Fatty acid binding protein (FABP) inhibitors, methods of use and methods of preparation

Abstract

Disclosed herein are FABP inhibitor compounds and their use in pharmaceutical compositions for the treatment of diseases, including cancers that highly express any of FABP, particularly triple negative breast cancer (TNBC), and other inflammation-inducing diseases, including cancer that highly express any of FABP. Comprise cardiovascular diseases, obesity or obesity-related diseases, diabetes, dyslipidemia, impaired blood glucose tolerance or impaired fasting blood glucose, vitiligo, psoriasis, autoimmune diseases, pain and dementia. Also disclosed herein are methods for preparing the disclosed compounds.

Description

Cross-reference to related applications

[0001] This application claims priority to U.S. Provisional Application No. 63 / 471,207, filed June 5, 2023, which is incorporated herein by reference. Technical Field

[0002] This disclosure relates to compounds that inhibit fatty acid-binding proteins (such as FABP3, FABP4, FABP5 and / or FABP7), pharmaceutical compositions containing these inhibitory compounds, and the use of these compounds and compositions for the treatment or prevention of cancers (particularly triple-negative breast cancer (TNBC)) that highly express any of these FABPs, autoimmune diseases and conditions, viral infections, and other diseases associated with chronic inflammation, including cardiovascular disease, obesity or obesity-related conditions, diabetes, dyslipidemia, impaired glucose tolerance or impaired fasting glucose, vitiligo, psoriasis, pain, and dementia. Background Technology

[0003] Fatty acid-binding proteins (FABPs) are members of a small (12-15 kDa) family of soluble proteins that facilitate the transport of fatty acids within the cytoplasmic compartments of cells. This family of proteins is highly conserved and lacks catalytic function, but can transport hydrophobic fatty acids to various destinations in the aqueous environment of the cytoplasm, thereby enabling fatty acid oxidation, membrane homeostasis, or nuclear signaling. Furthermore, they are involved in signal transduction processes, for which our understanding is currently limited [1-4]. Structurally, all members of the FABP family possess a β-barrel structure, consisting of a water-filled cavity and binding sites that are unique to each member for specific lipid ligands. Except for FABP5, which is universally expressed in most tissues, FABPs exhibit unique tissue expression profiles. However, in general, tissues with active lipid metabolism tend to express more than one subtype. FABP3 is primarily expressed in muscle tissue, particularly in the heart and neurons. FABP4 is highly expressed in adipose tissue, macrophages, and endothelial cells. FABP5 is also expressed in macrophages and endothelial cells, as well as in skin, adipocytes, neurons, glial cells, and several other tissues [2, 5, 6]. FABP7 is expressed in the brain, specifically in glial cells and astrocytes. Recent patient data have shown that FABP5 is highly upregulated in breast tumors, particularly in TNBC tumors. This protein has been reported to induce the growth and metastasis of TNBC cells, and high levels of this protein are associated with poor survival in TNBC patients. In the MMTV-NeuT mouse model of breast cancer, gene ablation of FABP5 significantly delayed tumor formation and inhibited its growth rate [7]. Similarly, chemoinhibition of FABP5 suppressed tumor growth in xenograft models [8, 9]. Data suggest that inhibition of FABP5 is a promising new approach for treating TNBC and potentially other cancers that highly express this protein.

[0004] TNBC is the most aggressive and deadliest subtype of breast cancer, accounting for 10-20% of all breast cancer cases. Women diagnosed with TNBC are four times more likely to have their cancer spread or metastasize to other organs within five years compared to patients with other types of breast cancer. TNBC is a heterogeneous group of breast tumors that are poorly characterized at the molecular level and lack clear prognostic biomarkers and selective therapeutic targets. This makes the treatment and management of TNBC a significant clinical issue, and there is an urgent need for novel targeted therapies for this disease. Current standard treatment regimens for TNBC include neoadjuvant systemic therapy, such as anthracyclines, taxanes, and cyclophosphamide. Platinum-based chemotherapy has been proposed, but it is not currently recommended in guidelines. Currently, there are no approved targeted therapies for TNBC in neoadjuvant therapy.

[0005] Deletion of the FABP4 and FABP5 genes in mice improves insulin sensitivity, lowers blood glucose, and prevents atherosclerosis. In clamp studies in ob / ob mice, a specific FABP4 inhibitor (BMS309403) showed reduced hepatic glucose production, increased glucose uptake in muscle and adipose tissue, and reduced hepatic steatosis, but no change in body weight or energy expenditure. In addition, this compound showed a reduction in atherosclerotic plaque formation in ApoE KO mice [2,3]. In humans, plasma FABP4 levels are elevated in patients with metabolic syndrome and atherosclerosis

[10] . Furthermore, there is increasing evidence that FABP4 is involved in angiogenesis

[11] and the growth of certain tumors

[12] . The global prevalence of obesity is growing in an epidemic manner. Obesity causes a range of health problems, reduces life expectancy, and costs more than $100 billion annually. More than a quarter of the population suffers from multiple comorbidities, including obesity, atherosclerosis, insulin resistance, dyslipidemia, coagulation disorders, hypertension, and a pro-inflammatory state known as metabolic syndrome. People with metabolic syndrome are at high risk of developing atherosclerosis, type 2 diabetes, and other health problems. Similar to obesity, treatment options for atherosclerosis are very limited.

[0006] In the United States, atherosclerosis is the leading cause of death. At the heart of this syndrome are dysregulations in lipid metabolism and abnormal inflammatory responses. While fatty acids have been proposed to play a mechanistic role in the development of obesity and diabetes by altering glucose and lipid metabolism and in inflammatory cascades, the mechanisms linking fatty acids or other lipid signaling to inflammatory responses and the formation of atherosclerotic lesions remain poorly understood.

[0007] The ability to modulate the immune system opens up possibilities for treating a wide range of conditions, including those caused by chronic inflammation and cancer. In addition to its established role in vaccine development, immune modulation has the potential to treat a variety of diseases, including autoimmune diseases and cancer, as well as inflammatory diseases, fibrotic diseases, and infectious diseases.

[0008] Immune cells play a crucial role in the tumor microenvironment (TME). Tumor-infiltrating immune cells are involved in the regulation of tumor development, progression, and treatment response. The cellular and molecular profiles of the immune TME influence the disease response to treatment and its outcome by regulating the balance between inhibitory and cytotoxic responses near the tumor

[13] . Certain immune cells, such as T cells and natural killer cells, can help suppress tumor growth and promote the clearance of cancer cells. Conversely, the accumulation of immunosuppressive immune cells, such as regulatory T cells and myeloid-derived suppressor cells, can create an environment conducive to tumor growth and progression. The balance between immunostimulatory and immunosuppressive cells in the tumor microenvironment is crucial for the success of immunotherapy and other cancer treatments. Therefore, the ability to manipulate the complex interactions between immune cells and the tumor microenvironment is essential for developing more effective cancer therapies.

[0009] Targeting immune cells in the tumor microenvironment is a promising strategy for improving the efficacy of cancer treatment [13-15]. This approach aims to shift the balance of immune cells in the microenvironment from pro-tumor to anti-tumor. One way to achieve this is by enhancing the activity of immune cells capable of recognizing and eliminating cancer cells, such as T cells, natural killer cells, and pro-inflammatory M1 macrophages (classical activated macrophages). This can be achieved by using immune checkpoint inhibitors, which can remove the "brakes" that normally prevent immune cells from attacking tumors. Another strategy is to reduce the number or activity of immune cells that suppress the immune response, such as regulatory T cells, myeloid-derived suppressor cells, or anti-inflammatory M2 macrophages (alternative to activated macrophages). This can be achieved by using drugs that specifically target these cell types. Combining these approaches with other methods, such as vaccines or CAR-T cell therapy, can provide a more comprehensive and effective way to target immune cells in the tumor microenvironment and improve cancer treatment outcomes.

[0010] One mechanism by which FABPs regulate immune cells is believed to be through their role in mediating immune cell metabolism, which is crucial for the normal functioning of the immune system. FABPs regulate energy production and participate in signaling pathways involved in immune cell activation and function by influencing fatty acid utilization. For example, FABP5 has been found to regulate lipid metabolism and function of T cells in the TME by mediating the uptake and oxidation of intracellular long-chain fatty acids. Activated T cells rely primarily on aerobic glycolysis to promote their proliferation and antitumor function. However, due to the limited availability of glucose and high levels of long-chain fatty acids, tumor-infiltrating T lymphocytes (TILs) expressing high levels of FABP5 often exhibit an exhausted phenotype and impaired antitumor activity. Therefore, inhibition of FABP5 in TILs is expected to activate cellular antitumor activity by shifting their energy balance [16, 17].

[0011] Another example of the regulation of immune cells by FABP5 and FABP4 is the regulation of tissue-resident T cells (Trm). Trm cells are a subset of memory T cells that remain in non-lymphoid tissues such as the intestine, lung, reproductive tract, and skin for extended periods without entering circulation, and provide the first line of defense against antigens and pathogens through rapid recall responses [18-20]. Trm cells have the ability to regulate local immune homeostasis in tissues and participate in immune responses mediated by pathogens, cancer, and possibly by self-antigens during autoimmune processes

[21] . It has recently been established that this unique population of T cells contributes to the pathogenesis of autoimmune diseases such as psoriasis, vitiligo, autoimmune hepatitis, and rheumatoid arthritis

[19] . Due to their tissue-specific functions and locations, Trm cells have distinct gene expression signatures and metabolic requirements compared to other types of T cells [20, 22-24]. One of the most distinctive features of Trm cells is their dependence on exogenous free fatty acids (FFAs), which are internalized from the surrounding environment and metabolized intracellularly to produce the ATP required for their maintenance and survival

[20] . It has been recently reported that Trm cells selectively express fatty acid-binding proteins FABP4 and FABP5, which are crucial for the uptake of FFA by cells

[20] . T cell-specific loss of FABP4 / 5 impairs the uptake of FFA by Trm cells and significantly reduces their lifespan and survival in vivo, without affecting the survival of central memory T (TCM) cells in lymph nodes

[20] . Therefore, FABP4 / 5 inhibitors could be used to specifically target Trm cells in autoimmune diseases.

[0012] FABP4 and FABP5 are expressed in macrophages and have been shown to regulate their function by promoting the uptake and metabolism of intracellular fatty acids and lipids

[25] . FABP4 in Ly6C - MHCII - CD36 +Highly expressed in circulating monocytes / macrophages, FABP5 promotes oxidative lipid uptake, foam cell formation, angiogenesis, tissue remodeling, and pro-tumor functions

[25] . FABP5 is highly expressed in Ly6C+MHCII+CD36- macrophages, in which it has been shown to participate in the formation of lipid droplets (LD) in macrophages

[26] ; and in CD11c+ macrophages, in which it has been shown to promote the secretion of the pro-inflammatory cytokine IL-1b [27, 28] and induce ER stress, exhaustion, and ferroptosis [28, 29], thereby affecting immune cell fate and disease progression. This suggests that inhibiting FABP4 / 5 in macrophages would be beneficial for the treatment of a variety of diseases. It is expected to enhance anti-tumor responses in cancer, prevent foam cell formation and chronic inflammation in obesity and atherosclerosis, and promote anti-inflammatory responses in inflammatory and autoimmune diseases as well as infections. In the case of viral infection, FABP4 has been shown to be involved in viral replication and transmission in SARS-CoV-2 (COVID-19) and OCT43 (common cold coronavirus)

[30] . FABP4 has been shown to be recruited to the ER membrane in infected cells, and inhibition of it has been shown to reduce viral replication and viral load and improve viral load in cell culture models. in vivo Disease symptoms

[30] . Overall, the role of FABP in immune cell regulation highlights the importance of lipid metabolism in immune system regulation and suggests that targeting FABP could be a promising strategy for improving immune cell function and treating immune-related diseases.

[0013] The description of FABP inhibitor compounds in this art includes the following. U.S. Patent No. 6,919,323 B2 (Sulsky) et al. This describes certain pyridazinone compounds that inhibit FABP aP2 (FABP4), and the use of these compounds in the treatment of type 2 diabetes and related diseases. US Patent No. 8,748,470 B2 (Lengyel) et al. This describes a method for alleviating or inhibiting cancer, comprising administering to a subject an inhibitor of FABP4 and / or FABP5, wherein said inhibitor is selected from a list of known compounds, including carbazole butyric acid, arylsulfonamide, sulfonylthiophene, 4-hydroxypyrimidine, 2,3-dimethylindole, benzoylbenzene, biphenyl-alkyl acid, 2-oxazol-alkyl acid, tetrahydropyrimidinone, pyridone, pyrazinone, arylcarboxylic acid, tetrazolium, triazolopyrimidinone, indole, or BMS480404. U.S. Patent No. 8,815,875 B2 (Shipps, Jr.). et al.This describes certain heterocyclic compounds that inhibit FABP, and the use of these compounds in the treatment of diseases or conditions including cardiovascular disease, metabolic disorders, obesity, diabetes, dyslipidemia, and impaired glucose tolerance. US Patent No. 9,278,918 B2 (Buettelmann) et al. This paper describes certain urea-derived compounds that inhibit FABP4 and / or FABP5, and their use in treating diseases or conditions including type 2 diabetes, atherosclerosis, chronic kidney disease, and cancer. PCT Publication No. WO2023043803A1 (Levi et al.) describes certain aniline-derived compounds that inhibit FABP4 and / or FABP5, and their use in treating diseases related to fatty acid metabolism, including cancer.

[0014] There is still a need for improved FABP inhibitor compounds, including compounds that inhibit one or more of FABP3, FABP4, FABP5, and FABP7, and for the use of these compounds in the treatment of diseases and conditions, including cancer. Summary of the Invention

[0015] This disclosure generally relates to compounds based on substituted 2-aminothiophene structures that are inhibitors of one or more of FABP3, FABP4, FABP5, and FABP7 (i.e., "FABP3 / 4 / 5 / 7 inhibitors"). ， This disclosure includes the uses of these inhibitors, methods for preparing them, and their use in pharmaceutical compositions for treating diseases related to fatty acid metabolism. The present invention is intended to introduce the subject matter of this disclosure but does not cover every embodiment, combination, or variation contemplated and described within this disclosure. Further embodiments are contemplated and described through the detailed description, drawings, and claims.

[0016] In at least one embodiment, this disclosure provides a FABP3 / 4 / 5 / 7 inhibitor compound of structural formula I or a pharmaceutically acceptable salt thereof:

[0017]

[0018] (I)

[0019] in,

[0020] R 1 Selected from hydrogen, cyano, and 5-membered heteroaryl rings;

[0021] R 2 and R 3 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, cyclopropyl, cyclobutyl, phenyl, trifluoromethyl and benzyl, or R2 and R 3 Together they form a 5- to 8-membered monocyclic, bicyclic, or spirocyclic carbocyclic or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein the carbocyclic, heterocyclic, aryl, or heteroaryl ring is optionally substituted by one or two substituents selected from C1-C4 alkyl, methoxy, or fluorine.

[0022] X is a part of the following formula:

[0023]

[0024] in,

[0025] Y is a heteroatom selected from –S– and –O– or –CR. 8 R 9 –, where R 8 and R 9 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, or R 8 and R 9 Together they form cyclopropyl, cyclobutyl, cyclopentyl, or oxetyl butyl rings;

[0026] R 4 R 5 R 6 and R 7 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, and / or

[0027] R 4 and R 5 Together or R 6 and R 7 Together they form a cyclopropyl ring or a cyclobutyl ring, or

[0028] R 5 and R 6 Together they form 5- to 6-membered carbon rings or heterocycles with Y as a ring member.

[0029] In at least one embodiment of the compounds of structural formula I disclosed herein, the compounds do not include the specific compounds shown in Table 1 (as shown in other parts of this document).

[0030] In at least one embodiment of the compound of structural formula I disclosed herein, R 1 The chemical group at the position is a cyano group, and the compound has the structural formula Ia:

[0031]

[0032] (Ia).

[0033] In at least one embodiment, the compound of structural formula Ia is a compound having structural formulas Ij, Ik, Il, Im, In, Io, Ip, Iq, Ir, Is, and It as shown in Table 2 (other parts of this document).

[0034] In at least one embodiment of the compound of structural formula I disclosed herein, R 2 and R 3 The chemical substituents at the positions together form a 5- to 8-membered aryl or heteroaryl ring, and the compound has the structural formula Ib.

[0035]

[0036] (Ib)

[0037] Where R 10 and R 11 Each is independently selected from hydrogen, halogen, C1-C4 straight-chain or branched alkyl, cyclopropyl and cyclobutyl.

[0038] In at least one embodiment, the compound of structural formula Ib is a compound having structural formulas Iu, Iv and Iw as shown in Table 3 (other parts of this document).

[0039] In at least one embodiment of the compound of structural formula I disclosed herein, R 1 The chemical group at the position is a 5-membered heteroaryl ring (e.g., a 3-substituted 1,2,4-oxadiazole), and the compound has a structural formula selected from Ic, Id, Ie, If, Ig, Ih, and Ii:

[0040]

[0041] Where R 12 It is selected from hydrogen, halogen, C1-C4 straight-chain or branched alkyl, cyclopropyl, cyclobutyl, and phenyl.

[0042] In at least one embodiment, compounds of the structural formulas Ic, Id, Ie, If, Ig, Ih, and Ii include, but are not limited to, compounds having the structural formulas Ix, Iy, Iz, Iaa, Ibb, Icc, Idd, Iee, Iff, Igg, Ihh, Iii, Ijj, Ikk, Ill, Imm, Inn, Ioo, Ipp, Iqq, and Irr as shown in Table 5 (other parts of this document).

[0043] In at least one embodiment of a compound of structural formula I, Ia, Ib, Ic, Id, Ie, If, Ig, Ih, Ii, Ij, Ik, Il, Im, In, Io, Ip, Iq, Ir, Is, It, Iu, Iv, Iw, Ix, Iy, Iz, Iaa, Ibb, Icc, Idd, Iee, Iff, Igg, Ihh, Iii, Ijj, Ikk, Ill, Imm, Inn, Ioo, Ipp, Iqq, and Irr, the X portion is selected from the exemplary portions shown in Tables 6, 7, or 8 (other parts of this document).

[0044] In at least one embodiment, this disclosure provides FABP3 / 4 / 5 / 7 inhibitor compounds of structural formula II and any pharmaceutically acceptable salt thereof:

[0045]

[0046] (II)

[0047] in,

[0048] R 1 Selected from hydrogen, cyano, and 5-membered heteroaryl rings;

[0049] R 2 and R 3 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, cyclopropyl, cyclobutyl, phenyl, trifluoromethyl and benzyl, or R 2 and R 3 Together they form a 5- to 8-membered monocyclic, bicyclic, or spirocyclic carbocyclic or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein the carbocyclic, heterocyclic, aryl, or heteroaryl ring is optionally substituted by one or two substituents selected from C1-C4 alkyl, methoxy, or fluorine.

[0050] Y is a heteroatom selected from –S– and –O– or –CR. 8 R 9 –, where R 8 and R 9 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, or R 8 and R 9 Together they form cyclopropyl, cyclobutyl, cyclopentyl, or oxetyl butyl rings;

[0051] R 4 R 5 R 6 and R 7 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, and / or

[0052] R 4 and R 5 Together or R6 and R 7 Together they form a cyclopropyl ring or a cyclobutyl ring, or

[0053] R 5 and R 6 Together they form 5- to 6-membered carbon rings or heterocycles with Y as a ring member.

[0054] In at least one embodiment of the compounds of structural formula II of this disclosure, the compounds do not include the compounds shown in Table 1.

[0055] In at least one embodiment of the compound of structural formula II, R 1 The chemical group at the position is a cyano group, and the compound has the structural formula IIa.

[0056]

[0057] (IIa)

[0058] Among them, the chemical group R 2 R 3 R 4 R 5 R 6 and R 7 As defined for compounds of structural formula II. In at least one embodiment, a compound having a substructure of structural formula IIa may have structural formulas IIj, IIk, IIl, IIm, IIin, IIo, IIp, IIq, IIr, IIs, or IIt as shown in Table 9 (other parts of this document).

[0059] In at least one embodiment of the compound of structural formula II, R 1 The chemical group at that position is a cyano group, R 2 and R 3 Together they form a 6-membered aryl ring, and the compound has the structural formula IIb.

[0060]

[0061] (IIb)

[0062] Among them, the chemical group R 2 R 3 R 4 R 5 R 6 and R 7 As defined for compounds of structural formula II, and R 10 and R 11Each is independently selected from hydrogen, halogen, C1–C4 straight-chain or branched alkyl, cyclopropyl, and cyclobutyl. In at least one embodiment, the compound of formula IIb may have the structural formulas IIu, IIv, or IIw as shown in Table 10 (other parts of this document).

[0063] In at least one embodiment of the compound of structural formula II, R 1 The chemical group at position R is a 5-membered heteroaryl ring. 2 and R 3 The chemical groups at each position are independently selected from hydrogen, C1-C4 straight-chain or branched alkyl groups, cyclopropyl, cyclobutyl, phenyl, and benzyl, and the compound has the structural formula IIc, IId, IIe, IIIf, IIg, IIh, or IIi.

[0064]

[0065] Among them, the chemical group R 4 R 5 R 6 and R 7 As defined for compounds of structural formula II, and with chemical group R 12 It is hydrogen, halogen, C1–C4 straight-chain or branched alkyl, cyclopropyl, cyclobutyl, or phenyl. In at least one embodiment, the compounds of structural formulas IIc, IId, IIe, IIf, IIg, IIh, and IIii are compounds having structural formulas IIx, IIy, IIz, IIaa, IIbb, IIcc, IIdd, IIee, IIff, IIgg, IIhh, IIii, IIjj, IIkk, IIll, IImm, IInn, IIoo, IIpp, IIqq, IIrr, and IIss as shown in Table 12 (other parts of this document).

[0066] In at least one embodiment of the compound of structural formula I or structural formula II of this disclosure, the compound is selected from any of the exemplary compounds 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, and 65 as shown in Table 13 (other parts of this document).

[0067] In another embodiment, this disclosure provides a pharmaceutical composition comprising a compound of formula I or formula II and one or more adjuvant ingredients.

[0068] In another embodiment, this disclosure provides the use of a FABP3 / 4 / 5 / 7 inhibitor compound of formula I or II in a method of preparing a medicament or pharmaceutical composition for treating diseases or ailments affected by FABP, namely FABP3, FABP4, FABP5 and FABP7 (i.e., "ailments affected by FABP3 / 4 / 5 / 7").

[0069] In another embodiment, this disclosure provides a method for treating a subject suffering from a disease or ailment affected by FABP3 / 4 / 5 / 7, comprising administering to the subject in need a therapeutically effective amount of a compound of formula I or II, or a pharmaceutical composition comprising a compound of formula I or II and one or more adjuvant ingredients.

[0070] In at least one embodiment, the compound of formula I or formula II may be used to treat diseases or disorders affected by FABP3 / 4 / 5 / 7 present in the subject, which may be selected from: atherosclerosis, coronary atherosclerosis, arterial fibrosis, pulmonary hypertension, heart failure, obesity, type 2 diabetes, type 1 diabetes, gestational diabetes, polycystic ovary syndrome, endometriosis, disorders affected by lipid metabolism and serum free fatty acid levels, metabolic disorders, fatty liver disease, renal fibrosis, systemic inflammation, acute inflammation, allergic inflammation, airway inflammation, viral infections (e.g., COVID-19, common cold), skin diseases (e.g., vitiligo, psoriasis, atopic dermatitis, allergic contact dermatitis, mycosis fungoides, alopecia areata, cicatricial alopecia, graft-versus-host disease (GvHD), contact dermatitis, chronic eczema, herpetic dermatitis, skin diseases). Lupus, scleroderma, dermatomyositis, vasculitis, pemphigus, epidermolysis bullosa, linear IgA, vesicular diseases), neurological disorders and diseases (e.g., pain, multiple sclerosis (MS), Parkinson's disease), autoimmune diseases (e.g., experimental autoimmune encephalomyelitis (EAE), asthma, type 1 diabetes, autoimmune lung disease, autoimmune hepatitis, rheumatoid arthritis (RA), spondyloarthritis, vesicular stomatitis virus infection, multiple sclerosis (MS), lupus nephritis, Crohn's disease, ulcerative colitis, and food allergies), ischemic stroke, graft-versus-host disease (GvHD), and cancers (e.g., breast cancer, prostate cancer, ovarian cancer, skin cancer, gastric cancer, glioma, bile duct cancer, bladder cancer, multiple myeloma, colorectal cancer, hepatocellular carcinoma, cervical cancer, oral squamous cell carcinoma, and / or non-small cell lung cancer (NSCLC)).

[0071] In another embodiment, this disclosure also provides a method for preparing compounds of formula I or II, the method comprising: (a) combining a substituted anhydride compound of formula III in a solvent:

[0072]

[0073] (III)

[0074] Among them, Y and R 4 R 5 R 6 and R 7 As defined above for compounds of formulas I and II; and for substituted 2-aminothiophene compounds of formula IV:

[0075]

[0076] (IV)

[0077] Among them, R 1 R 2 and R 3 As defined above for compounds of formula I and II, and (b) removing the solvent to obtain a compound having structural formula I or formula II.

[0078] In at least one embodiment of the compound of formula IV, the compound is a compound of structural formula IVa:

[0079]

[0080] (IVa)

[0081] Among them, the chemical group R 2 R 3 R 4 R 5 R 6 and R 7 As defined for compounds of structure II. In at least one embodiment, compounds of structure IVa are selected from compounds 4a, 4b, 4c, 4d, 4e, 4f, 4g, 4h, 4i, 4j and 4k shown in Tables 14 and 15 (other parts of this document).

[0082] In at least one embodiment of the compound of formula IV, the compound has the structural formula IVb:

[0083]

[0084] (IVb)

[0085] Among them, the chemical group R 2 R 3 R 4 R 5 R 6 and R 7 As defined for compounds of structural formula II, and where R10 and R 11 Each is independently selected from hydrogen, halogen, C1–C4 straight-chain or branched alkyl, cyclopropyl, and cyclobutyl. In at least one embodiment, the compound of formula IVb is selected from compounds 4l, 4m, and 4n shown in Table 16 (other parts of this document).

[0086] In at least one embodiment of the compound of formula IV, R 1 The chemical group at the location is a 5-membered heteroaryl ring (e.g., a 3-substituted 1,2,4-oxadiazole), and the compound is selected from compounds of structural formulas IVc, IVd, IVe, IVf, IVg, IVh, and IVi shown in Table 17 (other parts of this document). In at least one embodiment, the compounds of structural formulas IVc, IVd, IVe, IVf, IVg, IVh, and IVi are selected from compounds 4o, 4p, 4q, 4r, 4s, 4t, 4u, 4v, 4w, 4x, 4y, 4z, 4aa, 4bb, 4cc, 4dd, 4ee, 4ff, 4gg, 4hh, and 4ii shown in Table 18 (other parts of this document). Attached Figure Description

[0087] A better understanding of the novel features and advantages of this disclosure will be obtained by referring to the following detailed description of exemplary embodiments and the accompanying drawings (also referred to herein as “Figures”), in which the principles of this disclosure are utilized, as illustrated in the drawings:

[0088] Figure 1A , Figure 1B and Figure 1C The results were plotted, showing that the inhibitory compounds FTS005, FTS030, FTS031, FTS037, and FTS039 did not activate the transcription of PPARα, PPARγ, or PPARδ. This was achieved after co-transfection with PPARα-coding proteins. Figure 1A ), PPARγ ( Figure 1B ) or PPARδ ( Figure 1C Transcriptional activation assays were performed in COS7 cells containing a vector carrying PPAR response elements (PPREs) and a vector carrying β-galactosidase as a transfection control. Cells were treated with one of the PPAR-specific agonists Wy-134643 (5 μM), rosiglitazone (5 μM), and GW0742 (5 μM) or one of the compounds FTS005, FTS030, FTS031, FTS037, and FTS039 (10 μM). Data are the mean ± standard deviation of three independent experiments.

[0089] Figure 2A , Figure 2B and Figure 2CThe results are plotted, showing that the FABP3 / 4 / 5 / 7 inhibitor compound FTS005, which is specific for FABP4 / 5, inhibits TNBC cell growth in a FABP5-dependent manner and is more potent than the FABP5 / 7 inhibitor SBF-I26. Cells in all experiments were treated for 4 days with one of the indicated compounds at the specified concentration. Cell confluence was measured using Incucyte software. Figure 2A The results showed that different concentrations of the compound FTS005 inhibited the proliferation of MB-231 and BT-549 cell lines. The IC50 was calculated using the GraphPad fitting algorithm. 50 value. Figure 2B The results showed that different concentrations of the compound FTS005 inhibited the proliferation of wild-type MB-231 cell lines, but did not inhibit those cells that stably expressed FABP5 shRNA (“shF5”). Figure 2C The results show that different concentrations of the compound FTS005 inhibited the proliferation of the MB-231 cell line, and its effect was superior to that of the commercially available FABP5 / 7 inhibitor SBFI-26. Data are the mean ± standard deviation of three independent experiments.

[0090] Figure 3A , Figure 3B and Figure 3C The results are plotted, showing that the FABP3 / 4 / 5 / 7 inhibitor compound FTS005 inhibited the growth of mouse breast cancer cell lines, with high-level expression of FABP5 compared to low-level expression of the gene. Cells in all experiments were treated with the indicated compounds at specified concentrations for 4 days. Cell confluence was measured using Incucyte software. Figure 3A The level of FABP5 mRNA in the cell lines shown was measured by qPCR. Figure 3B The results showed that compound FTS005 inhibited the proliferation of MB-231 and 4T1 cell lines, and this inhibition was correlated with the expression level of FABP5 in the cells. The IC50 was calculated using the GraphPad fitting algorithm. 50 value. Figure 3C The results showed that compound FTS005 inhibited cell proliferation only in cells expressing FABP5. Data are the mean ± standard deviation of three independent experiments.

