Bicyclic pyridine compositions and methods of using same in the treatment of cancer - Patents.com

Abstract

Disclosed herein are bicyclic pyridine compounds, such as thienopyridine, pyrrolopyridine, and furopyridine compounds, and methods for treating cancer. The methods can include administering a therapeutically effective amount of any of the compositions described herein. In some embodiments, the cancer is prostate cancer, leukemia, breast cancer, colon cancer, ovarian cancer, pancreatic cancer, or melanoma.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Application No. 62 / 800,239, filed February 1, 2019, the contents of which are incorporated herein by reference in their entirety. [Background technology]

[0002] Two closely related transcriptional regulatory kinases, CDK8 and CDK19, have become targets for a burgeoning array of novel anticancer drugs (Philip, S. et al., J Med Chem 2018, 61, 5073-5092). In particular, CDK8 / 19 inhibitors have been shown to be effective in castration-resistant prostate cancer (CRPC) (Chen, Roninson, U.S. Patent No. 9,636,342), acute myeloid leukemia (Pelish et al., Nature. 2015 Oct 8;526(7572):273-276), colon cancer liver metastases (Liang et al., Cancer Res. 2018 Dec 1;78(23):6594-6606), estrogen receptor-positive breast cancer when combined with anti-estrogens (McDermott et al., Oncotarget. 2017 Feb 21;8(8):12558-12575), and HER2-positive breast cancer when combined with HER2-targeted agents (McDermott et al., WO 2016 / 018511). CDK8 / 19 inhibitors also prevent the induction of genes that promote metastasis and drug resistance in cancer cells of various tumor types treated with conventional DNA-damaging chemotherapy agents or radiation (Porter, DC, et al., Proc Natl Acad Sci USA 2012, 109, 13799-804). In vivo administration of CDK8 / 19 inhibitors also improved the efficacy of the chemotherapy drug doxorubicin in a lung cancer model (Porter et al., ibid.), demonstrating the utility of CDK8 / 19 inhibitors for the treatment of various cancers when combined with various DNA-damaging agents. Apart from cancer, CDK8 / 19 inhibitors show promise in inflammation-related diseases (US Patent Application Publication No. 2014 / 0309224, Porter, DC; Johnannessen, L., et al., Nat Chem Biol 2017, 13, 1102-1108); cardiovascular diseases (Hall, D., et al., JCI Insight 2017, 2; WO 2016 / 100782, Roninson, IB); ribosomal diseases; conditions characterized by reduced numbers of hematopoietic stem and / or progenitor cells; and bone anabolic disorders (WO 2017 / 076968, Flygare, J.).

[0003] Several CDK8 / 19 inhibitors have been reported (Philip et al., J Med Chem. 2018 Jun 28;61(12):5073-5092. doi: 10.1021 / acs.jmedchem.7b00901). These include certain quinazoline compounds that are highly selective for CDK8 / 19 developed by some of the present inventors, such as SNX2-1-53 (also known as Senexin A) (Porter, DC, et al., Proc Natl Acad Sci USA 2012, 109, 13799-804; U.S. Patent No. 8,598,344, Porter, DC) and SNX2-1-165 (also known as Senexin B) (U.S. Patent No. 9,321,737, Roninson, IB), as well as highly CDK8 / 19-selective quinoline compounds [U.S. Patent Application Nos. 62 / 720,774 and 62 / 720,776]. Other CDK8 / 19 inhibitors have been reported recently (Hatcher, JM et al., ACS Med Chem Lett 2018, 9, 540-545; Nakamura, A. et al., Oncotarget 2018, 9, 13474-13487; Han, X., et al., Bioorg Med Chem Lett 2017, 27, 4488-4492).

[0004] Thienopyridines are a group of compounds with bicyclic aromatic rings. Various thienopyridines are disclosed, for example, in U.S. Pat. No. 6,964,956, U.S. Pat. App. Pub. No. 2007 / 0219234, WO 2017 / 076968, Saito, K. et al., Bioorg Med Chem 2013, 21, 1628-42, and Saito et al., Bioorg Med Chem Lett 2019, 29, 1769-73. U.S. Pat. No. 6,964,956 discloses that some thienopyridines inhibit the IKB kinase (IKK) complex. Saito and U.S. Pat. App. Pub. No. 2007 / 021923 disclosed some thienopyridines with potential bone anabolic activity. Compound 15w was shown to have the highest bone anabolic activity in cell-based assays (Saito, 2013). Kinome profiling also showed that 15w (or DBA-7) and 15k (or DBA-6) are selective inhibitors of CDK8 and CDK19 (WO 2017 / 076968). Despite the high bone anabolic activity of 15w in vitro, 15w has a low C max It had poor pharmacokinetics (PK) (Saito, 2013).

[0005] The clinical efficacy of any CDK8 / 19 inhibitor has yet to be demonstrated, which is determined not only by the compound's ability to inhibit CDK8 / 19, but also by any off-target activity that may be therapeutically beneficial or cause adverse effects, as well as the pharmacokinetics (PK) of the compound. Summary of the Invention

[0006] Disclosed herein are bicyclic pyridine compounds for use in the treatment of cancer. The bicyclic pyridine compounds include compounds of formula 1:

[0007] [ka] (Formula 1) Q may be selected from sulfur, -NH-, or oxygen. X is -(CH2) n -, where n is selected from 0, 1, or 2. 4 R can be hydrogen, or saturated or unsaturated, branched or unbranched, substituted or unsubstituted C1-C6 alkyl. 3 R may be selected from hydrogen, cyano, halo, substituted or unsubstituted amino, substituted or unsubstituted amido; or substituted or unsubstituted sulfonamido. 2 R may be selected from hydrogen, cyano, halo, substituted or unsubstituted amino, substituted or unsubstituted amido; or substituted or unsubstituted sulfonamido. 1 may be selected from hydrogen, cyano, deuterated or non-deuterated hydroxyl, deuterated or non-deuterated carboxy, halo, substituted or unsubstituted deuterated or non-deuterated amino; substituted or unsubstituted deuterated or non-deuterated amide; substituted or unsubstituted deuterated or non-deuterated sulfonamide, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated or non-deuterated C1-C6 alkyl, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated or non-deuterated C1-C6 alkoxyl. In some embodiments, when Q is sulfur, n is 0 and R 4 is hydrogen, and R 3 is -C(O)NH2, R 2 is -NH2, R 1 is selected from deuterated hydroxyl, deuterated carboxy, substituted or unsubstituted deuterated amino; substituted or unsubstituted deuterated amide; substituted or unsubstituted deuterated or non-deuterated sulfonamide, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated C1-C6 alkyl, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated C1-C6 alkoxyl. In some embodiments, when at least one Q is not sulfur, n is not 0 and R 4 is not hydrogen, R 3 is not -C(O)NH2, but R 2 is not -NH2, but R 1is selected from cyano, deuterated or non-deuterated hydroxyl, deuterated or non-deuterated carboxy, halo, substituted or unsubstituted deuterated or non-deuterated amino; substituted or unsubstituted deuterated or non-deuterated amide; substituted or unsubstituted deuterated or non-deuterated sulfonamide, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated or non-deuterated C1-C6 alkyl, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated or non-deuterated C1-C6 alkoxyl. In certain embodiments, the compound has the formula

[0008] [ka] has. In some embodiments, R 1 teeth

[0009] [ka] W is -(CH2) m -or-(CD2) m -, and m may be selected from 0, 1, or 2. 5 and R 6 R may be independently selected from hydrogen, deuterium, deuterated or non-deuterated, saturated or unsaturated, branched or unbranched, substituted or unsubstituted C1-C6 alkyl. 7 and R 8 can be hydrogen, R 7 and R 8 can be deuterium, or R 7 and R 8 and taken together can be oxo. In certain embodiments, when Q is sulfur, n is 0 and R 4 is hydrogen, and R 3 is -C(O)NH2, R 2 is -NH2, R 1 contains at least one deuterium. In some embodiments, R 1 teeth

[0010] [ka] W is -(CH2) m -or-(CD2) m -, and m may be selected from 0, 1, or 2. 9 R may be selected from hydrogen, deuterium, or deuterated or non-deuterated, saturated or unsaturated, branched or unbranched, substituted or unsubstituted C1-C6 alkyl. 7 and R 8 can be hydrogen, R 7 and R 8 can be deuterium, or R 7 and R 8 together can be oxo. In some embodiments, the C4N2 heterocycle is deuterated.

