0009] BAD is divided into three types: Type I occurs in patients with ileal disease (e.g. Crohn's disease) or resection. Type II, also referred to as idiopathic or primary BAD, occurs spontaneously, and Type III is found in patients with gastrointestinal conditions other than ileal disease (e.g. celiac disease, cholecystectomy). Patient with hypertriglyceridemia and patients on metformin treatment may also develop BAD.

[0010] Mucosal damage and increased intestinal permeability (leaky gut) in BAD: BAs are detergents and play an important role in digestion and absorption of dietary lipids. Moreover, bile acids have signaling / enteroendocrine hormone functions and interact with bile acid receptors or gut microbiota, which are critical in homeostasis and maintenance of the intestinal barrier [Cells 2023;12:1888; Physiol. Rev. 2018;98:1983],

[0011] Tight junctions (TJs) formed between neighboring intestinal epithelial cells are essential for the establishment and maintenance of an intestinal barrier. To control the paracellular movement of nutrients, fluid, electrolytes and antimicrobial contents, TJs and its constituents the TJ proteins are highly regulated and represent dynamic structures / protein complexes. BAD Patients exhibit higher intestinal permeability [Gastroenterology 2022;162:1343], Ascending colon biopsies from patients with BAD showed an upregulation in the TJ gene CLDN2 [Gut 2023;72:54], encoding the TJ protein claudin-2, which is implicated in increased mucosal permeability and contributing to leak-flux diarrhea in many intestinal diseases [Tissue Barriers 2015;3:e977176],

[0012] Bile acids have been shown to weaken the intestinal barrier, increase the epithelial permeability and bacterial uptake in ex vivo cultures of human colon biopsies [Scand. J. Gastroenterol. 2007;42:1167], In line with that, treatment of monolayers of human intestinal epithelial cells (Caco-2) with bile acids results in increased transepithelial permeability via redistribution of the TJ protein occludin [Am. J. Physiol. Gastrointest. Liver Physiol. 2008;294:G906] and generation of reactive oxygen species [Free Radio. Biol. Med. 2005;39:769], A role of bile acids in regulation TJ permeability has been also demonstrated in high-fat diet fed mouse model, where intestinal hyperpermeability and decreased expression of TJ proteins (ZO-2 and JAM-A) are associated to increased bile acids in the colon [J. FoodSci. 2016;81:H216]. In another high-fat diet mouse model high levels of intestinal and circulating bile acids were shown to induce intestinal mucosal barrier damage by reducing the number of functional ileal intestinal stem cells (ISCs) and decreasing the TJ protein occludin in the ileum of these mice [Biochem. Biophys. Res. Commun. 2020;529:289], This suggests that dysregulated BA recycling also impacts intestinal barrier function by impairing epithelial regeneration and repair via ISCs (next to impact on TJ proteins).

[0013] Autoimmune enteropathy (AIE) is one of the differential diagnoses of chronic intractable diarrhea, malnutrition and weight loss. This mixed-type diarrhea with protein-losing enteropathy usually involves the small intestine but gastric and colonic involvements are not rare [Adv. Dig. Med. 2022;9:75], Small intestinal biopsy is characterized by villous atrophy of variable severity, crypt hyperplasia, and infiltration of mononuclear cell. It is usually diagnosed by presence of clinical features, circulating auto antibodies against enterocytes, presence of histological characters and executions of other causes of villous atrophy. Majority of AIE patients respond to corticosteroid therapy (e.g. budesonide) but patients who are not responding to steroid are treated by immunomodulators (e.g. azathioprine, cyclophosphamide, tacrolimus, cyclosporine, infliximab, ustekinumab, vedolizumab or abatacept). Addressing the tissue damage that occurs is of paramount importance. While immunosuppressive therapy plays a crucial role, a multifaceted approach that encompasses both the mitigation of tissue damage, strengthening the epithelial barrier in the gut and the modulation of the host's biological response is essential for achieving optimal therapeutic outcomes in patients with BAD or AIE.

[0014] Current therapy for primary BAD, or BAD caused by ileal resection or Crohn's disease, includes bile acid binders such as cholestyramine (A3384; Albiero), colestipol and colesevelam. Farnesoid X receptor (FXR) agonists such as obeticholic acid or UN452 as well as cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel inhibitor or fibroblast growth factor 19 (FGF19) analogues such as aldafermin are in development. For IBS-D, in which BAD is likely involved in pathogenesis in more than one-third of patients, FDA-approved therapies include the 5-HT3 antagonist alosetron, the mixed P opioid receptor agonist eluxadoline and the broad-spectrum gut-specific antibiotic rifaximin. Other commonly used therapies for IBS- D include loperamide, bile acid sequestrants, antispasmodics and tricyclic antidepressants. Some of these therapies are associated with significant side effects such as ischemic colitis with alosetron and pancreatitis with eluxadoline.

[0015] An unmet need for improved and alternative therapeutics for BAD or AIE persists.

[0016] DETAILED DESCRIPTION OF THE INVENTION

[0017] In one aspect, the present invention relates to a compound of Formula (I) Formula (I), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof, wherein:

[0018] R1is selected from the group consisting of hydrogen, carboxyl, cyano, fluorine, chlorine, methyl, isopropyl, t-butyl, trifluoromethyl, trifluoromethoxy, cyclopropylmethoxy, 1,1- difluoro-2-methylpropyl, l,l-difluoro-2,2-dimethylpropyl, 1-methyl-l-cyclobutyl, methoxycarbonyl, ethoxycarbonyl, isopropoxycarbonyl, 1-hydroxy-l-methylethyl, azetidine- 1-carbonyl, 3-methyloxetan-3-yl, 4,5-dihydrooxazol-2-yl and cyclopropylcarbonyl;

[0019] R2is hydrogen or methyl;

[0020] A1is =N- or =CH-;

[0021] X is benzene, pyridine or cyclohexane;

[0022] J is any ring selected from the group

[0023] Y is selected from the group consisting of phenyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl, piperidinyl, azetidinyl, tetrahydropyranyl, morpholinyl and tetrahydropyridinyl, optionally substituted with 1, 2 or 3 groups selected from Y1, or alternatively Y is amino, optionally substituted with 1 or 2 groups selected from Y1;

[0024] Y1is selected from the group consisting of hydroxy, cyano, fluorine, amino, methyl, ethyl, isopropyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl, wherein amino is optionally substituted with 1 or 2 groups selected from Y2and methyl, ethyl, isopropyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl, are optionally substituted with 1,

[0025] 2 or 3 groups selected from Y2;

[0026] Y2is selected from the group consisting of hydroxyl, fluorine, amino, methyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl, wherein amino is optionally substituted with 1 or 2 groups selected from Y3and methyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl, is optionally substituted with 1, 2 or 3 groups selected from Y3,

[0027] Y3is hydroxyl, fluorine, cyano, methyl or methoxy, wherein methyl or methoxy are optionally substituted with fluorine.

[0028] In some embodiments, the compound of Formula (I) is selected from the following group:

[0029] or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof.

[0030] In some embodiments, the compound of Formula (II) is Formula (II), or an isotopic variant, a pharmaceutically acceptable salt or solvate thereof.

[0031] In some embodiments, the compound of Formula (II) is

[0032] The compound of Formula (I), in particular the compound of Formula (II), targets sirtuin 6 (SIRT6), a member of a family of proteins related to the founding member of the family - the silent information regulator 2 protein (Sir2p) of Saccharomyces cerevisiae, a nicotinamide adenine dinucleotide (NAD+)-dependent histone deacetylase (HDAC) regulating chromatin silencing. Mammals contain at least seven sirtuin homologues numbered SIRT1 to SIRT7, characterised by significant sequence homology, particularly within their conserved NAD+- binding domains [Biochem. Biophys. Res. Commun. 2000;273:793], SIRT6 serves as a transcriptional regulator of intestinal barrier function and regeneration of bowel epithelium. Based on preclinical data, the compound represents a unique treatment approach, as the mechanism of action targets the restoration of the intestinal barrier function and bowel wall architecture in patients suffering from gastrointestinal diseases such as celiac disease, atypical wheat allergy, gluten intolerance (e.g., non-celiac gluten sensitivity (NCGS), gluten ataxia, dermatitis herpetiformis (DH) and wheat allergy), inflammatory bowel disease (IBD) (e.g., Crohn's disease, ulcerative colitis or IBD-associated diarrhea), irritable bowel syndrome (IBS), (especially irritable bowel syndrome with diarrhea (IBS-D)), microscopic colitis (e.g., collagenous colitis and lymphocytic colitis) and eosinophilic esophagitis (EoE).

[0033] Targeting SIRT6 with a compound of Formula (I), in particular the compound of Formula (II), prevents the deacetylation of one of its ligands, acetylated lysine (K) 56 of histone H3 (H3K56), in an in-vitro cell-free and cellular system and significantly accelerates the recovery of transepithelial electrical resistance (TEER) across a monolayer of colorectal adenocarcinoma- 2 (Caco-2) cells following inflammation with tumor necrosis factor (TNF-a) or interleukin (I L)- 6. In one TEER model, tight junction (TJ)-related proteins were investigated, and a normalisation of relevant TJ proteins (claudin (Cldn)-l and claudin-2) was observed. Interaction with a compound of Formula (I), in particular the compound of Formula (II), is highly selective for SIRT6 over other members of the human SIRT protein family and does not inhibit enzymatic activity of other histone deacetylases (HDACs).

[0034] Compounds of Formula (I), in particular the compound of Formula (II), has shown to increase expression of SIRT6 protein in patient samples / biopsies, cell cultures and organoids. At the same time, an increase in H3K56 acylation in cell culture and organoids can be observed. Both effects have an influence on target gene regulation and therefore cannot be isolated. A consistent upregulation of protective gastrointestinal genes in intestinal tissue after treatment with a compounds of Formula (I), in particular the compound of Formula (II), compared to vehicle control can be observed, while in non-stimulated intestinal cells (Caco- 2) no change in gene regulation was observed after treatment. This implies, that compounds of Formula (I) have a positive effect on the regulation of important target genes, specifically in the disease situation, especially enhancing the natural regenerative process (regeneration of crypts, where intestinal stem cells are located which undergo asymmetric division, thereby producing fully differentiated epithelial gut cells and renewing intestinal stem cells. These new epithelial cells renew the lining of crypts and villi and maintain / restore a healthy gut and proper intestinal barrier) in the gut, which may already been damaged since the outbreak of the disease BAD or AIE.

[0035] In summary, compounds of Formula (I), in particular the compound of Formula (II), is a highly selective and very potent biochemical inhibitor of the deacetylase and ribosyltransferase activity of SIRT6 and at the same time induces stability and enhanced expression of SIRT6 protein due to the unique irreversible binding mode called "ribosylation trapping" in the substrate binding site of the triazolo moiety in Formula (I).

[0036] In some embodiments, said compound of Formula (I), in particular the compound of Formula (II), is capable of restoring intestinal barrier function, inducing physiologic regeneration of the crypt cells.

[0037] In some embodiments, said subject is human.

[0038] In some embodiments, said subject suffers from a disease selected from bile acid diarrhea (BAD) or autoimmune enteropathy (AIE).

[0039] In some embodiments, said subject suffers from suffers from bile acid diarrhea (BAD).

[0040] In some embodiments, said subject suffers from suffers from bile acid diarrhea type I.

[0041] In some embodiments, said subject suffers from suffers from bile acid diarrhea type II.

[0042] In some embodiments, said subject suffers from suffers from bile acid diarrhea type III.

[0043] In some embodiments, said subject suffers from suffers from autoimmune enteropathy (AIE). In some embodiments, said subject is responsive to steroid treatment.

[0044] In some embodiments, said subject is not responsive to steroid treatment.

[0045] In some embodiments, said subject is responsive to treatment with ruxolitinib.

[0046] In some embodiments, said subject is not responsive to treatment with ruxolitinib.

[0047] In some embodiments, said subject is responsive to treatment with upadacitinib.

[0048] In some embodiments, said subject is not responsive to treatment with upadacitinib.

[0049] In some embodiments, said subject is a pediatric patient.

[0050] In some embodiments, said subject is an adult patient.

[0051] In some embodiments, said subject is a geriatric patient. In some embodiments, the present invention relates to a compound of Formula (I) Formula (I), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE), comprising administering to a subject in need thereof an effective amount of the compound according to Formula (I) or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof, optionally in combination with one or more additional therapeutically active compounds.