[0091] Figure 4A and Figure 4BThe results are plotted, showing that the FABP3 / 4 / 5 / 7 inhibitor compounds FTS005, FTS040, FTS041, FTS042, FTS043, FTS044, FTS045, and FTS049 inhibited the proliferation of ovarian cancer cells OVCAR8. Cells in all experiments were treated with the indicated compounds at the specified concentrations for 4 days. Cell confluence was measured using Incucyte software. Figure 4A The results showed that compounds FTS005, FTS040, FTS041, FTS042, FTS043, FTS044, FTS045 and FTS049 inhibited the proliferation of the OVCAR8 cell line. Figure 4B Calculate the IC50 of all compounds using the GraphPad fitting algorithm. 50 Values. Data are the mean ± standard deviation of three independent experiments.

[0092] Figure 5A , Figure 5B and Figure 5C The results were depicted, showing that the FABP3 / 4 / 5 / 7 inhibitor FTS005 inhibited the growth of neuroblastoma and sensitized cells to all-trans retinoic acid (atRA) treatment. NPG human neuroblastoma cells were treated with FTS005 at the specified concentration for 4 days in the presence or absence of retinoic acid (1 mM). Cell confluence was measured using Incucyte software. Figure 5A The results showed that compound FTS005 inhibited the proliferation of NPG cells, but exhibited a synergistic effect when combined with atRA. Figure 5B , Figure 5C The results showed that the combination of compound FTS005 and atRA was more effective in inhibiting cell proliferation than atRA alone. The IC50 was calculated using the GraphPad fitting algorithm. 50 Values. Data are the mean ± standard deviation of three independent experiments.

[0093] Figure 6A , Figure 6B , Figure 6C , Figure 6D , Figure 6E , Figure 6F and Figure 6G The results were described, showing that the FABP3 / 4 / 5 / 7 inhibitor FTS005 was effective in xenograft models. in vivo Inhibits tumor growth. Figure 6A Tumor growth in an MB-231 xenograft model. MB-231 cells (5x10⁻¹²) were transplanted into the tumor xenograft model. 6FTS005 was transplanted into the right abdomen of 7-week-old female NOD scid gamma (NSG) mice. Treatment began one day later with FTS005 via gavage five times weekly (20 or 40 mg / kg) or a carrier. Tumor growth was monitored twice weekly. Mean ± standard deviation (n=5) (by unpaired t-test). Figure 6B The data plotted represent the tumor weight of each mouse at the endpoint (day 24). Statistical significance between control and treatment mice was assessed using the Student's t-test in all experiments. Figure 6C Representative histological sections from paraffin-embedded sections of tumors, stained with antibodies against Ki67, VEGFA, and F4 / 80. Figure 6D , Figure 6E and Figure 6F The figure indicates the intensity of immunohistochemical staining for each sample. Figure 6G : Expression levels of PPARδ target genes shown in samples from collected tumors. Figure 5C The data in Figures 5D and 5G represent the mean ± standard deviation of three mice in each group. Statistical analysis was performed using a two-tailed Student's t-test. *p<0.05, **p<0.01.

[0094] Figure 7A , Figure 7B , Figure 7C , Figure 7D , Figure 7E and Figure 7F The results were described, showing that the FABP3 / 4 / 5 / 7 inhibitor FTS005 inhibited tumor growth in vivo in a homologous mouse model. Figure 7A Tumor growth in a 4T1 autologous xenograft model. 4T1 cells (1x10⁻¹²) were transplanted into the tumor xenograft model. 5 FTS005 was transplanted into the mammary fat pads of 7-week-old female BALB / c mice. Treatment began one day later by gavage five times weekly (40 mg / kg) or with a carrier. Tumor growth was monitored twice weekly. Mean ± standard deviation (n=4) (by unpaired t-test). Figure 7B The data plotted represent the weight of a single tumor in each mouse at the endpoint (day 32). Statistical significance between control and treatment mice was assessed using the Student's t-test in all experiments. Figure 7C Expression levels of VEGFA, ACSL1, and PLIN2, known direct targets of PPARδ, involved in tumor growth, as well as FA storage and oxidation, in tumor samples collected from treated and untreated mice. Figure 7D Representative histological sections from paraffin-embedded sections of treated and untreated tumors, stained with antibodies against the proliferation markers Ki67 and VEGFA. Figure 7E and Figure 7FThe figure indicates the Ki67 content in each sample. Figure 7E ) and VEGFA ( Figure 6F The percentage of total section area with positive immunohistochemical staining. *p<0.05.

[0095] Figure 8A , Figure 8B , Figure 8C and Figure 8D This study presents metabolomics analysis results of glycolysis, TCA cycle, FA oxidation, long-chain fatty acids, and ADP and ATP metabolites in tumor samples collected from treated and untreated mice, measured by LC / MS / MS. Figure 8A The amount of long-chain fatty acids in tumor cells. Figure 8B The amount of circulating TCA metabolites in treated and untreated tumors. Figure 8C The amount and calculated ratio of ADP and ATP in treated and untreated tumors. Figure 8D The amount of glycolytic metabolites in treated and untreated tumors. Data are the mean ± standard deviation of 3 mice in each group. Statistical analysis was performed using a two-tailed Student's t-test. *p<0.05, **p<0.01.

[0096] Figure 9A , Figure 9B , Figure 9C , Figure 9D , Figure 9E and Figure 9F The results were described, showing that the FABP3 / 4 / 5 / 7 inhibitor FTS005 modulates tumor-associated macrophages in the tumor microenvironment in a 4T1 homologous mouse model. Figure 9A , Figure 9B and Figure 9C Representative histological sections from paraffin-embedded sections of treated and untreated tumors, these sections were engraved with F4 / 80 ( Figure 9A CD68 Figure 9B ) and CD163 ( Figure 9C Antibody staining. Figure 9D , Figure 9E and Figure 9F The figure indicates the percentage of positive immunohistochemical staining for each antibody in each sample. Figure 9D , Figure 9E and Figure 9F The data in the table represent the mean ± standard deviation of three mice in each group. Statistical analysis was performed using a two-tailed Student's t-test. *p<0.05.

[0097] Figure 10A , Figure 10B , Figure 10C , Figure 10D , Figure 10E and Figure 10F The results were described, showing that the FABP3 / 4 / 5 / 7 inhibitor FTS005 modulates T cells in the tumor microenvironment in a 4T1 homologous mouse model. Figure 10A Representative histological sections from paraffin-embedded sections of treated and untreated tumors, stained with antibodies against CD3 (A), CD4 (B), and CD8 (C). Figure 10B The figure indicates the percentage of positive immunohistochemical staining for each antibody in each sample. Data are presented as mean ± standard deviation for each group of 3 mice. Statistical analysis was performed using a two-tailed Student's t-test. *p<0.05. Figure 10C , Figure 10D The figure shows CD4 and CD8 spleen T cells collected from untreated and treated homologous mice. Figure 10C ) and activated CD4 and CD8 T cells (TNFα+CD4 CD8 T cells) Figure 10D (frequency) Figure 10E The collected spleen cells were subjected to in vitro stimulation (IVS) with Luc2 peptide (2 μg / mL) in T cell culture medium supplemented with 20 ng / mL IL-7 and 20 U / mL IL-2 (5 x 10). 6 Cells / well). Cells were counted after 2 weeks of IVS (n=5). Figure 10F Two weeks after in vitro cytoscopy (IVS), splenic T cells were co-cultured overnight with 4T1Luc2-CFSE-high (target cells, Balb / c source) and F420Luc2-CFSE-low (control, B6 source) cells (target cells:effect cells = 1:5). The next day, viable CFSE+ cells were counted by flow cytometry, and the percentage of specific lysis was calculated using the following formula: (%, = 100(1 - viable CFSE-high / viable CFSE-low)), normalized to control. Statistical significance was measured by a two-way Student's t-test: ns, nonspecific, *p < 0.05.

[0098] Figure 11 The results of immune cell profiling analysis using the nCounter PanCancer Immune Profiling Panel are presented, showing that treatment with the FABP3 / 4 / 5 / 7 inhibitor FTS005 modulates multiple immune cells within the tumor microenvironment in a 4T1 homologous mouse model. Figure 11 Immune cell signature profile, indicating all significantly different immune cell types found in the analysis (P < 0.05, fold change > 1.5).

[0099] Figure 12A and Figure 12BThe results were described, showing that FTS005 inhibited lipid uptake in a culture model of adipocytes with fatty degeneration and in mature adipocytes. Figure 12A The figure indicates the Nile Red staining intensity in liver HepG2 cells treated with a specified concentration of FTS005 or the known FABP4 inhibitor BMS309403 for 4 h, followed by oleic acid treatment (OA) (1 mM, 24 h). Lipid accumulation was measured only in viable cells that were positive for Dapi staining. Total lipid uptake was quantified using a Biotek Cytation 5 microplate reader. Figure 12B Histogram: This plot depicts the color intensity of Nile Red staining in surviving mature adipocytes after FTS005 treatment. 3T3-L1 preadipocytes differentiated in culture. Cells were treated with FTS005 or the known FABP4 inhibitor BMS309403 (BMS) on day 6 and stained with Nile Red on day 12.

[0100] Figure 13A , Figure 13B , Figure 13C , Figure 13D , Figure 13E , Figure 13F , Figure 13G , Figure 13H , Figure 13I , Figure 13J and Figure 13K The results were described, showing that the FABP inhibitor FTS005 regulates macrophage differentiation in culture. Figure 13A , Figure 13C and Figure 13D After naïve macrophages differentiate into M1 or M2 macrophages, the M1 marker MHC-II ( Figure 13A ) and M2 marker CD36 ( Figure 13C ) and CD206 ( Figure 13D (frequency) Figure 13B , Figure 13E : The cytokine IL-12 secreted from M1 and M2 macrophages during differentiation ( Figure 13B ) and IL-10 ( Figure 13E )level. Figure 13F , Figure 13G , Figure 13H CD206 in macrophages that differentiated from naïve M2 macrophages under conditions of both absence and presence of the FABP4 / 5 inhibitor FTS005. Figure 13F The expression levels of IL-10 and IL-10 Figure 13G ) and IL-12 ( Figure 13H ) level. Figure 13I , Figure 13J , Figure 13KThe frequency of CD206 in M1 macrophages that differentiate into M2 macrophages under conditions of both absence and presence of FTS005. Figure 13I ) and level of expression ( Figure 13J and the level of IL-12 ( Figure 13K ). Detailed Implementation

[0101] In this specification and the following claims, several terms will be referenced, and these terms shall be defined to have the following meanings:

[0102] General definition

[0103] In this specification and the following claims, several terms will be referenced, and these terms shall be defined to have the following meanings:

[0104] Unless otherwise specified, all percentages, ratios, and proportions in this document are by weight. Unless otherwise specified, all temperatures are in degrees Celsius (°C).

[0105] Unless otherwise expressly provided in this disclosure, the terms “a” and “an” are defined as one or more.

[0106] A range may be expressed herein as from “about” a particular value and / or to “about” another particular value. When such a range is expressed, the other side includes from that one particular value and / or to that other particular value. Similarly, when a value is expressed as an approximation, by using the antecedent “about”, it will be understood that the particular value forms the other side. It should be further understood that each endpoint of a range is both significantly related to and independent of the other endpoint.

[0107] The terms “comprise” (and any form of inclusion, such as “comprises” and “comprising”), “have” (and any form of having, such as “has” and “having”), “include” (and any form of inclusion, such as “includes” and “including”), and “contain” (and any form of containing, such as “contains” and “containing”) are all open-ended copulas. Therefore, an apparatus that “comprises,” “has,” “includes,” or “contains” one or more elements has those elements, but is not limited to having only those elements. Similarly, a method that “comprises,” “has,” “includes,” or “contains” one or more steps has those steps, but is not limited to having only those steps.

[0108] Any embodiment of any disclosed method or composition may consist of or substantially consist of any of the described steps, elements, and / or features—but not include / comprise / contain / have any of the described steps, elements, and / or features. Therefore, in any claim, the scope of the given claim may be changed to differ from the scope when the open-ended copula is used by replacing any of the above-mentioned open-ended copulas with "consisting of" or "substantially consisting of".

[0109] Even if not described or illustrated, one or more features of one embodiment may be applied to other embodiments, unless expressly prohibited by this disclosure or the nature of the embodiment.

[0110] Any embodiment of any disclosed compound or method may consist of or substantially consist of any of the described steps, elements, and / or features—but not include / comprise / contain / have any of the described steps, elements, and / or features. Therefore, in any claim, the scope of the given claim may be changed to differ from the scope when the open-ended copula is used by replacing any of the above-mentioned open-ended copulas with "consisting of" or "substantially consisting of".

[0111] Even if not described or illustrated, one or more features of one embodiment may be applied to other embodiments, unless expressly prohibited by this disclosure or the nature of the embodiment.

[0112] As used herein, the term “subject” means a person or animal who could benefit from administration of the FABP3 / 4 / 5 / 7 inhibitor compounds discussed in this application, such as those suffering from (but not limited to) diseases affected by the expression of one or more of FABPs, namely FABP3, FABP4, FABP5 and FABP7, uncontrolled serum free fatty acid levels, cancer, metabolic syndrome or atherosclerosis.

[0113] As used herein, the terms “treat,” “treating,” and “treatment” refer to the reduction or improvement of a condition and / or related symptoms. It should be understood that, while not excluded, treating a condition or disease does not require the complete elimination of the condition, disease, or related symptoms.

[0114] As used in this article, the terms “prevent,” “preventing,” “prevention,” and “prophylactic treatment” are all included in the term “treating,” and refer to reducing the likelihood that a subject who does not have a disease or condition but is at risk or susceptible to develop a disease or condition will develop a disease or condition.

[0115] As used herein, “pharmaceutical acceptable” means physiologically tolerable, including for human or veterinary use. Furthermore, “pharmaceutical acceptable” means a material that is biologically or otherwise harmless, meaning that the material can be administered to a subject without causing any adverse biological effects or interacting harmfully with any other component of the pharmaceutical composition containing it. Essentially, pharmaceutically acceptable materials are non-toxic to the recipient. The choice of carrier should be natural to minimize any degradation of the active ingredient and any adverse side effects in the subject, as is well known to those skilled in the art. For a discussion of pharmaceutically acceptable carriers and other components of the pharmaceutical composition, see, for example, Remington's Pharmaceutical Sciences, 18th edition, Mack Publishing Company, 1990.

[0116] As used herein, “test agents” or “test compounds” refer to agents or compounds that will be screened in one or more assays described herein. Test agents include a wide variety of general types of compounds, including but not limited to small molecule organic compounds, known drugs, peptides, carbohydrates such as oligosaccharides and polysaccharides, polynucleotides, lipids or phospholipids, fatty acids, steroids, or amino acid analogs. Test agents can be obtained from libraries, such as natural product libraries and combinatorial libraries. Furthermore, methods for automating the screening of thousands of compounds are known to be feasible in a short time.

[0117] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. While any methods and materials similar to or equivalent to those described herein may be used in the practice or testing of this invention, preferred methods and materials are described hereafter. All references herein are incorporated by way of citation to disclose and describe the methods and / or materials associated with the cited references.

[0118] The relevant details of the implementation scheme described above and elsewhere are described below.

[0119] The aryl, heterocyclic, and heteroaryl units disclosed herein may have one or more hydrogen atoms substituted. Non-limiting examples of hydrogen substitution include the following:

[0120] Substituted and unsubstituted straight-chain, branched, or cyclic alkyl units include, but are not limited to, the following non-limiting examples: methyl (C1), ethyl (C2), n-propyl (C3), isopropyl (C3), cyclopropyl (C3), n-butyl (C4), sec-butyl (C4), isobutyl (C4), tert-butyl (C4), cyclobutyl (C4), cyclopentyl (C5), cyclohexyl (C6), etc.; while substituted straight-chain, branched, or cyclic alkyl units include, but are not limited to, hydroxymethyl (C1), chloromethyl (C1), trifluoromethyl (C1), aminomethyl (C1), 1-chloroethyl (C2), 2-hydroxyethyl (C2), 1,2-difluoroethyl (C2), 2,2,2-trifluoroethyl (C3), 3-carboxypropyl (C3), 2,3-dihydroxycyclobutyl (C4), etc.

[0121] Substituted and unsubstituted straight-chain, branched, or cyclic alkenyl groups include vinyl (C2), 3-propenyl (C3), 1-propenyl (also known as 2-methylvinyl) (C3), isopropenyl (also known as 2-methylvinyl-2-yl) (C3), buten-4-yl (C4), etc.; substituted straight-chain or branched alkenyl groups, non-limiting examples of which include 2-chloroethenyl (also known as 2-chlorovinyl) (…C2 ), 4-hydroxybuten-1-yl (C4), 7-hydroxy-7-methyloct-4-en-2-yl (C9), 7-hydroxy-7-methyloct-3,5-dien-2-yl (C9), etc.

[0122] Substituted and unsubstituted straight-chain or branched alkynyl groups include ethynyl (C2), propynyl-2-propynyl (also known as propynyl) (C3), propynyl-1-yl (C3), and 2-methyl-hexyl-4-yn-1-yl (C7); substituted straight-chain or branched alkynyl groups, non-limiting examples of which include 5-hydroxy-5-methylhexyl-3-ynyl (C7), 6-hydroxy-6-methylheptyl-3-yn-2-yl (C8), 5-hydroxy-5-ethylheptyl-3-ynyl (C9), etc.

[0123] As used herein, substituted and unsubstituted "alkoxy" refers to the compound with the general formula -OR 100的 Unit, where R 100 It is an alkyl, alkenyl, or alkynyl unit as defined above, such as methoxy, methoxymethyl, or methoxymethyl.

[0124] As used herein, substituted and unsubstituted “halogenated alkyl” refers to an alkyl unit having a hydrogen atom substituted by one or more halogen atoms, such as trifluoromethyl, 1,2-dichloroethyl and 3,3,3-trifluoropropyl.

[0125] As used herein, the term "aryl" refers to a cyclic organic unit comprising at least one benzene ring having a conjugated aromatic six-membered ring, and non-limiting examples include phenyl (C6), naphth-1-yl (C6), and so on. 10 ), Naphth-2-yl (C 10 The aryl ring may have one or more hydrogen atoms substituted by another organic or inorganic group. Non-limiting examples of substituted aryl rings include: 4-fluorophenyl (C6), 2-hydroxyphenyl (C6), 3-methylphenyl (C6), 2-amino-4-fluorophenyl (C6), 2-( N , N- Diethylaminophenyl (C6), 2-cyanophenyl (C6), 2,6-di-tert-butylphenyl (C6), 3-methoxyphenyl (C6), 8-hydroxynaphthyl-2-yl (C6) 10 ), 4,5-dimethoxynaphth-1-yl (C 10 ) and 6-cyanonaphth-1-yl (C 10 ).

[0126] The term "heteroaryl" refers to an organic unit comprising a five- or six-membered conjugated aromatic ring, wherein at least one ring atom is a heteroatom selected from nitrogen, oxygen, or sulfur. A heteroaryl ring may comprise a single ring, for example, a ring having 5 or 6 atoms, wherein at least one ring atom is a heteroatom not limited to nitrogen, oxygen, or sulfur, such as a pyridine ring, a furan ring, or a thiofuran ring. "Heteroaryl" can also be a fused polycyclic and heteroaryl ring system, wherein at least one ring is an aromatic ring, and at least one atom of the aromatic ring is a heteroatom comprising nitrogen, oxygen, or sulfur. According to this disclosure, the following are non-limiting examples of heteroaryl rings:

[0127] ; ; ; ;

[0128] ; ; ;;

[0129] ; ; ; ;

[0130] ; ;and .

[0131] The term "heterocycle" refers to a ring system having 3 to 10 atoms, wherein at least one ring atom is a heteroatom, not limited to nitrogen, oxygen, or sulfur. The ring can be a monocyclic, fused, or bicyclic ring. Non-limiting examples of heterocycles include:

[0132] ; ;and .

[0133] All of the aforementioned heteroaryl or heterocyclic groups may optionally be substituted with one or more hydrogen substituents as further described herein.

[0134] Throughout the description of this disclosure, the terms “thiophene-2-yl and thiophene-3-yl” are used to describe heteroaryl units having the following formulas:

[0135] ; ;

[0136] When naming the compounds disclosed herein, the chemical names of these portions are typically spelled as "thiophen-2-yl and thiophen-3-yl," respectively. Throughout this document, the terms "thiophen-2-yl and thiophen-3-yl" are used when describing these rings as units or portions constituting the compounds of this disclosure, solely to clarify for those skilled in the art which rings are referred to herein.

[0137] The following can replace hydrocarbon groups (C1-C) 20 Non-limiting examples of units with hydrogen atoms on a straight-chain, branched, or cyclic alkyl group, aryl group, heterocyclic group, or heteroaryl ring:

[0138] i) Straight-chain, branched, or cyclic alkyl, alkenyl, and alkynyl groups; for example, methyl (C1), ethyl (C2), n-propyl (C3), isopropyl (C3), cyclopropyl (C3), propenyl-2-yl (C3), propynyl (C3), n-butyl (C4), isobutyl (C4), sec-butyl (C4), tert-butyl (C4), cyclobutyl (C4), n-pentyl (C5), cyclopentyl (C5), n-hexyl (C6), and cyclohexyl (C6);

[0139] ii) Substituted or unsubstituted aryl groups; for example, phenyl, 2-fluorophenyl, 3-chlorophenyl, 4-methylphenyl, 2-aminophenyl, 3-hydroxyphenyl, 4-trifluoromethylphenyl and biphenyl-4-yl;

[0140] iii) Substituted or unsubstituted heterocycles; for example, those provided below;

[0141] iv) Substituted or unsubstituted heteroaryl groups; for example, those provided below;

[0142] v) Alkyl groups; for example, -OH, -CH2OH, -OCH3, -CH2OCH3, -OCH2CH3,

[0143] -CH2OCH2CH3, -OCH2CH2CH3 and -CH2OCH2CH2CH3;

[0144] vi) Ketone group; for example, –COCH3, –CH2COCH3, –OCH2CH3, –CH2COCH2CH3,

[0145] –COCH2CH2CH3 and –CH2COCH2CH2CH3;

[0146] vii) Alkyl carboxyl group; for example, –CO2CH3, –CH2CO2CH3, –CO2CH2CH3,

[0147] –CH2CO2CH2CH3, –CO2CH2CH2CH3 and –CH2CO2CH2CH2CH3;

[0148] viii) Alkylamide group; for example, –CONH2, –CH2CONH2, –CONHCH3,

[0149] –CH2CONHCH3, –CON(CH3)2 and –CH2CON(CH3)2;

[0150] ix) Alkyl carbamate group; for example, –OC(O)NH2, –CH2OC(O)NH2, –OC(O)NHCH3,

[0151] –CH2OC(O)NHCH3, –OC(O)N(CH3)2 and –CH2OC(O)N(CH3)2;

[0152] x) Alkylamino groups; for example, –NH2, –CH2NH2, –NHCH3, –N(CH3)2, –NH(CH2CH3).

[0153] –CH2NHCH3, –CH2N(CH3)2 and –CH2NH(CH2CH3);

[0154] xi) Halogens: –F, –Cl, –Br and –I;

[0155] xii) –CH m X n Where X is a halogen, m is 0 to 2, and m+n=3; for example, –CH2F,

[0156] –CHF2, –CF3, –CCl3, or –CBr3;

[0157] xiii) Alkyl cyano; for example, –CN, –CH2CN and –CH2CH2CN;

[0158] xiv) Alkyl nitro; for example, –NO2, –CH2NO2 and –CH2CH2NO2;

[0159] xv) alkylenesulfonylalkyl; for example, –SO2H, –CH2SO2H, –SO2CH3, –CH2SO2CH3,

[0160] –SO2C6H5 and –CH2SO2C6H5;

[0161] xvi) Alkyl sulfonic acids; for example, –SO3H, –CH2SO3H;

[0162] xvii) hydroxyl or thiol group, or

[0163] xviii) Amino, monosubstituted amino or disubstituted amino.

[0164] For the purposes of this disclosure, the terms “compound,” “analog,” and “composition of matter” are equally applicable to the HIF-1α prolyl hydroxylase inhibitors described herein, including all enantiomers, diastereomers, salts, etc., and the terms “compound,” “analog,” and “composition of matter” are used interchangeably in this specification.

[0165] The compounds disclosed herein include all salt forms, such as salts of basic groups, especially amines, and salts of acidic groups, especially carboxylic acids. The following are non-limiting examples of anions that can form pharmaceutically acceptable salts with basic groups: chloride, bromide, iodide, sulfate, bisulfate, carbonate, bicarbonate, phosphate, formate, acetate, propionate, butyrate, pyruvate, lactate, oxalate, malonic acid, maleate, succinate, tartrate, fumarate, citrate, etc. The following are non-limiting examples of cations that can form pharmaceutically acceptable salts with the anionic forms of the compounds described herein: sodium, lithium, potassium, calcium, magnesium, zinc, bismuth, etc.

[0166] FABP3 / 4 / 5 / 7 inhibitor compounds

[0167] This article discloses compounds that inhibit one or more of FABP3, FABP4, FABP5, and FABP7 (i.e., "FABP3 / 4 / 5 / 7 inhibitors"). 。 The FABP3 / 4 / 5 / 7 inhibitor compounds disclosed herein are based on substituted thiophene rings having the universal structural formula I:

[0168]

[0169] (I)

[0170] in,

[0171] R 1 Selected from hydrogen, cyano, and 5-membered heteroaryl rings;

[0172] R 2 and R 3 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, cyclopropyl, cyclobutyl, phenyl, trifluoromethyl and benzyl, or R 2 and R 3Together they form a 5- to 8-membered monocyclic, bicyclic, or spirocyclic carbocyclic or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein the carbocyclic, heterocyclic, aryl, or heteroaryl ring is optionally substituted by one or two substituents selected from C1-C4 alkyl, methoxy, or fluorine.

[0173] X is a part of the following formula:

[0174]

[0175] in,

[0176] Y is a heteroatom selected from –S– and –O– or –CR. 8 R 9 –, where R 8 and R 9 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, or R 8 and R 9 Together they form cyclopropyl, cyclobutyl, cyclopentyl, or oxetyl butyl rings;

[0177] R 4 R 5 R 6 and R 7 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, and / or

[0178] R 4 and R 5 Together or R 6 and R 7 Together they form a cyclopropyl ring or a cyclobutyl ring, or

[0179] R 5 and R 6 Together they form 5- to 6-membered carbon rings or heterocycles with Y as a ring member.

[0180] In at least one embodiment, the compound of structural formula I includes any pharmaceutically acceptable salt thereof.

[0181] In at least one embodiment, the compounds of structural formula I disclosed herein do not include the compounds shown in Table 1 below.

[0182] Table 1

[0183]

[0184] In at least one embodiment of a FABP3 / 4 / 5 / 7 inhibitor compound having structural formula I, R 1 The chemical group at that position is a cyano group. For example, a compound of formula I can have formula Ia:

[0185]

[0186] (Ia)

[0187] Exemplary compounds of formula Ia include, but are not limited to, compounds having the structural formulas Ij, Ik, Il, Im, In, Io, Ip, Iq, Ir, Is, and It as shown in Table 2 below.

[0188] Table 2

[0189]

[0190] In at least one implementation, R 2 and R 3 The chemical substituents at the positions together form a 5- to 8-membered aryl or heteroaryl ring. For example, compounds of formula I can have formula Ib.

[0191]

[0192] (Ib)

[0193] Where R 10 and R 11 Each is independently selected from hydrogen, halogen, C1-C4 straight-chain or branched alkyl, cyclopropyl and cyclobutyl.

[0194] Exemplary compounds of formula Ib include, but are not limited to, compounds having the structural formulas Iu, Iv, and Iw as shown in Table 3 below.

[0195] Table 3

[0196]

[0197] In at least one embodiment of a FABP3 / 4 / 5 / 7 inhibitor compound having structural formula I, R 1 The chemical group at the position is a 5-membered heteroaryl ring (e.g., a 3-substituted 1,2,4-oxadiazole). For example, compounds of formula I can be compounds having the structural formulas Ic, Id, Ie, If, Ig, Ih, or Ii as shown in Table 4 below.

[0198] Table 4

[0199]

[0200] Where R 12 It is selected from hydrogen, halogen, C1-C4 straight-chain or branched alkyl, cyclopropyl, cyclobutyl, and phenyl.

[0201] Exemplary compounds of the formula Ic, Id, Ie, If, Ig, Ih, or Ii, including but not limited to compounds having the structural formulas Ix, Iy, Iz, Iaa, Ibb, Icc, Idd, Iee, Iff, Igg, Ihh, Iii, Ijj, Ikk, Ill, Imm, Inn, Ioo, Ipp, Iqq, and Irr as shown in Table 5 below.