[0011] Another aspect of the present invention is a method of treating a subject with cancer. The method may include administering a therapeutically effective amount of any of the compounds described herein. In some embodiments, the cancer is prostate cancer, leukemia, breast cancer, colon cancer, ovarian cancer, pancreatic cancer, or melanoma. In certain embodiments, the cancer is prostate cancer, such as castration-resistant prostate cancer or prostate cancer resistant to androgen deprivation therapy. In other embodiments, the cancer is leukemia, such as acute myeloid leukemia. In yet other embodiments, the cancer is breast cancer, such as metastatic breast cancer or triple-negative breast cancer.

[0012] Non-limiting embodiments of the present invention are described, by way of example, with reference to the accompanying figures, which are schematic and not intended to be drawn to scale. In the figures, each identical or nearly identical element shown is typically designated by a single numeral. For the sake of clarity, not every element is shown in every figure, and not every element of each embodiment of the present invention is shown, unless explanation is necessary for a person skilled in the art to understand the invention. [Brief description of the drawings]

[0013] [Figure 1A-1B] FIG. 1A shows the effect of various concentrations of 15u (FIG. 1A) and 15w (FIG. 1B) in NFκB reporter assays in parental and CDK8 / 19 double knockout reporter cells. [Figure 1C] FIG. 1 compares IC50 values ​​for various thienopyridines measured in an NFκB reporter assay in the parental 293-derived reporter cell line with cell-based activity values ​​measured by Saito (2013) based on the effect of the same compounds on alkaline phosphatase (ALPase) in the mouse bone marrow stromal cell line ST2. [Figures 2A-2D] Figure 2A shows PK profiles and calculated parameters for 15k (Figure 2A), 15v (Figure 2B), 15u (Figure 2C), and Senexin B (Figure 2D) in female FVB mice, where 0.5 mg / kg of each compound was administered intravenously (iv). [Figure 3A-3E] Figure 3A shows PK curves and calculated parameters for 15k (Figure 3A), 15v (Figure 3B), 15u (Figure 3C), 15w (Figure 3D), and Senexin B (Figure 3E) where 1 mg / kg of each compound was administered orally to mice. [Figure 4A-4B] Figure 4 shows PK curves and calculated parameters for a mixture of 15u (Figure 4A) and 15w (Figure 4B) administered at 30 mg / kg of each compound to female CD1 mice. [Figure 5A] FIG. 1 shows the PK profiles of deuterated 15u_D6 and non-deuterated 15u administered at 30 mg / kg of each compound to female CD-1 mice. [Figure 5B] FIG. 1 shows the PK profiles of deuterated 15w_D2 and 15w_D6 and non-deuterated 15w administered at 16 or 18 mg / kg of each compound to female CD-1 mice. [Figure 6A-6C] FIG. 6 shows the effect of various concentrations of thienopyridine derivatives 15u (FIG. 6A) and 15w (FIG. 6B) and cenexin B (FIG. 6C) on PSA expression in cell culture supernatants of the CRPC cell line C4-2. [Fig. 6D-6F] Figure 6D shows the effect of a mixture of 15u and 15w on PSA serum protein fold changes (Figure 6D) and tumor sample PSA mRNA expression (Figures 6E and 6F) in male NSG mice bearing C4-2 xenografts after 4 days of treatment with 30 mg / kg qd of each compound. [Figure 7A] Figure 1 shows the effect of 15u on xenograft tumor growth of CRPC cell line 22rv1 (P-value style: (*) 0.05-0.01; (**) 0.01-0.001; (***) <0.001). [Figure 7B] FIG. 1 shows tumor mass at the end of the same study. [Figure 7C] FIG. 14. Body weight changes in control and 15u treated mice in the same study. [Figure 8A] FIG. 1 shows immunoblot analysis of CDK8 protein expression in murine 4T1 TNBC cells expressing CDK8 shRNA and their derivatives. [Figure 8B] FIG. 1 shows the mass of primary tumors formed by parental and CDK8 knockdown 4T1 cells. [Figure 8C] FIG. 1 shows mouse survival rates after resection of primary tumors formed by parental and CDK8 knockdown 4T1 cells. [Figure 8D] FIG. 1 shows the primary tumor volume formed by parental 4T1 cells in groups of mice subsequently treated with vehicle or 15u (25 mg / kg, bid). [Figure 8E] FIG. 1 shows the survival rate of mice treated with vehicle or 15u (25 mg / kg, bid) after resection of primary tumors. [Figure 8F] FIG. 1 shows the mass of primary tumors formed by parental 4T1 cells in groups of mice subsequently treated with vehicle or Senexin B (50 mg / kg qd+350 ppm SnxB in food). [Figure 8G]FIG. 1 shows survival rates of mice treated with vehicle or senexin B (50 mg / kg qd+350 ppm SnxB in food) following primary tumor resection. [Figure 9A] Figure 14: Examining the effect of combining either Senexin B (SnxB) or 15u with Enzalutamide (Enza) on the proliferation of MYC-CAP-CR cells in androgen-containing medium. The top panel shows the effect on cell proliferation as a function of Enza concentration. The middle panel shows the effect on cell proliferation as a function of SnxB concentration. The bottom panel shows the effect on cell proliferation as a function of 15u concentration. [Figure 9B] FIG. 1 shows the results of a clonogenesis assay comparing the effects of treatment with DMSO, 1 μM Senexin B (SnxB), 1 μM 15u, 5 μM Enzalutamide (Enza), a combination of 1 μM Senexin B and 5 μM Enzalutamide (Enza), and a combination of 1 μM 15u and 5 μM Enzalutamide (Enza). [Fig. 9C-9D] FIG. 9C compares the volume (FIG. 9D) and mass (FIG. 9D) of MYC-CaP-CR tumors growing subthelially during treatment with vehicle (veh), 15u, enzalutamide (Enza), or a combination of 15u and enzalutamide (Comb) in intact (non-castrated) FVB male mice. [Figure 10A] FIG. 1 shows the effect of various concentrations of 15u and Senexin B on the proliferation of luciferase-expressing MV4-11 cells as detected by bioluminescence imaging. [Figure 10B-10D] Figure 10 compares tumor growth in mice injected with 2x106 luciferase-expressing MV4-11 cells after treatment with vehicle by gavage, 15u of 30mg / kg suspended in vehicle by gavage twice a day, and 1g / kg of drug-added food containing 15u. Figure 10B shows the in vivo bioluminescence image of treated mice. Figure 10C shows a line graph of the bioluminescence signal as total flux in photons / second (p / s). Figure 10D shows the survival curve of treated mice. [Figures 11A-11C]Figure 11 shows the effect of 15u on the in vivo growth of MDA-MB-468 triple negative breast cancer (TNBC) xenografts. Figure 11A is a graph showing the kinetics of tumor volume in control and 15u treated mice. ***: p<0.02. Figure 11B is a bar graph showing the final tumor mass after treatment. Figure 11C is a graph showing the kinetics of mouse body weight. [Figure 12A-12B] Figure 12 shows the maximum tolerated dose (MTD) of 15u in CD-1 mice. Figure 12A shows the body weight kinetics in male and female CD-1 mice treated with 15u in solution formulation at various doses twice daily (bid) by gavage for 2 weeks. Figure 12B shows the body weight kinetics in male and female CD-1 mice treated with 15u in diet with various dose strengths of the drug for 4-5 weeks. [Figure 13] FIG. 1 shows the binding modes of 15u, 15w, 15w_APP, 15w_PP, and 15w_CN superimposed with CDK8. [Figures 14A-14C] Figures confirm the synthesis of 15u_D6 via LCMS. Figure 14A shows the UV chromatograph of 15u_D6. Figure 14B shows the ES+TIC chromatograph of 15u_D6. Figure 14C shows the parent ion of 15u_D6 as further confirmation of the synthesis of the compound. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0014] Disclosed herein are bicyclic pyridines, such as thienopyridines, pyrrolopyridines, furopyridines, and methods for treating cancer.The compositions can selectively inhibit kinases CDK8 and CDK19, and in some cases, RIOK2, CSNK1A1, and CSNK1E.Inhibition of each of these kinases is beneficial in treating conditions such as cancer.