[0052] In some embodiments, a compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof, wherein said compound or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject in a daily dose of 34 pmol to 1370 pmol.

[0053] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject in a daily dose of 34 pmol to 1370 pmol, preferably of about 68 pmol to about 411 pmol, more preferably of about 137 pmol to about 411 pmol, and even more preferably of about 256 pmol to about 411 pmol.

[0054] In some embodiments the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject in a daily dose of 68 pmol to 411 pmol, preferably of about 137 pmol to about 411 pmol, and more preferably of about 256 pmol to about 411 pmol.

[0055] In some embodiments the compound of Formula (II) is administered to said subject in a daily dose of about 68 pmol, about 102 pmol, about 137 pmol, about 256 pmol or about 411 pmol, preferably in a daily dose of about 256 pmol or about 411 pmol.

[0056] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject in a daily dose from about 20 mg to about 800 mg, or from about 80 mg to about 320 mg, from about 60 mg to about 240 mg, from about 150 mg to about 240 mg, or from about 60 mg to about 160 mg. Exemplary unit doses include about 20 mg, about 40 mg, about 60 mg, about 80 mg, about 100 mg, about 120 mg, about 150 mg, about 160 mg, about 200 mg, about 240 mg, about 280 mg, about 300 mg, about 320 mg, about 400 mg, about 480 mg, about 500 mg, or about 600 mg.

[0057] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof result in a plasma trough level which is above 20 ng / mL at a steady state. For example, administration the compound of Formula (II) may result in a plasma trough level above 20 ng / mL, above 40 ng / mL, above 60 ng / mL, above 80 ng / mL, above 100 ng / mL, above 120 ng / mL, above 140 ng / mL, above 160 ng / mL, above 180 ng / mL, above 200 ng / mL, above 240 ng / mL, above 280 ng / mL, above 320 ng / mL, above 400 ng / mL, or above 500 ng / mL.

[0058] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof result in a plasma trough level, which is above 20 ng / mL at a steady state. For example, administration of the compound of Formula (II) may result in a plasma trough level range from about 20 ng / mL to 1000 ng / mL, from about 40 ng / mL to 500 ng / mL, from about 60 ng / mL to 320 ng / mL, from about 60 ng / mL to 200 ng / mL, from about 60 ng / mL to 200 ng / mL, from about 160 ng / mL to 320 ng / mL, from about 60 ng / mL to 90 ng / mL, from about 70 ng / mL to 85 ng / mL, from about 140 ng / mL to 200 ng / mL, or from about 150 ng / mL to 190 ng / mL.

[0059] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof, the steady state is achieved between about 3 to 14 days, about 5 to 10 days, or about 8 days after periodic or regular administration to a optionally once-a-day dosing of the drug, therapeutic combination or pharmaceutical dosage form.

[0060] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof, the plasma concentration for the drug, therapeutic combination or pharmaceutical dosage form is: (a) the trough level or trough concentration (Ctrough), or the lowest concentration reached by the drug, therapeutic combination or pharmaceutical dosage form before a second or next dose is administered, or (b) determined from blood samples taken between about 2 hours to 24 hours, or 4 to 12 hours, or 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 or more hours, after the last dose or administration of the drug, therapeutic combination or pharmaceutical dosage form.

[0061] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof, wherein said compound or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least one additional therapeutic treatment.

[0062] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof, wherein said compound or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least one additional therapeutic treatment selected from an immunosupressive agent, a bile acid binder, farnesoid X receptor (FXR) agonist, a cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel inhibitor, a fibroblast growth factor 19 (FGF19) analogue, a 5-HT3 antagonist, a glucagon-like peptide-2 (GLP-2) agonist, an opioid receptor agonist, a mixed P opioid receptor agonist or a broad-spectrum gut-specific antibiotic.

[0063] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least one immunosupressive agent.

[0064] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least one immunosupressive or modulating agent which is selected from the group consisting of prednisone, prednisolone, methylprednisolone, budesonide, beclomethasone, dexamethasone, tacrolimus, sirolimus (rapamycin), cyclosporine, rifaximin, acalabrutinib, anlotinib, baricitinib, bortezomib, carfilzomib, entospletinib, fostamatinib, glasdegib, ibruitinib, imatinib, itacitinib, ixazomib (MLN9708), jaktinib, nilotinib, nintedanib, pacritinib, pimicotinib, ruxolitinib, upadacitinib, sonidegib, vismodegib, other JAK inhibitors (e.g. SHR0302), axatilimab, alemtuzumab, basiliximab, belimumab, brentuximab vedotin, daclizumab, efalizumab, gavilimomab (ABX- CBL), ibritumomab tiuxetan, infliximab, inolimomab, itolizumab, milatuzumab, muromonab- CD3, natalizumab, neihulizumab, obinutuzumab, ofatumumab, rituximab, siplizumab, tildrakizumab, tocilizumab, ustekinumab, vedolizumab, visilizumab, etanercept, alefacept, abatacept, IL-2, teduglutide, lenalidomide, pomalidomide, leflunomide, thalidomide, panobinostat (LBH589), alvelestat (MPH966), vorinostat, mycophenolate mofetil (MMF), methotrexate (MTX), decitabine, clofarabine, melphalan, thiotepa, pentostatin, palifermin, filgrastim, prochymal (remestemcel-L), cyclophosphamide, defibrotide, cannabidiol, VM-001 (from ViGenCell), RGI-2001, ASC930, TQ05105 (or other JAK / ROCK inhibitors), GDC-8264, ALPN-101, voriconazole, itraconazole, opebacan, belumosudil (KD025), efmarodocokin alfa, efavaleukin alfa (AMG 592), efprezimod alfa, romidepsin, busulfan, fludarabine phosphate, aldesleukin, thymoglobulin, methoxsalen, mesenchymal stromal cells (e.g. CYP-001, OTI-010 or MC0518), allogeneic faecal microbiota (e.g. MaaT013), fecal microbiota transplantation, BET inhibitor (e.g. PLX51107), C5a inhibitor (e.g. ALXN1007), sitagliptin, atorvastatin, clobetasol, maraviroc, ribaxamase (SYN-004), sargramostim, alpha 1-antitrypsin, mitoxantrone or combinations thereof.

[0065] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least one immunosupressive or modulating agent which is selected from the group consisting of prednisone, prednisolone, methylprednisolone, budesonide, beclomethasone, dexamethasone, ruxolitinib, upadacitinib, belumosudil or combinations thereof.

[0066] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least one steroid, e.g. prednisone, prednisolone, methylprednisolone, budesonide, beclomethasone or dexamethasone.

[0067] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least ruxolitinib.

[0068] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least upadacitinib.

[0069] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least belumosudil.

[0070] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least a bile acid binder.

[0071] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least cholestyramine.

[0072] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least colestipol.

[0073] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least colesevelam.

[0074] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with a farnesoid X receptor (FXR) agonist.

[0075] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with obeticholic acid.

[0076] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with UN452.

[0077] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with cilofexor.

[0078] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with a cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel inhibitor. In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least a fibroblast growth factor 19 (FGF19) analogue.

[0079] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with aldafermin.

[0080] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least a 5-HT3 antagonist.

[0081] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with alosetron.

[0082] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least a glucagon-like peptide- 2 (GLP-2) agonist.

[0083] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with teduglutide.

[0084] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with apraglutide.

[0085] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with glepaglutide.

[0086] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least an opioid receptor agonist.

[0087] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with loperamide.

[0088] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least a mixed P opioid receptor agonist.

[0089] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with eluxadoline. In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least a broad-spectrum gutspecific antibiotic.

[0090] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with rifaximin.

[0091] In some emdodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered orally.

[0092] In some embodiments, the compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof may be administered for a prolonged period, for example, for at least about 1 week, at least about 2 weeks, at least about 3 weeks, at least about 4 weeks, at least about 5 weeks, at least about 6 weeks, at least about 7 weeks, at least about 8 weeks, at least about 9 weeks, at least about 10 weeks, at least about 11 weeks, at least about 12 weeks, at least about 14 weeks or at least about 26 weeks. For example, the compound of Formula (II) may be administered for about 2 weeks, at least about 3 weeks, at least about 4 weeks, at least about 5 weeks, at least about 6 weeks, at least about 7 weeks at least about 8 weeks, at least about 9 weeks, at least about 10 weeks, at least about 11 weeks, at least about 14 weeks, or at least about 26 weeks. In some embodiments, the compound of Formula (II) is administered for at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 5 months, at least about 6 months, at least about 7 months, at least about 8 months, at least about 9 months, at least about 10 months, at least about 11 months, at least about 12 months, or longer.

[0093] In some emdodiments, the amount of a compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is effective to reduce intestinal fluid secretion resulting from the BID.

[0094] In some emdodiments, the intestinal fluid secretion is reduced by administering a compound of Formula (I), in particular the compound of Formula (II), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof effective to reduce said intestinal fluid secretion. As used herein, reduction of intestinal fluid secretion refers to a decrease in the amount of fluid that is secreted into the intestinal lumen relative to the amount of fluid that is secreted into the intestinal lumen in the absence of administration of a compound of Formula (I). Methods of measuring decrease in fluid secretion are known to those skilled in the art and include measuring the water content of the intestinal luminal contents, and diarrhea output [see e.g. Clinical Methods: The History, Physical, and Laboratory Examinations. 3rd edition (1990). Chapter 88 Diarrhea], In some embodiments, the reduction in the water content of the subjects's stool is demonstrated by measurement using a clinical instrument such as, for example, the Bristol Stool Form Scale (BSFS). The BSFS [see e.g. Aliment. Pharmacol. Ther. 2016;44:693], assigns the subject's stool a consistency score ranging from 1 (hard) to 7 (watery). Thus, in some embodiments, the reduction in the water content of the subject's stool using the present methods is demonstrated by a decrease in score using the BSFS. In some embodiments, the reduction in the water content of the subject's stool using the present methods is demonstrated by a decrease of 1 point (e.g., 7 to 6, 6 to 5, 5 to 4, 4 to 3, 3 to 2, 2 to 1) using the BSFS. In other embodiments, the reduction in the water content of the subject's stool using the present methods is demonstrated by a decrease of 2 points (e.g., 7 to 5, 6 to 4, 5 to 3, 4 to 2, 3 to 1) using the BSFS. In other embodiments, the reduction in the water content of the subject's stool using the present methods is demonstrated by a decrease of 3 points (e.g., 7 to 4, 6 to 3, 5 to 2, 4 to 1) using the BSFS. In other embodiments, the reduction in the water content of the subject's stool using the present methods is demonstrated by a decrease of more than 3 points using the BSFS. It will be understood by those skilled in the art that the reduction in the water content of the subject's stool can be demonstrated using other clinical scales known the art.

[0095] In some embodiments, the methods of the present invention result in a reduction in the subject's frequency of defecation. The extent of defecation frequency reduction can be determined by comparing the subject's defecation frequency before administering the compound of Formula (I) to the subject's defection frequency after administering the compound of Formula (I). Here, defection frequency may be measured by, for example, interviewing the subject, or administering to the subject an instrument designed to elicit this information.

[0096] In some embodiments, the methods of the present invention result in a reduction in the subject's stool output. The extent of stool output reduction can be determined by measuring the quantity of subject's stool before administering the compound of Formula (I) and comparing that amount to the quantity of the subject's stool after administering the compound of Formula (I). Here, the quantity may be measured by weight or by volume. Methods for measuring stool output are known to those in the art.

[0097] In some embodiments, the methods of the present invention result in a reduction in the subject's abdominal pain. The extent of abdominal pain reduction can be determined by comparing the subject's abdominal pain before administering the compound of Formula (I) with the subject's abdominal pain after administering the compound of Formula (I). Here, abdominal pain may be measured by, for example, interviewing the subject, or administering to the subject an instrument designed to elicit this information. Methods of determining abdominal pain are known to those in the art.

[0098] In some embodiments, the methods of the present invention result in a reduction in the subject's abdominal bloating. The extent of abdominal bloating reduction can be determined by comparing the subject's abdominal bloating before administering the compound of Formula (I) with the subject's abdominal bloating after administering the compound of Formula (I). Here, abdominal bloating may be measured by, for example, interviewing the subject, or administering to the subject an instrument designed to elicit this information. Methods of determining abdominal bloating are known to those in the art.