[0202] Table 5

[0203]

[0204]

[0205]

[0206] The FABP3 / 4 / 5 / 7 inhibitor compound of structural formula I disclosed herein comprises a portion X connected to an amino group of thiophene. The X portion is a chemical group of the following formula:

[0207]

[0208] in,

[0209] Y is a heteroatom selected from –S– and –O– or –CR. 8 R 9 –, where R 8 and R 9 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, or R 8 and R 9 Together they form cyclopropyl, cyclobutyl, cyclopentyl, or oxetyl butyl rings;

[0210] R 4 R 5 R 6 and R 7 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, and / or

[0211] R 4 and R 5 Together or R 6 and R 7 Together they form a cyclopropyl ring or a cyclobutyl ring, or

[0212] R 5 and R 6 Together they form 5- to 6-membered carbon rings or heterocycles with Y as a ring member.

[0213] In at least one embodiment of the compound of structural formula I, the chemical group Y of the X portion is selected from –S– or –O–, and R 4 R5 R 6 and R 7 Each is independently hydrogen or a C1-C4 straight-chain or branched alkyl group.

[0214] The exemplary X portion of Y, where Y is a sulfur atom (-S-), can include any portion shown in Table 6 below.

[0215] Table 6

[0216]

[0217] The exemplary X portion of Y, where Y is an oxygen atom (-O-), can include any portion shown in Table 7 below.

[0218] Table 7

[0219]

[0220] In at least one embodiment of the compound of structural formula I, the chemical group Y of the X portion is –CR 8 R 9 –, where R 8 and R 9 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, or R 8 and R 9 Together they form cyclopropyl, cyclobutyl, cyclopentyl, or oxetyl butyl rings, and R 4 R 5 R 6 and R 7 Each of the chemical groups at the position is independently hydrogen or a C1-C4 straight-chain or branched alkyl group.

[0221] Y is –CR 8 R 9 The exemplary X portion can include any of the portions shown in Table 8 below.

[0222] Table 8

[0223]

[0224] The various inhibitory compounds of structural formula I provided in this disclosure include a series of compounds having various substituted thiophene ring moieties linked to various X moieties. Various FABP3 / 4 / 5 / 7 inhibitory compounds can be represented as compounds of structural formula II and any pharmaceutically acceptable salt thereof:

[0225]

[0226] (II)

[0227] in,

[0228] R1 Selected from hydrogen, cyano, and 5-membered heteroaryl rings;

[0229] R 2 and R 3 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, cyclopropyl, cyclobutyl, phenyl, trifluoromethyl and benzyl, or R 2 and R 3 Together they form a 5- to 8-membered monocyclic, bicyclic, or spirocyclic carbocyclic or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein the carbocyclic, heterocyclic, aryl, or heteroaryl ring is optionally substituted by one or two substituents selected from C1-C4 alkyl, methoxy, or fluorine.

[0230] Y is a heteroatom selected from –S– and –O– or –CR. 8 R 9 –, where R 8 and R 9 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, or R 8 and R 9 Together they form cyclopropyl, cyclobutyl, cyclopentyl, or oxetyl butyl rings;

[0231] R 4 R 5 R 6 and R 7 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, and / or

[0232] R 4 and R 5 Together or R 6 and R 7 Together they form a cyclopropyl ring or a cyclobutyl ring, or

[0233] R 5 and R 6 Together they form 5- to 6-membered carbon rings or heterocycles with Y as a ring member.

[0234] In at least one embodiment, the compounds of structural formula II of this disclosure do not include the compounds in Table 1 (see above).

[0235] Similar to the FABP3 / 4 / 5 / 7 inhibitor compounds of Formula I, compounds of Formula II comprise a series of compounds represented by substructures. For example, in at least one embodiment of a FABP3 / 4 / 5 / 7 inhibitor compound having Formula II, R 1 The chemical group at that position is a cyano group, and the compound has the structural formula (IIa).

[0236]

[0237] (IIa)

[0238] Among them, the chemical group R 2 R 3 R 4 R 5 R 6 and R 7 As defined for compounds of structure II.

[0239] In at least one embodiment, the compound having the substructure of formula IIa may have the structural formulas IIj, IIk, IIl, IIm, IIin, IIo, IIp, IIq, IIr, IIs and IIt as shown in Table 9 below.

[0240] Table 9

[0241]

[0242]

[0243] Among them, the chemical group R 4 R 5 R 6 and R 7 As defined for compounds of structure II.

[0244] In at least one embodiment of a FABP3 / 4 / 5 / 7 inhibitor compound having structural formula II, R 1 The chemical group at that position is a cyano group, R 2 and R 3 Together they form a 6-membered aryl ring, and the compound has the structural formula (IIb).

[0245]

[0246] (IIb)

[0247] Among them, the chemical group R 2 R 3 R 4 R 5 R 6 and R 7 As defined for compounds of structural formula II, and where R 10 and R 11 Each is independently selected from hydrogen, halogen, C1–C4 straight-chain or branched alkyl, cyclopropyl and cyclobutyl.

[0248] In at least one embodiment, the compound having the substructure of formula IIb may have the structural formulas IIu, IIv and IIw as shown in Table 10 below.

[0249] Table 10

[0250]

[0251] In at least one embodiment of the compound of structural formula II, R 1 The chemical group at position R is a 5-membered heteroaryl ring (e.g., a 3-substituted 1,2,4-oxadiazole), and R 2 and R 3 The chemical groups at each location are independently selected from hydrogen, C1 - C4 Straight-chain or branched alkyl, cyclopropyl, cyclobutyl, phenyl, and benzyl groups. For example, in at least one embodiment, the compound may have the structural formulas IIc, IId, IIe, IIf, IIg, IIh, and IIi shown in Table 11 below.

[0252] Table 11

[0253]

[0254] Among them, the chemical group R 4 R 5 R 6 and R 7 As defined for compounds of structural formula II, and the chemical groups in IIc, IId, IIe, IIf, IIg, and IIh R12 It can be hydrogen, halogen, C1-C4 straight-chain or branched alkyl, cyclopropyl, cyclobutyl or phenyl.

[0255] In at least one embodiment, compounds having substructures of the structural formulas IIc, IId, IIe, IIf, IIg, and IIh can have the structural formulas IIx, IIy, IIz, IIaa, IIbb, IIcc, IIdd, IIee, IIff, IIgg, IIhh, IIii, IIjj, IIkk, IIll, IImm, IInn, IIoo, IIpp, IIqq, IIrr, and IIss, as shown in Table 12 below.

[0256] Table 12

[0257]

[0258]

[0259] In each different substructure embodiment of the compound of structural formula II, including compounds of structural formulas IIa, IIb, IIc, IId, IIe, IIf, IIg, IIh, IIi, IIj, IIk, IIl, IIm, IIn, IIo, IIp, IIq, IIr, IIs, IIt, IIu, IIv, IIw, IIx, IIy, IIz, IIaa, IIbb, IIcc, IIdd, IIee, IIff, IIgg, IIhh, IIIii, IIjj, IIkk, IIll, IImm, IInn, IIoo, IIpp, IIqq, IIrr, and IIss, it is envisioned that the atom or chemical group represented by Y can be -S- or -O-, and the chemical group R 4 R 5 R 6 and R 7 Each can be independently hydrogen or a C1-C4 straight-chain or branched alkyl group. Therefore, exemplary FABP3 / 4 / 5 / 7 inhibitor compounds of structures I and II include, but are not limited to, compounds 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, and 65.

[0260] Table 13

[0261]

[0262]

[0263]

[0264]

[0265]

[0266]

[0267] As described elsewhere in this document, those skilled in the art will understand that the FABP3 / 4 / 5 / 7 compounds provided herein can exist in a variety of well-known, closely related, and / or equivalent forms that are not explicitly described by chemical structures and formulas. The FABP3 / 4 / 5 / 7 inhibitor compounds of formulas I and II of this disclosure (including the compounds in Tables 2 to 5 and Tables 9 to 13, and those described in the examples) are intended to include these closely related forms of compounds as defined by chemical structures and formulas, including but not limited to pharmaceutically acceptable salts of the compound, mixtures of stereoisomers of the compound, single stereoisomers of the compound, tautomer forms of the compound, and / or prodrug forms of the compound.

[0268] Preparation of FABP3 / 4 / 5 / 7 inhibitor compounds

[0269] This disclosure also provides methods for preparing the FABP3 / 4 / 5 / 7 inhibitor compounds disclosed herein, including compounds of structural formulas I and II (as defined elsewhere herein), the methods being generally summarized in Scheme A and described in more detail below.

[0270] Option A

[0271]

[0272] A mixture of the substituted anhydride compound of formula III and the substituted 2-aminothiophene compound of formula IV (1:0.75 molar ratio) was purged with argon and then dissolved in an anhydrous solvent. The reaction mixture was then stirred at a temperature ranging from room temperature to the reflux temperature of the selected solvent. The progress of the reaction could then be monitored by one or more analytical methods, such as thin-layer chromatography (TLC), gas chromatography, etc. Once the starting material 2-aminothiophene IV or the substituted anhydride III was considered consumed, the solvent was removed under vacuum to obtain the desired FABP3 / 4 / 5 / 7 inhibitor of formula II.

[0273] In at least one embodiment, a method for preparing the disclosed FABP3 / 4 / 5 / 7 inhibitor compound of structural formula II includes:

[0274] (a) Assemblage of substituted acid anhydrides of formula III in solvent:

[0275]

[0276] (III)

[0277] Where Y is a heteroatom selected from –S– and –O– or –CR. 8 R 9 –, where R 8 and R 9Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, or R 8 and R 9 Together they form cyclopropyl, cyclobutyl, cyclopentyl, or oxetyl butyl rings; R 4 R 5 R 6 and R 7 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, phenyl and benzyl, and / or R. 4 and R 5 Together or R 6 and R 7 Together they form a cyclopropyl ring or a cyclobutyl ring, or R 5 and R 6 Together they form 5- to 6-membered carbon rings or heterocycles with Y as a ring member;

[0278] With substituted 2-aminothiophene compounds of formula IV:

[0279]

[0280] (IV)

[0281] Among them, R 1 Selected from hydrogen, cyano, and 5-membered heteroaryl rings; and R 2 and R 3 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, cyclopropyl, cyclobutyl, phenyl and benzyl, or R 2 and R 3 Together they form a 5- to 8-membered monocyclic, bicyclic, or spirocyclic carbocyclic or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein the carbocyclic, heterocyclic, aryl, or heteroaryl ring is optionally substituted by one or two substituents selected from C1-C4 alkyl, methoxy, or fluorine.

[0282] as well as

[0283] (b) Remove the solvent to obtain a compound having structural formula II:

[0284]

[0285] (II)

[0286] Where R 1 R 2 R 3 R 4 R 5 R 6 R 7 R 8 and R 9 As defined above.

[0287] In at least one embodiment of the substituted anhydride of Formula III, the chemical group Y is a sulfur atom. In another embodiment of the anhydride of Formula III, the chemical group Y is an oxygen atom. In a further embodiment of the anhydride of Formula III, the chemical group Y is –CR 5 R 6 –, where R 5 and R 6 Each is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl groups. In a further embodiment, when Y is R 5 and R 6 When each is independently selected from C1-C4 straight-chain alkyl groups, R 5 and R 6 They can together form a spiroring with 4 to 7 atoms. In a further embodiment, when Y is sulfur or oxygen, R 1 and R 4 They can together form heterocycles having 4 to 6 carbon atoms. The examples further describe a series of specific acid anhydride compounds of formula III that can be used to prepare formula II compounds.

[0288] In at least one embodiment of the substituted 2-aminothiophene compound of formula IV, the compound is a compound of structural formula IVa:

[0289]

[0290] (IVa)

[0291] Among them, R 2 and R 3 Each of the chemical groups at the position is independently selected from hydrogen, C1-C4 straight-chain or branched alkyl, cyclopropyl, cyclobutyl, phenyl, and benzyl, and optionally substituted by one or two substituents selected from C1-C4 alkyl, methoxy, chlorine, or fluorine. Exemplary compounds of this type with structural formula IVa include, but are not limited to, compounds 4a, 4b, 4c, 4d, 4e, and 4f shown in Table 14 below.

[0292] Table 14

[0293]

[0294] In at least one embodiment of the substituted 2-aminothiophene compound of formula IVa, R 2 and R 3 The chemical substituents at the positions together form 5 to 8-membered monocyclic, bicyclic, or spirocyclic carbocyclic or heterocyclic rings. Exemplary compounds of this type with structural formula IVa include, but are not limited to, compounds 4g, 4h, 4i, 4j, and 4k shown in Table 15 below.

[0295] Table 15

[0296]

[0297] In at least one embodiment of the substituted 2-aminothiophene compound of formula IVa, R 2 and R 3 The chemical substituents at the positions together form a 5- to 8-membered aryl or heteroaryl ring, wherein the carbocyclic, heterocyclic, aryl, or heteroaryl ring is optionally substituted by one or two substituents selected from C1-C4 alkyl, methoxy, or fluorine, as depicted in compounds of structural formula IVb:

[0298]

[0299] (IVb)

[0300] Where R 10 and R 11 Each compound is independently selected from hydrogen, halogen, C1-C4 straight-chain or branched alkyl, cyclopropyl, and cyclobutyl. Such compounds of structural formula IVb include, but are not limited to, compounds 4l, 4m, and 4n shown in Table 16 below.

[0301] Table 16

[0302]

[0303] In at least one embodiment of the substituted 2-aminothiophene compound of formula IV, R 1 The chemical group at the location is a 5-membered heteroaryl ring (e.g., a 3-substituted 1,2,4-oxadiazole), and the compound is selected from compounds with structural formulas IVc, IVd, IVe, IVf, IVg, IVh and IVi shown in Table 17 below.

[0304] Table 17

[0305]

[0306] Among them, R 2 and R 3 The chemical groups at each position are independently selected from hydrogen, C1-C4 straight-chain or branched alkyl groups, cyclopropyl, cyclobutyl, phenyl, and benzyl, or R 2 and R 3 Together they form a 5- to 8-membered monocyclic, bicyclic, or spirocyclic carbocyclic or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein the carbocyclic, heterocyclic, aryl, or heteroaryl ring is optionally substituted by one or two substituents selected from C1-C4 alkyl, methoxy, or fluorine groups; and R 12The chemical groups at the position are selected from hydrogen, halogens, C1–C4 straight-chain or branched alkyl groups, cyclopropyl, cyclobutyl, and phenyl. Compounds of this class with structural formulas IVc, IVd, IVe, IVf, IVg, IVh, and IVi include, but are not limited to, compounds 4o, 4p, 4q, 4r, 4s, 4t, 4u, 4v, 4w, 4x, 4y, 4z, 4aa, 4bb, 4cc, 4dd, 4ee, 4ff, 4gg, 4hh, and 4ii, as shown in Table 18 below.

[0307] Table 18

[0308]

[0309]

[0310] Synthetic methods for 2-aminothiophenes of structure IV have been previously reported, for example in U.S. Patent No. 9,353,102, U.S. Patent Publication No. 2015 / 0175594A1, and PCT Publication No. WO2014 / 040938, each of which is incorporated herein by reference. These previously reported synthetic methods can be used to prepare starting materials useful in the synthetic routes for preparing compounds of structure II as described herein and in the examples below. In particular, these starting materials are useful in the preparation of intermediate compounds of structures IVa, IVb, IVc, IVd, IVe, IVf, IVg, or IVh as shown above.

[0311] For example, compounds with the structure IVd can be synthesized using the route shown in Scheme B below.

[0312] Option B

[0313]

[0314] Another route for synthesizing compounds with the structure IVd has been proposed by J. Sarvanan et al. (Indian Journal of Heterocyclic Chemistry 1998). 7 As described in (285-288), and shown in Scheme C below.

[0315] Option C

[0316]

[0317] The route for synthesizing compounds of formula IVE from 2-amino-3-cyanothiophene has been described by RWSabnis et al. ( J. Het. Chem. 1992, 4, 285-288), and shown in Scheme D below.

[0318] Option D

[0319]

[0320] Another synthetic route for synthesizing compounds with the structural formula IVE has been described by JK Augustine et al. ( Tetrahedron 2009, 65 In , 9989-9996), and shown in the following scheme E.

[0321] Option E

[0322]

[0323] Another useful method for introducing 1,3,4-thiadiazole ring compounds with the structural formula IVie has been reported by V. Polshettiwar et al. Tetrahedron Lett ., 2008, 49 , 879 in.

[0324] An exemplary synthetic route for preparing compounds of structural formula IVf from 2-amino-3-cyanothiophene is illustrated in Scheme F below.

[0325] Option F

[0326]

[0327] Alternative synthetic routes for preparing compounds of formula IVf are described in scheme G below.

[0328] Option G

[0329]

[0330] Exemplary synthetic routes for preparing compounds of formula IVg or IVh from 2-amino-3-cyanothiophene are illustrated in Scheme H below (see also, for example, ZP Demko et al.). J. Org. Chem ., 2001, 66 , 7945-7950).

[0331] Option H

[0332]

[0333] As shown in the reaction scheme above, in some cases, it is advantageous to protect the 2-amino substituent of the 2-aminothiophene precursor with a Boc group using standard methods before proceeding with further reactions.

[0334] The examples illustrate a general synthetic route for preparing compounds of formula IVc (see, for example, scheme 21). Furthermore, the examples describe specific 2-aminothiophene compounds of formula VI that can be used to prepare compounds of formula II.

[0335] Non-limiting examples of a general procedure for preparing the disclosed FABP3 / 4 / 5 / 7 inhibitors. A mixture of substituted anhydride III and substituted 2-aminothiophene (1:0.75 molar ratio) was purged with argon and then dissolved in anhydrous dichloromethane. The reaction mixture was stirred at room temperature for 24 h. The solvent was removed using a rotary evaporator. The residue was then dissolved in 5 mL of ice-cold dichloromethane and transferred to a glass vanillin vial. The vial was then cooled on dry ice until visible crystals formed. The resulting crystals were separated by vacuum filtration and washed with ice-cold dichloromethane. The crystals were then dried in air for 30 min by vacuum filtration. A small amount of the separated crystalline sample was dissolved in acetone and its purity was verified by silica gel thin-layer chromatography using a solvent system of hexane containing 40% ethyl acetate and 0.1% acetic acid. The plate was stained with PMA as a universal staining agent, and the formation of carboxylic acids was verified by bromocresol green staining. 1 The structure of the purified crystals was verified by 1H NMR. The crystals were transferred to clean, pre-weighed glass vials, and the yield was calculated. The examples below provide specific synthetic procedures and reagents for the preparation of compounds of structures I and II (including the specific compounds in Table 1).

[0336] Uses and treatments

[0337] As described elsewhere herein, the FABP3 / 4 / 5 / 7 inhibitor compounds of Formulas I and II disclosed herein have shown potential therapeutic effects based on in vitro, preclinical, or clinical studies in a variety of diseases and ailments. Therefore, this disclosure contemplates the use of the inhibitor compounds of this disclosure in compositions and methods for treating diseases and / or ailments known to be affected by FABP3, FABP4, FABP5, and / or FABP7. Generally, methods of treating a subject suffering from a disease or ailment affected by FABP3, FABP4, FABP5, and / or FABP7 using the FABP3 / 4 / 5 / 7 inhibitor compounds of this disclosure include administering a therapeutically effective amount of a compound of Formulas I and II, or a pharmaceutical composition comprising such a compound and one or more pharmaceutically acceptable adjuvant ingredients, to a subject in need.

[0338] The following are known disorders and diseases that may be affected by one or more of FABP4, FABP5, FABP3, and FABP7, and are proposed for treatment with inhibitory compounds of FABP3 / 4 / 5 / 7 of structural formulas I and II, including but not limited to: atherosclerosis, coronary atherosclerosis, arterial fibrosis, pulmonary hypertension, heart failure, obesity, type 2 diabetes, type 1 diabetes, gestational diabetes, polycystic ovary syndrome, endometriosis, disorders affected by lipid metabolism and serum free fatty acid levels, metabolic disorders, fatty liver disease, renal fibrosis, systemic inflammation, acute inflammation, allergic inflammation, airway inflammation, viral infections (e.g., COVID-19, common cold), skin diseases (e.g., vitiligo, psoriasis, atopic dermatitis, allergic contact dermatitis, mycosis fungoides, alopecia areata, cicatricial alopecia, graft-versus-host disease (GvHD), contact dermatitis, chronic eczema, herpetic dermatitis, cutaneous lupus, scleroderma, dermatomyositis, vascular diseases). Inflammation, pemphigus, epidermolysis bullosa, linear IgA, vesicular diseases), neurological disorders and diseases (e.g., pain, multiple sclerosis (MS), Parkinson's disease), autoimmune diseases (e.g., experimental autoimmune encephalomyelitis (EAE), asthma, type 1 diabetes, autoimmune lung disease, autoimmune hepatitis, rheumatoid arthritis (RA), spondyloarthritis, vesicular stomatitis virus infection, multiple sclerosis (MS), lupus nephritis, Crohn's disease, ulcerative colitis) Colitis and food allergies), ischemic stroke, graft-versus-host disease (GvHD), and cancers (e.g., breast cancer, prostate cancer, ovarian cancer, skin cancer, gastric cancer, glioma, bile duct cancer, bladder cancer, multiple myeloma, colorectal cancer, hepatocellular carcinoma, cervical cancer, oral squamous cell carcinoma, and / or non-small cell lung cancer (NSCLC)). Further detailed descriptions of the various uses and therapeutic indications of the FABP3 / 4 / 5 / 7 inhibitor compounds of structures I and II are provided below.

[0339] A. Cancer Treatment

[0340] As described elsewhere in this paper, inhibition of FABP5 provides a method for inhibiting the metastasis of human cancer cells. Triple-negative breast cancer (TNBC) accounts for approximately 10-20% of all breast cancers. The term "triple-negative breast cancer" refers to a condition in which cancer cells do not produce sufficient estrogen or progesterone receptors, or insufficient amounts of human epidermal growth factor receptor 2 (HEGR-2) protein. Because TNBC tumors lack well-defined prognostic markers and selective therapeutic targets, the treatment and management of this disease are significant clinical issues, and there is an urgent need for direct methods to inhibit the biological processes that regulate tumorigenesis and metastasis. Not wishing to be limited by theory, the data disclosed in this paper demonstrate that the disclosed FABP inhibitor compounds provide inhibition of FABP5 and, consequently, modulate TNBC levels.

[0341] Therefore, in at least one embodiment, the FABP3 / 4 / 5 / 7 inhibitor compounds of structural formulas I and II can be used in a method of treating a subject's cancer, wherein the method includes administering a composition to a subject in need, the composition comprising: (a) an effective amount of one or more of the disclosed FABP3 / 4 / 5 / 7 inhibitors or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier or other adjuvant. In at least one embodiment of the method, the subject may be diagnosed with, have, and / or be treating one or more cancers selected from breast cancer, prostate cancer, ovarian cancer, skin cancer, gastric cancer, glioma, cholangiocarcinoma, bladder cancer, multiple myeloma, colorectal cancer, hepatocellular carcinoma, cervical cancer, oral squamous cell carcinoma, and / or non-small cell lung cancer (NSCLC). In one embodiment of the disclosed cancer treatment method, breast cancer is involved. In another embodiment of the disclosed method, the prevention of metastasis of TNBC cells in a subject diagnosed with cancer is involved.

[0342] Another further aspect of the disclosed method relates to a method for treating a subject with cancer, comprising administering a composition to a subject in need, said composition comprising: (a) an effective amount of one or more of the disclosed FABP3 / 4 / 5 / 7 inhibitors or pharmaceutically acceptable salts thereof; and (b) a pharmaceutically acceptable carrier or other adjuvant. In at least one embodiment of the cancer treatment method, the cancer is selected from breast cancer, prostate cancer, ovarian cancer, hepatocellular carcinoma, multiple myeloma, neuroblastoma, lung adenocarcinoma, or gastric cancer. In one example, the cancer is breast cancer. In a further example, the cancer is prostate cancer. In another example, the cancer is ovarian cancer. In yet another example, the cancer is hepatocellular carcinoma. In a further example, the cancer is multiple myeloma. In yet another example, the cancer is neuroblastoma. In yet another example, the cancer is lung adenocarcinoma. In yet another example, the cancer is gastric cancer.

[0343] In at least another embodiment, the FABP3 / 4 / 5 / 7 inhibitors of this disclosure are envisioned for use in methods of sensitizing cancer cells to other chemotherapeutic agents. Such agents may include standard chemotherapeutic compounds such as doxorubicin, gemcitabine, cisplatin, paclitaxel, all-trans retinoic acid (atRA), PARP inhibitor compounds, and immune checkpoint inhibitor compounds, including but not limited to antibodies targeting PD-1 or PD-L1. Therefore, in at least one embodiment, this disclosure provides a method of sensitizing cancer cells to other chemotherapeutic agents, said method comprising contacting cancer cells with one or more of the disclosed FABP3 / 4 / 5 / 7 inhibitors and contacting the cells with one or more chemotherapeutic agents. This method is envisioned to be performed by contacting the FABP3 / 4 / 5 / 7 inhibitors with the cancer cells before, in parallel with, or after contacting the cancer cells with the chemotherapeutic agents.

[0344] In at least one embodiment, this disclosure also provides the use of a FABP3 / 4 / 5 / 7 inhibitor compound of structural formula I or II, or a pharmaceutical composition comprising such a compound, in the manufacture of a medicament for treating a subject's cancer. In at least one embodiment, the cancer treated by said use or the medicament is selected from breast cancer, prostate cancer, ovarian cancer, hepatocellular carcinoma, multiple myeloma, neuroblastoma, lung adenocarcinoma, or gastric cancer.

[0345] B. Fatty acid control

[0346] One aspect of the disclosed uses and methods relates to a method for inhibiting one or more of FABP3, FABP4, FABP5 and FABP7 in a subject, comprising administering a composition to a subject in need, said composition comprising: (a) an effective amount of one or more FABP3 / 4 / 5 / 7 inhibitory compounds of structural formula I or II or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier or other adjuvant.

[0347] Another aspect of the disclosed uses and methods relates to a method for controlling serum free fatty acid levels in a subject, comprising administering a composition to a subject in need, said composition comprising: (a) an effective amount of one or more of the disclosed FABP3 / 4 / 5 / 7 inhibitors of formula I or II or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier or other adjuvant.

[0348] In at least one embodiment, this disclosure also provides the use of the FABP3 / 4 / 5 / 7 inhibitor compound of this disclosure or a pharmaceutical composition comprising a FABP3 / 4 / 5 / 7 inhibitor compound of structural formula I or II in the manufacture of a medicament for treating a disease or ailment affected by FABP3 / 4 / 5 / 7 in a subject. In at least one embodiment, the disease or ailment involves controlling the subject's serum free fatty acid levels.

[0349] C. Treatment of metabolic disorders

[0350] In addition, FABP4 and FABP5 are members of a small family of soluble proteins that facilitate the transport of fatty acids within the cytoplasmic compartments of cells. These proteins do not have catalytic functions; instead, they transport hydrophobic fatty acids to various destinations in the aqueous environment of the cytoplasm, thereby enabling fatty acid oxidation, membrane homeostasis, or nuclear signaling. Furthermore, they are involved in signal transduction processes, the details of which are poorly understood. FABP4 is highly expressed in adipose tissue, macrophages, and endothelial cells. FABP5 is also expressed in macrophages, adipocytes, and endothelial cells, as well as in skin and several other tissues.

[0351] Unwilling to be limited by theory, plasma FABP4 levels are elevated in humans with metabolic syndrome and atherosclerosis. Furthermore, there is evidence that FABP4 is involved in angiogenesis. More than a quarter of the population suffers from multiple comorbidities, including obesity, atherosclerosis, insulin resistance, dyslipidemia, coagulation disorders, hypertension, and a pro-inflammatory state known as metabolic syndrome. Patients with metabolic syndrome are at high risk of developing atherosclerosis, type 2 diabetes, and other health problems. Similar to obesity, treatment options for atherosclerosis are very limited.

[0352] A further aspect of the disclosed method relates to a method for modulating insulin sensitivity in a subject, comprising administering a composition to a subject in need, said composition comprising: (a) an effective amount of one or more FABP3 / 4 / 5 / 7 inhibitor compounds of structural formula I or II or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier or other adjuvant.

[0353] A further aspect of the disclosed method relates to a method for treating a subject with type 2 diabetes, comprising administering a composition to a subject in need, said composition comprising: (a) an effective amount of one or more FABP3 / 4 / 5 / 7 inhibitor compounds of formula I or II or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier or other adjuvant.

[0354] A further aspect of the disclosed method relates to a method for measuring plasma glucose levels in a subject, comprising administering a composition to a subject in need, the composition comprising: (a) an effective amount of one or more FABP3 / 4 / 5 / 7 inhibitor compounds of structural formula I or II or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier or other adjuvant.

[0355] A further aspect of the disclosed method relates to a method for treating atherosclerosis in a subject, comprising administering a composition to the subject in need, said composition comprising: (a) an effective amount of one or more FABP3 / 4 / 5 / 7 inhibitor compounds of structural formula I or II or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier or other adjuvant.

[0356] A further aspect of the disclosed method relates to a method for treating hepatic steatosis in a subject, comprising administering a composition to a subject in need, said composition comprising: (a) an effective amount of one or more FABP3 / 4 / 5 / 7 inhibitor compounds of structural formula I or II or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier or other adjuvant.

[0357] In at least one embodiment, this disclosure also provides the use of a FABP3 / 4 / 5 / 7 inhibitor compound of structural formula I or II, or a pharmaceutical composition comprising a FABP3 / 4 / 5 / 7 inhibitor compound of this disclosure, in the manufacture of a medicament for treating a subject’s metabolic disorder.