[0015] The examples described below demonstrate that these compounds are suitable for the preparation of pharmaceutical compositions based on pharmacokinetics and for the treatment of subjects suffering from cancer. Intravenous and oral administration of the compounds disclosed herein results in high AUC and very slow clearance, making these compounds suitable for use in the preparation of pharmaceutical compositions and for the treatment of cancer. 15u, deuterated compounds 15u_D6 and 15w_D6, and compound 6304 show surprisingly good PK. The mean serum concentration of 15u at late time points (8 hours) was C max 15u has high AUC and very slow clearance, since the AUC was 64.4% of that of 15u (Example 3). The deuterated analog 15u_D6 also had high AUC, which was comparable to or better than 15u (Examples 4 and 12). 15w_D6 not only had a greater inhibitory potency than its non-deuterated counterpart 15w, but also had a superior AUC (Examples 4 and 12). The compounds disclosed herein also specifically inhibit the kinases CDK8 and CDK19. For example, compounds 15u and 15u_D6 showed high specificity for these kinase targets (Example 3). The compounds disclosed herein exhibit the ability to treat or inhibit the progression of various cancers. For example, the compounds disclosed herein have shown efficacy in vivo against prostate cancer, breast cancer, and leukemia (Examples 5 and 9). Because the compounds disclosed herein possess favorable PK, in vivo activity against several different cancers, along with favorable kinome profiles, these compounds are effective CDK8 / 19 inhibitors for the treatment of cancers associated with CDK8 / 19 activity.

[0016] Bicyclic pyridine compounds Disclosed herein are bicyclic pyridine compounds, such as thienopyridine, pyrrolopyridine, and furopyridine compounds. The compounds are represented by formula 1

[0017] [ka] This includes compounds of the formula: In some embodiments, the compound has the formula

[0018] [ka] It is a compound of the formula:

[0019] Q may be selected from sulfur to give thienopyridine, -NH- to give pyrrolopyridine, or oxygen to give furopyridine, hi some embodiments, Q is sulfur.

[0020] X is -(CH2) n -, n is selected from 0, 1, or 2. Suitably X is a 7-membered ring and R 1 and the aryl substituted with is methylene (i.e., n=1), ethylene (i.e., n=2), or a covalent bond (i.e., n=0). In some embodiments, n is 0.

[0021] R 4 is hydrogen, or saturated or unsaturated, branched or unbranched, substituted or unsubstituted C1-C6 alkyl. 4 can be hydrogen or methyl. R 3 may be selected from hydrogen, cyano, halo, substituted or unsubstituted amino, substituted or unsubstituted amido or substituted or unsubstituted sulfonamido. Suitably R 3 may be selected from substituted or unsubstituted amides such as -C(O)NH2. R 2 may be selected from hydrogen, cyano, halo, substituted or unsubstituted amino, substituted or unsubstituted amido, or substituted or unsubstituted sulfonamido. Suitably, R 2 may be selected from substituted or unsubstituted amino, such as -NH2. R 1may be selected from hydrogen, cyano, deuterated or non-deuterated hydroxyl, deuterated or non-deuterated carboxy, halo, substituted or unsubstituted deuterated or non-deuterated amino; substituted or unsubstituted deuterated or non-deuterated amide; substituted or unsubstituted deuterated or non-deuterated sulfonamide, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated or non-deuterated C1-C6 alkyl, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated or non-deuterated C1-C6 alkoxyl. Exemplary compounds include, without limitation, those disclosed in Table 6.

[0022] In some embodiments, when Q is sulfur, n is 0 and R 4 is hydrogen, and R 3 is -C(O)NH2, R 2 is -NH2, R 1 is selected from deuterated hydroxyl, deuterated carboxy, substituted or unsubstituted deuterated amino; substituted or unsubstituted deuterated amide; substituted or unsubstituted deuterated or non-deuterated sulfonamide, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated C1-C6 alkyl, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated C1-C6 alkoxyl.

[0023] In some embodiments, when at least one of Q is not sulfur, n is not 0 and R 4 is not hydrogen, R 3 is not -C(O)NH2, but R 2 is not -NH2, but R 1is selected from cyano, deuterated or non-deuterated hydroxyl, deuterated or non-deuterated carboxy, halo, substituted or unsubstituted deuterated or non-deuterated amino; substituted or unsubstituted deuterated or non-deuterated amide; substituted or unsubstituted deuterated or non-deuterated sulfonamide, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated or non-deuterated C1-C6 alkyl, saturated or unsaturated, branched or unbranched, substituted or unsubstituted deuterated or non-deuterated C1-C6 alkoxyl. In some embodiments, R 1 teeth

[0024] [ka] W is -(CH2) m -or-(CD2) m -, and m may be selected from 0, 1, or 2. 5 and R 6 R may be independently selected from hydrogen, deuterium, deuterated or non-deuterated, saturated or unsaturated, branched or unbranched, substituted or unsubstituted C1-C6 alkyl. 7 and R 8 can be hydrogen, R 7 and R 8 can be deuterium, or R 7 and R 8 and taken together can be oxo. In certain embodiments, when Q is sulfur, n is 0 and R 4 is hydrogen, and R 3 is -C(O)NH2, R 2 is -NH2, R 1 contains at least one deuterium.

[0025] In some embodiments, W is -(CH2). m -or-(CD2) m -, and m is 0. In some embodiments, W is -(CH2). m-, and m is 1. In other embodiments, W is -(CD2) m -, and m is 1. In some embodiments, R 7 and R 8 and taken together are oxo. In some embodiments, R 7 and R 8 are each hydrogen. In some embodiments, R 5 and R 6 is each methyl. In other embodiments, is each methyl-d3. In some embodiments, R 5 and R 6 One of R is hydrogen and the other is methyl. 5 and R 6 One is deuterium and the other is methyl-d3. In some embodiments, R 5 or R 6 At least one of is a deuterated or non-deuterated, saturated or unsaturated, branched or unbranched, substituted or unsubstituted C1-C6 alkyl. Exemplary substitutions include, without limitation, hydroxyl, amino, or carbamate substitutions.

[0026] Example R 1 Examples of suitable carbamates include, without limitation, N,N-bis(methyl-d3)formamide, N,N-bis(methyl-d3)acetamide, N,N-dimethylacetamide-2,2-d2, N,N-dimethylformamide, N,N-dimethylacetamide, N-methylformamide, N-(3-hydroxypropyl)formamide, N-(3-aminopropyl)-N-methylformamide, or tert-butyl(3-(N-methylformamido)propyl)carbamate. In some embodiments, R 1 teeth

[0027] [ka] W is -(CH2) m -or-(CD2) m -, and m may be selected from 0, 1, or 2. 9 R may be selected from hydrogen, deuterium, or deuterated or non-deuterated, saturated or unsaturated, branched or unbranched, substituted or unsubstituted C1-C6 alkyl. 7 and R 8 can be hydrogen, R 7 and R 8 can be deuterium, or R 7 and R 8 together can be oxo. In some embodiments, the C4N2 heterocycle is deuterated.

[0028] In some embodiments, W is -(CH2). m -or-(CD2) m -, and m is 0. In some embodiments, W is -(CH2). m -, and m is 1. In other embodiments, W is -(CD2) m -, and m is 1. In some embodiments, R 7 and R 8 and taken together are oxo. In another embodiment, R 7 and R 8 are each hydrogen. In some embodiments, R 9 is hydrogen, methyl, or C(O)OC(CH3). Example R 1 include, without limitation, (4-methylpiperazin-1-yl)methylene, 4-methylpiperazine-1-carbaldehyde, piperazine-1-carbaldehyde, or tert-butyl 4-formylpiperazine-1-carboxylate. As shown in the Examples below, some of the bicyclic pyridine compounds have surprisingly good PK. For example, thienopyridines 15u, 15u_D6, 15w_D6, and 604 exhibit PK properties that are suitable for use in the preparation of pharmaceutical compositions and in the treatment of cancer. As shown in the Examples, intravenous and oral administration of these compounds results in high C0 and C1 in an in vivo mouse model. max Concentrations, slower elimination, and larger AUCs are obtained.

[0029] definition As used herein, an asterisk " * " or plus sign " + " may be used to indicate the point of attachment for any radical group or substituent. The term "alkyl" as contemplated by the present invention includes straight or branched alkyl groups in all their isomeric forms, such as those herein defined as C1-C6, respectively. 12 Alkyl, C1-C 10 -alkyl, and linear or branched groups of 1 to 12, 1 to 10, or 1 to 6 carbon atoms, referred to as C1-C6-alkyl.