[0099] In some embodiments, the methods of the present invention result in a reduction in the subject's nausea. The extent of nausea reduction can be determined by comparing the subject's nausea before administering the compound of Formula (I) with the subject's nausea after administering the compound of Formula (I). Here, nausea may be measured by, for example, interviewing the subject, or administering to the subject an instrument designed to elicit this information, such as the Nausea Questionnaire. Methods of measuring nausea are known to those in the art, e.g. in Can. Med. Assoc. J. 1985;133:755.

[0100] In another aspect, the present invention refers to a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) comprising administering the compound of Formula (I), in particular the compound of Formula (II), as defined in any of the above embodiments, to a subject in need thereof.

[0101] In another aspect, the present invention refers to a use of compound of Formula (I), in particular the compound of Formula (II), as defined in any of the above embodiments, for manufacturing a medicament for prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof.

[0102] In another aspect, the present invention refers to a use of compound of Formula (I), in particular the compound of Formula (II), as defined in any of the above embodiments, for prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof.

[0103] It will be recognized that features specified in each embodiment may be combined with other specified features to provide further embodiments of the present disclosure:

[0104] 1. A compound of Formula (I) Formula (I), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof, wherein:

[0105] R1is selected from the group consisting of hydrogen, carboxyl, cyano, fluorine, chlorine, methyl, isopropyl, t-butyl, trifluoromethyl, trifluoromethoxy, cyclopropylmethoxy, 1,1- difluoro-2-methylpropyl, l,l-difluoro-2,2-dimethylpropyl, 1-methyl-l-cyclobutyl, methoxycarbonyl, ethoxycarbonyl, isopropoxycarbonyl, 1-hydroxy-l-methylethyl, azetidine-l-carbonyl, 3-methylo 4,5-dihydrooxazol-2-yl and cyclopropylcarbonyl; R2is hydrogen or methyl;

[0106] A1is =N- or =CH-;

[0107] X is benzene, pyridine or cyclohexane;

[0108] J is any ring selected from the group

[0109] Y is selected from the group consisting of phenyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl, piperidinyl, azetidinyl, tetrahydropyranyl, morpholinyl and tetrahydropyridinyl, optionally substituted with 1, 2 or 3 groups selected from Yl, or alternatively Y is amino, optionally substituted with 1 or 2 groups selected from Yl;

[0110] Y1is selected from the group consisting of hydroxy, cyano, fluorine, amino, methyl, ethyl, isopropyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl, , wherein amino is optionally substituted with 1 or 2 groups selected from Y2and methyl, ethyl, isopropyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl, are optionally substituted with 1, 2 or 3 groups selected from Y2;

[0111] Y2is selected from the group consisting of hydroxyl, fluorine, amino, methyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl, wherein amino is optionally substituted with 1 or 2 groups selected from Y3and methyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl, is optionally substituted with 1, 2 or 3 groups selected from Y3,

[0112] Y3is hydroxyl, fluorine, cyano, methyl or methoxy, wherein methyl or methoxy are optionally substituted with fluorine. A compound of Formula (I) or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof according to embodiment 1, wherein the compound is selected from the following group:

[0113] A compound of Formula (I) or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof according to embodiment 1 or 2, wherein the compound is The compound of Formula (II), which is for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof. A compound of Formula (I) or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment in a subject in need thereof according to any of embodiments 1 to 4, wherein said prevention or treatment is bile acid diarrhea (BAD). A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to any of embodiments 1 to 5, wherein said compound or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject in a daily dose of 34 pmol to 1370 pmol. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to any of embodiments 1 to 6, wherein said compound or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least one additional therapeutic treatment. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to embodiment 7, wherein said at least one additional therapeutic treatment is an immunosupressive treatment. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to embodiment 8, wherein said immunosupressive treatment is selected from the group consisting of prednisone, prednisolone, methylprednisolone, budesonide, beclomethasone, dexamethasone, tacrolimus, sirolimus (rapamycin), cyclosporine, rifaximin, acalabrutinib, anlotinib, baricitinib, bortezomib, carfilzomib, entospletinib, fostamatinib, glasdegib, ibruitinib, imatinib, itacitinib, ixazomib (MLN9708), jaktinib, nilotinib, nintedanib, pacritinib, pimicotinib, ruxolitinib, upadacitinib, sonidegib, vismodegib, other JAK inhibitors (e.g. SHR0302), axatilimab, alemtuzumab, basiliximab, belimumab, brentuximab vedotin, daclizumab, efalizumab, gavilimomab (ABX-CBL), ibritumomab tiuxetan, infliximab, inolimomab, itolizumab, milatuzumab, muromonab- CD3, natalizumab, neihulizumab, obinutuzumab, ofatumumab, rituximab, siplizumab, tildrakizumab, tocilizumab, ustekinumab, vedolizumab, visilizumab, etanercept, alefacept, abatacept, IL-2, teduglutide, lenalidomide, pomalidomide, leflunomide, thalidomide, panobinostat (LBH589), alvelestat (MPH966), vorinostat, mycophenolate mofetil (MMF), methotrexate (MTX), decitabine, clofarabine, melphalan, thiotepa, pentostatin, palifermin, filgrastim, prochymal (remestemcel-L), cyclophosphamide, defibrotide, cannabidiol, VM-001 (from ViGenCell), RGI-2001, ASC930, TQ05105 (or other JAK / ROCK inhibitors), GDC-8264, ALPN-101, voriconazole, itraconazole, opebacan, belumosudil (KD025), efmarodocokin alfa, efavaleukin alfa (AMG 592), efprezimod alfa, romidepsin, busulfan, fludarabine phosphate, aldesleukin, thymoglobulin, methoxsalen, mesenchymal stromal cells (e.g. CYP-001, OTI-010 or MC0518), allogeneic faecal microbiota (e.g. MaaT013), fecal microbiota transplantation, BET inhibitor (e.g. PLX51107), C5a inhibitor (e.g. ALXN1007), sitagliptin, atorvastatin, clobetasol, maraviroc, ribaxamase (SYN-004), sargramostim, alpha 1-antitrypsin, mitoxantrone or combinations thereof. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to embodiment 9, wherein said agent is selected from prednisone, prednisolone, methylprednisolone, budesonide, beclomethasone, dexamethasone, ruxolitinib, upadacitinib, belumosudil or combinations thereof. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to embodiment 7, wherein said at least one additional therapeutic treatment is a bile acid binder, preferably cholestyramine, colestipol or colesevelam. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to embodiment 7, wherein said at least one additional therapeutic treatment is a broadspectrum gut-specific antibiotic, preferably rifaximin. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to embodiment 7, wherein said at least one additional therapeutic treatment is a mixed P opioid receptor agonist, preferably eluxadoline. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to embodiment 7, wherein said at least one additional therapeutic treatment is a 5-HT3 antagonist, preferably alosetron.

[0114] 15. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to embodiment 7, wherein said at least one additional therapeutic treatment is an opioid receptor agonist, preferably loperamide.

[0115] DEFINITIONS

[0116] An "isotopic variant" refers within the scope of the invention to deuterated variations of compounds of Formula (I) or (II), or a pharmaceutically acceptable salt or solvate thereof. Usually, each molecule in the proportion of the population is substituted with a deuterium atom at the same position. The proportion of molecules of the population that comprise a deuterium atom at a specific atomic position can be represented by an isotopic enrichment factor sp / s, where £p / s = (Rp / Rs - 1) x 1000, where Rp is the abundance of deuterium at that position within the population of molecules, and Rs is the natural abundance of deuterium at the position. The proportion of molecules of the population that comprise a deuterium atom at a specific atomic position can alternatively be represented by the molar percent of the population that is substituted with deuterium at that atomic position (i.e., % deuterium incorporation). Non-limiting examples of an isotopic enrichment factor include at least 835 (12.5% deuterium incorporation), at least 1670 (25% deuterium incorporation), at least 3500 (52.5% deuterium incorporation), at least 4500 (67.5% deuterium incorporation), at least 5000 (75% deuterium incorporation), at least 5500 (82.5% deuterium incorporation), at least 6000 (90% deuterium incorporation), at least 6333.3 (95% deuterium incorporation), at least 6466.7 (97% deuterium incorporation), at least 6600 (99% deuterium incorporation), or at least 6633.3 (99.5% deuterium incorporation). Non-limiting examples of an abundance of deuterium in a sample of a compound herein include a natural abundance, and abundance that is at least 3340 times of the natural abundance of deuterium, which is 0.015% (i.e., at least 50.1% incorporation of deuterium), at least 3500 times of the natural abundance of deuterium (52.5% deuterium incorporation), at least 4500 times of the natural abundance of deuterium (67.5% deuterium incorporation), at least 5000 (75% deuterium), at least 5500 times of the natural abundance of deuterium (82.5% deuterium incorporation), at least 6000 times of the natural abundance of deuterium (90% deuterium incorporation), at least 6333.3 times of the natural abundance of deuterium (95% deuterium incorporation), at least 6466.7 times of the natural abundance of deuterium (97% deuterium incorporation), at least 6600 times of the natural abundance of deuterium (99% deuterium incorporation), at least 6633.3 times of the natural abundance of deuterium (99.5% deuterium incorporation), or any other abundance. When a particular position in a compound of the invention (e.g., a compound represented by Formula (I) or (II), or a pharmaceutically acceptable salt and / or solvate thereof) is designated specifically by name or structure as "H" or "hydrogen", the position is understood to have hydrogen at its natural abundance isotopic composition. The percentage of deuterium incorporation can be obtained by quantitative analysis using, for example, mass spectroscopy (peak area) or by quantifying the remaining residual 1H-NMR signals of the specific deuteration site compared to signals from internal standards or other, nondeuterated 1H signals in the compound.

[0117] The term "pharmaceutically acceptable salts" refers to salts prepared from pharmaceutically acceptable non-toxic acids, including inorganic acids and organic acids. Thus, the compound of the present disclosure contains a basic group and can be used according to the disclosure, for example, as hydrochloride, aspartate, glutamate, L-tatrate, malonate, fumarate, citrate, malate, maleate, lactate, gluconate, benzoate, succinate, acetate, phosphate, sulfate, napsylate, besylate, tosylate or mesylate salt. The respective salts can be obtained by customary methods which are known to the person skilled in the art like, for example, by contacting these with an organic or inorganic acid in a solvent or dispersant, or by anion exchange with other salts. The present disclosure also includes all salts of the compounds of the present disclosure which, owing to low physiological compatibility, are not directly suitable for use in pharmaceuticals but which can be used, for example, as intermediates for chemical reactions or for the preparation of pharmaceutically acceptable salts.

[0118] "Pharmaceutically acceptable" means suitable for use in a human subject.

[0119] The term "solvate" refers to a crystalline form of a molecule that further comprises molecules of a solvent or solvents incorporated into the crystalline latice structure. Thus, the compounds of the present disclosure may be present in the form of solvates, such as those which include as solvate water, or pharmaceutically acceptable solvates, such as alcohols, in particular ethanol. A stoichiometric or non-stoichiometric amount of solvent is bound by non- covalent intermolecular forces. When the solvent is water, the "solvate" is a "hydrate." It is understood, that a "pharmaceutically acceptable salts" can in addition optionally contain a "solvate".

[0120] "A week" preferably refers to a period of time of or about 5, about 6 or about 7 days. It may be about 5-8 days.

[0121] "A month" preferably refers to a period of time of or about 28, about 29, about 30 or about 31 days. It may be about 26-33 days.

[0122] The term "treating" or "treatment" means an alleviation of symptoms associated with a disease, disorder or condition, or halt of further progression or worsening of those symptoms. Depending on the disease and condition of the subject, the term "treatment" as used herein may include one or more of curative and palliative treatment. Treatment can also include administering a pharmaceutical formulation of the present invention in combination with other therapies.

[0123] The term "prevention" or "preventing" refers to the administration or application of a compound, composition, or treatment regimen in a manner that inhibits the onset or reduces the risk of occurrence of a specific disease or pathological condition in a subject, e.g. BAD or AIE. This includes any intervention that precludes the initiation, progression, or recurrence of the disease, thereby maintaining the health and well-being of the subject. Prevention encompasses both prophylactic measures administered prior to any signs or symptoms of the disease and strategies aimed at preventing disease relapse or recurrence in individuals who have previously been treated for the condition.

[0124] "Daily dose" preferably refers to the total dose of a compound according to Formula (I) or (II) or an isotopic variant, a pharmaceutically acceptable salt or a solvate thereof, preferably orally administered to the subject or patient each day of administration. The daily dose can be reached through a single or several administrations per day, such as for example once a day, twice a day or three times a day. Preferably, it is reached or achieved by single administration per day, preferably consisting of one or more tablets or capsules, preferably tablets or capsules as described herein.