[0358] D. Regulation of immune cell activity and immune cell populations

[0359] As described elsewhere in this paper, FABPs are involved in the regulation of immune cell activity. While not intended to be mechanistically limited, it is believed that FABPs mediate immune cell metabolism, which is crucial for the normal functioning of the immune system. More specifically, FABP activity affects fatty acid utilization, thereby regulating energy production and signaling pathways involved in immune cell activation and function. For example, FABP5 has been found to regulate lipid metabolism and function of T cells in the tumor microenvironment (TME) by mediating the uptake and oxidation of intracellular long-chain fatty acids (FAs). Furthermore, due to limited glucose availability and high levels of long-chain FAs, tumor-infiltrating T lymphocytes (TILs) expressing high levels of FABP5 typically exhibit an exhausted phenotype and impaired antitumor activity. Therefore, inhibition of FABP5 in TILs is expected to activate cellular antitumor activity. FABP4 in Ly6C - MHCII - CD36 +High expression of FABP4 in circulating monocytes / macrophages promotes oxidized lipid uptake, foam cell formation, angiogenesis, tissue remodeling, and pro-tumorigenic functions. This high expression suggests that inhibiting FABP4 can enhance anti-tumor immune responses in cancer cells and may also prevent foam cell formation and chronic inflammation in obesity. Another function of FABP4 / 5 in immune cells is their role in maintaining CD8+ tissue-resident memory T cells (Trm). Specifically, according to other literature, skin TRMs that rely on fatty acids as an energy source express high levels of FABP4 / 5, which is necessary for fatty acid uptake by cells and transport to mitochondria for metabolism. The energy generated in this process is essential for TRM cell survival. Targeting TRM cells by inhibiting FABP4 / 5 is expected to have therapeutic effects in autoimmune diseases. Overall, the role of FABP in immune cell regulation highlights the importance of lipid metabolism in immune system regulation and suggests that targeting FABP could be a promising strategy for improving immune cell function and treating a variety of diseases and disorders caused by chronic inflammation and cancer.

[0360] A method for modulating the immune cell population and / or immune cell activity of a subject in need, comprising administering to the subject in need a therapeutically effective amount of a compound of formula I or II, or a pharmaceutical composition of such a compound. In at least one embodiment, the subject in need suffers from a disease or condition caused by, affected by, and / or characterized by the immune cell population and / or immune cell activity, for example, wherein the immune cells are M2 macrophages or tumor-associated macrophages (TAMs). In at least one embodiment, the subject requiring treatment to modulate the immune cell population and / or immune cell activity has been diagnosed with, is suffering from, and / or is being treated for cancer. In another embodiment, it is envisioned that the subject requiring treatment to modulate the immune cell population and / or immune cell activity suffers from an autoimmune disease or condition.

[0361] Pharmaceutical Composition

[0362] This disclosure also provides uses and methods for administering FABP4 / 5 inhibitor compounds, such as those of formula I or II, to a subject in the form of a pharmaceutical composition. In such embodiments, the pharmaceutical composition comprises a therapeutically effective amount of a FABP3 / 4 / 5 / 7 inhibitor compound (e.g., compounds in Table 11), or a pharmaceutically acceptable salt or ester of such a compound, and one or more pharmaceutically acceptable carriers. Such pharmaceutical compositions can be prepared using methods well-known in the pharmaceutical industry. For example , See Remington's Pharmaceutical Sciences Mace Publishing Co., Philadelphia, PA, 17th edition (1985) and Modern PharmaceuticsMarcel Dekker, Inc., 3rd Edition (edited by GS Banker and CT Rhodes). This disclosure describes methods for preparing pharmaceutical compositions of FABP3 / 4 / 5 / 7 inhibitor compounds, including the examples disclosed herein.

[0363] In at least one embodiment, this disclosure provides a pharmaceutical composition comprising: an effective amount of one or more disclosed FABP3 / 4 / 5 / 7 inhibitors, such as compounds of structural formula I or II; and one or more adjuvants, such as pharmaceutically acceptable carriers. The disclosed composition may comprise from about 10% to about 95% by weight of one or more disclosed FABP3 / 4 / 5 / 7 inhibitors. In another embodiment, the composition comprises from about 10% to about 80% by weight of one or more disclosed FABP3 / 4 / 5 / 7 inhibitors. In a further embodiment, the composition comprises from about 20% to about 50% by weight of one or more disclosed FABP3 / 4 / 5 / 7 inhibitors. In a still further embodiment, the composition comprises from about 50% to about 90% by weight of one or more disclosed FABP3 / 4 / 5 / 7 inhibitors. In yet another embodiment, the composition comprises from about 70% to about 90% by weight of one or more disclosed FABP3 / 4 / 5 / 7 inhibitors. In a further embodiment, the composition comprises about 80% to about 95% by weight of one or more of the disclosed FABP3 / 4 / 5 / 7 inhibitors. In a still further embodiment, the composition comprises about 90% to about 95% by weight of one or more of the disclosed FABP3 / 4 / 5 / 7 inhibitors.

[0364] Generally, pharmaceutical compositions can be prepared by diluting the active ingredient with an excipient and / or encapsulating it within a carrier in the form of capsules, pouches, paper, or other containers. When the excipient is used as a diluent, it can be a solid, semi-solid, or liquid material (as described above), acting as a medium, carrier, or mediator of the active ingredient. Therefore, pharmaceutical compositions suitable for administration in the methods of this disclosure can be tablets, pills, powders, lozenges, pouches, capsules, elixirs, suspensions, emulsions, solutions, syrups, aerosols (solid or liquid media), such as ointments containing up to 10% by weight of the active compound, gelatin soft and hard capsules, sterile injectable solutions, and sterile packaged powders.

[0365] The carrier used in the preparation of the pharmaceutical composition may include excipients such as inert solid diluents and fillers, diluents including sterile aqueous solutions and various organic solvents, penetration enhancers, solubilizers, and adjuvants. Suitable excipients used in the pharmaceutical compositions comprising triptolide derivatives disclosed herein are well known in the art and include lactose, glucose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginate, tragacanth gum, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, sterile water, syrup, and methylcellulose. The pharmaceutical composition may additionally include: lubricants such as talc, magnesium stearate, and mineral oil; wetting agents; emulsifiers and suspending agents; preservatives such as methylparaben and propylparaben; sweeteners; and flavoring agents.

[0366] In terms of use and treatment methods, it is envisioned that pharmaceutical compositions containing FABP3 / 4 / 5 / 7 inhibitory compounds (such as compounds of structural formula I) be administered as a single dose or multiple doses via any recognized mode of administration of an active ingredient with similar efficacy. For example, pharmaceutical compositions containing triptolide derivatives can be administered in a variety of different ways, including oral administration, intravenous administration, topical administration, parenteral administration, intraperitoneal administration, intramuscular administration, intrathecal administration, intrathecal administration, intralesional administration, intralesional administration, intracranial administration, intranasal administration, intraocular administration, intracardiac administration, intravitreal administration, intraosseous administration, intracerebral administration, intra-arterial administration, intra-articular administration, intradermal administration, transdermal administration, transmucosal administration, sublingual administration, enteral administration, sublipal administration, insufflation administration, suppository administration, inhalation administration, or subcutaneous administration.

[0367] The pharmaceutical compositions comprising FABP3 / 4 / 5 / 7 inhibitor compounds disclosed herein can be used in a range of treatment methods, and a range of dosages are envisioned to achieve pharmaceutically effective dosage. The dosage and frequency (single or multiple doses) of the pharmaceutical composition administered to a subject can vary depending on a range of factors, such as the route of administration; the subject's body type, age, sex, health status, weight, and / or diet; the state of the disease being treated; whether the subject has any other diseases; and any concurrent treatments being received. Those skilled in the art will understand that predetermined dosages (e.g., frequency and duration) may need to be adjusted based on the subject to achieve a therapeutically effective dosage. Typically, the amount of the pharmaceutical composition comprising FABP3 / 4 / 5 / 7 inhibitor compounds to be administered to a subject in a treatment method will be determined by the physician based on relevant circumstances of the subject being treated, the chosen route of administration, and of course, factors such as age, weight, symptom severity, and the individual subject's response to treatment.

[0368] Generally, a therapeutically effective amount is the amount of the composition applied, relative to the absence of the compound, sufficient to achieve the desired therapeutic purpose. For example, a therapeutically effective amount may be an amount determined to be sufficient to promote the treatment, prevention, or relief of one or more symptoms of a disease. Methods for determining the dosage of a compound that provides a therapeutically effective amount are well known to those skilled in the art and are typically based on analysis of amounts determined in cell assays and / or animal models. For example, a dosage for administration to humans may be formulated to achieve concentrations that have been observed to be therapeutically effective in animal models. The dosage in the pharmaceutical composition for humans can be further adjusted by monitoring therapeutic efficacy and making adjustments upward or downward. Those skilled in the art can use methods well known in the art to adjust the dosage in the pharmaceutical compositions of this disclosure to achieve maximum therapeutic efficacy in humans.

[0369] Generally, the development of methods for therapeutic treatment begins with pharmaceutical compositions containing FABP3 / 4 / 5 / 7 inhibitor compounds at doses below the optimum level. Subsequently, the dosage of the compound is gradually increased until optimal efficacy is achieved. A key factor to consider when developing the optimum dosage is the ratio between the toxicity of the active ingredient and its therapeutic efficacy. This ratio, known as the therapeutic index of the compound, is often described as the LD50 of the active ingredient. 50 (The amount of compound that is 50% lethal to a population) and its ED 50 The ratio of the amount of compound effective in 50% of the population. Generally, a higher therapeutic index is preferred for the compound. Therapeutic index data can be obtained from cell culture assays and / or animal model studies and are then used to determine the safe dose range of the active ingredient in the pharmaceutical composition for human administration. Ideally, the determined dose provides the effect of achieving ED in the subject. 50 The active ingredients are present at low levels, while having little or no toxicity.

[0370] Solid forms of pharmaceutical compositions may include, for example, powders, tablets, dispersible granules, capsules, flat capsules, and suppositories. A solid carrier may be one or more substances that can also function as a diluent, flavoring agent, solubilizer, lubricant, suspending agent, binder, or tablet disintegrant; it may also be an encapsulation material.

[0371] In powder form, the carrier is typically a finely dispersed solid mixed with a finely dispersed active ingredient (e.g., the disclosed FABP3 / 4 / 5 / 7 inhibitor). In tablet form, the active ingredient is mixed with a carrier having the necessary binding properties in a suitable proportion and compressed into the desired shape and size. Solid formulations of the pharmaceutical compositions disclosed herein may contain a binder from about 0.5% to about 10% by weight. Non-limiting examples of binders suitable for use in the disclosed compositions are selected from polyethylene glycol 1500, polyethylene glycol 2000, polyethylene glycol 3000, polyethylene glycol 4000, polyethylene glycol 6000, polyethylene glycol 8000, polyethylene glycol 10000, methylcellulose, ethylcellulose, hydroxymethylcellulose, hydroxyethylcellulose, hydroxypropyl methylcellulose, polyethylene oxide, polyethylene oxide-polypropylene copolymer, and mixtures thereof. In one embodiment, the binder is methylcellulose, ethylcellulose, hydroxymethylcellulose, or hydroxyethylcellulose. In a non-limiting example, the binder is ethylcellulose.

[0372] In some embodiments, the solid composition may comprise about 0.5% to about 10% by weight of a carrier. Non-limiting examples of solid carriers include: starches such as tapioca starch, corn starch, potato starch, gelatin, dextrin, inulin, cyclodextrin, oxidized starch, starch esters, starch ethers, cross-linked starch, α-starch, octenyl succinate, and processed starches obtained by acid, heat, or enzymatic treatment; or emulsifiers such as gum arabic, modified starch, pectin, xanthan gum, gum indica, tragacanth gum, fenugreek gum, mead gum, monoglycerides and diglycerides of long-chain fatty acids, sucrose monoesters, sorbitan esters, polyethoxylated glycerol, stearic acid, palmitic acid, monoglycerides, diglycerides, propylene glycol esters, lecithin, lactated monoglycerides and diglycerides, propylene glycol monoesters, polyglycerides, diacetylated tartrate esters of monoglycerides and diglycerides, citrate esters of monoglycerides, stearoyl-2-lactone... Esters, polysorbates, succinylated monoglycerides, acetylated monoglycerides, ethoxylated monoglycerides, soap gums, whey protein isolates, casein, soy protein, plant protein, pullulan, sodium alginate, guar gum, locust bean gum, tragacanth gum, tamarind gum, carrageenan, furcelellaran, gellan gum, psyllium gum, curdlan, konjac mannan, agar and cellulose derivatives, and combinations thereof, or optionally sugar alcohols with moisturizing properties, such as ethylene glycol, glycerin, erythritol, threitol, arabinitol, xylitol, ribitol, mannitol, sorbitol, galactitol, fructose, idutol, sucrose, fructose, isomaltitol, maltitol, lactitol, sorbitol, glucose or inositol, and combinations thereof.

[0373] The disclosed compositions may contain one or more of the disclosed FABP3 / 4 / 5 / 7 inhibitors, from about 25 mg to about 1200 mg. In one aspect, the disclosed single-dose composition of the disclosed FABP3 / 4 / 5 / 7 inhibitor may contain any amount from about 25 mg to about 500 mg.

[0374] In a further aspect, the disclosed single-dose composition of the disclosed FABP3 / 4 / 5 / 7 inhibitor may contain any amount from about 100 mg to about 500 mg. In an even further aspect, the disclosed single-dose composition of the disclosed FABP3 / 4 / 5 / 7 inhibitor may contain any amount from about 500 mg to about 1000 mg.

[0375] A single-dose composition may contain any amount of FABP3 / 4 / 5 / 7 inhibitors, ranging from about 25 mg to about 250 mg. For example, the disclosed compositions may contain 25 mg, 26 mg, 27 mg, 28 mg, 29 mg, 30 mg, 31 mg, 32 mg, 33 mg, 34 mg, 35 mg, 36 mg, 37 mg, 38 mg, 39 mg, 40 mg, 41 mg, 42 mg, 43 mg, 44 mg, 45 mg, 46 mg, 47 mg, 48 mg, 49 mg, 50 mg, 51 mg, 52 mg, 53 mg, 54 mg, 55 mg, 56 mg, 57 mg, 58 mg, 59 mg, 60 mg, 61 mg, 62 mg, 63 mg, 64 mg, 65 mg, 66 mg, 67 mg, 68 mg, 69 mg, 70 mg, 71 mg, 72mg, 73 mg, 74 mg, 75 mg, 76 mg, 77 mg, 78 mg, 79 mg, 80 mg, 81 mg, 82 mg, 83 mg, 84 mg, 85mg, 86 mg, 87 mg, 88 mg, 89 mg, 90 mg, 90 mg, 91 mg, 92 mg, 93 mg, 94 mg, 95 mg, 96 mg, 97mg, 98 mg, 99 mg, 100 mg, 101 mg, 102, mg, 103, mg, 104 mg, 105 mg, 106 mg, 107 mg, 108mg, 109 mg, 120 mg, 121 mg, 122 mg, 123 mg, 124 mg, 125 mg, 126 mg, 127 mg, 128 mg, 129mg, 120 mg, 1 21 mg, 122 mg, 123 mg, 124 mg, 125 mg, 126 mg, 127 mg, 128 mg, 129 mg, 130mg 31 mg, 132 mg, 133 mg, 134 mg, 135 mg, 136 mg, 137 mg, 138 mg, 139 mg, 140 mg, 141mg, 142 mg, 143 mg, 144 mg, 145 mg, 146 mg, 147 mg, 148 mg, 149 mg, 150 mg, 151 mg, 152mg, 153 mg, 154 mg, 155 mg, 156 mg, 157 mg, 158 mg, 159 mg, 160 mg, 161 mg, 1 62 mg, 163 mg, 164 mg, 165 mg, 166 mg, 167 mg, 168 mg, 169 mg, 170mg, 171 mg, 172 mg, 173 mg, 174mg, 175 mg, 176 mg, 177 mg, 178 mg, 179 mg, 180 mg, 181 mg, 182 mg, 183 mg, 184 mg, 185mg, 186 mg, 187 mg, 188 mg, 189 mg, 190 mg, 190 mg, 191 mg, 192 mg 193 mg 194 mg 195 mg 196 mg 197 mg 198 mg 199 mg 200 mg 201 mg 202 mg mg, 216 One or more of the disclosed FABP3 / 4 / 5 / 7 inhibitors, in mg, 217 mg, 218 mg, 219 mg, 220 mg, 221 mg, 222 mg, 223 mg, 224 mg, 225 mg, 226 mg, 227 mg, 228 mg, 229 mg, 230 mg, 231 mg, 232 mg, 233 mg, 234 mg, 235 mg, 236 mg, 237 mg, 238 mg, 239 mg, 240 mg, 241 mg, 242 mg, 243 mg, 244 mg, 245 mg, 246 mg, 247 mg, 248 mg, 249 mg, or 250 mg.

[0376] Liquid forms of the pharmaceutical composition may include, for example, solutions suitable for oral or parenteral administration, suspensions suitable for oral administration, and emulsions. Examples of liquid compositions suitable for parenteral administration include sterile aqueous solutions of the active ingredient or sterile solutions of the active ingredient in solvents comprising water, buffered water, physiological saline, PBS, ethanol, or propylene glycol. The composition may contain pharmaceutically acceptable excipients, such as pH adjusters and buffers, osmotic regulators, wetting agents, detergents, etc., to approximate physiological conditions. In at least one embodiment, the disclosed liquid composition may contain about 5% to about 25% by weight of a liquid carrier.

[0377] For liquid embodiments of the compositions disclosed herein, target cells (e.g., cancer cells or tumor cells) may be contacted with an aqueous solution containing about 0.5 μg / mL to about 250 μg / mL. In one embodiment, the composition may contain about 1 μg / mL to about 100 μg / mL. In another embodiment, the composition may contain about 10 μg / mL to about 100 μg / mL. In a further embodiment, the composition may contain about 5 μg / mL to about 20 μg / mL. In yet another embodiment, the composition may contain about 1 μg / mL to about 50 μg / mL. In yet another embodiment, the composition may contain about 1 μg / mL to about 10 μg / mL. In a further still embodiment, the composition may contain about 15 μg / mL to about 50 μg / mL. In yet another embodiment, the composition may contain about 20 μg / mL to about 200 μg / mL.

[0378] The disclosed composition can deliver a single dose of the disclosed FABP3 / 4 / 5 / 7 inhibitor based on the weight of the subject being treated. Therefore, a single dose of the disclosed FABP3 / 4 / 5 / 7 inhibitor is in the range of about 0.35 mg / kg to about 20 mg / kg based on the subject's weight. In one embodiment, the amount of the disclosed FABP3 / 4 / 5 / 7 inhibitor in a single dose is about 1 mg / kg to about 8 mg / kg based on the subject's weight. In another embodiment, the amount of the disclosed FABP3 / 4 / 5 / 7 inhibitor in a single dose is about 2 mg / kg to about 5 mg / kg based on the subject's weight. In a further embodiment, the amount of the disclosed FABP3 / 4 / 5 / 7 inhibitor in a single dose is about 1.5 mg / kg to about 4 mg / kg based on the subject's weight. In a still further embodiment, the amount of the disclosed FABP3 / 4 / 5 / 7 inhibitor in a single dose is about 4 mg / kg to about 10 mg / kg based on the subject's weight. In a further embodiment, the amount of the disclosed FABP3 / 4 / 5 / 7 inhibitor in a single dose is approximately 5 mg / kg to approximately 8 mg / kg based on the subject's body weight.

[0379] For example, the dosage, based on the weight of the subject being treated, can include any amount from about 0.5 mg / kg to about 10 mg / kg.For example, based on body weight, 0.5 mg / kg, 0.6 mg / kg, 0.7 mg / kg, 0.8 mg / kg, 0.9 mg / kg, 1 mg / kg, 1.1 mg / kg, 1.2 mg / kg, 1.3 mg / kg, 1.4 mg / kg, 1.5 mg / kg, 1.6 mg / kg, 1.7 mg / kg, 1.8 mg / kg, 1.9 mg / kg, 2.0 mg / kg, 2.1 mg / kg, 2.2 mg / kg, 2.3 mg / kg, 2.4 mg / kg, 2.5 mg / kg, 2.6 mg / kg, 2.7 mg / kg, 2.8 mg / kg, 2.9 mg / kg, 3.0 mg / kg, 3.1 mg / kg, 3.2 mg / kg, 3.3 mg / kg, 3.4 mg / kg, 3.5 mg / kg, 3.6 mg / kg, 3.7 mg / kg, 3.8 mg / kg, 3.9 mg / kg, 4.0 mg / kg, 4.1 mg / kg, 4.2 mg / kg, 4.3 mg / kg, 4.4 mg / kg, 4.5 mg / kg, 4.6 mg / kg, 4.7 mg / kg, 4.8 mg / kg, 4.9 mg / kg, or 5.0 mg / kg, 5.1 mg / kg, 5.2 mg / kg, 5.3 mg / kg, 5.4 mg / kg, 5.5 mg / kg, 5.6 mg / kg, 5.7 mg / kg, 5.8 mg / kg, 5.9 mg / kg, 6.0 mg / kg, 6.1 mg / kg, 6.2 mg / kg, 6.3 mg / kg, 6.4 mg / kg, 6.5 mg / kg, 6.6 mg / kg, 6.7 mg / kg, 6.8 mg / kg, 6.9 mg / kg, 7.0 mg / kg, 7.1 mg / kg, 7.2 mg / kg, 7.3 mg / kg, 7.4 mg / kg, 7.5 mg / kg, 7.6 mg / kg, 7.7 mg / kg, 7.8 mg / kg, 7.9 mg / kg, 8.0 mg / kg, 8.1 mg / kg, 8.2 mg / kg, 8.3 mg / kg, 8.4 mg / kg, 8.5 mg / kg, 8.6 mg / kg, 8.7 mg / kg, 8.8 mg / kg, 8.9 mg / kg, 9.0 mg / kg, 9.1 mg / kg, 9.2 mg / kg, 9.3 mg / kg, 9.4 mg / kg, 9.5 mg / kg, 9.6 mg / kg, 9.7 mg / kg, 9.8 mg / kg, 9.9 mg / kg or 10.0 mg / kg.

[0380] Example

[0381] Various features and embodiments of this disclosure are illustrated in the following representative examples, which are intended to be illustrative and not restrictive. Those skilled in the art will readily recognize that the specific embodiments are merely illustrative of the invention, as more fully described in the following claims. Each embodiment and feature described in this application should be understood to be interchangeable and combinable with each embodiment contained therein.

[0382] The following examples illustrate the method for synthesizing the compound of Formula 1:

[0383] Example 1: 2-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[ b Preparation of thiophene-2-yl)amino)-2-oxoethyl)thio)acetic acid (compound FTS001)

[0384]

[0385] FTS001 (CAS 41922-19-1)

[0386] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS001 using the synthetic method described in Scheme 1 below.

[0387] Option 1

[0388]

[0389] Materials and methods

[0390] 1,4-oxothiacyclohexane-2,6-dione (0.552 mmol) and 2-amino-4,5,6,7-tetrahydrobenzo[ b Thiophene-3-carboxynitrile (0.552 mmol) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 20 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 2-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Thiophene-2-yl)amino)-2-oxoethyl)thio)acetic acid (137 mg, 80%).

[0391] NMR analysis confirmed the preparation of the desired product compound. 1H NMR (400 MHz, DMSO-d6) δ: 1.75ppm (4H, t, J= 6.0 Hz), 2.59 ppm (2H, m, J=6.0Hz), 3.41 ppm (2H,s), 3.55 ppm(2H,s) 11.68 (1H, s), 12.66 (1H, s).

[0392] Example 2: 2-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[ b Preparation of thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (compound FTS003)

[0393]

[0394] FTS003

[0395] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS003 using the synthetic method shown in Scheme 2 below.

[0396] Option 2

[0397]

[0398] Materials and methods

[0399] 3,3-Dimethyl-1,4-oxothiacyclohexane-2,6-dione (2.13 mmol) and 2-amino-4,5,6,7-tetrahydrobenzo[ b Thiophene-3-carboxynitrile (1.58 mmol, 0.75 equivalents) was added to a 100 mL round-bottom flask. The flask was purged with argon, and 50 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 2-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[]] as a white powder. b Thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (355mg, 66.4%).

[0400] NMR analysis confirmed the preparation of the desired product compound. 1H NMR (400 MHz, DMSO-d6) δ: 1.43(s,4H), 1.75 (m,4H), 2.59 (m, 4H), 3.70 (s, 2H), 11.71 (s, 1H), 12.66 (s,1H). 13 C NMR (125MHz): 175.3, 167.7, 146.7, 131.2, 128, 114.5, 93.2, 47.43,33.59, 25.95, 23.95, 23.77, 23.05, 22.16.

[0401] Example 3: 2-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[ b Preparation of thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (compound FTS005)

[0402]

[0403] FTS005

[0404] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS005 using the synthetic method described in Scheme 3 below.

[0405] Option 3

[0406]

[0407] Materials and methods

[0408] 3,3-Dimethyl-1,4-oxothiacyclohexane-2,6-dione (2.13 mmol) and 2-amino-4,5,6,7-tetrahydrobenzo[ b Thiophene-3-carboxynitrile (1.58 mmol, 0.75 equivalents) was added to a 100 mL round-bottom flask. The flask was purged with argon, and 50 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 2-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[]] as a white powder. b Thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (355 mg, 66.4%).

[0409] NMR analysis confirmed the preparation of the desired product compound. 1H NMR (400 MHz, DMSO-d6) δ: 1.43(s,4H), 1.75 (m,4H), 2.59 (m, 4H), 3.70 (s, 2H), 11.71 (s, 1H), 12.66 (s,1H). 13 C NMR (125MHz): 175.3, 167.7, 146.7, 131.2, 128, 114.5, 93.2, 47.43,33.59, 25.95, 23.95, 23.77, 23.05, 22.16.

[0410] Example 4: 2-((1-((3-cyano-4,5,6,7-tetrahydrobenzo[ b Preparation of thiophene-2-yl)amino)-1-oxopropane-2-yl)thio)propionic acid (compound FTS007)

[0411]

[0412] FTS007

[0413] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS007 using the synthetic method shown in Scheme 4 below.

[0414] Option 4

[0415]

[0416] Materials and methods

[0417] 0.357 mmol of 3,5-dimethyl-1,4-oxothiacyclohexane-2,6-dione and 2-amino-4,5,6,7-tetrahydrobenzo[ b Thiophene-3-carboxynitrile (0.226 mmol, 0.75 equivalents) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 20 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 2-((1-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Thiophene-2-yl)amino)-1-oxopropane-2-yl)thio)propionic acid (26 mg, 34%).

[0418] NMR analysis confirmed the preparation of the desired product compound. 1H NMR (500 MHz, DMSO-d6) δ: 1.33(d, 3H, J =7.13MHz), 1.43 (d, 3H, J= 6.99 MHz), 1.75 (s, 4H), 2.36-2.63 (m, 4H), 3.56 (q, 1H, J =7.12 MHz), 4.05 (q, 1H, J= 6.97 MHz), 11.74 (s, 1H), 12.65 (s, 1H)

[0419] Example 5: 5-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Preparation of thiophene-2-yl)carbamoyl)tetrahydrothiophene-2-carboxylic acid (compound FTS009)

[0420]

[0421] FTS009

[0422] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS009 using the synthetic method shown in Scheme 5 below.

[0423] Option 5

[0424]

[0425] Materials and methods

[0426] 0.312 mmol of 3-oxa-8-thiabicyclo[3.2.1]octane-2,4-dione and 2-amino-4,5,6,7-tetrahydrobenzo[ b Thiophene-3-carboxynitrile (0.219 mmol, 0.75 equivalents) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 20 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 5-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Thiophene-2-yl)carbamoyl)tetrahydrothiophene-2-carboxylic acid (20 mg, 27.2%).

[0427] NMR analysis confirmed the preparation of the desired product compound. 1H NMR (400 MHz, DMSO-d6) δ:1.75(s, 4H), 2.05 (m, 2H), 2.38 (m, 2H), 2.58 (m, 2H), 4.04 (t, 1H), 4.30 (t,1H), 11.70 (s, 1H), 12.67 (s, 1H)

[0428] Example 6: 2-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Preparation of thiophene-2-yl)amino)-2-oxoethoxy)acetic acid (compound FTS011)

[0429]

[0430] FTS011 (CAS 74195-03-8)

[0431] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS011 using the synthetic method described in Scheme 6 below.

[0432] Option 6

[0433]

[0434] Materials and methods

[0435] 1,4-dioxane-2,6-dione (0.968 mmol) and 2-amino-4,5,6,7-tetrahydrobenzo[ b Thiophene-3-carboxynitrile (0.907 mmol) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 20 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 2-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Thiophene-2-yl)amino)-2-oxoethoxy)acetic acid (152 mg, 57%).

[0436] NMR analysis confirmed the preparation of the desired product compound. 1H NMR (400 MHz, DMSO-d6) δ: 1.72ppm (2H, dt, J=5.7 Hz), 1.76 (2H, dt, J=5.7 Hz), 2.60 ppm (4H, m, J=5.7Hz), 4.19 (2H, s), 4.35 (2H, s), 11.41 ppm (1H, s), 12.81 ppm (1H, s).

[0437] Example 7: 5-((3-cyano-4,5,6,7-tetrahydrobenzo[ b Preparation of thiophene-2-yl)carbamoyl)tetrahydrofuran-2-carboxylic acid (compound FTS013)

[0438]

[0439] FTS013 (CAS 2191241-83-3)

[0440] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS013 using the synthetic method described in Scheme 7 below.