[0030] The term "alkylene" refers to a diradical of an alkyl group. An exemplary alkylene group is -CHCH-. The term "haloalkyl" refers to an alkyl group substituted with at least one halogen, for example, -CH2F, -CHF2, -CF3, -CH2CF3, -CF2CF3, and the like.

[0031] As used herein, the term "heteroalkyl" refers to an "alkyl" group in which at least one carbon atom has been replaced with a heteroatom (e.g., an O, N, or S atom). Suitably, a heteroalkyl group can be an "alkoxyl" group, an "amino" group, or a "sulfanyl". As used herein, the term "alkenyl" refers to an unsaturated linear or branched hydrocarbon having at least one carbon-carbon double bond, e.g., C2-C6, respectively, as defined herein. 12 -Alkenyl, C2-C 10C2-C6-alkenyl refers to linear or branched groups of 2 to 12, 2 to 10, or 2 to 6 carbon atoms, such as C2-C6-alkenyl, C2-C6-alkenyl, and C2-C6-alkenyl. As used herein, the term "alkynyl" refers to an unsaturated linear or branched hydrocarbon having at least one carbon-carbon triple bond, e.g., C2-C3, respectively, as defined herein. 12 -Alkynyl, C2-C 10 C2-C6-alkynyl refers to linear or branched groups of 2-12, 2-10, or 2-6 carbon atoms, such as C2-C6-alkynyl, C2-C6-alkynyl, and C2-C6-alkynyl.

[0032] The term "cycloalkyl" refers to a group derived from a cycloalkane, e.g., as used herein, "C 4-8 "-cycloalkyl" refers to a monovalent saturated cyclic, bicyclic, or bridged cyclic (e.g., adamantyl) hydrocarbon group of 3 to 12, 3 to 8, 4 to 8, or 4 to 6 carbons. Unless otherwise specified, a cycloalkyl group can be optionally substituted at one or more ring positions with, for example, alkanoyl, alkoxy, alkyl, haloalkyl, alkenyl, alkynyl, amido, amidino, amino, aryl, arylalkyl, azide, carbamate, carbonate, carboxy, cyano, cycloalkyl, ester, ether, formyl, halogen, haloalkyl, heteroaryl, heterocyclyl, hydroxyl, imino, ketone, nitro, phosphate, phosphonato, phosphinato, sulfate, sulfide, sulfonamido, sulfonyl, or thiocarbonyl. In certain embodiments, a cycloalkyl group is unsubstituted, i.e., unsubstituted. The term "cycloalkylene" refers to a diradical of a cycloalkyl group.

[0033] The term "partially unsaturated carbocyclyl" refers to a monovalent cyclic hydrocarbon containing at least one double bond between ring atoms, and at least one ring of the carbocyclyl is not aromatic. Partially unsaturated carbocyclyls can be characterized by the number of ring carbon atoms. For example, partially unsaturated carbocyclyls can contain 5-14, 5-12, 5-8, or 5-6 ring carbon atoms, and thus can be referred to as 5-14, 5-12, 5-8, or 5-6 membered partially unsaturated carbocyclyls, respectively. Partially unsaturated carbocyclyls can be in the form of monocyclic carbocycles, bicyclic carbocycles, tricyclic carbocycles, bridged carbocycles, spirocyclic carbocycles, or other carbocyclic ring systems. Exemplary partially unsaturated carbocyclyl groups include partially unsaturated cycloalkenyl groups and bicyclic carbocyclyl groups. Unless otherwise specified, a partially unsaturated carbocyclyl group may be substituted at one or more ring positions, for example, by alkanoyl, alkoxy, alkyl, haloalkyl, alkenyl, alkynyl, amido, amidino, amino, aryl, arylalkyl, azide, carbamate, carbonate, carboxy, cyano, cycloalkyl, ester, ether, formyl, halogen, haloalkyl, heteroaryl, heterocyclyl, hydroxyl, imino, ketone, nitro, phosphate, phosphonato, phosphinato, sulfate, sulfide, sulfonamide, sulfonyl, or thiocarbonyl. In certain embodiments, the partially unsaturated carbocyclyl is unsubstituted, i.e., unsubstituted.

[0034] The term "aryl" is art-recognized and refers to a carbocyclic aromatic group. Representative aryl groups include phenyl, naphthyl, anthracenyl, and the like. The term "aryl" includes polycyclic ring systems having two or more carbocyclic rings having two or more carbons common to two adjacent rings (the rings are "fused rings"), where at least one of the rings is aromatic, e.g., the other rings may be cycloalkyl, cycloalkenyl, cycloalkynyl, and / or aryl. Unless otherwise specified, the aromatic ring may be substituted at one or more ring positions with, for example, halogen, azide, alkyl, aralkyl, alkenyl, alkynyl, cycloalkyl, hydroxyl, alkoxyl, amino, nitro, sulfhydryl, imino, amide, carboxylic acid, -C(O)alkyl, -CO2alkyl, carbonyl, carboxyl, alkylthio, sulfonyl, sulfonamido, sulfonamide, ketone, aldehyde, ester, heterocyclyl, aryl or heteroaryl moiety, -CF3, -CN, and the like. In certain embodiments, the aromatic ring is substituted at one or more ring positions with halogen, alkyl, hydroxyl, or alkoxyl. In certain other embodiments, the aromatic ring is unsubstituted, i.e., unsubstituted. In certain embodiments, the aryl group is a 6-10 membered ring structure.

[0035] The terms "heterocyclyl" and "heterocyclic group" are art-recognized and refer to saturated, partially unsaturated, or aromatic 3- to 10-membered ring structures, or 3- to 7-membered rings, whose ring structures contain from 1 to 4 heteroatoms, such as nitrogen, oxygen, and sulfur. The number of ring atoms in a heterocyclyl group can be specified using the Cx-Cx rearrangement system, where x is an integer that specifies the number of ring atoms. For example, a C3-C7 heterocyclyl group refers to a saturated or partially unsaturated 3- to 7-membered ring structure that contains from 1 to 4 heteroatoms, such as nitrogen, oxygen, and sulfur. The designation "C3-C7" indicates that the heterocyclic ring contains a total of 3 to 7 ring atoms, including any heteroatoms occupying ring atom positions. The terms "amine" and "amino" are art-recognized and refer to both unsubstituted and substituted amines, where the substituents may include, for example, alkyl, cycloalkyl, heterocyclyl, alkenyl, and aryl. Suitably, the amino may be unsubtitued (i.e., -NH2) or substituted amino of formula -NHR or -NRR', where R and R' are C1-C 12 alkyl or C1-C6 alkyl, such as methyl or ethyl. The terms "alkoxyl" or "alkoxy" are art-recognized and refer to an alkyl group, as defined above, having an oxygen radical attached thereto. Representative alkoxyl groups include methoxy, ethoxy, tert-butoxy, and the like.

[0036] An "ether" is two hydrocarbons covalently linked by an oxygen. Thus, the substituent of an alkyl that makes it an ether is or resembles an alkoxyl and can be represented, for example, by one of -O-alkyl, -O-alkenyl, -O-alkynyl, etc. An "epoxide" is a cyclic ether having a three-atom ring, usually containing two carbon atoms, and approximating an isosceles triangle in shape. Epoxides can be produced by oxidation of a double bound carbon atom, which forms the epoxide with an oxygen atom.

[0037] As used herein, the term "carbonyl" refers to the group -C(O)-. As used herein, the term "carboxamide" refers to the group -C(O)NRR', where R and R' may be the same or different. R and R' may independently be alkyl, aryl, arylalkyl, cycloalkyl, formyl, haloalkyl, heteroaryl, or heterocyclyl. Suitably, a carboxamide may comprise -C(O)NRR', where R and R' are C1-C 12 alkyl or C1-C6 alkyl, such as methyl or ethyl.