[0125] As used herein, the term "effective amount" includes a dosage sufficient to produce a desired result with respect to the indicated disorder, condition, or mental state. The desired result may comprise a subjective or objective improvement in the recipient of the dosage.

[0126] As used herein, the term "administering" includes activities associated with providing a patient an amount of a compound according to Formula (I) or (II) or an isotopic variant, a pharmaceutically acceptable salt or a solvate thereof. Administering includes providing unit dosages of compositions set forth herein to a patient in need thereof. Administering includes providing effective amounts of compounds, e.g. the compound of Formula (II) or an isotopic variant, a pharmaceutically acceptable salt or a solvate thereof, for a specified period of time, e.g. for about 6, 9, 12, 15 or more months, or about 1, 2, 3, 4, 5 or more years.

[0127] As used herein, the term "patient" or "subject" refers to refers to a living mammalian organism. In one embodiment the patient is a human subject.

[0128] As used herein, the term "administered as adjunct therapy" or "administered with at least one additional therapeutic target" includes sequential or simultaneous administration of two or more structurally different compounds. For example, two or more structurally different pharmaceutically active compounds can be co-administered by administering a pharmaceutical composition adapted for oral administration that contains two or more structurally different active pharmaceutically active compounds. As another example, two or more structurally different compounds can be co-administered by administering one compound and then administering the other (second) compound. In some instances, the coadministered compounds are administered by the same route. In other instances, the coadministered compounds are administered via different routes. For example, one compound can be administered orally, and the other compound can be administered, e.g. sequentially or simultaneously, via intravenous or intraperitoneal injection.

[0129] The term "about" as used herein with respect to numbers, figures, ranges and / or amounts is preferably meant to mean "circa" and / or "approximately". The meaning of those terms is well known in the art and preferably includes a variance, deviation and / or variability of the respective number, figure, range and / or amount of plus / minus 15% and especially of plus / minus 10%.

[0130] In the context of SIRT6 modulation within the SIRT family, "selectively modulating" refers to an selectivity index of SIRT6 towards one or more members of the SIRT family of about 5-, about 10-, about 20-, about 50-, about 100-fold, about 500-fold, about 1000-fold, or more. The selectivity index can be determined e.g. by measuring the respective SIRT modulation at the same concentration for the respective member and generation of the quotient as outlined in Example 5.

[0131] Compositions may further comprise one or more pharmaceutically acceptable additional ingredient(s) such as alum, stabilizers, antimicrobial agents, buffers, coloring agents, flavoring agents, adjuvants, and the like.

[0132] Compositions may be in the form of tablets or lozenges formulated in a conventional manner. For example, tablets and capsules for oral administration may contain conventional excipients including, but not limited to, binding agents, fillers, lubricants, disintegrants and weting agents. Binding agents include, but are not limited to, syrup, acacia, gelatine, sorbitol, tragacanth, mucilage of starch and polyvinylpyrrolidone. Fillers include, but are not limited to, lactose, sugar, microcrystalline cellulose, maize starch, calcium phosphate, and sorbitol. Lubricants include, but are not limited to, magnesium stearate, stearic acid, talc, polyethylene glycol, and silica. Disintegrants include, but are not limited to, potato starch and sodium starch glycollate. Weting agents include, but are not limited to, sodium lauryl sulfate). Tablets may be coated according to methods well known in the art. Compositions may also be liquid formulations including, but not limited to, aqueous or oily suspensions, solutions, emulsions, syrups, and elixirs. The compositions may also be formulated as a dry product for constitution with water or other suitable vehicle before use, such liquid preparations may contain additives including, but not limited to, suspending agents, emulsifying agents, nonaqueous vehicles and preservatives. Suspending agent include, but are not limited to, sorbitol syrup, methyl cellulose, glucose / sugar syrup, gelatine, hydroxyethylcellulose, carboxymethyl cellulose, aluminium stearate gel, and hydrogenated edible fats. Emulsifying agents include, but are not limited to, lecithin, sorbitan monooleate, and acacia. Nonaqueous vehicles include, but are not limited to, edible oils, almond oil, fractionated coconut oil, oily esters, propylene glycol, and ethyl alcohol. Preservatives include, but are not limited to, methyl or propyl p-hydroxybenzoate and sorbic acid.

[0133] Combination treatments:

[0134] As already indicated, in all of the therapeutic methods or indications disclosed above, the compound of the present invention may be administered alone. However, it may also be administered in combination with one or more additional therapeutically active agents, substances or compounds, either sequentially or concomitantly.

[0135] As described above, compounds of the present invention may be administered or applied in combination with one or more additional therapeutically active compounds, substances or treatments, and suitable additional compounds, substances or treatments may be selected, for example, from bile acid binder (e.g. cholestyramine (A3384; Albiero), colestipol and colesevelam); farnesoid X receptor (FXR) agonist (e.g. obeticholic acid, UN452, cilofexor); cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel inhibitor (e.g. glibenclamide); fibroblast growth factor 19 (FGF19) analogue (e.g. aldafermin);

[0136] 5-HT3 antagonist (e.g. alosetron); glucagon-like peptide-2 (GLP-2) agonist (e.g. teduglutide, apraglutide, glepaglutide); opioid receptor agonist (e.g. loperamide, diphenoxylate, difenoxin); natural opioids (e.g. tincture of opium, paregoric, codeine); mixed P opioid receptor agonist (e.g. eluxadoline); broad-spectrum gut-specific antibiotic (e.g. rifaximin); systemic antibiotic (e.g. cephalosporin antibiotics (cefalexin, cefuroxime, cefadroxil, cefazolin, cefalotin, cefaclor, cefamandole, cefoxitin, cefprozil and ceftobiprole), fluoroquinolone antibiotics (ciprofloxacin, levofloxacin, moxifloxacin, gatifloxacin and norfloxacin), tetracycline antibiotics (tetracycline, minocycline, oxytetracycline and doxycycline), penicillin antibiotics (amoxicillin, ampicillin, phenoxymethylpenicillin, dicloxacillin, carbenicillin, vancomycin and meticillin), monobactam antibiotics (aztreonam), and carbapenem antibiotics (ertapenem, doripenem, imipenem / cilastatin and meropenem)); dihydroorotate dehydrogenase inhibitor (e.g. teriflunomide, vidofludimus calcium); compound that modifies gluten or neutralizes gluten (e.g. latiglutenase, zamaglutenase, AGY 010, GliadinX®); transglutaminase 2 inhibitor (e.g. ZED1227 / TAK-227, GSK3915393); compound suitable for tolerance inducing strategy in celiac disease (e.g. TAK-101, TPM502, KAN-101);

[0137] IL-15 modulator (e.g. ordesekimab, CALY-002, TEV-53408, EQ102);

[0138] T-cell activation inhibitor (e.g. DONQ52); lymphocyte trafficking modulator (e.g. PTG-100); tight junction modulator (e.g. larazotide actetate);

[0139] COX2 inhibitor (e.g. celecoxib); steroidal anti-inflammatory agent (e.g. hydroxyltriamcinolone, alpha-methyl dexamethasone, beta-methyl betamethasone, beclomethasone dipropionate, clobetasol valerate, desonide, desoxymethasone, dexamethasone, diflorasone diacetate, diflucortolone valerate, fluadrenolone, fluclorolone acetonide, flumethasone pivalate, fluosinolone acetonide, fluocinonide, flucortine butylester, fluocortolone, fluprednidene (fluprednylidene) acetate, flurandrenolone, halcinonide, methylprednisolone, triamcinolone acetonide, cortisone, cortodoxone, flucetonide, fludrocortisone, difluorosone diacetate, fluradrenolone acetonide, medrysone, amcinafel, amcinafide, betamethasone, betamethasone esters, chloroprednisone, clocortelone, clescinolone, dichlorisone, difluprednate, flucloronide, flunisolide, fluoromethalone, fluperolone, fluprednisolone, hydrocortisone, meprednisone, paramethasone, prednisolone, prednisone, budesonide); nonsteroidal anti-inflammatory agent (e.g. salicylic acid, acetyl salicylic acid, methyl salicylate, glycol salicylate, salicylmides, benzyl-2,5-diacetoxybenzoic acid, ibuprofen, fulindac, naproxen, ketoprofen, etofenamate, phenylbutazone and indomethacin); immunosuppressive agent (e.g. azathioprine);

[0140] DPP-IV inhibitor (e.g. sitagli ptin, vi Idagli ptin, saxagliptrin, omarigliptin); glutamine; bismuth subsalicylate; lanreotide; somatostatin analog (e.g. vapreotide, octreotide); traditional antidiarrheal remedies (e.g. kaolin, pectin, berberine, muscarinic agents); probiotic (e.g. Saccharomyces boulardii; Lactobacillus rhamnosus GG; Lactobacillus plantarum 299v; Clostridium butyricum M588; Clostridium difficile VP20621 (non-toxigenic C. difficile strain); combination of Lactobacillus casei, Lactobacillus acidophilus (Bio-K + CL1285); combination of Lactobacillus casei, Lactobacillus bulgaricus, Streptococcus thermophilus (Actimel); combination of Lactobacillus acidophilus, Bifidobacterium bifidum (Florajen3); combination of Lactobacillus acidophilus, Lactobacillus bulgaricus delbrueckii subsp. bulgaricus, Lactobacillus bulgaricus casei, Lactobacillus bulgaricus plantarum, Bifidobacterium longum, Bifidobacterium infantis, Bifidobacterium breve, and Streptococcus salivarius subsp. thermophilus). vitamins and dietary supplements; anti-thymocyte globulin; alpha 1-antitrypsin; calcineurin inhibitor (e.g. cyclosporine, tacrolimus) antimetabolite (e.g. methotrexate, mycophenolate mophetyl); anti-CD20 monoclonal antibody (e.g. rituximab, ofatumumab, obinutuzumab);

[0141] CSF-1R blocking monoclonal antibody (e.g. axatilimab);

[0142] CD52-binding monoclonal antibody (e.g. alemtuzumab);

[0143] CD26-binding monoclonal antibody (e.g. begelomab); anti-TNFa antibody (e.g. infliximab); anti-CD30 antibody-drug conjugate (e.g. brentuximab vedotin); deacetylase inhibitor (e.g. panobinostat); mTOR inhibitor (e.g. sirolimus, everolimus, temsirolimus, ridaforolimus); cytokine IL-2 or IL-2 mutein (e.g. AMG 592); tyrosine kinase inhibitor (e.g. imatinib, nilotinib, ibrutinib, fostamatinib, entospletinib), and the sub-group JAK1 / 2 inhibitors (e.g. tofacitinib, oclacitinib, filgotinib, momelotinib, ilginatinib, BMS-911543, INCB52793, peficitinib, WP1066, ivarmacitinib, brepocitinib, ritlecitinib, abrocitinib, itacitinib, ruxolitinib, baricitinib, upadacitinib) or hedgehog inhibitors (e.g. vismodegib, sonidegib); proteasome inhibitor (e.g. ixazomib, bortezomib, carfilzomib); rho-kinase 2 inhibitor (rho-associated protein kinase 2 inhibitor or ROCK2 inhibitor, e.g. belumosudil, SR3677, GV101) or Rho-kinase 1 / 2 inhibitors; mitogen activated extracellular signal regulated kinases 1 and 2 (MEK1 and MEK2) inhibitor (e.g. trametinib);

[0144] PI3K inhibitor (e.g. idelalisib, GS-649443); fusion protein inhibiting the CD28 signaling pathway (e.g. Abatacept), inhibits T cell activation and proliferation (e.g. alefacept); fusion protein inhibiting TNFa (e.g. etanercept); hydroxychloroquine; cytoprotective adjuvant or cytoprotectant (e.g. amifostine, mesna, palifermin, dexrazoxane, / V-acetylcysteine, glutamine, ursodeoxycholic acid); or chemotherapeutic (e.g. pentostatin). DESCRIPTION OF FIGURES

[0145] Figure 1A shows the single-cycle-kinetic analysis of SIRT6 binding to immobilized compound 2 (without NAD) according to Example 3.

[0146] Figure IB illustrates the SIRT6 binding to immobilized compound 2 from Example 3 in absence and presence of NAD.