[0441] Option 7

[0442]

[0443] Materials and methods

[0444] 0.191 mmol of 3,8-dioxabicyclo[3.2.1]octane-2,4-dione and 2-amino-4,5,6,7-tetrahydrobenzo[ b Thiophene-3-carboxynitrile (0.176 mmol, 0.75 equivalents) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 20 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 5-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Thiophene-2-yl)carbamoyl)tetrahydrofuran-2-carboxylic acid (30 mg, 53.2%).

[0445] NMR analysis confirmed the preparation of the desired product compound. 1H NMR (400 MHz, DMSO-d6) δ: 2.65-2.76 ppm,( 6H, m), 3.0- 3.14 ppm (6H, m), 3.34 ppm (2H, t, J=4.98 Hz), 5.41-5.47 ppm (2H, dt, J=8.45 Hz), 12.25 ppm (1H, s), 14.40 ppm (1H, s).

[0446] Example 8: 5-((3-cyano-4,5,6,7-tetrahydrobenzo[ b Preparation of thiophene-2-yl)amino)-5-oxovaleric acid (compound FTS015)

[0447]

[0448] FTS015 (CAS 313230-00-1)

[0449] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS015 using the synthetic method described in Scheme 8 below.

[0450] Option 8

[0451]

[0452] Materials and methods

[0453] dihydro-2 H -pyran-2,6(3) H )-Diketone (1.07 mmol) and 2-amino-4,5,6,7-tetrahydrobenzo[ b Thiophene-3-carboxynitrile (0.75 mmol, 0.75 equivalents) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 20 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 5-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Thiophene-2-yl)amino)-5-oxovalerate (83 mg, 37.8%).

[0454] NMR analysis confirmed the preparation of the desired product compound. 1H NMR (400 MHz, DMSO-d6) δ: 1.68-1.82 ppm (6H, m), 2.22-2.28 (4H, dt, J= 5.8 Hz), 2.58 ppm (4H, t, J=4.45Hz), 11.53 ppm (1H, s), 12.38 ppm (1H, s).

[0455] Example 9: Preparation of 2-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethoxy)-2-methylpropionic acid (compound FTS027)

[0456]

[0457] FTS027

[0458] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS027 using a four-step synthetic method as shown in Scheme 9 below.

[0459] Option 9

[0460] Step 1

[0461]

[0462] Step 2

[0463]

[0464] Step 3

[0465]

[0466] Step 4

[0467]

[0468] Materials and methods

[0469] Step 1 - 2-Amino-4,5,6,7-Tetrahydrobenzo[ b Synthesis of thiophene-3-carboxynitrile:In a dry 250 mL single-necked round-bottom flask under a nitrogen atmosphere, cyclohexanone (5.0 g, 50.9 mmol) was dissolved in dioxane (100 mL), followed by the addition of malononitrile (3.37 g, 50.9 mmol) and sulfur (1.633 g, 50.9 mmol). The mixture was heated to 50 °C, and morpholine (4.44 g, 50.9 mmol) was added. The mixture was stirred at the same temperature for 16 h. The reaction progress was monitored by LCMS and TLC. After the reaction was complete, the reaction mixture (RM) was concentrated on a rotary evaporator, and the resulting residue was diluted with water (80 mL) and extracted with ethyl acetate (2 × 50 mL). The combined organic layers were washed with aqueous brine, dried over sodium sulfate, and concentrated to give the crude compound. The crude product was purified by column chromatography (Isolera) using EtOAc and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 10% EtOAc) to obtain pure 2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxynitrile as a pale yellow crystalline solid.

[0470] NMR analysis confirmed the preparation of the intermediate compound required in step 1. 1 H NMR: 400 MHz DMSO- d6 δ:6.94 (s, 2H), 2.42-2.39 (m, 2H), 2.35-2.32 (m, 2H), 1.73-1.68 (m, 4H).

[0471] Step 2 - 2-Chloro- N -(3-cyano-4,5,6,7-tetrahydrobenzo[ b Synthesis of thiophene-2-yl)acetamide Under a nitrogen atmosphere, 2-chloroacetyl chloride (1.521 g, 13.46 mmol) was added to a stirred solution of 2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxynitrile (2.0 g, 11.22 mmol) in dioxane (25 mL) at a dry single-necked round-bottom flask. The mixture was stirred at room temperature (RT) for 16 h. The reaction progress was monitored by TLC (hexane containing 20% ​​EtOAc, 0.7 rf). After the reaction was complete, hexane was added to the reaction mixture (RM) and the mixture was stirred for 10 min. The resulting solid was filtered through a Buchner funnel and washed with hexane to give pure 2-chloro-N-(3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)acetamide.

[0472] NMR analysis confirmed the preparation of the intermediate compound required in step 2. 1 H NMR: 400 MHz DMSO- d6δ:11.91 (s, 1H), 4.46 (s, 2H), 2.61-2.51 (m, 2H), 2.51-2.50 (m, 2H), 1.77-1.75 (m, 4H).

[0473] Step 3 - 2-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[ b ]Thiophene-2-yl)amino)-2-oxoethoxy Synthesis of methyl 2-methylpropionate Under a nitrogen atmosphere, in a dry, single-necked round-bottom flask, methyl 2-hydroxy-2-methylpropionate (0.464 g, 3.93 mmol) and 2-chloro-N-(3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)acetamide (0.500 g, 1.963 mmol) were added to a stirred solution of methyl 2-hydroxy-2-methylpropionate (0.464 g, 3.93 mmol) in THF (25 mL) and heated at 70 °C for 16 h. The reaction progress was monitored by TLC (hexane containing 20% ​​EtOAc, 0.4 rf). After the reaction was complete, the reaction mixture was diluted with water (80 mL) and extracted with ethyl acetate (2 × 100 mL). The combined organic layers were washed with aqueous brine, dried over sodium sulfate, and concentrated to give the crude compound. The crude product was purified by column chromatography (Isolera) using EtOAc and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 15% EtOAc) to give pure methyl 2-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethoxy)-2-methylpropionate as an orange solid.

[0474] NMR analysis confirmed the preparation of the intermediate compound required in step 3. 1 H NMR: 400 MHz DMSO- d6 δ:10.96 (s, 1H), 4.25 (s, 2H), 3.68 (s, 3H), 2.61-260 (m, 2H), 2.51-2.50 (m,2H), 1.76 (s, 4H), 1.42 (s, 6H).

[0475] Step 4 - 2-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[ b ]Thiophene-2-yl)amino)-2-oxoethoxy Synthesis of 2-methylpropionic acid:Methyl 2-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethoxy)-2-methylpropionate (0.180 g, 0.535 mmol) was dissolved in THF (5 mL) in a dry 100 mL single-necked round-bottom flask under a nitrogen atmosphere. Water (3 mL) and lithium hydroxide hydrate (0.067 g, 1.605 mmol) were added, and the mixture was stirred at room temperature for 1 h. The progress of the reaction was monitored by TLC (DCM containing 0.5% MeOH). When the reaction was complete, the reaction mixture was diluted with water (5 mL) and washed with ethyl acetate (25 mL). The aqueous layer was acidified with citric acid and extracted with DCM containing 10% MeOH (3 x 20). The organic layer was dried with sodium sulfate and concentrated on a rotary evaporator to obtain pure 2-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethoxy)-2-methylpropionic acid as a grayish-white solid.

[0476] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR: 400 MHz DMSO- d6 δ: 12.95 (s,1H), 11.17 (s, 1H), 4.23 (s, 2H), 2.70-2.51 (m, 4H), 1.76 (s, 4H), 1.40 (s,6H).

[0477] Example 10: Preparation of 1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)cyclopropanecarboxylic acid (compound FTS028)

[0478]

[0479] FTS028

[0480] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS028 via a seven-step synthetic method as shown in Scheme 10 below.

[0481] Option 10

[0482] Step 1

[0483]

[0484] Step 2

[0485]

[0486] Step 3

[0487]

[0488] Step 4

[0489]

[0490] Step 5

[0491]

[0492] Step 6

[0493]

[0494] Materials and methods

[0495] Step 1 – Synthesis of 2-((bis(4-methoxyphenyl)(phenyl)methyl)thio)ethyl acetate: To a solution of ethyl 2-thioalkylbenzene (10.0 g, 83.22 mmol, 1.0 equivalent) in DCM (120 mL), NaHCO3 (8.3 g, 99.86 mmol, 1.2 equivalent) was added. The mixture was stirred at 25 °C for 20 min. 1-[chloro-(4-methoxyphenyl)-phenyl-methyl]-4-methoxybenzene (28.20 g, 83.22 mmol, 1 equivalent) was added, and the mixture was stirred at 25 °C for 3 h. The reaction mixture was quenched by adding H2O (120 mL) and extracted with DCM (120 mL x 2). The combined organic layers were dried over Na2SO4, filtered, concentrated, and purified by rapid silica gel chromatography (petroleum ether containing ethyl acetate = 0 to 10%) to give ethyl 2-((bis(4-methoxyphenyl)(phenyl)methyl)thio)ethyl acetate (10.0 g, 23.67 mmol, yield 58.8%) as a grayish-white oil.

[0496] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, CDCl3) δ = 7.34(d, J = 8.0 Hz, 2H), 7.26-7.16 (m, 6H), 7.15-7.14 (m, 1H), 6.76-6.73 (m, 4H), 4.00-3.95 (m, 2H), 3.72 (s, 6H), 2.90 (s, 2H), 1.31 (t, J = 6.8 Hz, 3H).

[0497] Step 2 - Synthesis of 1-((bis(4-methoxyphenyl)(phenyl)methyl)thio)cyclopropanecarboxylate ethyl ester: At -60°C, ethyl 2-[bis(4-methoxyphenyl)-phenyl-methyl]thioethyl acetate (10.0 g, 23.67 mmol, 1.0 μL) was added. when quantity LDA (2 M, 29.58 mL, 2.5 mL) was added dropwise to a 100 mL THF solution. equivalent After stirring at -60°C for 1.5 h, 2,2-dioxide containing 1,3,2-dioxane (4.41 g, 35.50 mmol, 1.5 g) was added dropwise at -60°C. equivalent 17 mL of THF was added, followed by dropwise addition of DMPU (4.55 g, 35.50 mmol, 4.29 mL, 1.5 mL) at -60°C. equivalent The mixture was stirred at 25°C for 16 h. After cooling to -60°C, saturated NH4Cl (200 mL) was added. The mixture was extracted with EtOAc (400 mL). The organic layer was washed with brine (400 mL), dried over Na2SO4, filtered, concentrated, and purified by rapid silica gel chromatography (petroleum ether containing ethyl acetate = 0% to 10%) to give ethyl 1-((bis(4-methoxyphenyl)(phenyl)methyl)thio)cyclopropanecarboxylate (7.6 g, 16.94 mmol, yield 73.6%) as a yellow oil.

[0498] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, CDCl3) δ = 7.36(d, J = 7.2 Hz, 2H), 7.26-7.20 (m, 3H), 7.19-7.17 (m, 4H), 6.72 (d, J = 9.2 Hz,4H), 3.73 (s, 3H), 3.60 (m, 2H), 1.29 (m, 2H), 1.09 (m, 2H), 0.94 (t, J = 7.2Hz, 3H).

[0499] Step 3 - Synthesis of ethyl 1-mercaptocyclopropanecarboxylate:At 0 °C, triethylsilane (933.2 mg, 8.03 mmol, 1.28 mL, 1.2 equivalent) was added to a DCM (30 mL) solution of ethyl 1-[bis(4-methoxyphenyl)-phenyl-methyl]thioalkylcyclopropane carboxylate (3.0 g, 6.69 mmol, 1.0 equivalent). After the addition was complete, TFA (762.5 mg, 6.69 mmol, 495.16 μL, 1.0 equivalent) was added, and the mixture was stirred at 25 °C for 16 h. The resulting solution was used directly in the next step.

[0500] Step 4 - Synthesis of ethyl 1-((2-(tert-butoxy)-2-oxoethyl)thio)cyclopropanecarboxylate: At 0°C, THF (300 mL), K₂CO₃ (4.4 g, 32.49 mmol, 5.0 equivalents), and tert-butyl 2-bromoacetate (1.3 g, 6.50 mmol, 960.12 uL, 1.0 equivalents) were added to the above solution. The mixture was stirred at 25°C for 16 h, filtered, and concentrated to obtain ethyl 1-((2-(tert-butoxy)-2-oxoethyl)thio)cyclopropanecarboxylate (1.6 g, 6.49 mmol, crude product), which was a brown oil and could be used in the next step without further purification.

[0501] Step 5 - Synthesis of 2-((1-(ethoxycarbonyl)cyclopropyl)thio)acetic acid: TFA (15.4 g, 135.06 mmol, 10 mL, 20.8 equivalents) was added to a DCM (10 mL) solution of ethyl 1-(2-tert-butoxy-2-oxoethyl)thioalkylcyclopropanecarboxylate (1.6 g, 6.49 mmol, 1.0 equivalent). The mixture was stirred at 25 °C for 1 h, filtered, and concentrated to give 2-((1-(ethoxycarbonyl)cyclopropyl)thio)acetic acid (1.3 g, 6.51 mmol, crude) as a yellow oil, which could be used in the next step without further purification.

[0502] Step 6 - 2-(1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[ b ]Thiophene-2-yl)amino)-2-oxoethyl Synthesis of ethyl 2-oxopropyl (2-ethyl 2-oxopropyl)-2-oxopropyl:DIEA (1.16 g, 8.98 mmol, 1.56 mL, 8.0 equivalent) was added to a DMF (2.0 mL) solution of 2-(1-ethoxycarbonylcyclopropyl)thioalkylacetic acid (1.15 g, 5.61 mmol, 5.0 equivalent). After stirring at 25 °C for 10 min, HATU (639.9 mg, 1.68 mmol, 1.5 equivalent) and 2-amino-4,5,6,7-tetrahydrobenzothiophene-3-carboxynitrile (200 mg, 1.12 mmol, 1.0 equivalent) were added. The mixture was stirred at 50 °C for 16 h. The reaction mixture was quenched by adding H₂O (20 mL) and extracted with DCM (20 mL). The organic layer was washed with brine (25 mL x 3), dried over Na2SO4, filtered, concentrated, and purified by silica gel rapid chromatography (ethyl acetate / petroleum ether = 0 to 10%) to produce ethyl 1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)cyclopropanecarboxylate (405.0 mg, 1.11 mmol, yield 99.0%) as a yellow solid, which was used directly in the next step.

[0503] LCMS analysis confirmed the preparation of the desired intermediate compound. LCMS R t = 0.454 min, 0.8 min chromatography, 5-95AB, ESI calculated value, C 17 H 20 N₂O₃S₂Na [M+Na] + 387.1, measured value 387.0.

[0504] Step 7 - 1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl) Synthesis of thiocyclopropanecarboxylic acid (FTS028): Ethyl 1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)cyclopropanecarboxylate (400 mg, 1.10 mmol, 1.0 mmol) equivalentLiOH·H2O (230.2 mg, 5.49 mmol, 5.0 equivalent) was added to a mixed solvent solution of MeOH (5 mL) and H2O (1 mL). The mixture was stirred at 25 °C for 16 h, filtered, concentrated, and purified by preparative HPLC (column: Phenomenex luna C18 150*25mm*10 μm; mobile phase: [water(FA)-ACN]; B%: 35%-65%, 8 min) to give 1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)cyclopropanecarboxylic acid (78.5 mg, 233.33 μmol, yield 21.2%) as a white solid.

[0505] NMR, LCMS, and HPLC analyses confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO-d6)δ = 3.66 (s, 2H), 2.68-2.53 (m, 4H), 1.75-1.72 (m, 4H), 1.46-1.43 (m, 2H), 1.19-1.12 (m, 2H). LCMS R t = 1.455 min, 3 min chromatography, 5-95 AB, ESI calculated value, C 15 H 16 N₂O₃S₂Na [M+Na] + 359.1, measured value 358.9. HPLC purity 98.8%, R t = 2.967 min, 6.0 min chromatography, 10-80AB_6min.

[0506] Example 11: 2-(1-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Preparation of thiophene-2-yl)amino)-2-oxoethyl)cyclopropyl)acetic acid (compound FTS029)

[0507]

[0508] FTS029

[0509] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS029 using the synthetic method described in Scheme 11 below.

[0510] Option 11

[0511]

[0512] Materials and methods

[0513] 6-oxaspiro[2,5]octane-5,7-dione (0.638 mmol) and 2-amino-4,5,6,7-tetrahydrobenzo[ b Thiophene-3-carboxynitrile (0.488 mmol, 0.75 equivalents) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 20 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 2-(1-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)cyclopropyl)acetic acid (23 mg, 14.8%) as an amorphous white powder.

[0514] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (500 MHz, DMSO-d6) δ = 12.13(bs, 1H), 11.41 (s, 1H), 2.73 (s, 2H), 2.44 (m, 2H), 2.34 (m, 2H), 2.27 (s,2H), 1.77-1.65 (m, 4H), 0.53 (d, 2H), 0.43 (d, 2H)

[0515] Example 12: Preparation of 1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)cyclobutanecarboxylic acid (compound FTS030)

[0516]

[0517] FTS030

[0518] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS030 via a four-step synthetic method as shown in Scheme 12 below.

[0519] Option 12

[0520] Step 1

[0521]

[0522] Step 2

[0523]

[0524] Step 3

[0525]

[0526] Step 4

[0527]

[0528] Materials and methods

[0529] Step 1 - Synthesis of ethyl 1-((2-(tert-butoxy)-2-oxoethyl)thio)cyclobutane carboxylate : To ethyl 1-bromocyclobutanecarboxylate (2.0 g, 9.66 mmol, 1.56 mL, 1.0 equivalent Add tert-butyl 2-thioalkyl acetate (1.43 g, 9.66 mmol, 1.0 equivalent) and KOH (541.9 mg, 9.66 mmol, 1.0 equivalent) to a 10 mL THF solution. equivalent The mixture was stirred at 25°C for 2 h. The reaction mixture was quenched with saturated NH4Cl (20 mL) and extracted with EtOAc (20 mL × 2 times). The combined organic layers were washed with brine (20 mL × 2), dried over Na2SO4, filtered, and concentrated to give ethyl 1-((2-(tert-butoxy)-2-oxoethyl)thio)cyclobutanecarboxylate (2.6 g, 9.48 mmol, quantitative) as a grayish-white liquid, which could be used in the next step without further purification.

[0530] Step 2 - 2-((1-(ethoxycarbonyl)cyclobutyl)thio)acetic acid TFA (5.9 g, 51.94 mmol, 3.85 mL, 14.2 equivalents) was added to a DCM (15 mL) solution of ethyl 1-(2-tert-butoxy-2-oxoethyl)thioalkylcyclobutane carboxylate (1.0 g, 3.64 mmol, 1.0 equivalent). The mixture was stirred at 25 °C for 16 h. The reaction mixture was concentrated to give 2-((1-(ethoxycarbonyl)cyclobutyl)thioacetic acid (0.79 g, 4.58 mmol, quantitative) as a grayish-white liquid, which could be used in the next step without further purification.

[0531] Step 3 - 1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl) Thioyl cyclobutanecarboxylateDIEA (2.22 g, 17.18 mmol, 2.99 mL, 5.0 equivalent) was added to a DMF (1 mL) solution of 2-(1-ethoxycarbonylcyclobutyl)thioalkylacetic acid (0.75 g, 3.44 mmol, 1.0 equivalent). After stirring at 25 °C for 0.5 h, HATU (1.96 g, 5.15 mmol, 1.5 equivalent) and 2-amino-4,5,6,7-tetrahydrobenzothiophene-3-carboxynitrile (612.5 mg, 3.44 mmol, 1.0 equivalent) were added. The mixture was stirred at 50 °C for 16 h. The reaction mixture was quenched by adding H2O (20 mL) and extracted with DCM (20 mL). The organic layer was washed with brine (25 mL x 3), dried over Na2SO4, filtered, concentrated, and purified by silica gel rapid chromatography (ethyl acetate / petroleum ether = 0 to 10%) to give ethyl 1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)cyclopropanecarboxylate (0.8 g, 2.11 mmol, yield 61.5%) as a yellow solid, which was used directly in the next step.

[0532] LCMS analysis confirmed the preparation of the desired intermediate compound. (LCMS / LCMS analysis confirmed the preparation of the intermediate compound.) t = 0.475 min, 0.8 min chromatography, 5-95AB, ESI calculated value, C 18 H 22 N₂O₃S₂[M+H] + 379.1, measured value 379.1.

[0533] Step 4 - 1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophen-2-yl)amino)-2-oxoethyl) Synthesis of thiocyclobutanecarboxylic acid Add LiOH·H2O (443.4 mg, 10.57 mmol, 5.0 equivalent) to a mixed solvent solution of 1-[2-[(3-cyano-4,5,6,7-tetrahydrobenzothiophene-2-yl)amino]-2-oxoethyl]thioalkylcyclobutanecarboxylate (800.0 mg, 2.11 mmol, 1.0 equivalent) in MeOH (10 mL) and H2O (2 mL). The mixture was stirred at 50 °C for 2 h, filtered, concentrated, and purified by preparative HPLC (column: Phenomenex luna C18 150 x 25 mm x 10 μm; mobile phase: [water (FA)-ACN]; B%: 40%-70%, 10 min) to obtain 1-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)cyclobutanecarboxylic acid (63.6 mg, 181.48 μmol, yield 8.5%) as a yellow solid.

[0534] NMR, LCMS, and HPLC analyses confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, CDCl3) δ= 9.98 (s, 1H), 3.56 (s, 2H), 2.80-2.70 (m, 2H), 2.65-2.54 (m, 4H), 2.32-2.14(m, 3H), 2.04-1.92 (m, 1H), 1.87-1.77 (m, 4H). LCMS R t = 1.548 min, 3 min chromatography, 5-95 AB, ESI calculated value, C 16 H 18 N₂O₃S₂Na [M+Na] + 373.1, measured value 373.0. HPLC purity 97.6%, R t =3.008 min, 6.0 min chromatography, 10-80AB_6min.

[0535] Example 13: Preparation of 3-((2-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)oxetane-3-carboxylic acid (FTS035)

[0536]

[0537] FTS035

[0538] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS035 via a four-step synthetic method as shown in Scheme 13 below.

[0539] Option 13

[0540] Step 1

[0541]

[0542] Step 2

[0543]

[0544] Step 3

[0545]

[0546] Step 4

[0547]

[0548] Materials and methods

[0549] FTS035 was prepared using the same general synthetic method for preparing FTS030 as described in Example 12, except that the starting compound ethyl 1-bromocyclobutane carboxylate was replaced by the starting compound ethyl 3-bromooxetane-3-carboxylate, as shown in Scheme 13.

[0550] Example 14: 2-(1-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Preparation of thiophene-2-yl)amino)-2-oxoethyl)cyclobutyl)acetic acid (compound FTS031)

[0551]

[0552] FTS031

[0553] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS031 using the synthetic method described in Scheme 14 below.

[0554] Option 14

[0555]

[0556] Materials and methods

[0557] 0.275 mmol of 7-oxaspiro[3.5]nonane-6,8-dione and 2-amino-4,5,6,7-tetrahydrobenzo[ b Thiophene-3-carboxynitrile (0.236 mmol, 0.75 equivalents) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 10 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. The solvent was removed by rotary evaporation. The crude mixture was purified by preparative TLC in 2.5:1 hexane: EtOAc + 0.1% acetic acid to produce 2-(1-(2-((3-cyano-4,5,6,7-tetrahydrobenzo[] b Thiophene-2-yl)amino)-2-oxoethyl)cyclobutyl)acetic acid (15 mg, 19.1%).

[0558] Example 15: Preparation of (1s,3s)-3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)cyclobutane-1-carboxylic acid (compound FTS032)

[0559]

[0560] FTS032

[0561] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS032 via a two-step synthetic method as shown in Scheme 15 below.

[0562] Option 15

[0563] Step 1

[0564]

[0565] Step 2

[0566]

[0567] Materials and methods

[0568] Step 1 - (1 s ,3 s )-3-((3-cyano-4,5,6,7-tetrahydrobenzo[ b ]Thiophen-2-yl)carbamoyl)cyclobutane Synthesis of methyl alkyl-1-carboxylate: Under a nitrogen atmosphere, 2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxynitrile (200 mg, 1.122 mmol) was dissolved in DCM (40 mL) in a 100 mL dry double-necked round-bottom flask. Under a nitrogen atmosphere, (1s,3s)-3-(methoxycarbonyl)cyclobutane-1-carboxylic acid (266 mg, 1.683 mmol) and DIPEA (1.176 mL, 6.73 mmol) were added to this reaction mixture at 25 °C. The reaction mixture was then cooled to 0 °C, and POCl3 (0.314 mL, 3.37 mmol) was added dropwise, with stirring at 25 °C for 16 h under a nitrogen atmosphere. The mixture was then analyzed by TLC (petroleum ether containing 20% ​​EtOAc, 0.3 mL). rf The reaction progress was monitored. After the reaction was complete, the reaction mixture was quenched with 100 mL of NaHCO3 solution. The reaction mixture was extracted with DCM (3 x 100 mL). The combined organic layers were dried over sodium sulfate and concentrated by rotary evaporation under reduced pressure (bath temperature 45 °C) to obtain a crude product. The crude product was purified by column chromatography (Isolera) using ethyl acetate and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 18% ethyl acetate) to give a pure (1S,3S)-3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)cyclobutane-1-carboxylate as a yellow solid.

[0569] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, DMSO- d6) δ =11.46 (s, 1H), 3.61 (s, 3H), 3.39 (dd, J = 9.60, 18.00 Hz, 1H), 3.17 (t, J =8.80 Hz, 1H), 2.68 (s, 4H), 2.36 (t, J = 8.80 Hz, 4H), 1.75 (s, 4H).

[0570] Step 2 - (1) s ,3 s )-3-((3-cyano-4,5,6,7-tetrahydrobenzo[ b ]Thiophen-2-yl)carbamoyl)cyclobutane Synthesis of alkyl-1-carboxylic acid: Under a nitrogen atmosphere, methyl (1s,3s)-3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)cyclobutane-1-carboxylate (150 mg, 0.471 mmol) was dissolved in THF (10 mL) and water (5 mL) in a 100 mL dry single-necked round-bottom flask. Under a nitrogen atmosphere, lithium hydroxide monohydrate (59.4 mg, 1.413 mmol) was added to this reaction mixture at 25 °C. The reaction mixture was stirred for 1 h at 25 °C under a nitrogen atmosphere. The mixture was analyzed by TLC (DCM containing 10% MeOH, 0.2 μL). rf The reaction progress was monitored. After the reaction was complete, the reaction mixture was concentrated by rotary evaporation under reduced pressure (bath temperature 45°C). The reaction mixture was extracted with ethyl acetate (30 mL). The aqueous layer was then acidified with citric acid (pH = 5-6). The reaction mixture was extracted with DCM containing 10% MeOH (3 x 50 mL). The combined organic layers were dried over sodium sulfate, concentrated by rotary evaporation under reduced pressure (bath temperature 45°C), and lyophilized to give pure (1s,3s)-3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)cyclobutane-1-carboxylic acid as a white solid.

[0571] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO- d6 ) δ = 12.11(s, 1H), 11.50 (s, 1H), 3.52 (t, J = 7.60 Hz, 1H), 3.05-3.01 (m, 1H), 2.68(t, J = 1.60 Hz, 2H), 2.51 (t, J = 1.60 Hz, 2H), 2.42-2.34 (m, 4H), 1.75 (s, 4H).

[0572] Example 16: Preparation of (1r,3r)-3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)cyclobutane-1-carboxylic acid (compound FTS033)

[0573]

[0574] FTS033

[0575] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS033 via a two-step synthetic method as shown in Scheme 16 below.

[0576] Option 16

[0577] Step 1

[0578]

[0579] Step 2

[0580]

[0581] Materials and methods

[0582] Step 1 - (1 r ,3 r )-3-((3-cyano-4,5,6,7-tetrahydrobenzo[ b ]Thiophen-2-yl)carbamoyl)cyclobutane Synthesis of methyl alkyl-1-carboxylate: Under nitrogen atmosphere, 2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxynitrile (400 mg, 2.244 mmol) was dissolved in DCM (15 mL) in a 100 mL dry double-necked round-bottom flask. Under nitrogen atmosphere, (1r,3r)-3-(methoxycarbonyl)cyclobutane-1-carboxylic acid (426 mg, 2.69 mmol) and DIPEA (2.352 mL, 13.46 mmol) were added to this reaction mixture at 25 °C. The reaction mixture was then cooled to 0 °C, and Pocl3 (0.629 mL, 6.73 mmol) was added dropwise, followed by stirring at 25 °C for 16 h under nitrogen atmosphere. The mixture was analyzed by TLC (petroleum ether containing 20% ​​EtOAc, 0.6 mL). rfThe reaction progress was monitored. After the reaction was complete, the reaction mixture was quenched with 100 mL of NaHCO3 solution. The reaction mixture was extracted with DCM (3 x 100 mL). The combined organic layers were dried over sodium sulfate and concentrated by rotary evaporation under reduced pressure (bath temperature 45 °C) to give the crude product. The crude product was purified by column chromatography (Isolera) using ethyl acetate and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 15% ethyl acetate) to give pure (1r,3r)-3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)cyclobutane-1-carboxylate as a yellow solid.