[0038] As used herein, the term "carboxy" refers to the group -COOH or its corresponding salts, such as -COONa. As used herein, the term "amide" or "amido" refers to -R 1 C(O)N(R 2 )-, -R 1 C(O)N(R 2 )R 3 -, -C(O)NR 2 R 3 or a group of the form -C(O)NH, where R 1 , R 2 and R 3 are each independently alkoxy, alkyl, alkenyl, alkynyl, amido, amino, aryl, arylalkyl, carbamate, cycloalkyl, ester, ether, formyl, halogen, haloalkyl, heteroaryl, heterocyclyl, hydrogen, hydroxyl, ketone, or nitro. Suitably, the amide is -C(O)NR 2 R 3 where R 2 and R 3 is hydrogen or C1-C 12 alkyl or C1-C6 alkyl. Suitably R 2 and R 3 may be independently selected from hydrogen, methyl, or ethyl. As used herein, the term "sulonamide" refers to -R 1 C(S)2N(R 2 )-, -R 1 C(S)2N(R 2 )R 3 -, -C(S)2NR 2 R 3 or a group of the form -C(S)2NH2, where R 1 , R 2 and R 3are each independently alkoxy, alkyl, alkenyl, alkynyl, amido, amino, aryl, arylalkyl, carbamate, cycloalkyl, ester, ether, formyl, halogen, haloalkyl, heteroaryl, heterocyclyl, hydrogen, hydroxyl, ketone, or nitro. Suitably, the sulfonamide is -C(S)NR 2 R 3 where R 2 and R 3 is hydrogen or C1-C 12 alkyl or C1-C6 alkyl. Suitably R 2 and R 3 may be independently selected from hydrogen, methyl, or ethyl.

[0039] The compounds of the present disclosure may contain one or more chiral centers and / or double bonds and therefore exist as stereoisomers, e.g., geometric isomers, enantiomers or diastereomers. The term "stereoisomer" as used herein consists of any geometric isomer, enantiomer or diastereomer. These compounds may be designated by the symbols "R" or "S" depending on the arrangement of the substituents around the stereogenic carbon atom. The present invention encompasses various stereoisomers of these compounds and mixtures thereof. Stereoisomers include enantiomers and diastereomers. Mixtures of enantiomers or diastereomers may be designated by "(±)" in a postscriptive manner, although the structure may implicitly indicate chiral centers as will be recognized by those skilled in the art. It is understood that a chemical structure, e.g., a schematic representation of a general chemical structure, encompasses all stereoisomeric forms of the defined compound unless otherwise indicated. Compositions comprising substantially purified stereoisomers, epimers, or enantiomers, or analogs or derivatives thereof (e.g., compositions comprising at least about 90%, 95%, or 99% pure stereoisomers, epimers, or enantiomers) are contemplated by the present invention.

[0040] Pharmaceutical Compositions The compounds utilized in the methods disclosed herein may be formulated as pharmaceutical compositions comprising (a) a therapeutically effective amount of one or more compounds disclosed herein, and (b) one or more pharma- ceutically acceptable carriers, excipients, or diluents. The pharmaceutical compositions may comprise a range of about 0.1-2000 mg (preferably about 0.5-500 mg, more preferably about 1-100 mg) of the compound. The pharmaceutical compositions may be administered to provide a daily dose of the compound of about 0.1-100 mg / kg body weight (preferably about 0.5-20 mg / kg body weight, more preferably about 0.1-10 mg / kg body weight). In some embodiments, after the pharmaceutical composition is administered to the patient (e.g., about 1, 2, 3, 4, 5, or 6 hours after administration), the concentration of the compound at the site of action is about 1 nM-100 μM.

[0041] The compound utilized in the methods disclosed herein can be formulated as a pharmaceutical composition in solid dosage form, but any pharma- ceutically acceptable dosage form can be utilized.Exemplary solid dosage forms include, but are not limited to, tablets, capsules, sachets, lozenges, powders, pills, or granules, and the solid dosage form can be, for example, a fast-dissolving dosage form, a controlled release dosage form, a lyophilized dosage form, a delayed release dosage form, a sustained release dosage form, a pulsed release dosage form, a mixed immediate release and controlled release dosage form, or a combination thereof.

[0042] The compounds utilized in the methods disclosed herein may be formulated as pharmaceutical compositions containing a carrier, for example, the carrier may be selected from the group consisting of proteins, carbohydrates, sugars, talc, magnesium stearate, cellulose, calcium carbonate, and starch-gelatin paste.

[0043] The compounds utilized in the methods disclosed herein may be formulated as pharmaceutical compositions that include one or more binders, fillers, lubricants, suspending agents, sweeteners, flavoring agents, preservatives, buffers, wetting agents, disintegrants, and effervescent agents. Fillers include lactose monohydrate, anhydrous lactose, and various starches. Examples of binders are various celluloses and cross-linked polyvinylpyrrolidone, microcrystalline cellulose, such as Avicel® PH101 and Avicel® PH102, microcrystalline cellulose, and silicified microcrystalline cellulose (ProSolv SMCC™). Suitable lubricants, including agents that affect the flowability of the powder to be compressed, include colloidal silicon dioxide, such as Aerosil® 200, talc, stearic acid, magnesium stearate, calcium stearate, and silica gel. Examples of sweeteners may include any natural or artificial sweetener, such as sucrose, xylitol, sodium saccharin, cyclamate, aspartame, and acesulfame. Examples of flavoring agents are Magnasweet® (a trademark of MAFCO), bubble gum flavor, and fruit flavors. Examples of preservatives may include potassium sorbate, methylparaben, propylparaben, benzoic acid and its salts, other esters of parahydroxybenzoic acid, such as butylparaben, alcohols, such as ethyl or benzyl alcohol, phenolic compounds, such as phenol, or quaternary compounds, such as benzalkonium chloride. Suitable diluents may include pharma- ceutically acceptable inert fillers, such as microcrystalline cellulose, lactose, dibasic calcium phosphate, saccharides, and any mixtures of the above. Examples of diluents include microcrystalline cellulose, such as Avicel® PH101 and Avicel® PH102, lactose, such as lactose monohydrate, anhydrous lactose, and Pharmatose® DCL21, dibasic calcium phosphate, such as Emcompress®, mannitol, starch, sorbitol, sucrose, and glucose.

[0044] Suitable disintegrants include lightly cross-linked polyvinylpyrrolidone, corn starch, potato starch, maize starch, and modified starches, croscarmellose sodium, crospovidone, sodium starch glycolate, and mixtures thereof. Examples of effervescent agents are effervescent couples, such as organic acids and carbonates or bicarbonates.Suitable organic acids include, for example, citric acid, tartaric acid, malic acid, fumaric acid, adipic acid, succinic acid, and alginic acid and anhydrides and acid salts.Suitable carbonates and bicarbonates include, for example, sodium carbonate, sodium bicarbonate, potassium carbonate, potassium bicarbonate, magnesium carbonate, sodium glycine carbonate, L-lysine carbonate, and arginine carbonate.Alternatively, only the sodium bicarbonate component of the effervescent couple may be present.

[0045] The compound utilized in the method disclosed herein can be formulated as a pharmaceutical composition for delivery by any suitable route.For example, pharmaceutical composition can be administered via oral, intravenous, intramuscular, subcutaneous, topical, and pulmonary routes.Examples of pharmaceutical composition for oral administration include capsules, syrups, concentrates, powders, and granules. The compounds utilized in the methods disclosed herein may be administered in conventional dosage forms prepared by combining the active ingredients with standard pharmaceutical carriers or diluents following conventional procedures well known in the art, which may involve mixing, granulating and compressing, or dissolving the ingredients as appropriate to the desired preparation.

[0046] Pharmaceutical compositions containing the compounds may be adapted for administration by any suitable route, for example, oral (including buccal or sublingual), rectal, nasal, topical (including buccal, sublingual or transdermal), vaginal or parenteral (including subcutaneous, intramuscular, intravenous or intradermal) routes. Such formulations may be prepared by any method known in the art of pharmacy, for example by combining the active ingredient with the carriers or excipients. Pharmaceutical compositions adapted for oral administration may be presented as discrete units such as capsules or tablets, powders or granules, solutions or suspensions in aqueous or non-aqueous liquids, edible foams or whips, or oil-in-water or water-in-oil liquid emulsions.

[0047] Pharmaceutical compositions adapted for transdermal administration may be presented as discrete patches intended to remain in intimate contact with the epidermis of the recipient for a prolonged period of time, for example, the active ingredient may be delivered from the patch by iontophoresis. Pharmaceutical compositions adapted for topical administration may be formulated as ointments, creams, suspensions, lotions, powders, solutions, pastes, gels, impregnated dressings, sprays, aerosols or oils and may contain suitable conventional additives such as preservatives, solvents to aid penetration of the drug and, in ointments and creams, emollients.