[0147] Figure 2 illustrates that compound 1 is a potent and highly selective inhibitor of SIRT6. Each recombinant human sirtuin protein was incubated with the corresponding biotinylated peptide, NAD+and indicated compound 1 concentrations at room temperature for 1 h. Changes in peptide modification were measured by AlphaLISA assay. The % inhibition was calculated from individual values of enzyme activities. The data are shown as the mean ± standard deviation (n=4).

[0148] EXAMPLES

[0149] Example 1: Chemical synthesis of the compound according to Formula (II)

[0150] Step 1: Methyl trans-4-(methylcarbamoyl)cyclohexane-l-carboxylate (II) trans-4-( Methoxycarbonyl)cyclohexane carboxylic acid (25.3 g) was dissolved in CH2CI2 (400 mL). To this, NEts (56.6 mL), methylamine hydrochloride (18.4 g), 3 / 7-l,2,3-triazolo[4,5- b] pyridin-3-ol (0.93 g) and l-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (29.5 g) were added and the mixture was stirred at room temperature (rt) for 70 h. To the mixture, waterand IN HCI was added, and the mixture was extracted with CFhCL The organic layer was dried over anhydrous MgSC . The resultant solution was concentrated under reduced pressure and the precipitated solid was diluted with hexane. The obtained solid was collected by filtration to yield compound II as a white solid.1H NMR (CDCI3) 6 (ppm) 1.38- 1.58 (m, 4H), 1.92-1.98 (m, 2H), 2.01-2.09 (m, 3H), 2.27-2.34 (m, 1H), 2.81 (d, J = 4.9 Hz, 3H), 3.67 (s, 3H), 5.40-5.48 (m, 1H).

[0151] Step 2: Methyl trans-4-(methylcarbamothioyl)cyclohexane-l-carboxylate (lk)

[0152] Compound II (19 g) was dissolved in toluene (400 mL). To this, Lawesson's reagent (21.3 g) was added and the mixture was stirred at 90°C for 8 h. To the mixture, saturated aqueous sodium bicarbonate was added and the mixture was extracted with EtOAc. The organic layer was washed with brine and dried over anhydrous MgSC . The residue obtained by concentration under reduced pressure was purified by NH silica gel column chromatography (CH2Cl2 / EtOAc = 9:1 to 1:1 (v / v)) to obtain compound lk as a white solid.TH NMR (CDCI3) 6 (ppm) 1.44-1.54 (m, 2H), 1.69-1.79 (m, 2H), 1.93-1.98 (m, 2H), 2.07-2.12 (m, 2H), 2.31-2.38 (m, 1H), 2.41-2.48 (m, 1H), 3.19 (d, J = 4.9 Hz, 3H), 3.67 (s, 3H), 7.26-7.31 (m, 1H).

[0153] Step 3: 2-((4-(Trifluoromethyl)pyridin-2-yl)oxy)acetohydrazide (lj)

[0154] Methyl 2-((4-(trifluoromethyl)pyridin-2-yl)oxy)acetate (111 g) was dissolved in EtOH (2.4 L). To this, hydrazine monohydrate (177 g) was added and the mixture was stirred at 90°C for 3 h. The resultant solution was concentrated under reduced pressure to obtain compound lj as a white solid.

[0155] Step 4: Methyl trans-4-((methylimino)(methylthio)methyl)cyclohexane-l-carboxylate (li)

[0156] Compound lk (1.5 g) was dissolved in acetone (400 mL). To this, K2CO3 (26.5 g) and methyl iodide (15.9 mL) were added and the mixture was stirred for 5 h under reflux with heating. After the reaction temperature was allowed to reach rt, concentrated under reduced pressure and diluted CH2CI2. The precipitated solid was collected by filtration to obtain compound li as a white solid.XH NMR (CDCI3) 6 (ppm) 1.44-1.56 (m, 4H), 1.7 9-1.85 (m, 0.7H), 1.88-1.97 (m,1.3H), 2.04-2.11 (m, 2H), 2.20 (s, 1H), 2.29-2.36 (m, 1H), 2.42 (s, 2H), 2.47-2.53 (m, 0.7H), 2.80-2.87 (m, 0.3H), 3.17 (s, 2H), 3.26 (s, 1H), 3.67 (s, 1.3H), 3.67 (s, 0.7H).

[0157] Step 5: Methyl trans-4-(4-methyl-5-(((4-(trifluoromethyl)pyridin-2-yl)oxy)methyl)-4 / 7-l,2,4- triazol-3-yl)cyclohexane-l-carboxylate (lh)

[0158] Compound li (20 g) was dissolved in EtOH (200 mL). To this, compound lj (20.5 g) was added and the mixture was stirred at 90°C for 4 h. The residue obtained by concentration under reduced pressure was purified by silica gel column chromatography (MeOH / CH2Cl2 = 0:1 to 1:9 (v / v)) to obtain compound lh as a white solid.1H NMR (CDCI3) 6 (ppm) 1.52-1.64 (m, 2H), 1.87-1.98 (m, 2H), 2.06-2.14 (m, 2H), 2.16-2.23 (m, 2H), 2.47 (dt, J = 12.2, 3.4 Hz, 1H), 2.75 (dt, J = 12.2, 3.4 Hz, 1H), 3.70 (s, 3H), 3.78 (s, 3H), 5.40 (s, 2H), 7.22-7.32 (m, 3H), 7.45 (t, J = 7.8 Hz, 1H). Step 6: (trans-4-(4-methyl-5-(((4-(trifluoromethyl)pyridin-2-yl)oxy)methyl)-4 / 7-l,2,4-triazol-

[0159] 3-yl)cyclohexyl)methanol (lg)

[0160] Compound lh (19.7 g) was dissolved in THF (250 mL). To this, a UAIH4-THF solution (2.5M, 20 mL) was added under ice-cooling and the mixture was stirred at rt for 25 minutes. To the mixture, water (1.9 mL), a 15% aqueous NaOH solution (1.9 mL) and water (5.7 mL) were sequentially added under ice-cooling and the mixture was diluted with EtOAc and then filtered with celite. The filtrate was concentrated under reduced pressure to obtain compound lg as a light yellow oily substance.1H NMR (CDCI3) 6 (ppm) 1.08-1.20 (m, 2H), 1.34 (d, J = 5.4 Hz, 1H), 1.55-1.72 (m, 1H), 1.76-1.89 (m, 2H), 1.94-2.11 (m, 4H), 2.62 (dt, J = 12.2, 3.4 Hz, 1H), 3.53 (t, J = 5.9 Hz, 2H), 3.66 (s, 3H), 5.29 (s, 2H), 7.22-7.29 (m, 3H), 7.42 (t, J = 7.8 Hz, 1H).

[0161] Step 7: trans-4-(4-Methyl-5-(((4-(trifluoromethyl)pyridin-2-yl)oxy)methyl)-4 / 7-l,2,4-triazol-3- yl)cyclohexane-l-carbaldehyde (If)

[0162] Compound lg (17.9 g) was dissolved in CH2CI2 (300 mL). To this, Dess-Martin periodinane (22.4 g) was added under ice-cooling and the mixture was stirred for 5 minutes under icecooling and at rt for40 minutes. The solution was ice-cooled. To this, a IN NaOH solution (230 mL) and a 10% aqueous sodium thiosulfate solution were added and stirred, and then the mixture was extracted with CH2CI2. The organic layer was dried over anhydrous Na2SO4. The residue obtained by concentration under reduced pressure was purified by silica gel column chromatography (MeOH / EtOAc = 0:1 to 1:9 (v / v)) to obtain compound If as a white solid.TH NMR (CDCh) 6 (ppm) 1.36-1.47 (m, 2H), 1.82-1.92 (m, 2H), 2.09-2.16 (m, 2H), 2.18-2.25 (m, 2H), 2.36-2.44 (m, 1H), 2.64 (dt, J = 12.2, 3.4 Hz, 1H), 3.67 (s, 3H), 5.30 (s, 2H), 7.22-7.31 (m, 3H), 7.43 (t, J = 7.8 Hz, 1H), 9.70 (s, 1H).

[0163] Step 8: 2-(trans-4-(4-Methyl-5-(((4-(trifluoromethyl)pyridin-2-yl)oxy)methyl)-4 / 7-l,2,4- triazol-3-yl)cyclohexyl)acetaldehyde (le)

[0164] (Methoxymethyl)triphenylphosphonium chloride (13.2 g) was suspended in THF (100 mL). To this, potassium tert-butoxide (4.31 g) was added under ice-cooling and the mixture was stirred for 20 minutes. The resultant solution was added dropwise using a cannula to a solution of compound If (9.44 g) in THF (150 mL). After 45 minutes, to a suspension of (methoxymethyl)triphenylphosphonium chloride (8.79 g) in THF (100 mL), potassium tert- butoxide (2.88 g) was added under ice-cooling and the mixture was stirred for 5 minutes. The obtained solution was added dropwise again to the reaction solution using a cannula. After 10 minutes, water was added to the reaction mixture and the mixture was concentrated under reduced pressure and extracted with EtOAc. The organic layer was washed with brine and then dried over anhydrous Na2SO4. After filtration, the residue obtained by concentration under reduced pressure was purified by silica gel column chromatography (EtOAc / hexane / MeOH = 1:1:1 to 1:0:0 to 9:0: 1 (v / v / v)). The obtained methoxyethenyl intermediate was dissolved by adding THF (100 mL) and IN HCI (70 mL) was added. Then the mixture was stirred at rt for 2.5 h. The solution was ice-cooled and 2N NaOH (35 mL) was added thereto and the mixture was extracted with EtOAc. The organic layer was washed with brine and then dried over anhydrous Na2SO4. After filtration, the residue obtained by concentration under reduced pressure was purified by silica gel column chromatography (MeOH / EtOAc = 0:1 to 1:9 (v / v)) to obtain compound le as a colorless oily substance.TH NMR (CDCI3) 6 (ppm) 1.13-1.22 (m, 2H), 1.61-2.06 (m, 7H), 2.36 (dd, J = 6.6, 2.2 Hz, 2H), 2.57-2.64 (m, 1H), 3.61 (s, 3H), 5.55 (s, 2H), 6.98-6.99 (m, 1H), 7.12-7.14 (m, 1H), 8.33 (d, J = 5.4 Hz, 1H), 9.76 (t, J = 2.2 Hz, 1H).

[0165] Step 9: 2-(trans-4-(4-Methyl-5-(((4-(trifluoromethyl)pyridin-2-yl)oxy)methyl)-4 / 7-l,2,4- triazol-3-yl)cyclohexyl)acrylaldehyde (Id)

[0166] Compound le (6.87 g) was dissolved in / V, / V-dimethylformamide (40 mL). To this, a formaldehyde solution (37%) (6.6 mL) and L-proline (618 mg) were added and the mixture was stirred at rt for 14 h. The mixture was diluted with EtOAc and washed with water and brine. The organic layer was dried over anhydrous Na2SO4. After filtration, the residue obtained by concentration under reduced pressure was purified by silica gel column chromatography (EtOAc / hexane / MeOH = 1:2:0 to 1:0:0 to 9:0:1 (v / v / v)) to obtain compound Id as a white solid.XH NMR (CDCI3) 6 (ppm) 1.34-1.44 (m, 2H), 1.85-1.98 (m, 4H), 2.03-2.08 (m, 2H), 2.56-2.63 (m, 1H), 2.64-2.71 (m, 1H), 3.62 (s, 3H), 5.55 (s, 2H), 5.98 (s, 1H), 6.25 (s, 1H), 6.99 (s, 1H), 7.13 (d, J = 5.4 Hz, 1H), 8.33 (d, J = 5.4 Hz, 1H), 9.52 (s, 1H).

[0167] Step 10: 2-(trans-4-(4-Methyl-5-(((4-(trifluoromethyl)pyridin-2-yl)oxy)methyl)-4 / 7-l,2,4- triazol-3-yl)cyclohexyl)oxirane-2-carbaldehyde (lc)

[0168] Compound Id (3.68 g) was dissolved in MeOH (60 mL). To this, a hydrogen peroxide solution (30%) (1.33 mL) and a 5N aqueous NaOH solution (0.47 mL) were sequentially added under ice-cooling and the mixture was stirred for 1 h. To the mixture, a 10% aqueous sodium thiosulfate solution was added and the reaction mixture was extracted with EtOAc. The organic layer was washed with brine, and then dried over anhydrous Na2SO4. The mixture was concentrated under reduced pressure, then CH2CI2 was added. The mixture was azeotropically concentrated to obtain compound lc as a white solid.TH NMR (CDCI3) 6 (ppm) 1.30-1.39 (m, 1H), 1.44-1.53 (m, 1H), 1.76-1.93 (m, 4H), 2.00-2.12 (m, 3H), 2.58-2.64 (m, 1H), 2.98 (d, J = 4.4 Hz, 1H), 3.10 (d, J = 4.4 Hz, 1H), 3.61 (s, 3H), 5.55 (s, 2H), 6.98-6.99 (m, 1H), 7.12-7.14 (m, 1H), 8.33 (d, J = 4.9 Hz, 1H), 8.85 (s, 1H).