[0583] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, DMSO- d6 ) δ=11.50 (s, 1H), 4.09-4.03 (m, 1H), 3.76 (dt, J = 4.80, Hz, 1H), 3.74 (s, 3H), 3.73 (t, J = 4.80 Hz, 1H), 3.16-3.13 (m, 1H), 2.68 (t, J = 2.00 Hz, 2H), 2.51-2.43 (m, 4H), 1.75 (s, 4H).

[0584] Step 2 - (1) r ,3 r )-3-((3-cyano-4,5,6,7-tetrahydrobenzo[ b ]Thiophen-2-yl)carbamoyl)cyclobutane Synthesis of alkyl-1-carboxylic acid: Under a nitrogen atmosphere, methyl (1r,3r)-3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)cyclobutane-1-carboxylate (48 mg, 0.151 mmol) was dissolved in THF (2 mL) and water (1 mL) in a 50 mL dry single-necked round-bottom flask. Under a nitrogen atmosphere, LiOH monohydrate (19 mg, 0.452 mmol) was added to this reaction mixture at 25 °C. The reaction mixture was stirred for 1 h at 25 °C under a nitrogen atmosphere. The mixture was analyzed by TLC (petroleum ether containing 20% ​​EtOAc, 0.1 mL). rfThe reaction progress was monitored. After the reaction was complete, the reaction mixture was concentrated by rotary evaporation under reduced pressure (bath temperature 45°C). The reaction mixture was extracted with ethyl acetate (30 mL). The combined aqueous layers were then acidified with citric acid (2 gm, pH = 1-2). The reaction mixture was extracted with DCM containing 10% MeOH (3 x 30 mL). The combined organic layers were dried over sodium sulfate, concentrated by rotary evaporation under reduced pressure (bath temperature 45°C), and lyophilized to give pure (1r,3r)-3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)cyclobutane-1-carboxylic acid as a white solid.

[0585] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO- d6 ) δ= 12.21(s, 1H), 11.53 (s, 1H), 3.52 (t, J = 7.60 Hz, 1H), 3.05-3.01 (m, 1H), 2.68(t, J = 1.60 Hz, 2H), 2.51 (t, J = 1.60 Hz, 2H), 2.42-2.34 (m, 4H), 1.75 (s, 4H).

[0586] Example 17: Preparation of 3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)bicyclo[1.1.1]pentane-1-carboxylic acid (compound FTS034)

[0587]

[0588] FTS034

[0589] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS034 via a two-step synthetic method as shown in Scheme 17 below.

[0590] Option 17

[0591] Step 1

[0592]

[0593] Step 2

[0594]

[0595] Materials and methods

[0596] Step 1 - 3-((3-cyano-4,5,6,7-tetrahydrobenzo[ b [1.1.1] Thiophene-2-yl)carbamoyl)bicyclic [1.1.1] Synthesis of methyl pentane-1-carboxylate: In a dry, double-necked round-bottom flask under nitrogen atmosphere, 2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxynitrile (200 mg, 1.122 mmol) was dissolved in CH₂Cl₂ (10 mL). Under nitrogen atmosphere, 3-(methoxycarbonyl)bicyclo[1.1.1]pentane-1-carboxylic acid (229 mg, 1.346 mmol) and DIPEA (1.176 mL, 6.73 mmol) were added to this reaction mixture at 25 °C. The reaction mixture was cooled to 0 °C, and Pocl₃ (0.315 mL, 3.37 mmol) was added dropwise, followed by stirring at 25 °C for 16 h under nitrogen atmosphere. The mixture was analyzed by TLC (petroleum ether containing 20% ​​EtOAc, 0.6 mL). rf The reaction progress was monitored. After the reaction was complete, the reaction mixture was quenched with 100 mL of NaHCO3 solution. The reaction mixture was extracted with DCM containing 10% MeOH (3 x 50 mL). The combined organic layers were dried over sodium sulfate and concentrated by rotary evaporation under reduced pressure (bath temperature 45 °C) to give the crude product. The crude product was purified by column chromatography (Isolera) using ethyl acetate and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 10% ethyl acetate) to give pure methyl 3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)bicyclo[1.1.1]pentane-1-carboxylate as a white solid.

[0597] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, DMSO- d6 ) δ=11.30 (s, 1H), 3.74 (s, 3H), 2.76 (s, 2H), 2.68 (s, 2H), 2.40 (s, 2H), 2.35(d, J = 6.40 Hz, 2H), 2.25 (s, 2H), 1.78 (d, J = 14.40 Hz, 4H).

[0598] Step 2 - 3-((3-cyano-4,5,6,7-tetrahydrobenzo[ b [1.1.1] Thiophene-2-yl)carbamoyl)bicyclic [1.1.1] Synthesis of pentane-1-carboxylic acid:Under a nitrogen atmosphere, methyl 3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)bicyclo[1.1.1]pentane-1-carboxylate (100 mg, 0.303 mmol) was dissolved in THF (3.00 mL), water (3 mL), and MeOH (1 mL) in a dry 25 mL single-necked round-bottom flask. Under a nitrogen atmosphere, LiOH (21.74 mg, 0.908 mmol) was added to this reaction mixture at 25 °C. The reaction mixture was stirred for 1 h at 25 °C under a nitrogen atmosphere. The mixture was analyzed by TLC (DCM containing 10% MeOH, 0.4 μmol / L). rf The reaction progress was monitored. After the reaction was complete, the reaction mixture was concentrated by rotary evaporation under reduced pressure (bath temperature 45°C). The reaction mixture was washed with ethyl acetate (10 mL), the aqueous layer was acidified with 1.5N HCl solution (pH=1-2), and extracted with DCM containing 10% MeOH (3*20 mL). The combined organic layers were dried over sodium sulfate, concentrated by rotary evaporation under reduced pressure (bath temperature 45°C), and lyophilized to give pure 3-((3-cyano-4,5,6,7-tetrahydrobenzo[b]thiophene-2-yl)carbamoyl)bicyclo[1.1.1]pentane-1-carboxylic acid as a white solid.

[0599] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO- d6 ) δ= 12.43(bs, 1H), 11.27 (bs, 1H), 2.6 (dd, J = 5.60, 12.80 Hz, 4H), 2.29 (s, 6H), 1.75 (t, J = 2.40 Hz, 4H).

[0600] Example 18: Preparation of 1-((2-((3-cyano-4,5-dimethylthiophen-2-yl)amino)-2-oxoethyl)thio)cyclopropanecarboxylic acid (compound FTS036)

[0601]

[0602] FTS036

[0603] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS036 via a two-step synthetic method as shown in Scheme 18 below.

[0604] Option 18

[0605] Step 1

[0606]

[0607] Step 2

[0608]

[0609] Materials and methods

[0610] Step 1 - 1-((2-((3-cyano-4,5-dimethylthiophen-2-yl)amino)-2-oxoethyl)thio)cyclopropane Synthesis of ethyl alkyl carbamate: DIEA (891.5 mg, 6.90 mmol, 1.20 mL, 7.0 equivalent) was added to a DMF (2 mL) solution of 2-(1-ethoxycarbonylcyclopropyl)thioalkylacetic acid (1.0 g, 4.93 mmol, 5.0 equivalent). After stirring at 25 °C for 10 min, HATU (562.0 mg, 1.48 mmol, 1.5 equivalent) and 2-amino-4,5-dimethylthiophene-3-carboxynitrile (150 mg, 985.44 μmol, 1.0 equivalent) were added. The mixture was stirred at 50 °C for 16 h. The reaction mixture was quenched by adding H2O (20 mL) and extracted with DCM (20 mL). The organic layer was washed with brine (25 mL x 3), dried over Na2SO4, filtered, concentrated, and purified by silica gel rapid chromatography (ethyl acetate / petroleum ether = 0 to 10%) to give ethyl 1-((2-(((3-cyano-4,5-dimethylthiophene-2-yl)amino)-2-oxoethyl)thio)cyclopropanecarboxylate (290.0 mg, 103.68 μmol, yield 10.5%) as a yellow solid, which was used directly in the next step.

[0611] LCMS analysis confirmed the preparation of the desired intermediate compound. LCMS R t = 0.438 min, 0.8 min chromatography, 5-95AB, ESI calculated value, C 15 H 18 N₂O₃S₂Na [M+Na] + 361.1, measured value 361.0.

[0612] Step 2 - 1-((2-((3-cyano-4,5-dimethylthiophen-2-yl)amino)-2-oxoethyl)thio)cyclopropane Synthesis of alkylcarboxylic acid (FTS036): Add LiOH·H2O (179.8 mg, 4.28 mmol, 5.0 mmol) to a mixed solvent solution of ethyl 1-[2-[(3-cyano-4,5-dimethyl-2-thienyl)amino]-2-oxoethyl]thioalkylcyclopropanecarboxylate (290 mg, 856.86 μmol, 1.0 equivalent) in MeOH (5 mL) and H2O (1 mL). equivalentThe mixture was stirred at 25 °C for 16 h, filtered, concentrated, and purified by preparative HPLC (column: Phenomenex luna C18 150x25mmx10um; mobile phase: [water (FA)-ACN]; B%: 40%-70%, 10 min) to give the desired compound as a yellow solid (14.4 mg, 46.39 μmol, yield 5.4%).

[0613] NMR, LCMS, and HPLC analyses confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO-d6)δ = 3.70-3.62 (s, 2H), 2.23 (s, 3H), 2.10 (s, 3H), 1.48-1.42 (m, 2H), 1.19-1.11 (m, 2H). LCMS R t = 1.300 min, 3 min chromatography, 5-95 AB, ESI calculated value, C 13 H 14 N₂O₃S₂Na [M+Na] + 333.0, measured value 332.9. HPLC purity 98.8%, R t = 2.568 min, 6.0 min chromatography, 10-80AB_6min.

[0614] Example 19: Preparation of 1-((2-((3-cyano-4,5-dimethylthiophen-2-yl)amino)-2-oxoethyl)thio)cyclobutanecarboxylic acid (compound FTS037)

[0615]

[0616] FTS037

[0617] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS037 via a two-step synthetic method as shown in Scheme 19 below.

[0618] Option 19

[0619] Step 1

[0620]

[0621] Step 2

[0622]

[0623] Materials and methods

[0624] Step 1 - 1-((2-((3-cyano-4,5-dimethylthiophen-2-yl)amino)-2-oxoethyl)thio)cyclobutane Synthesis of ethyl alkyl carbamate To a DMF (1 mL) solution of 2-(1-ethoxycarbonylcyclobutyl)thioalkylacetic acid (358.4 mg, 1.64 mmol, 5.0 equivalents), DIEA (212.26 mg, 1.64 mmol, 286.07 μL, 5.0 equivalents), HATU (187.35 mg, 492.72 μmol, 1.5 equivalents), and 2-amino-4,5-dimethylthiophene-3-carboxynitrile (50.0 mg, 328.4 μmol, 1.0 equivalents) were added. The mixture was stirred at 50 °C for 16 h. The reaction mixture was quenched by adding H₂O (20 mL) and extracted with DCM (20 mL). The organic layer was washed with brine (25 mL x 3), dried over Na2SO4, filtered, concentrated, and purified by silica gel rapid chromatography (ethyl acetate / petroleum ether = 0 to 16%) to give ethyl 1-((2-((3-cyano-4,5-dimethylthiophene-2-yl)amino)-2-oxoethyl)thio)cyclobutane carboxylate (70.0 mg, 198.6 μmol, yield 60.4%) as a yellow solid, which was used directly in the next step.

[0625] LCMS analysis confirmed the preparation of the desired intermediate compound. LCMS R t = 0.445 min, 0.8 min chromatography, 5-95AB, ESI calculated value, C 16 H 21 N₂O₃S₂[M+H] + 353.1, measured value 352.9.

[0626] Step 2 -1-((2-((3-cyano-4,5-dimethylthiophen-2-yl)amino)-2-oxoethyl)thio)cyclobutane Synthesis of alkylcarboxylic acid (FTS037) To a mixed solvent solution of ethyl 1-[2-[(3-cyano-4,5-dimethyl-2-thienyl)amino]-2-oxoethyl]thioalkylcyclobutane carboxylate (70.0 mg, 198.6 μmol, 1.0 equivalent) in MeOH (1 mL) and H₂O (0.2 mL), LiOH·H₂O (41.6 mg, 992.9 μmol, 5.0 equivalent) was added. The mixture was stirred at 50 °C for 2 h, filtered, concentrated, and purified by preparative HPLC (column: Phenomenex luna C18 150x25 mm x 10 μm; mobile phase: [water (FA)-ACN]; B%: 28%-58%, 10 min) to give the desired compound as a yellow solid (25.3 mg, 77.99 μmol, yield 39.2%).

[0627] NMR, LCMS, and HPLC analyses confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, CDCl3) δ= 9.95 (s, 1H), 3.55 (s, 2H), 2.79-2.72 (m, 2H), 2.81-2.71 (m, 1H), 2.33-2.27(m, 1H), 2.26 (s, 3H), 2.22 (m, 1H), 2.20-2.17 (m, 1H), 2.16 (s, 3H), 2.02-1.95 (m, 1H). LCMS R t = 1.406 min, 3 min chromatography, 5-95 AB, ESI calculated value, C 14 H 16 N₂O₃S₂Na [M+Na] + 347.1, measured value 347.0. HPLC purity 99.4%, R t = 2.640 min, 6.0 min chromatography, 10-80AB_6min.

[0628] Example 20: Preparation of 2-((2-((3-cyano-4,5-dimethylthiophen-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (compound FTS038)

[0629]

[0630] FTS038

[0631] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS038 using the synthetic method shown in Scheme 20 below.

[0632] Option 20

[0633]

[0634] Materials and methods

[0635] 0.281 mmol of 3,3-dimethyl-1,4-oxothiacyclohexane-2,6-dione and 0.213 mmol (0.75 equivalents) of 2-amino-4,5-dimethylthiophene-3-carboxynitrile were added to a 50 mL round-bottom flask. The flask was purged with argon and 15 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. The crude mixture was purified by dissolving impurities in ethyl acetate and acetone. The resulting white powder was filtered and washed with more ethyl acetate to yield 2-((2-((3-cyano-4,5-dimethylthiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid as a pale yellow powder.

[0636] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (500MHz), DMSO-d6δ: 12.65 (s,1H), 11.66 (s, 1H), 4.15, 3.69 (s, 2H), 2.25 (s, 3H), 2.10 (s, 3H), 1.42 (s,6H)

[0637] Example 21: Preparation of 2-((2-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (compound FTS039)

[0638]

[0639] FTS039

[0640] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS039 via a three-step synthetic method as shown in Scheme 21 below.

[0641] Option 21

[0642] Step 1

[0643]

[0644] Step 2

[0645]

[0646] Step 3

[0647]

[0648] Materials and methods

[0649] Step 1: 0.680 mmol of 2-butanone, 1.36 mmol (2 equivalents) of 2-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)acetonitrile, and 1.36 mmol (2 equivalents) of ammonium acetate were added to a 100 mL double-necked round-bottom flask. The flask was purged with argon and 10 mL of anhydrous toluene was added. The reaction mixture was heated under reflux for 18 h. The reaction mixture was cooled and poured into 30 mL of 10% NaHCO3 aqueous solution and 30 mL of EtOAc. The aqueous layer was separated and extracted again with 30 mL of EtOAc. The combined organic layers were washed with brine and dried over anhydrous sodium sulfate. Excess solvent was removed by rotary evaporation. The crude mixture was purified by column chromatography in ethyl acetate containing 90% hexane to produce ( Z )-2-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-3-methylpent-2-enonitrile and ( E A mixture of 2-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-3-methylpent-2-enonitriles.

[0650] Step 2: Will( Z )-2-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-3-methylpent-2-enonitrile and ( E A mixture of 1.12 mmol of 2-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-3-methylpentan-2-enonitriles, 2.8 mmol (2.5 equivalents) of DBU, and 1.12 mmol (1 equivalent) of elemental sulfur was added to a 100 mL double-necked round-bottom flask. The flask was purged with argon and 20 mL of anhydrous EtOH. The reaction mixture was heated to 65 °C for 2 hours under reflux. The reaction mixture was cooled and then poured into 30 mL of 10% NaHCO3 aqueous solution and 30 mL of EtOAc. The aqueous layer was separated and extracted again with 30 mL of EtOAc. The organic layers were combined, washed with brine, and dried over anhydrous sodium sulfate. Excess solvent was removed by rotary evaporation. The crude mixture was purified by passing it through a silica gel column in ethyl acetate containing 90% hexane to produce 3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-amine (42 mg, 15.9%) as a pale yellow powder.

[0651] Step 3:0.227 mmol of 3,3-dimethyl-1,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-amine (0.170 mmol, 0.75 equivalents) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 15 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. The crude mixture was purified by preparative TLC in ethyl acetate containing 2.5 hexane and 0.1% acetic acid to give 2-((2-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (5 mg, 7.4%) as a pale yellow powder.

[0652] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (500 MHz, DMSO-d6) δ=12.67(bs),3.64 (s,2H),1.83 (m,3H),1.73 (m,3H),1.52 (m,1H),1.24 (m,2H),0.83 (m,2H)

[0653] Example 22: Preparation of (1s,3s)-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)carbamoyl)cyclobutane-1-carboxylic acid (compound FTS040)

[0654]

[0655] FTS040

[0656] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS040 via a four-step synthetic method as shown in Scheme 22 below.

[0657] Option 22

[0658] Step 1

[0659]

[0660] Step 2

[0661]

[0662] Step 3

[0663]

[0664] Step 4

[0665]

[0666] Materials and methods

[0667] Step 1 - Synthesis of 2-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)acetonitrile: Under a nitrogen atmosphere, in a dry, single-necked round-bottom flask, N-hydroxycyclopropanemidine (0.368 g, 3.68 mmol) was added to a stirred solution of 3-(3,5-dimethyl-1H-pyrazol-1-yl)-3-oxopropionitrile (0.5 g, 3.06 mmol) in dioxane (25 mL), and the mixture was heated to 105 °C for 3 h. The mixture was then analyzed by TLC (petroleum ether containing 10% EtOAc, 0.5 mL). Rf The reaction progress was monitored. After the reaction was complete, the reaction mixture was concentrated under reduced pressure using a rotary evaporator to give crude 2-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)acetonitrile as an orange liquid. TLC (petroleum ether containing 10% EtOAc) R f =0.5. LCMS method C, 1.44 min 19.12% of the measured value [MH]148.1.

[0668] Step 2 - 3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-amine Synthesis: Under a nitrogen atmosphere, in a dry single-necked round-bottom flask, sulfur (0.097 g, 3.06 mmol) was added to a stirred solution of 2-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)acetonitrile (1 g, crude) in ethanol (30 mL) and but-2-one (0.218 g, 3.06 mmol), and the mixture was heated to 50 °C. Morpholine (0.263 mL, 3.06 mmol) was then added at 50 °C, and the mixture was stirred at 50 °C for 16 h. The mixture was analyzed by TLC (petroleum ether containing 10% EtOAc, 0.7 μL). Rf The reaction progress was monitored. After the reaction was complete, the reaction mixture was concentrated under reduced pressure using a rotary evaporator, and the resulting residue was diluted with water (80 mL) and extracted with ethyl acetate (2 × 50 mL). The organic layer was washed with a brine solution, dried over sodium sulfate, and concentrated to give the crude compound. The crude product was purified by column chromatography (Isolera) using EtOAc and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 10% EtOAc) to give 3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-amine as a grayish-white solid.

[0669] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, DMSO- d6) δ =7.41 (s, 2H), 2.19 (s, 3H), 2.14-2.09 (m, 4H), 2.14-2.09 (m, 4H).

[0670] Step 3 - ( 1s , 3s )-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2- Synthesis of methyl carbamoyl cyclobutane-1-carboxylate: Under a nitrogen atmosphere, at 25 °C, in a dry, double-necked round-bottom flask, (1s,3s)-3-(methoxycarbonyl)cyclobutane-1-carboxylic acid (202 mg, 1.275 mmol) and DIPEA (0.891 mL, 5.10 mmol) were added to a stirred solution of 3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-amine (200 mg, 0.850 mmol) dissolved in DCM (40 mL). The reaction mixture was cooled to 0 °C, and POCl3 (0.238 mL, 2.55 mmol) was added dropwise at 0 °C, with stirring at 25 °C for 16 h under a nitrogen atmosphere. The mixture was then analyzed by TLC (petroleum ether containing 20% ​​EtOAc, 0.5 mL). rf ) Monitor the progress of the reaction. After the reaction was complete, the reaction mixture was quenched with 100 mL of NaHCO3 solution. The reaction mixture was extracted with DCM (3 x 100 mL). The combined organic layers were dried over sodium sulfate and concentrated by rotary evaporation under reduced pressure (bath temperature 45 °C) to obtain the crude product. The crude product was purified by column chromatography (Isolera) using EtOAc and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 10% EtOAc) to give pure (1S,3S)-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-yl)carbamoyl)cyclobutane-1-carboxylate as a brown solid.

[0671] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, DMSO- d6 ) δ=11.12 (s, 1H), 3.62 (s, 3H), 3.42-3.35 (m, 1H), 3.28-3.21 (m, 1H), 2.51 (s,5H), 2.37-2.27 (m, 5H), 2.26-2.22 (m, 1H), 1.22-1.16 (m, 2H), 1.13-1.04 (m, 2H).

[0672] Step 4 - (1 s ,3 s )-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2- Synthesis of (carbamoyl)cyclobutane-1-carboxylic acid:In a dry, single-necked round-bottom flask, methyl (1s,3s)-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-yl)carbamoyl)cyclobutane-1-carboxylate (0.120 g, 0.320 mmol) dissolved in THF (6 mL) and water (5 mL) with stirring, lithium hydroxide monohydrate (0.040 g, 0.959 mmol) was added, and the mixture was stirred at room temperature for 1 h. The reaction progress was monitored by TLC. The TLC met the requirements. Upon completion of the reaction, the reaction mixture was diluted with water (5 mL) and extracted with ethyl acetate (15 mL). The aqueous layer was acidified with citric acid and extracted with 10% MeOH / DCM. The organic layer was dried with sodium sulfate and concentrated under reduced pressure using a rotary evaporator to obtain pure (1s,3s)-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)carbamoyl)cyclobutane-1-carboxylic acid (0.080 g, 0.221 mmol, 69.1% yield) as a white solid.

[0673] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO- d6 ) δ = 12.24(s, 1H), 11.13 (s, 1H), 3.37 (s, 1H), 3.35-3.10 (m, 1H), 2.51-2.47 (m, 4H), 2.46-2.41 (m, 6H), 2.21 (s, 1H), 1.17-1.14 (m, 2H), 1.06-1.03 (m, 2H).

[0674] Example 23: Preparation of (1r,3r)-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)carbamoyl)cyclobutane-1-carboxylic acid (compound FTS041)

[0675]

[0676] FTS041

[0677] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS041 via a two-step synthetic method as shown in Scheme 23 below.

[0678] Option 23

[0679] Step 1

[0680]

[0681] Step 2

[0682]

[0683] Materials and methods

[0684] Step 1 - (1 r ,3 r )-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2- Synthesis of methyl carbamoyl cyclobutane-1-carboxylate In a dry, double-necked round-bottom flask under a nitrogen atmosphere, 3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-amine (0.170 g, 0.722 mmol) was dissolved in DCM (10 mL). Under a nitrogen atmosphere, (1r,3r)-3-(methoxycarbonyl)cyclobutane-1-carboxylic acid (0.137 g, 0.867 mmol) and DIPEA (0.560 g, 4.33 mmol) were added to this reaction mixture at 25 °C. The reaction mixture was cooled to 0 °C, and POCl3 (0.332 g, 2.167 mmol) was added dropwise, followed by stirring at 25 °C for 16 h under a nitrogen atmosphere. The mixture was analyzed by TLC (using petroleum ether containing 20% ​​EtOAc). 0.6 rf The reaction progress was monitored. After the reaction was complete, the reaction mixture was quenched with NaHCO3 solution (50 mL). The reaction mixture was extracted with DCM (3 x 50 mL). The combined organic layers were dried over sodium sulfate and concentrated by rotary evaporation under reduced pressure (bath temperature 45 °C) to give the crude product. The crude product was purified by column chromatography (Isolera) using ethyl acetate and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 15% ethyl acetate) to give pure (1r,3r)-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-yl)carbamoyl)cyclobutane-1-carboxylate (0.060 g, 0.160 mmol, yield 22.12%).

[0685] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, DMSO- d6 ) δ=11.13(s,1H),3.66 (s,3H),3.40-3.50 (m,1H),3.12-3.25 (m,1H),2.52-2.50 (m,4H),2.30-2.29 (m,7H),1.16-1.14 (m,2H),1.05-1.03 (m,2H).

[0686] Step 2 - (1) r ,3 r )-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2- Synthesis of (carbamoyl)cyclobutane-1-carboxylic acid Under a nitrogen atmosphere, methyl (1r,3r)-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)carbamoyl)cyclobutane-1-carboxylate (0.050 g, 0.133 mmol) was dissolved in THF (3 mL), water (2 mL), and lithium hydroxide (9.57 mg, 0.400 mmol) was added. The mixture was stirred at room temperature for 1 h. The progress of the reaction was monitored by TLC (DCM containing 5% MeOH). Upon completion of the reaction, the reaction mixture was diluted with water (5 mL), extracted with ethyl acetate (15 mL), and the aqueous layer was acidified with citric acid and extracted with 10% MeOH / DCM. The organic layer was dried with sodium sulfate and concentrated under reduced pressure using a rotary evaporator to obtain pure (1r,3r)-3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)carbamoyl)cyclobutane-1-carboxylic acid as a white solid.

[0687] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO- d6 ) δ = 11.06(s, 1H), 3.44-3.40 (m, 1H), 3.08 (s, 1H), 2.49 (s, 4H), 2.34-2.33 (m, 6H), 2.30-2.22 (m, 1H), 1.17-1.15 (m, 2H), 1.05-1.03 (m, 2H).

[0688] Example 24: Preparation of 3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)carbamoyl)-bicyclo[1.1.1]pentane-1-carboxylic acid (compound FTS042)

[0689]

[0690] FTS042

[0691] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS042 using the synthetic method shown in Scheme 24 below.

[0692] Option 24

[0693] Step 1

[0694]

[0695] Step 2

[0696]

[0697] Materials and methods

[0698] Step 1 - 3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)carbamoyl Synthesis of methyl 1-pentane-1-carboxylate (B2)bicyclo[1.1.1]pentane-1-carboxylate: In a dry, double-necked round-bottom flask under a nitrogen atmosphere, 3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-amine (500 mg, 2.125 mmol) was dissolved in DCM (15 mL). Under a nitrogen atmosphere, 3-(methoxycarbonyl)bicyclo[1.1.1]pentane-1-carboxylic acid (542 mg, 3.19 mmol) and DIPEA (1.856 mL, 10.62 mmol) were added to this reaction mixture at 25 °C. The reaction mixture was cooled to 0 °C, and Pocl3 (0.596 mL, 6.37 mmol) was added dropwise, followed by stirring at 25 °C for 16 h under a nitrogen atmosphere. The progress of the reaction was monitored by TLC (petroleum ether containing 20% ​​EtOAc, 0.4 rf).

[0699] After the reaction was complete, the reaction mixture was quenched with NaHCO3 solution (250 mL). The reaction mixture was extracted with DCM (3 x 200 mL). The combined organic layers were dried over sodium sulfate and concentrated by rotary evaporation under reduced pressure (bath temperature 45 °C) to obtain the crude product. The crude product was purified by column chromatography (Isolera) using EtOAC and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 15% EtOAC) to give pure methyl 3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-yl)carbamoyl)bicyclo[1.1.1]pentane-1-carboxylate as a pale yellow solid.

[0700] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO- d6 ) δ = 11.29(s, 1H), 3.67 (s, 3H), 2.46 (s, 6H), 2.30 (s, 6H), 2.28 (t, J = Hz, 1H), 1.14(d, J = 8.00 Hz, 2H), 1.12 (d, J = 6.00 Hz, 2H).

[0701] Step 2 - 3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)carbamoyl Synthesis of 1-pentane-1-carboxylic acid (BOC)In a dry, single-necked round-bottom flask under a nitrogen atmosphere, methyl 3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)carbamoyl)bicyclo[1.1.1]pentane-1-carboxylate (250 mg, 0.645 mmol) was dissolved in water (5 mL), THF (5 mL), and MeOH (1 mL). Lithium hydroxide monohydrate (81 mg, 1.936 mmol) was added to this reaction mixture at 25 °C under a nitrogen atmosphere. The reaction mixture was stirred at 25 °C for 1 h under a nitrogen atmosphere. The progress of the reaction was monitored by TLC (petroleum ether containing 20% ​​EtOAc, 0.2 rf). After the reaction was complete, the reaction mixture was concentrated by rotary evaporation under reduced pressure (bath temperature 45 °C). The reaction mixture was washed with ethyl acetate (10 mL). The aqueous layer was then acidified with citric acid (pH=5-6). The reaction mixture was extracted with DCM (3 x 20 mL) containing 10% MeOH. The combined organic layers were dried over sodium sulfate, concentrated by rotary evaporation under reduced pressure (bath temperature 45 °C), and lyophilized to give pure 3-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)carbamoyl)-bicyclo[1.1.1]pentane-1-carboxylic acid as a white solid.

[0702] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO- d6 ) δ = 12.73(s, 1H), 11.30 (s, 1H), 2.35 (s, 3H), 2.22 (s, 1H), 2.13 (s, 6H), 1.93 (s,3H), 1.21-1.17 (m, 2H), 1.10-1.06 (m, 2H).