[0048] For application to the eye or other external tissues, such as mouth and skin, the pharmaceutical composition is preferably applied as a topical ointment or cream.When formulated into ointment, the compound can be used with a paraffinic or water-miscible ointment base.Alternatively, the compound can be formulated into cream with an oil-in-water oil cream base or a water-in-oil base.The pharmaceutical composition adapted for topical administration to the eye includes eye drops, in which active ingredient is dissolved or suspended in a suitable carrier, particularly an aqueous solvent. Pharmaceutical compositions adapted for topical administration in the mouth include lozenges, pastilles and mouthwashes. Pharmaceutical compositions adapted for rectal administration may be presented as suppositories or enemas. Pharmaceutical compositions adapted for nasal administration wherein the carrier is a solid include coarse powders having a particle size (for example in the range 20 to 500 μm) which are administered in the way that a snuff is taken (i.e. by rapid inhalation through the nasal passage from a container of the powder held close to the nose). Suitable formulations wherein the carrier is a liquid for administration as a nasal spray or nasal drops include aqueous or oily solutions of the active ingredient.

[0049] Pharmaceutical compositions adapted for administration by inhalation include fine particle dusts or mists, which may be generated by means of various types of metered dose pressurized aerosols, nebulizers or inhalers. Pharmaceutical compositions adapted for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or spray formulations. Pharmaceutical compositions adapted for parenteral administration include aqueous and non-aqueous sterile injection solutions, which may contain antioxidants, buffers, bacteriostats and solutes that render the formulation isotonic with the blood of the intended recipient, and aqueous and non-aqueous sterile suspensions, which may contain suspending agents and thickening agents.The formulations may be presented in unit-dose or multi-dose containers, such as sealed ampoules and vials, and may be stored in a freeze-dried (lyophilized) condition, requiring only the addition of a sterile liquid carrier, such as water for injection, immediately prior to use.Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets.

[0050] Tablets and capsules for oral administration may be in unit dose presentation form and may contain conventional excipients such as binders, e.g., syrup, acacia, gelatin, sorbitol, tragacanth, or polyvinylpyrrolidone; fillers, e.g., lactose, sugar, corn starch, calcium phosphate, sorbitol, or glycine; tableting lubricants, e.g., magnesium stearate, talc, polyethylene glycol, or silica; disintegrants, e.g., potato starch; or acceptable wetting agents, e.g., sodium lauryl sulfate. Tablets may be coated according to methods well known in normal pharmaceutical practice. Oral liquid preparations may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups, or elixirs, or may be presented as a dry product for reconstitution with water or other suitable vehicle before use. Such liquid preparations may contain conventional additives, such as suspending agents, for example, sorbitol, methylcellulose, glucose syrup, gelatin, hydroxyethylcellulose, carboxymethylcellulose, aluminum stearate gel or hydrogenated edible fats, emulsifying agents, for example, lecithin, sorbitan monooleate, or acacia; non-aqueous vehicles (which may include edible oils), for example, almond oil, oily esters, for example, glycerin, propylene glycol, or ethyl alcohol; preservatives, for example, p-hydroxybenzoate or methyl or propyl sorbate, and, if desired, conventional flavors or colorants.

[0051] The compounds used in the compositions and methods disclosed herein can be administered as pharmaceutical compositions, and therefore pharmaceutical compositions incorporating the compounds are considered to be embodiments of the compositions disclosed herein. Such compositions can take any pharmaceutically acceptable physical form; illustratively, they can be orally administered pharmaceutical compositions. Such pharmaceutical compositions contain an effective amount of the disclosed compounds, which is related to the daily dose of the compound administered. Each dosage unit can contain a daily dose of a given compound, or each dosage unit can contain a portion of the daily dose, such as a half or a third of the dose. The amount of each compound contained in each dosage unit can depend, in part, on the identity of the particular compound selected for treatment and other factors, such as the condition for which it is administered. The pharmaceutical compositions disclosed herein can be formulated to provide a quick, sustained, or delayed release of the active ingredient to the patient after administration by using well-known techniques.

[0052] The compounds used according to the methods disclosed herein can be administered as a single compound or a combination of compounds.For example, a compound that treats cancer activity can be administered as a single compound or in combination with another compound that treats cancer or has a different pharmacological activity. As indicated above, pharma- ceutically acceptable salts of compounds are contemplated and may also be utilized in the disclosed methods. The term "pharma- ceutically acceptable salts" as used herein refers to salts of compounds that are substantially non-toxic to living organisms. Exemplary pharma- ceutical acceptable salts include salts prepared by reacting the compounds disclosed herein with pharma- ceutical acceptable mineral or organic acids or organic or inorganic bases. Such salts are known as acid addition and base addition salts. As the skilled reader will appreciate, most or all of the compounds disclosed herein can form salts, and drug salts are commonly used because they are often easier to crystallize and purify than free acids or bases.

[0053] Acids commonly used to form acid addition salts may include inorganic acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, phosphoric acid, and the like, and organic acids such as p-toluenesulfonic acid, methanesulfonic acid, oxalic acid, p-bromophenylsulfonic acid, carbonic acid, succinic acid, citric acid, benzoic acid, acetic acid, and the like. Examples of suitable pharma- ceutically acceptable salts include sulfate, pyrosulfate, bisulfate, sulfite, bisulfate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate, bromide, iodide, acetate, propionate, decanoate, caprylate, acrylate, formate, hydrochloride, dihydrochloride, isobutyrate, caproate, heptanoate, propiolate, oxalate, malonate, succinate, suberate, sebacate, fumarate, malate, and the like. at-), butyne-1,4-dioate, hexyne-1,6-dioate, benzoate, chlorobenzoate, methylbenzoate, hydroxybenzoate, methoxybenzoate, phthalate, xylenesulfonate, phenylpropionate, phenylbutyrate, lactate, alpha-hydroxybutyrate, glycolate, tartrate, methanesulfonate, propanesulfonate, naphthalene-1-sulfonate, naphthalene-2-sulfonate, mandelate, and the like. Base addition salts include those derived from inorganic bases, such as ammonium or alkali or alkaline earth metal hydroxides, carbonates, bicarbonates, etc. Bases useful in preparing such salts include sodium hydroxide, potassium hydroxide, ammonium hydroxide, potassium carbonate, sodium bicarbonate, potassium bicarbonate, calcium hydroxide, calcium carbonate, and the like.

[0054] The particular counterion forming part of any salt of a compound disclosed herein may not be critical to the activity of the compound, so long as the salt as a whole is pharmacologically acceptable and so long as the counterion does not contribute undesirable qualities to the salt as a whole, which may include undesirable solubility or toxicity. Pharmaceutically acceptable esters and amides of the compounds can also be used in the compositions and methods disclosed herein. Examples of suitable esters include alkyl, aryl, and aralkyl esters, such as methyl esters, ethyl esters, propyl esters, dodecyl esters, benzyl esters, and the like. Examples of suitable amides include unsubstituted amides, monosubstituted amides, and disubstituted amides, such as methyl amides, dimethyl amides, methylethyl amides, and the like. Additionally, the methods disclosed herein may be practiced with solvated forms of the compounds or their salts, esters, and / or amides. Solvated forms may include ethanol solvates, hydrates, and the like.

[0055] Treatment The compositions described herein are useful for treating subjects.As used herein, the term "treat" or "treating" refers to temporarily or permanently relieving symptoms, eliminating the cause of resulting symptoms, and / or preventing or slowing the appearance of, or reversing the progression or severity of, the resulting symptoms of the disease or disorder named.Therefore, the methods disclosed herein encompass both therapeutic and prophylactic administration.

[0056] As used herein, "subject" may be synonymous with "patient" or "individual" and refers to an animal, which may be a human or non-human animal, that requires treatment. "Subject in need of treatment" may include a subject with a disease, disorder, or condition that responds to treatment with the pyridine compounds disclosed herein. For example, "subject in need of treatment" may include a subject with a CDK8 / 19-related disease, disorder, or condition, such as cancer, inflammation-related disease, cardiovascular disease, ribosomal disease, a condition characterized by a reduced number of hematopoietic stem and / or progenitor cells, and bone anabolic disorder. CDK8 / 19-related disease, disorder, or condition includes any disease, disorder, or condition that the subject can be treated by inhibiting CDK8 or CDK19.