[0169] Step 11: Benzyl (trans-4-(4-(trans-4-(4-methyl-5-(((4-(trifluoromethyl)pyridin-2- yl)oxy)methyl)-4 / 7-l,2,4-triazol-3-yl)cyclohexyl)-l / 7-pyrazol-l-yl)cyclohexyl)carbamate (lb)

[0170] Benzyl (trans-4-hydrozinylcyclohexyl)carbamate hydrochloride (442 mg) was suspended in EtOH (10 mL). To this, compound lc (504 mg) was added and the mixture stirred at rt for 43 h. The solvent was distilled away under reduced pressure and the resultant residue was purified by NH silica gel column chromatography (hexane / CFhC^ / EtOAc = 1:4:0 to 0:1:4 (v / v / v)). The resultant crude product was washed with EtOAc to afford compound lb as a white solid.XH NMR (CDCI3) 6 (ppm) 1.29-1.39 (m, 2H), 1.42-1.53 (m, 2H), 1.80-1.99 (m, 4H), 2.03-2.25 (m, 9H), 2.58-2.73 (m, 2H), 3.66 (s, 3H), 4.02-4.11 (m, 1H), 4.61-4.69 (m, 1H), 5.10 (s, 2H), 5.59 (s, 2H), 7.02 (s, 1H), 7.16 (d, J = 5.4 Hz, 1H), 7.22 (s, 1H), 7.30-7.39 (m, 6H), 8.36 (d, J = 5.4 Hz, 1H).

[0171] Step 12: trans-4-(4-(trans-4-(4-Methyl-5-(((4-(trifluoromethyl)pyridin-2-yl)oxy)methyl)-4 / 7- l,2,4-triazol-3-yl)cyclohexyl)-l / 7-pyrazol-l-yl)cyclohexan-l-amine (la)

[0172] Compound lb (300 mg) was dissolved in meOH (6 mL) and CH2CI2 (1 mL). To this, acetic acid (0.14 mL) and 10% palladium on carbon (150 mg) were added at room temperature and the mixture was stirred at rt for 4 h under a H2 atmosphere. Then the catalyst was filtered off and the mixture was washed with CH2CI2. The resultant filtrate was concentrated under reduced pressure and the solvent was azeotropically concentrated with ethanol and toluene. The resultant residue was purified by silica gel column chromatography (MeOH / CH2Cl2 = 0:1 to 1:19 (v / v)) to obtain compound la as a white solid.TH NMR (CDCI3) 6 (ppm) 1.25-1.35 (m, 2H), 1.43-1.54 (m, 2H), 1.75-2.24 (m, 14H), 2.59- 2.73 (m, 3H), 3.66 (s, 3H), 4.02-4.09 (m, 1H), 5.59 (s, 2H), 7.02 (s, 1H), 7.16 (d, J = 5.4 Hz, 1H), 7.23 (s, 1H), 7.38 (s, 1H), 8.36 (d, J = 5.4 Hz, 1H). Step 13: 6-(trans-4-(4-(trans-4-(4-Methyl-5-(((4-(trifluoromethyl)pyridin-2-yl)oxy)methyl)-

[0173] 4 / 7-l,2,4-triazol-3-yl)cyclohexyl)-l / 7-pyrazol-l-yl)cyclohexyl)-l-oxa-6-azaspiro[3.3]heptane

[0174] (1)

[0175] Compound of oxetane-2,2-diyldimethanol (75 mg) and / V, / V-diisopropylethylamine (0.72 mL) were dissolved in CH2CI2 (1.5 mL). The mixture was cooled to -78°C, then trifluoromethanesulfonic acid anhydride (0.21 mL) was added dropwise thereto. The mixture was stirred at -78°C for 15 minutes. To this, a solution of compound la (213 mg) in CH2CI2 (5 mL) was added at 0°C and the mixture was stirred at 0°C for 15 minutes and then stirred at rt for 18 h. The residue obtained by concentration under reduced pressure was purified by NH silica gel column chromatography (MeOH / CH2Cl2 = 0:1 to 1:99 (v / v)). To the obtained crude product, diethyl ether was added, and the precipitated solid was collected by filtration to obtain title compound 1 as a white solid.1H NMR (CDCI3) 6 (ppm) 1.16-1.26 (m, 2H), 1.42-1.53 (m, 2H), 1.70-1.80 (m, 2H), 1.89-2.19 (m, 11H), 2.58-2.72 (m, 2H), 2.88 (t, J = 7.5 Hz, 2H), 3.11- 3.16 (m, 2H), 3.64-3.68 (m, 2H), 3.65 (s, 3H), 3.99-4.07 (m, 1H), 4.53 (t, J = 7.5 Hz, 2H), 5.59 (s, 2H), 7.01-7.02 (m, 1H), 7.15-7.17 (m, 1H), 7.21 (s, 1H), 7.37 (s, 1H), 8.36 (d, J = 5.1 Hz, 1H). MS (APCI) m / z: 586 [M+H]+.

[0176] Example 2: Chemical synthesis of linkeable tool compound

[0177] Step 1: Benzyl 4-(4-(trans-4-(4-methyl-5-(((4-(trifluoromethyl)pyridin-2-yl)oxy)methyl)-4 / 7- l,2,4-triazol-3-yl)cyclohexyl)-l / 7-pyrazol-l-yl)piperidine-l-carboxylate (2b)

[0178] To a solution of compound lc (891 mg) in EtOH (8 mL) was added benzyl 4- hydrazineylpiperidine-l-carboxylate hydrochloride (546 mg). The mixture was stirred at rt for 16 h, concentrated and purified by reversed-phase chromatography (C18) (MeCN:0.1% NH4HCO3= 10 to 100%) to give compound 2b as a white solid. LCMS (ESI): m / z = 590.4 (M+H)+.

[0179] Step 2: 2-((4-Methyl-5-(trans-4-(l-(piperidin-4-yl)-l / 7-pyrazol-4-yl)cyclohexyl)-4 / 7-l,2,4- triazol-3-yl)methoxy)-4-(trifluoromethyl)pyridine (2a) To a solution of compound lb (876 mg) in CH2CI2 (8 mL) was added TFA (1 mL). The mixture was stirred at rt for 2 h, concentrated and purified by reversed-phase chromatography (C18) (MeCN:0.1% NH4HCO3 = 10 to 100%) to give compound 2a as a white solid.1H NMR (500 MHz, DMSO-d6) 6 8.50 (d, J = 5.0 Hz, 1H), 7.55 (s, 1H), 7.41 (d, J = 5.0 Hz, 1H), 7.32 (d, J = 4.5 Hz, 2H), 5.52 (s, 2H), 4.13-4.08 (m, 1H), 3.63 (s, 3H), 3.03 (d, J = 12.0 Hz, 2H), 2.84 (t, J = 12.0 Hz, 1H), 2.58 (t, J = 12.0 Hz, 2H), 2.52-2.51 (m, 1H), 2.04-1.90 (m, 6H), 1.78-1.64 (m, 4H), 1.51- 1.41 (m, 2H). LCMS (ESI): m / z = 245.8 (M / 2+H)+, 490.3 (M+H)+.

[0180] Step 3: 2-((4-Methyl-5-(trans-4-(l-(l-(2-(prop-2-yn-l-yloxy)ethyl)piperidin-4-yl)-l / 7-pyrazol- 4-yl)cyclohexyl)-4 / 7-l,2,4-triazol-3-yl)methoxy)-4-(trifluoromethyl)pyridine (2)

[0181] To a solution of compound 2a (114 mg) and 2-(prop-2-yn-l-yloxy)ethyl 4-methylbenzene- sulfonate (200 mg) in MeCN (8 mL) was added Nal (12 mg) and K2CO3 (170 mg). The mixture was stirred at 60°C for 16 h, cooled to rt, concentrated and purified by reversed-phase chromatography (C18) (MeCN:0.1% NH4HCO3 = 10 to 100%) to give compound 2 as a white solid.TH NMR (500 MHz, MeOD-d4) 6 8.42 (d, J = 5.0 Hz, 1H), 7.55 (s, 1H), 7.38 (s, 1H), 7.28 (d, J = 4.5 Hz, 1H), 7.18 (s, 1H), 5.61 (s, 2H), 4.18 (d, J = 2.5 Hz, 2H), 4.15-4.08 (m, 1H), 3.76 (s, 3H), 3.70 (t, J = 5.5 Hz, 2H), 3.13-3-11 (m, 2H), 2.94-2.85 (m, 2H), 2.67 (t, J = 5.5 Hz, 2H), 2.66- 2.59 (m, 1H), 2.31-2.25 (m, 2H), 2.14-1.99 (m, 8H), 1.83-1.75 (m, 2H), 1.61-1.52 (m, 2H). LCMS (ESI): m / z = 286.9 (M / 2+H)+, 572.3 (M+H)+.

[0182] Example 3: Content of different doses of compound 1

[0183] As described above, compound 1 can be dosed as free base or as a pharmaceutically acceptable salt and / or solvate thereof. The preferred polymorph of compound 1 (as free base) is described in WO2019 / 054427, i.e., is characterized by an X-ray powder diffraction pattern having characteristic peaks at 2 theta (±0.2°) of 13.2°, 15.8°, 16.5°, 17.8°, 18.1°, 20.3°, 20.8°, 21.4° and 27.9°. The quantity stated in the embodiments and claims relates to the amount of active moiety, i.e., free base. For optional salt and / or solvate the amount has to be adjusted. In the following table the amount (in mg) of active moiety of the compound is converted into pmol.

[0184] Example 4: Binding characterization to SIRT6 with compound 2 using surface plasmon resonance

[0185] Step 1: Immobilization of l-amino-ll-azido-3,6,9-trioxaundecane l-Amino-ll-azido-3,6,9-trioxaundecane was used to link the alkyne analogue of a compound according Formula (I) (compound 2) with the dextran matrix of the CM5 sensor chip. To exclude an unspecific binding of SIRT6 to this linker only the l-amino-ll-azido-3,6,9- trioxaundecane without compound 2 was coupled onto the sensor chip surface and a possible interaction with SIRT6 as analyte was tested.

[0186] • Running buffer: HBS-P (0.01 M HEPES pH 7.4, 0.15 M NaCI, 0.05% v / v Tween-20) + 1% (v / v) DMSO

[0187] • Activation surface with EDC / NHS (10 pL / min, 600 sec contact time)

[0188] • 100 pL (=110 mg) l-amino-ll-azido-3,6,9-trioxaundecane were taken from the original flask under argon atmosphere and transferred into an argon rinsed 0.5 mL tube

[0189] • Dilution in 10 mM sodium phosphate pH 6.5: 990 pL sodium phosphate pH 6.5 + 9.09 pL l-amino-ll-azido-3,6,9-trioxaundecane 10 mg / mL

[0190] • Injection of 10 mg / mL l-amino-ll-azido-3,6,9-trioxaundecane: Flow rate: 10 pL / min

[0191] Injection of 300 pL of 10 mg / mL solution, then flow rate: 5 pL / min Injection of 325 pL of 10 mg / mL solution

[0192] • Deactivation of sensor chip surface: Flow rate: 10 pL / min, injection of 100 pL 1 M ethanolamine (contact time 600 sec)

[0193] • Rinsing the sensor chip overnight with 10 pL / min running buffer

[0194] A response increase of ~480 to ~590 resonance units (RU) was obtained (depending on the experiment) with this linker element coupled to the dextran matrix.