[0703] Example 25: Preparation of 2-(1-(2-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)amino)-2-oxoethyl)cyclopentyl)acetic acid (compound FTS043)

[0704]

[0705] FTS043

[0706] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS043 by the synthetic method shown in Scheme 25 below.

[0707] Option 25

[0708]

[0709] Materials and methods

[0710] Under a nitrogen atmosphere, in a dry, single-necked round-bottom flask, 0.100 g (0.425 mmol) of 3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophene-2-amine (10 mL) dissolved in DCM was added to a stirred solution, and the mixture was stirred at room temperature for 48 h. The progress of the reaction was monitored by TLC (hexane containing 20% ​​EtOAc, 0.3 rf). The TLC indicated the completion of the reaction. The reaction mixture was concentrated under reduced pressure to give the crude compound. The crude product was purified by column chromatography (Isolera) using EtOAc and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 15% EtOAc). The product peaks were collected together and concentrated to obtain pure 2-(1-(2-((3-(3-cyclopropyl-1,2,4-oxadiazol-5-yl)-4,5-dimethylthiophen-2-yl)amino)-2-oxoethyl)cyclopentyl)acetic acid as a white solid.

[0711] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO- d6 ) δ = 12.05(s, 1H), 11.16 (s, 1H), 2.69 (s, 2H), 2.38 (s, 2H), 2.29 (d, J = 7.20 Hz,6H), 2.26-2.21 (m, 1H), 1.67-1.58 (m, 8H), 1.15-1.12 (m, 4H).

[0712] Example 26: Preparation of 2-((2-((3-cyanobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (compound FTS044)

[0713]

[0714] FTS044

[0715] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS044 via a two-step synthetic method as shown in Scheme 26 below.

[0716] Option 26

[0717] Step 1

[0718]

[0719] Step 2

[0720]

[0721] Materials and methods

[0722] Step 1 - 2-Chloro- N -(3-cyanobenzo[ b Synthesis of thiophene-2-yl)acetamide: Under a nitrogen atmosphere, in a dry, single-necked round-bottom flask, 0.200 g (1.148 mmol) of 2-aminobenzo[b]thiophene-3-carboxynitrile was dissolved in 10 mL of dioxane, and 0.130 g (1.148 mmol) of 2-chloroacetyl chloride was added at room temperature. The mixture was stirred for 16 h at the same temperature. The reaction was monitored by TLC. 10 mL of hexane was added to the reaction mixture and stirred for 10 min. A solid was observed in the reaction mixture, and the solid was filtered through a Buchner funnel and washed with hexane. The solid was dried under reduced pressure to obtain a pure, off-white solid of 2-chloro-N-(3-cyanobenzo[b]thiophene-2-yl)acetamide.

[0723] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, DMSO- d6 ) δ =12.43 (s, 1H), 8.04 (d, J = 8.00 Hz, 1H), 7.71 (d, J = 8.00 Hz, 1H), 7.56-7.51 (m, 1H), 7.45-7.41 (m, 1H), 4.58 (s, 2H).

[0724] Step 2 - 2-((2-((3-cyanobenzo[ b ]Thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methyl Synthesis of propionic acid:Under a nitrogen atmosphere, in a dry 50 mL single-necked round-bottom flask, methyl 2-mercapto-2-methylpropionate (0.141 g, 1.053 mmol) and cesium carbonate (0.572 g, 1.755 mmol) were added to a stirred solution of 2-chloro-N-(3-cyanobenzo[b]thiophene-2-yl)acetamide (0.220 g, 0.878 mmol), and the mixture was stirred at room temperature for 16 h. The completion of the reaction was monitored by TLC (petroleum ether containing 20% ​​EtOAc). Then, THF (5 mL), water (5.00 mL), and lithium hydroxide (0.017 g, 0.717 mmol) were added to this reaction mixture, and the mixture was stirred at room temperature for 16 h. The completion of the reaction was monitored by TLC. The reaction mixture was diluted with water (5 mL) and washed with ethyl acetate (15 mL). The aqueous layer was acidified with citric acid (pH ~4) and extracted with 10% MeOH / DCM. The organic layer was dried over sodium sulfate and concentrated under reduced pressure by rotary evaporation to obtain pure 2-((2-((3-cyanobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid as an off-white solid.

[0725] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO- d6 ) δ = 12.69(s, 1H), 12.20 (s, 1H), 8.00 (d, J = 8.00 Hz, 1H), 7.69 (d, J = 8.00 Hz, 1H),7.52 (t, J = 7.20 Hz, 1H), 7.41 (t, J = 7.20 Hz, 1H), 3.83 (s, 2H), 1.46 (s, 6H).

[0726] Example 27: Preparation of 2-((2-((6-chloro-3-cyanobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (compound FTS045)

[0727]

[0728] FTS045

[0729] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS045 via a three-step synthetic method as shown in Scheme 27 below.

[0730] Option 27

[0731] Step 1

[0732]

[0733] Step 2

[0734]

[0735] Materials and methods

[0736] Step 1 - 2-Amino-6-chlorobenzo[ b Synthesis of thiophene-3-carboxynitrile: Under N2, sodium hydride (0.660 g, 16.51 mmol) was added dropwise to a solution of 2-(4-chloro-2-fluorophenyl)acetonitrile (2.0 g, 11.79 mmol) in DMSO (30.0 mL) at room temperature (slightly exothermic). After 30 min, the reaction mixture was cooled to 15 °C in a water bath, and O-ethyl carbonisothiocyanate (1.530 mL, 12.97 mmol) was added dropwise. After 1 h, the reaction mixture was heated at 100 °C for 2 h. The progress of the reaction was monitored by TLC (petroleum ether containing 30% EtOAc) and LC-MS. After the reaction was complete, the reaction mixture was quenched with water to obtain a solid. The solid was filtered, washed with water, and dried under vacuum to yield the target compound as a coarse yellow solid. The crude product was purified by column chromatography (Isolera) using EtOAc and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 30% EtOAc) to give solid 3-amino-6-chlorobenzo[b]thiophene-2-carboxynitrile.

[0737] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, DMSO- d6 ) δ =7.96 (br s, 2H), 7.83 (d, J = 2.00 Hz, 1H), 7.32 (dd, J = 2.00, 8.40 Hz, 1H), 7.26 (d, J = 8.40 Hz, 1H).

[0738] Step 2 - 2-Chloro -N- (6-Chloro-3-cyanobenzyl[ b Synthesis of thiophene-2-yl)acetamide:2-Chloroacetyl chloride (379 mg, 3.35 mmol) was added to a solution of 2-amino-6-chlorobenzo[b]thiophene-3-carboxynitrile (350 mg, 1.677 mmol) in dioxane (10.0 mL) at room temperature. The resulting mixture was stirred at room temperature for 16 h. The progress of the reaction was monitored by TLC (petroleum ether containing 30% EtOAc) and LCMS. After the reaction was complete, the mixture was diluted with hexane. The resulting solid was filtered, washed with hexane, and dried under vacuum to give the target compound 2-chloro-N-(6-chloro-3-cyanobenzo[b]thiophene-2-yl)acetamide (400 mg, 1.019 mmol, yield 60.8%) as a white solid.

[0739] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, DMSO- d6 ) δ =12.53 (s, 1H), 8.22 (d, J = 2.0 Hz, 1H), 7.70 (d, J = 8.80 Hz, 1H), 7.55 (dd,J = 2.00, 8.80 Hz, 1H), 4.59 (s, 2H).

[0740] Step 3 - 2-((2-((6-chloro-3-cyanobenzo[ b ]Thiophene-2-yl)amino)-2-oxoethyl)thio)-2- Synthesis of methylpropionic acid:Methyl 2-mercapto-2-methylpropionate (176 mg, 1.315 mmol) was added to a solution of 2-chloro-N-(6-chloro-3-cyanobenzo[b]thiophene-2-yl)acetamide (250 mg, 0.877 mmol) and cesium carbonate (428 mg, 1.315 mmol) in ACN (15.0 mL). The resulting mixture was stirred at room temperature for 16 h. The progress of the reaction was monitored by TLC and LC-MS, showing the formation of intermediate 7. THF (10.0 mL) was added to the same reaction mixture, followed by water (3.0 mL) containing lithium hydroxide monohydrate (184 mg, 4.38 mmol). The resulting mixture was stirred at room temperature for another 16 h. The reaction was monitored by TLC (petroleum ether containing 30% EtOAc + 0.5 mL acetic acid) and LC-MS. After the reaction was complete, the reaction mixture was quenched with water. The reaction mixture was extracted with ethyl acetate (2 x 30 mL). The separated aqueous layer was acidified with 6N HCl aqueous solution and extracted with ethyl acetate (3 x 30 mL). The combined organic layers were washed with brine, dried over sodium sulfate, and concentrated under reduced pressure by rotary evaporation to obtain the crude product. The crude product was purified by column chromatography (Isolera) using DCM and methanol as the elution solvent system (the product was eluted in DCM containing 3-5% MeOH) to give pure 2-((2-((6-chloro-3-cyanobenzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (28.6 mg, 0.877 mmol, yield 8.84%) as a white solid.

[0741] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO- d6 ) δ = 12.70(br s, 1H), 12.31 (br s, 1H), 8.18 (s, 1H), 7.67 (d, J = 8.40 Hz, 1H), 7.54(d, J = 8.40 Hz, 1H), 3.82 (s, 2H), 1.46 (s, 6H).

[0742] Example 28: 2-((2-((3-cyano-5,6-dihydro-4- H -Cyclopenta[ b Preparation of thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (compound FTS046)

[0743]

[0744] FTS046

[0745] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS046 using the synthetic method shown in Scheme 28 below.

[0746] Option 28

[0747]

[0748] Materials and methods

[0749] 3,3-Dimethyl-1,4-oxothiacyclohexane-2,6-dione (0.911 mmol) and 2-amino-5,6-dihydro-4 H -Cyclopenta[ b Thiophene-3-carboxynitrile (0.684 mmol, 0.75 equivalents) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 15 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 2-((2-((3-cyano-5,6-dihydro-4-)) as a gray powder. H -Cyclopenta[ b Thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (95 mg, 42.8%).

[0750] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400MHz), DMSO-d6δ: 1.43 (s,6H), 2.36 (quintet, 2H), 2.72 (t, 2H), 2.82 (t, 2H), 3.70 (s, 2H), 11.71 (s,1H), 12.66 (s, 1H).

[0751] Example 29: Preparation of 2-((2-((3-cyano-4,5,6,7-tetrahydro-4,7-methylenebridged benzo[b]thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (compound FTS047)

[0752]

[0753] FTS047

[0754] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS047 using the synthetic method shown in Scheme 29 below.

[0755] Option 29

[0756]

[0757] Materials and methods

[0758] 3,3-Dimethyl-1,4-oxothiacyclohexane-2,6-dione (0.697 mmol) and 2-amino-4,5,6,7-tetrahydro-4,7-methylbridged benzo[ b Thiophene-3-carboxynitrile (0.523 mmol, 0.75 equivalents) was added to a 50 mL round-bottom flask. The flask was purged with argon, and 20 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. The reaction mixture was extracted with saturated NaHCO3. Concentrated HCl was added to the aqueous layer, and the mixture was then extracted with CH2Cl2, washed with brine, and dried over anhydrous sodium sulfate. Excess solvent was removed by rotary evaporation to produce 2-((2-((3-cyano-4,5,6,7-tetrahydro-4,7-methylbridged benzo[] as a white powder. b Thiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (19 mg, 10.4%).

[0759] Example 30: Preparation of 2-((2-((3-cyano-4-cyclopropylthiophen-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (compound FTS048)

[0760]

[0761] FTS048

[0762] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS048 using the synthetic method described in Scheme 30 below.

[0763] Option 30

[0764]

[0765] Materials and methods

[0766] 3,3-Dimethyl-1,4-oxothiacyclohexane-2,6-dione (0.787 mmol) and 2-amino-4-cyclopropylthiophene-3-carboxynitrile (0.590 mmol, 0.75 equivalents) were added to a 50 mL round-bottom flask. The flask was purged with argon, and 15 mL of anhydrous dichloromethane was added. The mixture was stirred overnight under positive argon pressure. A white powder was formed by rapid cooling and recrystallization, which was separated by vacuum filtration and washed with cold CH2Cl2. The residual solvent was removed by rotary evaporation to produce 2-((2-(((3-cyano-4-cyclopropylthiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (45 mg, 23.4%) as a white powder.

[0767] NMR analysis confirmed the preparation of the desired product compound. ¹H NMR (500 MHz), DMSO-d⁶ δ: 0.66 (dd, 2H), 0.917 (dd, 2H), 1.437 (s, 6H), 1.84 (m, 1H), 3.72 (s, 2H), 6.65 (s, 1H), 11.76 (s, 1H), 12.67 (s, 1H).

[0768] Example 31: Preparation of 2-((2-((3-cyano-4-methyl-5-phenylthiophen-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (compound FTS049)

[0769]

[0770] FTS049

[0771] This embodiment illustrates the preparation of the FAB3 / 4 / 5 / 7 inhibitor compound FTS049 using the synthetic method described in Scheme 31 below.

[0772] Option 31

[0773] Step 1

[0774]

[0775] Materials and methods

[0776] Step 1 - 2-Chloro- N Synthesis of 3-cyano-4-methyl-5-phenylthiophen-2-yl)acetamide:Et3N (826 mg, 8.17 mmol) was added to a THF (15 mL) solution of 2-amino-4-methyl-5-phenylthiophene-3-carboxynitrile (500 mg, 2.333 mmol) at 0 °C, followed by the addition of 2-chloroacetyl chloride (659 mg, 5.83 mmol). The resulting mixture was stirred at room temperature for 2 h. The progress of the reaction was monitored by TLC (petroleum ether containing 10% EtOAc) and LCMS. After the reaction was complete, the reaction mixture was quenched with water. The reaction mixture was extracted with ethyl acetate (3 x 20 mL). The combined organic layers were separated and washed with brine, dried over sodium sulfate, and concentrated in a rotary evaporator under reduced pressure to give the crude product. The crude product was purified by column chromatography (Isolera) using EtOAc and petroleum ether as the elution solvent system (the product was eluted in petroleum ether containing 5-10% EtOAc) to give pure 2-chloro-N-(3-cyano-4-methyl-5-phenylthiophene-2-yl)acetamide (290 mg, 0.997 mmol, yield 43%) as a brown solid.

[0777] NMR analysis confirmed the preparation of the desired intermediate compound. 1 H NMR (400 MHz, DMSO-) d6 ) δ =12.16 (s, 1H), 7.39-7.51 (m, 5H), 4.52 (s, 2H), 2.30 (s, 3H).

[0778] Step 2 - 2-((2-((3-cyano-4-methyl-5-phenylthiophen-2-yl)amino)-2-oxoethyl)thio)- Synthesis of 2-methylpropionic acid:Methyl 2-mercapto-2-methylpropionate (242 mg, 1.806 mmol) was added to a solution of 2-chloro-N-(3-cyano-4-methyl-5-phenylthiophene-2-yl)acetamide (350 mg, 1.204 mmol) and cesium carbonate (588 mg, 1.806 mmol) in ACN (15.0 mL). The resulting mixture was stirred at room temperature for 16 h. The formation of intermediate 5 was observed by TLC and LCMS. THF (10.0 mL) was added to the same reaction mixture, followed by water (3.0 mL) containing lithium hydroxide monohydrate (253 mg, 6.02 mmol). The resulting mixture was stirred at room temperature for another 16 h. The progress of the reaction was monitored by TLC [petroleum ether containing 30% (10 mL) EA + 0.5 mL acetic acid] and LCMS. After the reaction was complete, the reaction mixture was quenched with water. The reaction mixture was extracted with ethyl acetate (2 x 30 mL). The separated aqueous layer was acidified with 6N HCl aqueous solution and extracted with ethyl acetate (3 x 30 mL). The combined organic layers were washed with brine, dried over sodium sulfate, and concentrated under reduced pressure by rotary evaporation to obtain the crude product. The crude product was purified by column chromatography (Isolera) using DCM and methanol as the elution solvent system (the product was eluted in DCM containing 3-5% MeOH) to give pure 2-((2-((3-cyano-4-methyl-5-phenylthiophene-2-yl)amino)-2-oxoethyl)thio)-2-methylpropionic acid (48.10 mg, 0.128 mmol, yield 10.67%) as a white solid.

[0779] NMR analysis confirmed the preparation of the desired product compound. 1 H NMR (400 MHz, DMSO-) d6 ) δ = 12.67(br s, 1H), 11.97 (br s, 1H), 7.44-7.51 (m, 4H), 7.38-7.42 (m, 1H), 3.76 (s, 2H), 2.29 (s, 3H), 1.44 (s, 6H).

[0780] Example 32: Screening of FABP3 / 4 / 5 / 7 inhibitor compounds

[0781] This example illustrates a two-step fluorescence binding assay used to determine the binding affinity of the various inhibitor compounds disclosed herein to various FABPs (FABP3, FABP4, FABP5, and FABP7). Further secondary screening was conducted on compounds exhibiting high binding affinity to FABPs and based on their ability to activate nuclear receptors PPARα, PPARγ, or PPARδ.

[0782] Materials and methods

[0783] A. Combined with determination

[0784] The binding of FABP3, FABP4, FABP5, and FABP7 was determined by fluorescent titration. His-tagged FABPs in... E. coli Bacterial expression was performed, purified using Ni Sepharose beads, and the equilibrium dissociation constant (Kd) characterizing their interactions with different inhibitory compounds was measured by a fluorescence competitive assay. The method comprises two steps, as described, for example, in Lin, Q. et al., “Ligand selectivity of the peroxisome proliferator-activated receptor alpha,” Biochemistry 38, 185-190, doi:10.1021 / bi9816094bi9816094 [pii] (1999). The first step involves measuring the Kd of protein association with the fluorescent fatty acid probe ANS. The protein (2 μM) was titrated from a concentrated solution in DMSO with ANS. Ligand binding was monitored by tracking the increase in fluorescence intensity after ligand binding to the protein, and the Kd of association between ANS and each FABP was calculated according to titration curves described, for example, Norris, AW & Li, E., “Fluorometrictitration of the CRABPs,” Methods Mol Biol 89, 123-139 (1998)). In a second step, the binding Kd of non-fluorescent ligands was measured by monitoring their ability to displace ANS in the protein binding pocket. Each FABP was pre-compounded with ANS in a 1:1 molar ratio and titrated with different compounds; the binding of the compounds was reflected by a decrease in probe fluorescence intensity. EC50 values ​​from the competition curves were used to determine the binding Kd of the non-fluorescent ligands. 50 Kd is extracted from the measurement Kd of ANS.

[0785] B. Transcription activation assay

[0786] COS-7 cells were cultured in 6-well plates and co-transfected with an expression vector for PPARδ, PPARα, or PPARγ driven by 3 copies of PPRE, and a vector carrying β-galactosidase cDNA as a transfection control. To test whether FABP4 or FABP5 mediates the activation of their respective receptors PPARγ and PPARδ, cells were also co-transfected with plasmids containing the FABP4 or FABP5 sequences. Eighteen hours post-transfection, cells were placed in serum-free medium and treated with an agonist / compound. After 18 hours, cells were lysed, and luciferase activity was analyzed (Promega, WI, USA), with transfection efficiency corrected for by β-galactosidase activity.

[0787] result

[0788] Table 19 below summarizes the results of binding assays of FABP5, FABP4, FABP3, and FABP7 with the disclosed compounds FTS001, FTS003, FTS005, FTS007, FTS009, FTS011, FTS013, FTS017, FTS019, FTS026, FTS029, FTS030, FTS031, FTS032, FTS033, FTS034, FTS037, FTS039, FTS040, FTS041, FTS042, FTS043, FTS044, FTS045, FTS046, FTS048, and FTS049. None of these tested compounds activated PPARα, PPARγ, or PPARδ.

[0789] Table 19

[0790]

[0791]

[0792]

[0793]

[0794]

[0795]

[0796]

[0797] Example 33: Biological study of FABP3 / 4 / 5 / 7 inhibitor compounds FTS005, FTS030, FTS031, FTS037 and FTS039

[0798] This example illustrates the biological functions of the FABP3 / 4 / 5 / 7 inhibitor compounds FTS005, FTS030, FTS031, FTS037, and FTS039 in cancer and metabolic disease models.

[0799] Materials and methods

[0800] A. Cells

[0801] COS-7, MDA-MB-231, NPG, HepG2, and 3T3-L1 cells were cultured in Dulbecco modified Eagle medium (DMEM) supplemented with 10% fetal bovine serum (Invitrogen Life Sciences, Carlsbad, CA). 4T1, 4T01, NR67, MDA-MB-157, MB-549, and OVCAR8 cells were cultured in RPMI medium containing L-glutamine supplemented with 10% fetal bovine serum (Invitrogen Life Sciences, Carlsbad, CA).

[0802] B. Transcription activation assay

[0803] As described above, transcriptional activation assays were performed using COS7 cells.

[0804] C. Real-time PCR

[0805] Cells were treated with the compound for 6 hours, then lysed, and RNA was extracted using Trizol according to the manufacturer's instructions. cDNA was synthesized using GeneAmp RNA PCR (Applied Biosystems). qPCR was performed using TaqMan chemistry and Assays-on-Demand probes (Applied Biosystems). Normalization was performed using 18S (4352930) rRNA. Relative expression levels were calculated as 2. -DDCT .

[0806] D. Proliferation assay

[0807] 2000 cells were seeded in each well of a 96-well plate. The cells were treated with the compound the following day and incubated in Incucyte for 4 days. Images taken every 4 hours throughout the period were analyzed to calculate the percentage of confluence in response to treatment. Growth inhibition rate was calculated as 1 - (percentage of surviving cells out of untreated cells).

[0808] E. Lipid uptake assay

[0809] 1) HepG2 cells: 5000 cells were seeded in 96-well plates. The next day, the cells were treated with the test compound for 4 hours, followed by an additional 24 hours of treatment with 1 mM oleic acid. The cells were then stained with Nile Red and Dapi, and lipid content was quantified using Cytation5 (Agilent).

[0810] 2) 3T3-L1: Mouse 3T3-L1 cells differentiated into mature adipocytes in culture according to the ATCC protocol. Compound treatment began on day 6 of differentiation. Cells were stained with Nile Red on the same day, and the color intensity of each cell relative to Dapi stain was measured.

[0811] F. Xenotransplantation experiments

[0812] 5 x 10 6 MB-231 cells were mixed with matrix gel at a 1:1 ratio and subcutaneously injected into the right abdomen of 7-week-old female NSG mice. Treatment began the day after cell injection and continued for 5 days a week. Mice were treated orally by gavage with the medium (sesame oil) or 20 mg / kg or 40 mg / kg FTS005 dissolved in the medium. Tumor size was measured twice weekly using digital calipers. Tumor volume was determined by measuring the length of the tumor. l ) and width ( w And calculate the volume () V = lw 2 The determination was made using / 2). Mice were sacrificed 24 days after injection. In both experiments, student... t The statistical significance between control and treated mice was assessed using laboratory tests. Mouse experiments were conducted with approval from the Case Western Reserve University Institutional Animal Care and Use Committee.

[0813] G. TNBC homologous mouse model

[0814] 1 x 10 5 Four T1 cells were mixed with matrix gel at a 1:1 ratio and subcutaneously injected into the mammary fat pads of 7-week-old female BALB / c mice. Treatment began the day after cell injection and continued for 5 days a week. Mice were treated by gavage with the medium (sesame oil) or orally with 40 mg / kg FTS005 dissolved in the medium. Tumor size was measured twice weekly using digital calipers. Tumor volume was determined by measuring the length of the tumor. l ) and width ( w And calculate the volume () V = lw 2 The determination was made using / 2). Mice were sacrificed 32 days after injection. In both experiments, student... tThe statistical significance between control and treated mice was assessed using laboratory tests. Mouse experiments were conducted with approval from the Case Western Reserve University Institutional Animal Care and Use Committee.

[0815] H. Isolation of primary T cells

[0816] T cells were collected from spleen cells of control BALB / c mice carrying 4T1 tumors or mice treated with FABP3 / 4 / 5 / 7 inhibitors.

[0817] I. Metabolomics analysis of tumor samples

[0818] 50 mg tumor samples were lysed in 500 μL PBS, and protein concentrations were measured. The samples were then analyzed by LC / MS / MS using a C18 (Gemini 5 μM, 2 x 150 mm, Phenomnex) column. The mobile phases used were: 1) Detection of TCA metabolites: A) water + 5 mM AmAc and B) methanol + 5 mM AmAc, at a flow rate of 0.3 mL / min; 2) Detection of glycolysis, fatty acid, and arachidonic acid oxidation metabolites: A) water + 0.1% acetic acid and B) methanol / CAN (1 / 1) + 0.1% acetic acid, at a flow rate of 0.3 mL / min.

[0819] result

[0820] A. Compounds FTS005, FTS030, FTS031, FTS037, and FTS039 do not activate PPAR transcription.

[0821] To eliminate the possibility that thiophene compounds FTS005, FTS030, FTS031, FTS037, and FTS039 are ligands of nuclear receptors PPARα, PPARγ, or PPARδ, thereby activating the transcription of these transcription factors, transcriptional activation assays were performed using COS7 cells. For comparison, known specific PPARα, PPARγ, or PPARδ agonist compound Wy-14643 was also analyzed. Figure 1A ), Rosiglitazone ( Figure 1B ) and GW0742 ( Figure 1C (All are 5 mM). For example... Figure 1A , Figure 1B and Figure 1C As shown in the data graph, unlike these known agonist compounds that can induce activation of their respective receptors, the tested compounds FTS005, FTS030, FTS031, FTS037 and FTS039 did not activate transcription.

[0822] B. Compound FTS005 more effectively inhibited FABP5-expressing TNBC cells compared to the commercially available inhibitor SBF-I26. growth

[0823] The efficacy of compound FTS005 in inhibiting cancer cell proliferation was calculated using TNBC cell lines MB-231 and BT-549. Cells were treated with serial dilutions of the compound, and proliferation was measured in Incucyte by calculating the percentage of confluence every 4 hours over 4 days. Figure 2A As shown, FTS005 can very effectively inhibit the proliferation of two human TNBC cell lines, MB-231 and BT-549, with calculated values ​​of 0.145 mM. IC50 and 0.25 mM IC50 .

[0824] To verify that the inhibitory effect of FTS005 on TNBC cells is mediated by FABP5, the MB-231 cell line stably expressing FABP5 shRNA was used. Figure 2B As shown, FTS005 only inhibited the proliferation of WT MB-231 and BT-549 cell lines expressing FABP5, but did not affect the proliferation of F5_KD MB-231 cell lines with low FABP5 levels.

[0825] The efficiency of FTS005 in inhibiting MB-231 cell proliferation was also compared with that of the known FABP5 / 7 inhibitor compound SBF-I26. Figure 2C As shown, the data clearly demonstrate that treatment of MB-231 cells with thiophene compounds is 10 to 20 times more effective than SBF-I26 in inhibiting the proliferation of TNBC cells.

[0826] In addition, the effect of FTS005 on the proliferation of mouse breast cancer cell lines 67NR, 4T07, and 4T1, which can serve as human TNBC models, was tested. Both 4T07 and 4T1 cell lines are derived from NR67, with 4T1 being the most metastatic and invasive of all. The expression levels of FABP5 in all cell lines were measured and compared with the human cell line MB-231. It was found that FABP5 levels were positively correlated with cell invasiveness, with the highest FABP5 levels in MB-231 cells, followed by 4T1 and 4T07. Figure 3A As shown, FABP5 was not detected in 67NR cells. As expected, treatment of mouse cancer cells with FTS005 inhibited their proliferation, but with less efficiency than in MB-231 cells (see [link to FTS005]). Figure 3B and Figure 3C The results showed that the inhibitor's effect depended on the expression level of FABP5. 67NR cells were completely unaffected by FTS005 ( Figure 3C ).

[0827] C. Compounds FTS005, FTS040, FTS041, FTS042, FTS043, FTS044, FTS045 and FTS049 inhibited Growth of human ovarian cancer cells OVCAR8

[0828] The efficacy of compounds FTS005, FTS040, FTS041, FTS042, FTS043, FTS044, FTS045, and FTS049 in inhibiting ovarian cancer cell proliferation was calculated using the ovarian cancer cell line OVCAR8. Cells were treated with serial dilutions of the compounds, and proliferation was measured in Incucyte by calculating the percentage of confluence every 4 hours over 4 days. Figure 4A As shown, all compounds can very effectively inhibit the proliferation of cancer cell lines. Figure 4B The calculated IC is shown. 50 This indicates that all compounds have similar efficacy, ranging from 0.602 μM (FTS044) to 1.1 μM (FTS042).

[0829] D. Compound FTS005 inhibits the growth of neuroblastoma (NB) cells in combination therapy and enhances resistance to all-reactive protein (ARP) antibodies. Sensitivity to atRA (atRA).

[0830] When FABP5 is available in cells, it is shown to bind to atRA, transduce it to PPARδ, and activate the nuclear receptor. Activation of PPARδ by atRA diverts the signaling of this vitamin from its corresponding receptor RAR (known for its anticancer activity in various cancers) to the pro-cancer PPARδ. Therefore, inhibition of FABP5 is expected to sensitize cancer cells to atRA by diverting their signaling back to RAR. Therefore, note (NB) that human NPG cells were used to test the effect of combining a FABP5 inhibitor with atRA on cell proliferation. Cells were treated with FTS005 in the presence or absence of atRA.