[0057] The term "effective amount" as used herein refers to an amount or dose of a compound that, upon administration to a subject in single or multiple doses, provides a desired effect to a subject under diagnosis or treatment. The disclosed methods can include administering an amount of the disclosed compounds (e.g., present in a pharmaceutical composition) effective to treat a disease associated with CDK8 / 19.

[0058] Effective amounts can be easily determined by the attending diagnostician, as one skilled in the art, by using known techniques and observing results obtained under similar circumstances. In determining the effective amount or dosage of the compound to be administered, many factors can be taken into consideration by the attending diagnostician, such as: the race of the subject; its size, age, and general health; the degree or severity of the involvement of the disease or disorder involved; the response of the individual subject; the individual compound to be administered; the method of administration; the bioavailability characteristics of the administered formulation; the selected dosage regimen; the use of concomitant drugs; and other relevant circumstances. A typical daily dose can contain from about 0.01 mg / kg to about 100 mg / kg (eg, from about 0.05 mg / kg to about 50 mg / kg and / or from about 0.1 mg / kg to about 25 mg / kg) of each compound used in the present methods of treatment. The compositions can be formulated in unit dosage forms, with each dosage containing about 1 to about 500 mg of each compound individually or in a single unit dosage form, e.g., about 5 to about 300 mg, about 10 to about 100 mg, and / or about 25 mg. The term "unit dosage form" refers to physically discrete units suitable as single dosages for a patient, each unit containing a predetermined amount of active material calculated to produce a desired therapeutic effect, together with a suitable pharmaceutical carrier, diluent, or excipient.

[0059] In some embodiments, the disease related to CDK8 / 19 is prostate cancer, suitably castration-resistant prostate cancer or prostate cancer that is resistant to androgen deprivation therapy.As used herein, " castration-resistant prostate cancer " or " castration-resistant prostate cancer " or " CRPC " is prostate cancer that continues to grow even when the amount of testosterone in the body is reduced to very low levels.Many early stage prostate cancers require substantially normal levels of testosterone to grow, but CRPC does not.

[0060] Androgen deprivation therapy (or androgen suppression therapy) is a prostate cancer hormone therapy. Androgen deprivation therapy may include treatments to reduce androgen levels, such as surgical or chemical castration, or treatments to inhibit the activity of androgen's cancer-promoting activity. Reducing androgen levels or inhibiting androgen activity may result in slowing the growth of prostate tumors and in some cases tumor shrinkage. Suitable treatments to inhibit the activity of androgens that promote cancer include administration of antiandrogens that can bind to the androgen receptor. Antiandrogens include, without limitation, cyproterone acetate, megestrol acetate, chlormadinone acetate, spironolactone, oxendolone, flutamide, bicalutamide, nilutamide, topilutamide, enzalutamide, abiraterone, or apalutamide. The method of the present disclosure can be useful for treating subjects who do not respond to androgen deprivation therapy.Some prostate cancers, such as CRPC, can be unresponsive or resistant to androgen deprivation therapy.As shown in the examples, 15u is effective in suppressing the growth of prostate tumors in CRPC.As a result, 15u can be administered to subjects who have previously undergone androgen deprivation therapy or subjects who do not respond to androgen deprivation therapy. The method of the present disclosure can also be useful for treating subjects who are currently undergoing androgen deprivation therapy.As shown in the examples, 15u is effective in suppressing the growth of prostate tumors in CRPC when administered together with antiandrogens.As a result, 15u can be administered to subjects who are currently undergoing androgen deprivation therapy.

[0061] In some embodiments, the CDK8 / 19 associated disease is leukemia, suitably acute myeloid leukemia. In some embodiments, the CDK8 / 19 associated disease is breast cancer, suitably metastatic breast cancer.

[0062] Methods for inhibiting CDK8 or CDK19 The compositions described are useful for inhibiting CDK8 and / or CDK19. As used herein, "inhibiting CDK8" or "inhibiting CDK19" means inhibiting the activity of CDK8 or CDK18, respectively, by any suitable mechanism, such as competitive binding. Methods for inhibiting CDK8 and / or CDK19 may include contacting any of the compounds or compositions described herein with CDK8 or CDK19. The degree of inhibition may be measured by the assays taught in the Examples herein, such as the assay conditions used by the service provider employed in the present invention. The results of these assays are generally expressed herein as percent of control (POC) using a control in which no compound is present. Alternatively, the results may be expressed as IC50. In some embodiments, the POC is less than 35%, suitably less than 30%, 25%, 20%, 15%, 10%, 5%, or 1% for an effective dose of any of the compounds of the compositions described herein. In some embodiments, the IC50 is less than 2000nM, 1500nM, 1000nM, 750nM, 500nM, 250nM, 200nM, 150nM, 100nM, 75nM, 50nM, 40nM, 30nM, or 25nM.

[0063] In some embodiments, the compounds and compositions disclosed herein specifically inhibit CDK8 or CDK19.As used herein, a compound or composition that "specifically inhibits CDK8" or "specifically inhibits CDK19" is a compound or composition that inhibits one or more CDK8 or CDK19 to a greater extent than it inhibits certain other CDKs.In some embodiments, such compounds also inhibit CDK8 and / or CDK19 to a greater extent than CDK2, CDK3, CDK4, CDK5, CDK7, CDK9, CDK11A, CDK11B, CDK13, CDK14, CDK15, CDK16, CDK17, CDK18, CDKL1, CDKL3, or CDKL5. In preferred embodiments, such greater degree is at least two-fold greater, or at least three-fold greater, than CDK2, CDK3, CDK4, CDK5, CDK7, CDK9, CDK11A, CDK11B, CDK13, CDK14, CDK15, CDK16, CDK17, CDK18, CDKL1, CDKL3, or CDKL5.

[0064] Miscellaneous Unless otherwise specified or indicated by context, the terms "a," "an," and "the" mean "one or more." For example, "a molecule" should be interpreted as meaning "one or more molecules." As used herein, the terms "about," "approximately," "substantially," and "substantially" will be understood by those of skill in the art and will vary to some extent depending on the context in which they are used. If any term is used which is not clear to persons of skill in the art given the context in which it is used, "about" and "approximately" will mean plus or minus ≦10% of the particular term, and "substantially" and "substantially" will mean plus or minus >10% of the particular term.

[0065] As used herein, the terms "include" and "including" have the same meaning as the terms "comprise" and "comprising." The terms "comprise" and "comprising" should be interpreted as "open" transition terms that allow for the inclusion of additional elements in addition to those recited in the claims. The terms "consist" and "consisting of" should be interpreted as "closed" transition terms that do not allow for the inclusion of additional elements other than those recited in the claims. The term "consisting essentially of" should be interpreted as partially closed, allowing only the inclusion of additional elements that do not fundamentally change the essence of the claimed subject matter.

[0066] All methods described herein can be performed in any suitable order unless otherwise indicated herein or clearly contradicted by context. The use of any and all illustrative or exemplary language (e.g., "such as") recited herein is intended merely to facilitate easier understanding of the invention and does not impose limitations on the scope of the invention unless otherwise recited in the claims. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention. All references cited in this specification, including publications, patent applications, and patents, are incorporated herein by reference to the same extent as if each reference was individually and specifically indicated to be incorporated by reference in its entirety.

[0067] Preferred embodiments of the present invention are described herein, including the best mode known to the inventors for carrying out the invention. Variations of those preferred embodiments will be apparent to those of skill in the art upon reading the foregoing description. The inventors anticipate that such variations will be employed by those of skill in the art as appropriate, and the inventors intend that the invention be practiced other than as specifically described herein. Accordingly, the present invention includes all modifications and equivalents permitted by applicable law as set forth in the appended claims. Also, any combination of the above-described elements in all possible variations thereof is encompassed by the present invention unless otherwise indicated herein or clearly contradicted by context. EXAMPLES

[0068] Example 1 Thienopyridine derivatives inhibit CDK8 / 19 activity in cell-based assays. NFκB activity assay. A cell-based assay was used to measure the inhibition of CDK8 / 19 activity by thienopyridine derivatives. Based on the role of CDK8 / 19 in NFκB-driven transcription (Li et al., Characterizing CDK8 / 19 Inhibitors through a NFκB-Dependent Cell-Based Assay, Cells 2019, 8(10), 1208), the assay measures the effect of CDK8 / 19 on the expression of a firefly luciferase reporter from an NFκB-dependent promoter in 293 cells. The lentiviral vector pHAGE-NFKB-TA-LUC-UBC-dTomato-W (Addgene #49335) was transfected into 293 cells, and the clonal cell line showing the strongest induction of luciferase expression upon TNFα treatment was established and used as the reporter cell line. As a control for the CDK8 / 19 dependence of NFκB inhibition, the same reporter construct was also introduced into 293 cells harboring CRISPR / CAS9 knockouts of both CDK8 and CDK19.