[0195] Step 2: Immobilization of compound 2 with amine-azide-linker via Copper-catalysed click chemistry onto a CM5 chip

[0196] • Dissolving of compound 2 in DMSO to a concentration of 50 mM

[0197] • Dissolving of 0.49 g sodium ascorbate in 4.9 mL water 504.77 mM

[0198] • Dissolving of 0.49 g CuSO4-5H2O in 4.711 mL water 416.51 mM

[0199] • Dilution of sodium ascorbate down to 50.477 mM in HBS-P

[0200] • Dilution of CuSO4 down to 4.1651 mM in HBS-P

[0201] • Pipetting of 50 pL 50 mM compound 2 into a 2 mL tube

[0202] • Fast Addition of 933 pL HBS-P and fast mixing to prevent precipitation of compound

[0203] 2 • Addition of 4.95 pL 50.4 mM sodium ascorbate

[0204] • Addition of 12 pL 4.16 mM CuSO4

[0205] • Mixing again 2.5 mM MWT-S-01343 in HBS-P with 5% DMSO

[0206] • Flow rate down to 2 pL / min

[0207] • Injection of 325 pL 2.5 mM compound 2 in HBS-P with 5% DMSO

[0208] • Injection of 170 pL 2.5 mM compound 2 in HBS-P with 5% DMSO

[0209] • Rinsing the sensor chip overnight with 10 pL / min running buffer

[0210] Coupling of compound 2 via the linker onto chip surface gave an increase of ~20 RU (the usage of 5% DMSO (according CM5 chip specification a 1-minute pulse with max. 10% DMSO is emphasized) has possibly led to washing out the dextran matrix during the coupling). The successful coupling was proven in the next step by SIRT6 binding.

[0211] Step 3: Interaction analysis between compound 2 and SIRT6 (without NAD)

[0212] A control experiment with linker alone (without coupled compound 2) bound to the chip surface revealed, that injection of SIRT6 (without or with 500 pM NAD) can be performed without organic solvents (DMSO, methanol) minimizing bulk effects and resulted in a good and stable baseline without a strong drift. Only a weak unspecific interaction of SIRT6 with the amine-azide linker was observed. Now the interaction of SIRT6 to compound 2 bound to the chip was analyzed in absence and presence of NAD. This measurement was performed with following running setup:

[0213] • Running buffer: 20 mM Tris, 150 mM NaCI, 0.05% Tween-20, 1 mM DTT (freshly dissolved) pH 8.0

[0214] • SIRT6 (1 mg / mL equal to 27.5 pM), stored in 150 mM NaCI, 20 mM Tris, 0,5 mM TCEP pH 7.6 at -80°C was thawed and following dilutions concentrations were generated by serial dilutions in running buffer: 2187 nM, 729 nM, 243 nM, 81 nM, 27 nM, 9 nM, 3 nM and 1 nM

[0215] • Starting the Sensorgram recording

[0216] • Flowpath: 1,2; Detection: 2-1; Flow: 30 pL / min

[0217] • Kinject: 240 pL (contact time: 480 sec), 600 sec dissociation time

[0218] • Injections: 1 nM SIRT6, 3 nM SIRT6, 9 nM SIRT6, 27 nM SIRT6, 81 nM SIRT6, 243 nM SIRT6, 729 nM SIRT6 and finally 2187 nM SIRT6

[0219] The resulting sensorgram is shown in Figure 1A. The sensorgram shows a very good and stable baseline. Only weak signals were observed which were evaluated by steady-state analysis obtaining a KD value of 2.16 pM, however, due to the weak curvage of the fit the calculated constants are error-prone. This finding indicates that in absence of NAD the interaction of SIRT6 to compound 2 is weak.

[0220] Step 4: Interaction analysis between compound 2 and SIRT6 (with NAD)

[0221] Afterwards the interaction was analyzed in presence of NAD. This binding experiment was performed with following running setup: • Running buffer: 20 mM Tris, 150 mM NaCI, 0.05% Tween-20, 1 mM DTT (freshly dissolved) pH 8.0

[0222] • To 4 mL running buffer 500 pM NAD was added

[0223] • SIRT6 (1 mg / mL equal to 27.5 pM), stored in 150 mM NaCI, 20 mM Tris, 0,5 mM TCEP pH 7.6 at -80°C was thawed and following dilutions concentrations were generated by serial dilutions in running buffer with 500 pM NAD: 2187 nM, 729 nM, 243 nM, 81 nM, 27 nM, 9 nM, 3 nM andl nM

[0224] • Starting the Sensorgram recording

[0225] • Flowpath: 1,2, 3, 4; Detection: 2-1, 3-1, 4-1; Flow: 30 pL / min

[0226] • Kinject: 240 pL (contact time: 480 sec), 300 sec dissociation time

[0227] • Injections: running buffer with 500 pM NAD: 1 nM SIRT6, 3 nM SIRT6, 9 nM SIRT6, 27 nM SIRT6, 81 nM SIRT6, 243 nM SIRT6, 729 nM SIRT6 and finally 2187 nM SIRT6

[0228] The resulting sensorgram is shown in Figure 2A. Control experiments indicate that the binding of SIRT6 is strongly mediated by the co-substrate NAD. The evaluation of the binding data shows a very strong interaction of SIRT6 and compound 2 in presence of NAD with a KD value of 99 pM, which is mainly driven by a very slow off-rate (kd):

[0229] After analyzing the interaction, the chip surface with immobilized compound 2 was initially regenerated with mobile compound 1 to compete the binding of SIRT6 to compound 2, however this approach failed, which indicated that the complex of compound 2 and NAD is strongly anchored in the active site of SIRT6 that compound 1 alone cannot displace it. This indicates that also the binding of compound 1 to SIRT6 requires NAD, otherwise the replacement should occur.

[0230] Conclusions:

[0231] • SIRT6 and compound 2 (an analogue representing Formula (I) and Formula (II)) can interact only in presence of NAD very tightly.

[0232] • SIRT6 catalyzes the transfer of the ADP moiety of NAD to compound 2 and forms a covalent conjugate that blocks the active site of SIRT6 very tightly.

[0233] • This binding mode is unique to SIRT6 and was confirmed by X-ray crystallography of a compound fo Formula (I) to SIRT6.

[0234] Example 5: Selectivity of compound 1 for SIRT6 over other members of the sirtuin family

[0235] Inhibition of the enzymatic activity mediated by compound 1 is highly selective for SIRT6 over other members of the human sirtuin protein family as well as members of other classes of histone deacetylases (HDACs). Compound 1 did not inhibit the enzyme activities of other tested sirtuin family members (IC50 >1000 nM for hSIRTl, hSIRT2, hSIRT3, hSIRT5 and hSIRT7) and compound 1 was determined to be highly selective for SIRT6 over other SIRT family members (Figure 2).

[0236] The modulatory potential of compound 1 on SIRT6 was assessed in vitro using an AlphaLISA immunodetection assay. In a cell free system, the half-maximal inhibitory concentration (IC50) was 15 nM for the human SIRT6. IC50 values for mouse, monkey and rat SIRT6 were 41 nM, 31 nM and 28 nM, respectively.

[0237] Besides deacetylation, compound 1 effectively inhibited SIRT6's deacylase activity (demyristoylation of H3K9) with an IC50 value of 41 nM in vitro. Modulation of SIRT6 was also tested in a cellular system in HT-29 cells. The half-maximum effectivity concentration (EC50) in these cells was 4.3 nM. In addition, treatment of Caco-2 cells and human intestinal organoids with compound 1 stabilizes SIRT6 protein and increases SIRT6 protein levels.

[0238] Experimental details of the in vitro enzymatic sirtuin activity assay shown in Figure 2:

[0239] The effects of compound 1 on the deacetylase activities of human sirtuin family members SIRT6, SIRT1, SIRT2, SIRT3, and SIRT7 as well as the desuccinylase activity of human SIRT5 were measured using an AlphaLISA immunodetection assay.

[0240] In detail, recombinant sirtuin proteins, biotinylated peptides, and compound 1 or the respective vehicle (1% DMSO) were incubated with or without p-nicotinamide adenine dinucleotide (NAD+, Sigma-Aldrich Co. LLC.) in 10 pL assay buffer [10 mM Tris-HCI pH 8.0 (Wako Pure Chemical Industries, Ltd.), 0.1% (w / v) bovine serum albumin (Sigma-Aldrich Co. LLC.), 0.01% Tween 20 (Bio-Rad Laboratories), 1 mM dithiothreitol solution (DTT, Sigma- Aldrich Co. LLC.), and 12.5% glycerol (Wako Pure Chemical Industries, Ltd.)] in a 384-well Alpha plate (PerkinElmer Inc.) at rt for 1 h. After the enzymatic reactions, 5 pL of AlphaLISA acceptor beads mixture [40 pg / mL AlphaLISA Protein A Acceptor beads (PerkinElmer Inc.), detection antibodies for each substrate and 8 mM nicotinamide (NAM, Sigma-Aldrich Co. LLC.) in AlphaLISA Epigenetics Buffer (PerkinElmer Inc.)] were added and incubated at rt for 1 h. Thereafter, 5 pL of 40 pg / mL AlphaScreen Streptavidin Donor beads (PerkinElmer Inc.) in AlphaLISA Epigenetic Buffer were added and incubated at rt for 30 min, shielded from light. The AlphaLISA signal was measured on the EnVision Multilabel Plate Reader (PerkinElmer Inc.). The values for enzyme activity for each sample were calculated using Microsoft Excel as below:

[0241] • Enzyme activity (%) = 100 - 100 x (Signal in the presence of NAD+) - (Signal in the absence of NAD+)

[0242] • Inhibition (%) = 100 - 100 x (Enzyme activity of test sample) / (Enzyme activity of vehicle control)

[0243] The 50% inhibitory concentration (IC50) values were determined using SAS system Release 9.2 software (SAS Institute, Inc.) according to the Sigmoid Emax model as follows:

[0244] • Inhibition (%) = [(Emax - Eo) x O] / [( logi0IC5o)Y_+ O'] + Eo

[0245] • C: Common logarithmic concentration of the compound (nM)

[0246] • y: Sigmoidicity factor Emax and Eo were fixed at 100% and 0%, respectively.

[0247] The table below summarizes the conditions of the enzymatic activity assay used for evaluating selectivity of compound 1 towards other sirtuin family members. sirtuin protein NAD+ Peptide

[0248] Detection antibody

[0249] (pg / mL) (pM) (nM)

[0250] SIRT6 0.1 15 H3K9ac 1 Anti-H3K9ac 0.5 pg / mL

[0251] SIRT1 0.01 15 H3K9ac 1 Anti-H3K9ac 0.5 pg / mL

[0252] SIRT2 2 50 H3K18ac 3 Anti-H3K18ac 0.1%

[0253] SIRT3 0.1 50 LCAD_K318ac 100 Anti-AcK 0.1%

[0254] SIRT5 0.1 15 CPSl_K1297suc 3 Anti-SucK 0.1%

[0255] SIRT7 0.3 100 H3K18ac 3 Anti-H3K18ac 0.1%

[0256] Conclusion:

[0257] Compound 1 is a potent SIRT6 inhibitor with high selectivity for SIRT6 over other human sirtuin family proteins.

[0258] Example 6: Selectivity of reference compounds on SIRT6 and other members of the sirtuin family

[0259] Notably, SIRT6 inhibitors described in the literature often lack the high selectivity and potency compared to compound 1, and exert substantial inhibitory effects on other sirtuin members, especially SIRT1, SIRT2 and SIRT3, or their selectivity towards other sirtuin members has not been tested so far [J. Med. Chem. 2021;64:9732], Reference compounds can be tested, e.g. in the SIRT6 activity assay kit from Abeam Ltd, which detects deacetylase activity of recombinant human SIRT6 (Fluorometric, catalogue# abl56068). The assay is also available for SIRTl, SIRT2 and SIRT3.

[0260] Example 7: Selectivity of compound 1 in the SafetyScreen87™

[0261] In vitro pharmacological profiling was performed with compound 1 against a broad range of targets (receptors, transporters, ion channels, and enzymes) in the SafetyScreen87™ (Eurofins Discovery) to screen for potential undesirable molecular interactions that may cause adverse drug reactions in humans. Compound 1 was tested, in duplicate, at a concentration of 10 pM against this panel of 87 targets. Test compound binding was calculated as a percentage inhibition of the binding of a radioactively labelled ligand specific for each target. Test compound enzyme inhibition effect was calculated as a percentage inhibition of control enzyme activity. Inhibition or stimulation higher than 50% represented a response that was considered significant. No significant results were noted with responses to all targets less than 40%.