[0831] like Figure 5A As shown, treatment with FTS005 significantly inhibited cell proliferation, and this trend was enhanced when the compound was combined with atRA. Furthermore, combination therapy with FTS005 and atRA significantly improved cell sensitivity to atRA (see [link to FTS005]). Figure 5B and Figure 5C (Results)

[0832] E. Compound FTS005 in in vivo In a xenograft model, tumor growth was inhibited and macrophages were restricted from entering TNBC tumors. Tumor infiltration

[0833] Using the TNBC xenograft model to test FTS005 in in vivo The effect of inhibiting tumor growth. (5×10) 6 MB-231 cells were subcutaneously injected into the right abdomen of NSG mice. Mice were treated five weeks weekly by gavage with FTS005 (20 mg / kg or 40 mg / kg) or the drug, and tumor growth was monitored. Figure 6A As shown, FTS005 significantly inhibited the growth of MB-231 tumors, as determined by tumor volume and tumor weight. Figure 6A and Figure 6B Molecular analysis of tumors showed that the level of the proliferation marker Ki67 was significantly reduced in tumors of mice treated with FTS005. Figure 6C and Figure 6D Similarly, in treated tumors, the level of the angiogenesis marker VEGFA (also a known target gene of PPARδ) was significantly reduced. Figure 6C , Figure 6E and Figure 6G Surprisingly, staining for the macrophage marker F4 / 80 in tumors showed a significant reduction in the total number of tumor-associated macrophages (TAMs) in treated tumors compared to untreated controls. Figure 6C and Figure 6F This indicates that inhibition of FABP5 affects the immune cell population in the tumor microenvironment. The levels of ACSL1 and PLIN2, genes involved in FA metabolism and lipid accumulation and also known target genes of δPPAR, were measured by qPCR, and their levels were found to be significantly reduced in treated tumors. Figure 6G ).

[0834] F. Compound FTS005 inhibited the growth of TNBC tumors in vivo in a homologous mouse model.

[0835] In the TNBC homology model, FTS005 was tested using 4T1 cells in an immunocompetent model. in vivo Its effect of inhibiting tumor growth. 1x10 5 Four T1 cells were injected into the mammary fat pads of BALB / c mice. Mice were treated with FTS005 (40 mg / kg) or the drug via gavage five times a week, and tumor growth was monitored.

[0836] like Figure 7A As shown, FTS005 significantly inhibited the growth of 4T1 tumors, as determined by tumor volume and tumor weight (see [reference]). Figure 7A and Figure 7B The expression level of the angiogenesis marker VEGFA (also a known target gene of PPARδ) was measured by qPCR, and it was found that the expression level was significantly reduced in treated tumors. Figure 7C Similarly, the genes ACSL1 and PLIN2, which are involved in FA metabolism and lipid accumulation and are also known target genes of PPARδ, were significantly reduced in treated tumors. Figure 7C Immunohistochemical staining of tumors showed that in mouse tumors treated with FTS005, the protein levels of the proliferation markers Ki67 and VEGFA were significantly reduced (see [link to relevant documentation]). Figure 7D , Figure 7E and Figure 7F ).

[0837] G. Fatty acid metabolism in the tumor microenvironment (TME) reprogrammed by compound FTS005

[0838] For 4T1 tumor samples treated or untreated with FTS005, the amounts of different fatty acid-related metabolites were analyzed. More specifically, using LC / MS / MS analysis, metabolites of the TCA cycle, glycolysis, long-chain fatty acids, arachidonic acid oxidation, and ATP / ADP were measured and quantified based on protein concentration in each sample. As shown in Figure 8, the metabolite profiles measured in FTS005-treated tumors changed significantly after treatment. The amount of long-chain fatty acids measured in treated tumors ( Figure 8A The levels of TCA circulating metabolites were significantly lower than in untreated tumors, and similarly, the levels of these metabolites were also lower. Figure 8B Simultaneously, the amount of glycolytic metabolites increased after treatment. Figure 8D This indicates that FTS005 treatment triggered a reprogramming of fatty acid metabolism in tumors, shifting energy utilization from fatty acids to glycolysis. The amount of ADP measured in treated tumors was significantly higher, and although no difference was observed in ATP levels, the significantly higher ADP levels resulted in a lower ATP / ADP ratio. Figure 8C This is a known marker of low proliferation rate and cell death.

[0839] H. Compound FTS005 modulates immune cell populations in the tumor microenvironment (TME).

[0840] To further investigate the effects of FTS005 treatment on immune cells in the tumor mesenteric endometrium (TME), samples from tumors established in immunocompetent BALB / c mice injected with 4T1 cells were used. To assess the effect of FTS005 on tumor-infiltrating macrophages (TAMs), tumors were stained with the TAM markers F4 / 80 and CD68, as well as the specific M2 marker CD163. Figure 9A , Figure 9B and Figure 9C Although FTS005 treatment was not found to modulate total macrophage levels, as indicated by F4 / 80 and CD68 staining ( Figure 9A , Figure 9B , Figure 9D and Figure 9E However, the level of the M2 marker CD163 was significantly reduced in the treated tumors. Figure 9C and Figure 9F Data indicate that the FABP5 inhibitor FTS005 restricts immunosuppressive M2 macrophages in tumors, thereby triggering an autoimmune response against tumor cells.

[0841] To assess the effect of FTS005 on tumor-infiltrating T cells, tumor samples were stained with T cell markers CD3, CD4, and CD8. Notably, the levels of CD3, CD4, and CD8 were significantly higher in treated tumors than in untreated controls. Figure 10A and Figure 10B The number of CD4 and CD8 T cells isolated from spleens collected from mice was also measured by flow cytometry. Although no significant difference was observed in the frequency of CD4 or CD8 cells in spleen cells collected from treated and untreated mice (…),… Figure 10C However, TNFα activation was observed in spleen cells collected from treated mice. + The number of CD8 T cells was significantly higher. Figure 10D The 4T1 cells used in this experiment stably expressed Luc2, and therefore, to stimulate an immune response from the collected spleen cells, the cells were treated with the Luc2 peptide in growth medium for 2 weeks, and the cell count was measured. Figure 10E As shown, the number of T cells collected from the spleen treated with FTS005 was significantly higher than that collected from untreated mice, indicating that the treated T cells had a stronger proliferative capacity when exposed to the Luc2 antigen. To test the cytotoxic activity of T cells, spleen T cells were co-cultured overnight with 4T1 Luc2-CFSE high (target cells, Balb / c source) and F420 Luc2-CFSE low (control, B6 source) cells (target cells:effect cells = 1:5). The next day, viable CFSE+ cells were counted by flow cytometry, and the percentage of lysed cells was calculated. Figure 10F As shown, incubation with T cells collected from treated mice resulted in a significant increase in the number of lysed cells. Therefore, the data indicate that treatment with the FABP3 / 4 / 5 / 7 inhibitor FTS005 can stimulate the formation of memory T cells.

[0842] To assess the effects of FTS005 on various immune cells and tumor immune function, RNA samples extracted from untreated and treated tumors were used, and the nCounter PanCancer Immune Profiling Panel was employed to perform a comprehensive characterization analysis of the immune response. Figure 11As shown, treatment of tumor-bearing mice with FTS005 altered the characteristic profile of immune cells in the tumor. Based on this analysis, higher numbers of CD45 cells, macrophages, B cells, dendritic cells, cytotoxic cells, T cells, CD8 T cells, NK cells, and NK CD56 dim cells were found in the treated tumors, indicating higher immune activity in the TME. Taken together, data collected from mice treated with FTS005 suggest that inhibition of FABP3 / 4 / 5 / 7 modulates immune cells in the TME in two ways: 1) inhibiting immunosuppressive M2 tumor-associated macrophages; and 2) stimulating tumor-infiltrating lymphocytes (TILs) to enter the tumor, including activated CD4 and CD8, NK cells, cytotoxic cells, and dendritic cells, all of which are known to activate immunosuppressive tumors and transform them into “hot” tumors that can be recognized by the immune system.

[0843] F.FTS005 in in vitro Inhibition of lipid uptake by hepatocytes in a model of hepatic steatosis

[0844] In an in vitro model of hepatic steatosis, the effect of FTS005 on lipid uptake by hepatocytes was tested using HepG2 cells. Cells were treated with oleic acid (OA) (1 mM) in the presence or absence of aniline compounds or the known FABP4 inhibitor BMS-309403 (BMS), and lipid accumulation in the cells was quantified using Nile Red. Lipid uptake by hepatocytes treated with FTS005 was significantly inhibited. Figure 12A Compared to BMS, FTS005 was more effective at inhibiting lipid uptake. For comparison, the amount of lipids accumulated in cells treated with 5 μM FTS005 was comparable to that accumulated after treatment with 25 μM BMS. Figure 12A This indicates that thiophene compounds are more efficient than BMS.

[0845] G.FTS005 inhibits lipid uptake by mature adipocytes.

[0846] Mouse 3T3-L1 cells differentiated into mature adipocytes in culture. Starting on day 6 of differentiation, different concentrations of the compound FTS005 or the known FABP4 inhibitor BMS were added to the cells. On day 12, lipid droplets were stained with Nile Red and quantified. Figure 12B Hepatocytes treated with FTS005 showed significantly inhibited lipid uptake. Figure 12A Compared to BMS, FTS005 was more effective at inhibiting lipid uptake. For comparison, the amount of lipids accumulated in cells treated with 10 μM FTS005 was comparable to that accumulated after treatment with 30 μM BMS. Figure 12B This indicates that thiophene compounds are more efficient than BMS.

[0847] Example 34: Biological study on the effects of FABP3 / 4 / 5 / 7 inhibitor compounds on immune cell populations

[0848] This example illustrates the biological effects of the FABP inhibitor compound FTS005 on immune cell populations in a cancer model.

[0849] Materials and methods

[0850] A. Isolation and differentiation of mouse bone marrow-derived macrophages (BMDM)

[0851] BMDM were isolated from C57BL6 mice. Briefly, bone marrow was washed with RPMI medium and cells were lysed using AKL buffer. The collected cells were seeded in growth medium (DMEM, 10% HI FBS, 1% penicillin / streptomycin, 25 ng / mL M-CSF) for 7 days (Mφ macrophages). To promote macrophage differentiation in culture, Mφ cells were treated with LPS (10 pM) and INFg (20 ng / mL) to promote differentiation into M1 macrophages, or with IL-4 (20 ng / mL) and IL-13 (20 ng / mL) to promote differentiation into M2 macrophages. Cells were incubated with cytokines for 3 days in the presence or absence of FABP3 / 4 / 5 / 7 inhibitors, then lysed and immunostained for further analysis.

[0852] B. Fluorescence-activated cell sorting (FACS)

[0853] Macrophages were fixed and then stained. Viable cells were identified using the LIVE / DEAD™ Fixable Aqua Dead Cell Stain Kit according to the manufacturer's protocol. To measure macrophage frequency, cells were stained with markers F4 / 80 (total macrophages), CD11b (total macrophages), MHC-II (M1 macrophages), CD36 (M2 macrophages), and CD206 (M2 macrophages). To assess T cell frequency, the following markers were used: CD4, CD8, CD25, and FoxP3 (Treg), as well as TNFα (activated T cells). Data were analyzed using FlowJo software.

[0854] C. Quantification of IL-10 and IL-12

[0855] Using an ELISA assay kit, the levels of the cytokines interleukin-10 (IL-10) and interleukin-12 p70 (IL-12) secreted by macrophages in the culture medium of treated cells were measured according to the manufacturer's protocol.

[0856] result

[0857] A. Treatment of macrophages with FABP3 / 4 / 5 / 7 inhibitors during differentiation supports the M1 phenotype, while inhibiting the M2 phenotype.

[0858] Bone marrow-derived macrophages (BMDM) were isolated from mice and differentiated in culture. Cells treated with M-CSF for 7 days were designated as naïve macrophages (Mφ). These naïve macrophages were then treated with LPS and IFNg to promote differentiation into M1 macrophages, or with IL-4 and IL-13 to promote differentiation into M2 macrophages. This experiment was performed in the absence or presence of FABP3 / 4 / 5 / 7 inhibitors. The marker MHC-II (Mφ) was used at high frequency. Figure 13A ) and high levels of IL-12 ( Figure 13B To verify M1 polarization in macrophages, the high-frequency marker CD36 was used. Figure 13C ) and CD206 ( Figure 13D ) and high levels of IL-10 ( Figure 13E This study aimed to verify M2 polarization in macrophages. In macrophages differentiated into the M2 state in the presence of either FABP inhibitor FTS005 or the commercially available inhibitor BMS480404, the expression level of CD206 was significantly lower than that in the M2 control group. Figure 13F Similarly, in cells differentiated in the presence of FABP3 / 4 / 5 / 7 inhibitors, the level of IL-10 secreted by M2 macrophages is known to be significantly reduced. Figure 13G The level of IL-12, known to be secreted by M1 macrophages, is significantly elevated. Figure 13H Therefore, the data indicate that FABP3 / 4 / 5 / 7 inhibitors can be used to prevent macrophages from differentiating from their initial state to the M2 state, as evidenced by both cell surface markers and secreted cytokines. To test the effect of FABP3 / 4 / 5 / 7 inhibitors on macrophage differentiation from the M1 to the M2 state, cells were first differentiated into M1 macrophages, and then differentiated into M2 macrophages in the presence of FABP3 / 4 / 5 / 7 inhibitors. The frequency of CD206+ macrophage populations in differentiated cells in the presence of FABP3 / 4 / 5 / 7 inhibitors ( Figure 13I ) and the expression level of CD206 ( Figure 13J ) decreased significantly, while IL-12 levels increased significantly. Figure 13K In summary, the data indicate that FABP3 / 4 / 5 / 7 inhibitors can be used to regulate M1 and M2 macrophage populations.

[0859] While the above disclosure of the present invention has been described in detail by way of example and illustration for clarity and understanding, this disclosure, including the embodiments, descriptions, and implementations herein, is for illustrative purposes and is intended to be exemplary, and should not be construed as limiting the scope of this disclosure. Those skilled in the art will appreciate that various modifications or changes can be made to the embodiments, descriptions, and implementations described herein, and that such modifications or changes are included within the spirit and scope of this disclosure and the appended claims. Furthermore, those skilled in the art will recognize many methods and procedures equivalent to those described and performed herein. All such equivalents should be understood to be within the scope of this disclosure and covered by the appended claims.

[0860] Further embodiments of the invention are set forth in the following claims.

[0861] All publications, patent applications, patents, or other documents mentioned herein are expressly incorporated in their entirety by reference for all purposes, as if each such individual publication, patent, patent application, or other document were individually and specifically cited in its entirety for all purposes and set forth herein. In case of conflict, this specification (including designated terminology) shall prevail.

[0862] References

[0863]

[0864]

[0865]

Claims

1. A compound of structural Formula I, or a pharmaceutically acceptable salt thereof, (I) wherein, R 1 selected from hydrogen, cyano and a 5-membered heteroaryl ring; R 2 and R 3 are each independently selected from hydrogen, C1-C4 linear or branched alkyl, cyclopropyl, cyclobutyl, phenyl, trifluoromethyl and benzyl, or R 2 and R 3 form together a 5- to 8-membered monocyclic, bicyclic or spiro carbocyclic or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein said carbocyclic, heterocyclic, aryl or heteroaryl ring is optionally substituted with one or two substituents selected from C1-C4 alkyl, methoxy or fluoro; X is a moiety of the formula: wherein, Y is a heteroatom selected from -S- and -O- or is -CR 8 R 9 -; wherein R 8 and R 9 are each independently selected from the group consisting of hydrogen, C1-C4 straight chain or branched alkyl, phenyl and benzyl, or R 8 and R 9 together form a cyclopropyl, cyclobutyl, cyclopentyl or oxetanyl ring; R 4 , R 5 , R 6 and R 7 are each independently selected from the group consisting of hydrogen, C1-C4 linear or branched alkyl, phenyl and benzyl, and / or R 4 and R 5 together or R 6 and R 7 form a cyclopropyl ring or a cyclobutyl ring, or R 5 and R 6 together form a 5- to 6-membered carbocyclic or heterocyclic ring having Y as a member of the ring; with the proviso that the compound of structural Formula I does not include the following compounds: 。 2. The compound of claim 1, wherein R 1 is cyano.

3. The compound of claim 1, wherein the compound has the structural Formula la: (Ia).

4. The compound of claim 3, wherein the compound has a structural formula selected from Ij, Ik, Il, Im, In, Io, Ip, Iq, Ir, Is, and It: 。 5. The compound of claim 1, wherein the compound has a structural formula selected from Ib: (Ib) wherein R 10 and R 11 each independently is selected from hydrogen, halogen, C1-C4 linear or branched alkyl, cyclopropyl, and cyclobutyl.

6. The compound of claim 5, wherein the compound has a structural formula selected from Iu, Iv, and Iw: 。 7. The compound of claim 1, wherein R 1 is a 5-membered heteroaryl ring, and R 2 and R 3 are each independently selected from hydrogen, C1-C4 straight or branched chain alkyl, cyclopropyl, cyclobutyl, phenyl, trifluoromethyl, and benzyl.

8. The compound of claim 1, wherein the compound has a structural formula selected from Ic, Id, Ie, If, Ig, Ih, and Ii: wherein R 12 is selected from hydrogen, halogen, C1-C4 linear or branched alkyl, cyclopropyl, cyclobutyl, phenyl.

9. The compound of claim 8, wherein the compound has a structural formula selected from Ix, Iy, Iz, Iaa, Ibb, Icc, Idd, Iee, Iff, Igg, Ihh, III, Ijj, Ikk, Iii, Imm, Inn, Ioo, Ipp, Iqq, and Irr: 。 10. The compound of any one of claims 1-9, wherein Y is selected from -S- or -O-, and R 4 , R 5 , R 6 , and R 7 are each independently hydrogen or C1-C4 straight chain or branched alkyl.

11. The compound of any one of claims 1-9, wherein the X moiety is selected from: 。 12. The compound of any one of claims 1-9, wherein the X moiety is selected from: 。 13. The compound of any one of claims 1-9, wherein the X moiety is selected from: 。 14. A compound of structural Formula II, or a pharmaceutically acceptable salt thereof, (II) wherein, R 1 selected from hydrogen, cyano and a 5-membered heteroaryl ring; R 2 and R 3 are each independently selected from the group consisting of hydrogen, C1-C4 linear or branched alkyl, cyclopropyl, cyclobutyl, phenyl, trifluoromethyl, and benzyl, or R 2 and R 3 form together a 5- to 8-membered monocyclic, bicyclic or spiro carbocyclic or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein said carbocyclic, heterocyclic, aryl or heteroaryl ring is optionally substituted with one or two substituents selected from C1-C4 alkyl, methoxy or fluoro; Y is a heteroatom selected from -S- and -O- or is -CR 8 R 9 -; wherein R 8 and R 9 are each independently selected from hydrogen, C1-C4 straight chain or branched alkyl, phenyl and benzyl, or R 8 and R 9 together form a cyclopropyl, cyclobutyl, cyclopentyl or oxetanyl ring; R 4 , R 5 , R 6 and R 7 are each independently selected from the group consisting of hydrogen, C1-C4straight chain or branched alkyl, phenyl and benzyl, and / or R 4 and R 5 together or R 6 and R 7 together form a cyclopropyl ring or a cyclobutyl ring, or R 5 and R 6 together form a 5- to 6-membered carbocyclic or heterocyclic ring having Y as a member of the ring; with the proviso that the compound of structural Formula II does not include the following compounds: 。 15. The compound of claim 13, wherein R 1 is cyano.

16. The compound of claim 13, wherein the compound has the structural Formula Ila: (IIa) wherein the chemical groups R 2 , R 3 , R 4 , R 5 , R 6 and R 7 are as defined for the compounds of the structural formula II.

17. The compound of claim 16, wherein the compound has a structural formula selected from IIj, IIk, IIl, IIm, IIn, IIo, IIp, IIq, IIr, IIs, and IIt: 。 18. The compound of claim 13, wherein the compound has the structural Formula IIb: (IIb) wherein the chemical groups R 2 , R 3 , R 4 , R 5 , R 6 and R 7 are as defined for the compounds of the structural formula II, and R 10 and R 11 are each independently selected from the group consisting of hydrogen, halogen, C1–C4 linear or branched alkyl, cyclopropyl and cyclobutyl.

19. The compound of claim 18, wherein the compound has a structural formula selected from IIu, IIv, and IIw 。 20. The compound of claim 13, wherein the compound has a structural formula selected from IIc, IId, IIe, IIf, IIg, IIh, and IIi: wherein R 2 , R 3 , R 4 , R 5 , R 6 and R 7 are as defined for the compounds of structural formula II, and R 12 is hydrogen, halogen, C1-C4 straight or branched chain alkyl, cyclopropyl, cyclobutyl or phenyl.

21. The compound of claim 20, wherein the compound has a structural formula selected from IIx, IIy, IIz, IIaa, IIbb, IIcc, IIdd, IIee, IIff, IIgg, IIhh, IIii, IIjj, IIkk, IIll, IImm, IInn, IIoo, IIpp, IIqq, IIrr, and IIss: 。 22. The compound of any one of claims 1-21, wherein the compound is selected from Compounds 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, and 65: 。 23. A pharmaceutical composition comprising the compound of any one of claims 1-22 and one or more auxiliary ingredients.

24. A method for treating a subject having a disease or condition affected by FABP3 / 4 / 5 / 7, comprising administering to a subject in need thereof a therapeutically effective amount of the compound of any one of claims 1-22 or the pharmaceutical composition of claim 23.

25. The method of claim 24, wherein the disease or condition affected by FABP3 / 4 / 5 / 7 is selected from the group consisting of: atherosclerosis, coronary atherosclerosis, arterial fibrosis, pulmonary arterial hypertension, heart failure, obesity, type 2 diabetes, type 1 diabetes, gestational diabetes, polycystic ovary syndrome, endometriosis, conditions affected by lipid metabolism and serum free fatty acid levels, metabolic disorders, fatty liver disease, renal fibrosis, systemic inflammation, acute inflammation, allergic inflammation, airway inflammation, viral infection (e.g., COVID-19, common cold), skin diseases (e.g., vitiligo, psoriasis, atopic dermatitis, allergic contact dermatitis, mycosis fungoides, alopecia areata, cicatricial alopecia, graft versus host disease (GvHD), contact dermatitis, chronic eczema, dermatitis herpetiformis, cutaneous lupus, scleroderma, dermatomyositis, vasculitis, pemphigus, epidermolysis bullosa, linear IgA, blistering diseases), neurological conditions and diseases (e.g., pain, multiple sclerosis (MS), Parkinson’s disease, autoimmune diseases (e.g., experimental autoimmune encephalomyelitis (EAE), asthma, type 1 diabetes, autoimmune lung disease, autoimmune hepatitis, rheumatoid arthritis (RA), spondyloarthropathies, vesicular stomatitis virus infection, multiple sclerosis (MS), lupus nephritis, Crohn’s disease, ulcerative colitis, and food allergies), ischemic stroke, graft versus host disease (GvHD), and cancer (e.g., breast cancer, prostate cancer, ovarian cancer, skin cancer, gastric cancer, glioma, cholangiocarcinoma, bladder cancer, multiple myeloma, colorectal cancer, hepatocellular carcinoma, cervical cancer, oral squamous cell carcinoma, and / or non-small cell lung cancer (NSCLC)).

26. A method for controlling serum free fatty acid levels in a subject in need thereof, comprising administering to a subject in need thereof a therapeutically effective amount of a compound of any one of claims 1-22 or a pharmaceutical composition of claim 23.

27. The method of claim 26, wherein the subject has a disease or condition caused by, affected by, and / or characterized by uncontrolled serum free fatty acid levels in the subject.

28. A method for treating a subject having cancer or diagnosed with cancer, comprising administering to a subject in need thereof a therapeutically effective amount of a compound of any one of claims 1-22 or a pharmaceutical composition of claim 23.

29. The method of claim 28, wherein the cancer is selected from breast cancer, prostate cancer, ovarian cancer, hepatocellular carcinoma, multiple myeloma, neuroblastoma, lung adenocarcinoma, or gastric cancer.

30. The method of any one of claims 28-29, wherein the cancer is characterized by metastasis of TNBC cells.

31. A method of sensitizing cancer cells in a subject having cancer to an additional therapy, comprising administering to a subject in need thereof a therapeutically effective amount of a compound of any one of claims 1-22 or a pharmaceutical composition of claim 23.

32. The method of claim 31, wherein the additional therapy comprises administration of a chemotherapeutic agent; optionally, wherein the chemotherapeutic agent is selected from doxorubicin, gemcitabine, cisplatin, paclitaxel, a PARP inhibitor compound, all-trans retinoic acid (atRA), and an immune checkpoint inhibitor, such as an anti-PD-1 or anti-PD-Ll antibody.

33. A method for treating a subject diagnosed with metabolic syndrome and / or atherosclerosis, comprising administering to a subject in need thereof a therapeutically effective amount of a compound of any one of claims 1-22 or a pharmaceutical composition of claim 23.

34. The method of claim 33, wherein the subject is diagnosed with type 2 diabetes.

35. A method for modulating an immune cell population and / or immune cell activity in a subject in need thereof, comprising administering to a subject in need thereof a therapeutically effective amount of a compound of any one of claims 1-22 or a pharmaceutical composition of claim 23.

36. The method of claim 35, wherein the subject has a disease or disorder caused by, affected by, and / or characterized by an immune cell population and / or immune cell activity.

37. The method of claim 36, wherein the immune cell is an M2 macrophage.

38. The method of claim 36, wherein the disease or disorder is cancer.

39. The method of claim 38, wherein the immune cell is a tumor-associated macrophage (TAM).

40. The method of claim 36, wherein the disease or disorder is an autoimmune disease or disorder.

41. Use of a compound of any one of claims 1-22 or a pharmaceutical composition of claim 23 in the manufacture of a medicament for treating a subject according to any one of claims 24-40.

42. A method for preparing a compound of structural formula II (II) wherein, R 1 selected from hydrogen, cyano and a 5-membered heteroaryl ring; R 2 and R 3 are each independently selected from hydrogen, C1-C4 linear or branched alkyl, cyclopropyl, cyclobutyl, phenyl, trifluoromethyl and benzyl, or R 2 and R 3 form together a 5- to 8-membered monocyclic, bicyclic or spiro carbocyclic or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein said carbocyclic, heterocyclic, aryl or heteroaryl ring is optionally substituted with one or two substituents selected from C1-C4 alkyl, methoxy or fluoro; Y is a heteroatom selected from -S- and -O- or is -CR 8 R 9 –, wherein R 8 and R 9 are each independently selected from the group consisting of hydrogen, C1-C4 straight chain or branched alkyl, phenyl and benzyl, or R 8 and R 9 together form a cyclopropyl, cyclobutyl, cyclopentyl or oxetanyl ring; R 4 , R 5 , R 6 and R 7 are each independently selected from the group consisting of hydrogen, C1-C4straight chain or branched alkyl, phenyl and benzyl, and / or R 4 and R 5 together or R 6 and R 7 form a cyclopropyl ring or a cyclobutyl ring, or R 5 and R 6 together form a 5- to 6-membered carbocyclic or heterocyclic ring with Y as a member of said ring; the method comprising: (a) combining a substituted anhydride compound of formula III: (III) wherein Y, R 4 , R 5 , R 6 and R 7 are as defined above; with a substituted 2-aminothiophene compound of formula IV: (IV) wherein R 1 , R 2 and R 3 are as defined above; and (b) removing the solvent to obtain a compound having the structural formula II.

43. The method of claim 42, wherein the compound is a compound of structural formula (IVa) (IVa) wherein R 2 and R 3 each independently is selected from hydrogen, C1-C4 linear or branched alkyl, cyclopropyl, cyclobutyl, phenyl and benzyl, or R 2 and R 3 together form a 5- to 8-membered monocyclic, bicyclic or spiro carbon or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein the carbon or heterocyclic, aryl or heteroaryl ring is optionally substituted with one or two substituents selected from C1-C4 alkyl, methoxy or fluoro.

44. The method of claim 43, wherein the compound is selected from compounds 4a, 4b, 4c, 4d, 4e, 4f, 4g, 4h, 4i, 4j, and 4k: 。 45. The method of claim 42, wherein the compound is a compound of structural formula IVb: (IVb) wherein R 10 and R 11 each independently is selected from hydrogen, halogen, C1-C4linear or branched alkyl, cyclopropyl, and cyclobutyl.

46. The method of claim 45, wherein the compound is selected from compounds 4l, 4m, and 4n: 。 47. The method of claim 42, wherein the compound is selected from compounds of structural formulae IVc, IVd, IVe, IVf, IVg, and IVh: wherein, R 2 and R 3 each independently is selected from hydrogen, C1-C4 linear or branched alkyl, cyclopropyl, cyclobutyl, phenyl, and benzyl, or R 2 and R 3 together form a 5- to 8-membered monocyclic, bicyclic, or spiro carbon or heterocyclic ring, or a 5- to 8-membered aryl or heteroaryl ring, wherein the carbon or heterocyclic ring, aryl or heteroaryl ring is optionally substituted with one or two substituents selected from C1-C4 alkyl, methoxy, or fluoro; and R 12 the chemical group at the 4-position is selected from the group consisting of hydrogen, halogen, C1-C4 linear or branched alkyl, cyclopropyl, cyclobutyl, and phenyl.

48. The method of claim 47, wherein the compound is selected from compounds 4o, 4p, 4q, 4r, 4s, 4t, 4u, 4v, 4w, 4x, 4y, 4z, 4aa, 4bb, 4cc, 4dd, 4ee, 4ff, 4gg, 4hh, and 4ii: 。

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