[0262] Example 8: In vitro intestinal barrier function model Caco-2 cell line is derived from human epithelial cells from the colon. They are frequently used to mimic intestinal barrier function to either look at permeability of drugs or to look at is functional resistance. For the present experiment, the Caco-2 cells were used to mimic the loss of its barrier function upon challenging the cells with 2,4,6-trinitrobenzene sulfonic acid (TNBS) and investigate the effect of compound 1 on this by measuring the electrical transepithelial resistance (TEER). TNBS is a chemical that in in vivo models is often used to induce a disease that shows similarity's with Crohn's disease in humans. For this Caco-2 cells, cultivated in a 6-well trans well plate on top of the membrane that has small pores that prevent the cells from going through but allow molecules and nutrients to pass. After 3 weeks, when the TEER is stable, the monolayer is challenged with 0.25% TNBS stimulation for 4 h. Then, cells were washed and fresh medium with vehicle or compound 1 was given. TEER was measured before and after challenge at different time points. For correct measurement, the shorter electrode is inserted into the transwell chamber (without touching the cells), and the longer electrode is inserted in the outer well.

[0263] The results show a higher TEER and a higher recovery rate for cells treated with compound 1 compared to vehicle control. This indicate that treatment with compound 1 induces a tightening of the barrier and that therefore, compound 1 can be active in diseases that show a reduced intestinal barrier function like in BAD or autoimmune enteropathy.

[0264] Example 9: In vitro intestinal barrier function model

[0265] The Caco-2 cell line is derived from human colonic epithelial cells. Monolayers of Caco-2 cells are widely used in transepithelial electrical resistance (TEER) assay to investigate epithelial barrier function and paracellular permeability.

[0266] Cytokines such as tumour necrosis factor-alpha (TNF-a) and interleukin-6 (IL-6) have been shown to reduce TEER by increasing paracellular permeability of epithelial cells via modulation of TJ proteins [Am. J. Physiol. Gastrointest. Liver Physiol. 2005;288:G422; J. Biol. Chem. 2011;286:31263],

[0267] To evaluate the effect of compound 1 on paracellular permeability, TEER was measured across a monolayer of Caco-2 cells cultured in a trans well plate. Once the RCELL value (TEER across the cell layer) had reached 1700±300 Q, cells were challenged with the cytokines TNF-a or IL- 6 for 48 hours to weaken the epithelial barrier. After 48 hours, cells were washed and recovery medium containing compound 1 (0.1 to 3 pM) or vehicle control (0.3% DMSO) was added to the cells. The TEER was measured every 24 hours up to 120 hours for TNF-a stimulated cells and 168 hours for IL-6 stimulated cells.

[0268] The TEER recovery rate following both TNF-a and IL-6 stimulation was significantly higher in cells treated with compound 1 compared to vehicle treated cells. This indicates that compound 1 enhances the recovery of the epithelial barrier and can be active in BAD exhibiting increased intestinal permeability. To determine whether compound 1 regulates TJ proteins, Caco-2 cells were cultured and treated with IL-6 as described above. After IL-6 stimulation cells were treated with compound 1 (1 pM or 3 pM) or DMSO (vehicle control) for 144 h. TEER was measured every 24 h to monitor changes in epithelial barrier integrity of the Caco-2 cell monolayer. After 144 h of treatment with compound 1, cells were harvested, and protein levels of TJ proteins and histone acetylation levels were analyzed by Western bloting.

[0269] The TEER recovery rate following IL-6-mediated epithelial barrier disruption was again significantly higher in cells treated with compound 1 (1 pM and 3 pM) compared to vehicle treated cells. Furthermore, treatment of Caco-2 cells with compound 1 significantly increased levels of the barrier strengthening TJ protein, claudin-1, and decreased levels of the barrier weaking TJ protein, claudin-2. This seems to be modulated through modulation of SIRT6, the acetylation of H3K56 was higher in the compound 1 treated versus vehicle-treated cells.

[0270] In summary, these results demonstrate that compound 1 restores epithelial barrier integrity via modulation of TJ proteins critical in gastrointestinal diseases including BAD and autoimmune enteropathy.

[0271] Example 10: In vitro intestinal barrier function model in context with bile acids

[0272] Bile acids have been shown to modulate intestinal barrier function by rearranging TJ proteins and increasing paracellular permeability [Am. J. Physiol. Gastrointest. Liver Physiol. 2008;294:G906],

[0273] In vitro model to investigate modulation of intestinal barrier function through bile acids:

[0274] (a) Monolayers of differentiated human intestinal epithelial cells (Caco-2, T84) will be challenged with different bile acids (cholic acid, deoxycholic acid (DCA) and / or chenodeoxycholic acid (CDCA)) to induce TJ-redistribution and increased paracellular permeability.

[0275] (b) Effects of subsequent treatment with compound 1 on regeneration of epithelial barrier function will be monitored by measurring the transepithelial electrical resistance (TEER).

[0276] (c) Effects on TJ protein composition will be assessed by Western blot analyses quantifying protein levels of TJ proteins critical for barrier integrity (such as claudin-1, claudin-2 or occludin).

[0277] Example 11: Therapeutic dextran sodium sulfate (DSS)-induced colitis mouse model

[0278] For this model, which is an often-used in vivo model for IBD, male C57BL / 6 mice were treated with 2.8% DSS in drinking water for 5 days. Treatment with either vehicle (PEG400), 60 mg / kg sulfasalazine (positive control) or 1 mg / kg of compound 1 was given at disease onset (day 4 after start of DSS treatment) per oral gavage once daily for 5 days. Diarrhea score (normal consistency; soft; pasty, unshaped, not sticking to anus; diarrhea, sticking to anus; diarrhea with macroscopic bleeding) was assessed daily. At the end of the study, mice were sacrificed and the colon was collected. Colon histology score consisting of an architecture score (0-3) and an infiltration score (0-3) was assessed based on hematoxylin and eosin-stained tissues slices.

[0279] This model showed that drinking of DSS induced an upregulation of the diarrhea score and this was also reflected in an increased histological score. The positive control showed a reduction in the diarrhea score while no improvement on the histology score was observed. Compound 1 also showed an improvement of the diarrhea score which was similar or even better than the positive control. In addition, compound 1 was able to also show a reduction in the histology score.

[0280] These results show that compound 1 has activity in the DSS-colitis model, which indicates its potential activity in human diseases in which a defective intestinal barrier function and intestinal inflammation play an important role such as in BAD.

Claims

I75301WO BOEHMERT & BOEHMERTCLAIMS1. A compound of Formula (I)Formula (I), or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof, wherein:R1is selected from the group consisting of hydrogen, carboxyl, cyano, fluorine, chlorine, methyl, isopropyl, t-butyl, trifluoromethyl, trifluoromethoxy, cyclopropylmethoxy, 1,1- difluoro-2-methylpropyl, l,l-difluoro-2,2-dimethylpropyl, 1-methyl-l-cyclobutyl, methoxycarbonyl, ethoxycarbonyl, isopropoxycarbonyl, 1-hydroxy-l-methylethyl, azetidine-l-carbonyl, 3-methyloxetan-3-yl, 4,5-dihydrooxazol-2-yl and cyclopropylcarbonyl;R2is hydrogen or methyl;A1is =N- or =CH-;X is benzene, pyridine or cyclohexane;J is any ring selected from the groupY is selected from the group consisting of phenyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl,piperidinyl, azetidinyl, tetrahydropyranyl, morpholinyl and tetrahydropyridinyl, optionally substituted with 1, 2 or 3 groups selected from Yl, or alternatively Y is amino, optionally substituted with 1 or 2 groups selected from Yl;Y1is selected from the group consisting of hydroxy, cyano, fluorine, amino, methyl, ethyl, isopropyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl,, wherein amino is optionally substituted with 1 or 2 groups selected from Y2and methyl, ethyl, isopropyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl,are optionally substituted with 1, 2 or 3 groups selected from Y2;Y2is selected from the group consisting of hydroxyl, fluorine, amino, methyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl,wherein amino is optionally substituted with 1 or 2 groups selected from Y3and methyl, methoxy, azetidinyl, pyrrolidinyl, morpholinyl,is optionally substituted with 1, 2 or 3 groups selected from Y3,Y3is hydroxyl, fluorine, cyano, methyl or methoxy, wherein methyl or methoxy are optionally substituted with fluorine.

2. A compound of Formula (I) or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof according to claim 1, wherein the compound is selected from the following group:A compound of Formula (I) or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof according to claim 1 or 2, wherein the compound isFormula (II).

4. The compound of Formula (II), which isfor use in a method of prevention or treatment of bile acid diarrhea (BAD) or autoimmune enteropathy (AIE) in a subject in need thereof.

5. A compound of Formula (I) or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment in a subject in need thereof according to any of claims 1 to 4, wherein said prevention or treatment is of bile acid diarrhea (BAD).

6. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to any of claims 1 to 5, wherein said compound or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject in a daily dose of 34 pmol to 1370 pmol.

7. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to any of claims 1 to 6, wherein said compound or an isotopic variant, a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof is administered to said subject with at least one additional therapeutic treatment.

8. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to claim 7, wherein said at least one additional therapeutic treatment is an immunosupressive treatment.

9. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of preventionor treatment of bile acid diarrhea (BAD) in a subject in need thereof according to claim 8, wherein said immunosupressive treatment is selected from the group consisting of prednisone, prednisolone, methylprednisolone, budesonide, beclomethasone, dexamethasone, tacrolimus, sirolimus (rapamycin), cyclosporine, rifaximin, acalabrutinib, anlotinib, baricitinib, bortezomib, carfilzomib, entospletinib, fostamatinib, glasdegib, ibruitinib, imatinib, itacitinib, ixazomib (MLN9708), jaktinib, nilotinib, nintedanib, pacritinib, pimicotinib, ruxolitinib, upadacitinib, sonidegib, vismodegib, other JAK inhibitors (e.g. SHR0302), axatilimab, alemtuzumab, basiliximab, belimumab, brentuximab vedotin, daclizumab, efalizumab, gavilimomab (ABX-CBL), ibritumomab tiuxetan, infliximab, inolimomab, itolizumab, milatuzumab, muromonab-CD3, natalizumab, neihulizumab, obinutuzumab, ofatumumab, rituximab, siplizumab, tildrakizumab, tocilizumab, ustekinumab, vedolizumab, visilizumab, etanercept, alefacept, abatacept, IL-2, teduglutide, lenalidomide, pomalidomide, leflunomide, thalidomide, panobinostat (LBH589), alvelestat (MPH966), vorinostat, mycophenolate mofetil (MMF), methotrexate (MTX), decitabine, clofarabine, melphalan, thiotepa, pentostatin, palifermin, filgrastim, prochymal (remestemcel-L), cyclophosphamide, defibrotide, cannabidiol, VM-001 (from ViGenCell), RGI-2001, ASC930, TQ05105 (or other JAK / ROCK inhibitors), GDC-8264, ALPN-101, voriconazole, itraconazole, opebacan, belumosudil (KD025), efmarodocokin alfa, efavaleukin alfa (AMG 592), efprezimod alfa, romidepsin, busulfan, fludarabine phosphate, aldesleukin, thymoglobulin, methoxsalen, mesenchymal stromal cells (e.g. CYP-001, OTI-OIO or MC0518), allogeneic faecal microbiota (e.g. MaaT013), fecal microbiota transplantation, BET inhibitor (e.g. PLX51107), C5a inhibitor (e.g. ALXN1007), sitagliptin, atorvastatin, clobetasol, maraviroc, ribaxamase (SYN-004), sargramostim, alpha 1-antitrypsin, mitoxantrone or combinations thereof.

10. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to claim 9, wherein said agent is selected from prednisone, prednisolone, methylprednisolone, budesonide, beclomethasone, dexamethasone, ruxolitinib, upadacitinib, belumosudil or combinations thereof.

11. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to claim 7, wherein said at least one additional therapeutic treatment is a bile acid binder, preferably cholestyramine, colestipol or colesevelam.

12. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of preventionor treatment of bile acid diarrhea (BAD) in a subject in need thereof according to claim 7, wherein said at least one additional therapeutic treatment is a broad-spectrum gutspecific antibiotic, preferably rifaximin.

13. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to claim 7, wherein said at least one additional therapeutic treatment is a mixed P opioid receptor agonist, preferably eluxadoline.

14. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to claim 7, wherein said at least one additional therapeutic treatment is a 5-HT3 antagonist, preferably alosetron.

15. A compound of Formula (I) an isotopic variant, or a pharmacologically acceptable salt thereof or a solvate thereof or a solvate of a salt thereof for use in a method of prevention or treatment of bile acid diarrhea (BAD) in a subject in need thereof according to claim 7, wherein said at least one additional therapeutic treatment is an opioid receptor agonist, preferably loperamide.

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