[0069] NFκB activity results. Figures 1A and 1B show the effect of various concentrations of 15u and 15w on NFκB reporter activity in parental 293 and CDK8 / 19-deficient (double knockout) reporter cells. These compounds inhibited reporter induction with IC50 values ​​of 10 and 4 nM, respectively, but had no effect on NFκB activation in CDK8 / 19-deficient cells, indicating that the inhibitory effect of both compounds was dependent on the presence of CDK8 / 19 and not on other determinants of NFκB activity such as IKK. Figure 1C and Table 1 compare IC50 values ​​for various thienopyridines measured in an NFκB reporter assay in the parental 293-derived reporter cell line with cell-based activity values ​​measured for the same compounds by Saito (2013) based on their effect on alkaline phosphatase (ALPase), an indicator of osteoblastic differentiation in the mouse bone marrow stromal cell line ST2. The latter effect is measured using the EC50, the concentration that increases ALPase activity to 200% of control. 200 The IC50 values ​​in the CDK8 / 19 NFkB assay are expressed as ALPase EC 200 values ​​(Fig. 1B), indicating that the ALPase effect is most likely mediated via CDK8 / 19 inhibition.

[0070] [Table 1]

[0071] Example 2 Kinome profiling of thienopyridine derivatives. Table 2 shows the kinome profile of 15u_D6 and 15u measured by KINOMEscan™ site-specific competitive binding assay at 2000 nM concentration. Compounds that bind to the kinase active site and directly (sterically) or indirectly (allosterically) prevent kinase binding to the immobilized ligand reduce the amount of kinase captured on the solid support. Conversely, test molecules that do not bind to the kinase have no effect on the amount of kinase captured on the solid support. Screening "hits" are confirmed by measuring the amount of kinase captured in test versus control samples by using a quantitative, precise, ultrasensitive qPCR method that detects bound DNA label. Similarly, the dissociation constant (Kd) for the test compound-kinase interaction is calculated by measuring the amount of kinase captured on the solid support as a function of test compound concentration. A detailed description of the assay technique can be found in Fabian, MA et al. A small molecule-kinase interaction map for clinical kinase inhibitors. Nat. Biotechnol. 23, 329-336 (2005).

[0072] Percent Control (%Ctrl). Compounds were screened at 10 nM concentration and results for primary screen binding interactions are reported as "%Ctrl" or "POC", where lower numbers indicate stronger hits in the matrix. %Ctrl is defined as (eqn 1). %Ctrl=100×(TS-CPOS) / (CNEG-CPOS)(eqn 1) Here, TS is the signal of the test compound, CPOS is the positive control signal (0% Ctrl), and CNEG is the DMSO negative control signal (100% Ctrl).

[0073] Results. Table 2 compares the kinome profiling results between 15u and 15u_D6. Both 15u and 15u_D6 are highly selective for CDK8 and CDK19. Although 15u_D6 showed somewhat greater inhibition of most of the off-target kinases, the effect of 15u_D6 on CDK8 and CDK19 was much greater than that of 15u. The %Ctrl of 15u on CDK8 and CDK19 is 2.6 and 13, respectively. The %Ctrl of 15u_D6 on CDK8 and CDK19 is 0.25 and 0, respectively. Hence, the structural differences between 15u and 15u_D6 result in large differences in target selectivity.

[0074] [Table 2] TIFF0007628252000011.tif218145 TIFF0007628252000012.tif218143 TIFF0007628252000013.tif213143 TIFF0007628252000014.tif208142 TIFF0007628252000015.tif213145 TIFF0007628252000016.tif192137 TIFF0007628252000017.tif177134

[0075] The effect on all kinases that showed >65% inhibition by 2,000 nM of 15u in this screen (CDK8, CDK19, RIOK2, CSNK1A1, CSNK1E, SCNK1D, HASPIN, GSK3A) was then further investigated by measuring the Kd values ​​of 15u in the DiscoverX assay. Kd assays were performed in duplicate and the results are shown in Table 3. The table also shows the results of the Kd determination for 15w versus CDK8, CDK19 and RIOK2.

[0076] [Table 3]

[0077] In particular, CDK8 and CDK19 K for 15u and 15w d The IC values ​​are roughly an order of magnitude higher than their IC values ​​for CDK8 / 19 inhibition in cell-based assays (Figures 1A and 1B), indicating that competition for ATP analog binding does not adequately reflect the inhibitory activity of these compounds. d The other kinase that is predominantly inhibited by 15u at this value is RIOK2 (also strongly inhibited by 15w). CSNK1A1 and CSNK1E were not tested against 15w.

[0078] Remarkably, the reported evidence suggests that inhibition of these three kinases may be beneficial rather than detrimental in the treatment of cancer. Thus, RIOK2, a prototypical kinase regulating ribosome biogenesis, was identified as a target for compounds that selectively inhibited the proliferation of prostate cancer cell lines carrying oncogenic gene fusions that activate the ERG gene in many prostate cancers. The same RIOK2-binding compounds had minimal effects on normal prostate or endothelial cells or ERG-negative tumor cell lines (Mohamed, AA et al., Cancer Res. 2018 Jul 1;78(13):3659-3671. doi: 10.1158 / 0008-5472.CAN-17-2949). CSNK1A1 has been implicated as an oncogenic factor in various leukemias and solid tumors (Mannis, S. et al. J Hematol Oncol. 2017 Oct 2;10(1):157. doi: 10.1186 / s13045-017-0529-5; Richter, J. et al., BMC Cancer. 2018 Feb 6;18(1):140. doi: 10.1186 / s12885-018-4019-0), and CSNK1A1 inhibitors synergized with lysosomotropic agents to inhibit proliferation and promote tumor cell death in KRAS-driven cancers (Cheong, JK et al., J Clin Invest. 2015 Apr;125(4):1401-18. doi: 10.1172 / JCI78018).CSNK1E inhibition has been reported to have selective antiproliferative activity in several types of tumor cells (Yang, WS, et al., Genome Biol. 2008;9(6):R92. doi: 10.1186 / gb-2008-9-6-r92; Kim, SY et al., PLoS One. 2010 Feb 1;5(2):e8979. doi: 10.1371 / journal.pone.0008979; Toyoshima, M., et al., Proc Natl Acad Sci US A. 2012 Jun 12;109(24):9545-50. doi: 10.1073 / pnas.1121119109; Varghese, RT, et al., Sci Rep. 2018 Sep 11;8(1):13621. doi: 10.1038 / s41598-018-31864-x.) Hence, 15u has unexpected activity for cancer therapy in addition to CDK8 / 19 inhibition.

[0079] Example 3 Pharmacokinetics of thienopyridine derivatives. Pharmacokinetic (PK) assay. To measure mouse pharmacokinetics (PK), the thienopyridine derivatives were dissolved in 5% dextrose and administered in female FVB mice at different dosing conditions, blood samples were collected at various time points, and the compound concentrations in serum were measured by LC / MS / MS.

[0080] PK Results. Figures 2A-2D and Table 4 show the PK curves and calculated parameters for 15k, 15v, and 15u mixed and administered intravenously (iv) to mice at 0.5 mg / kg of each compound. Area under the curve (AUC) and elimination half-life (t 1 / 2 ) values, 15u had the highest iv availability and 15k the lowest in this assay.

[0081] [Table 4]

[0082] Figures 3A-3C and Table 5 show the PK curves and calculated parameters for the same mixture of 15k, 15v, and 15u administered orally (by gavage) at 1 mg / kg of each compound. In a separate study shown in Figure 3D, 15w was also administered orally at 1 mg / kg. In these assays, 15u showed by far the highest availability (AUC value), followed by 15w, 15v, and 15k. SnxB showed a similar AUC to 15w (Figure 3E).

[0083] [Table 5]

[0084] Oral PK was also determined for a mixture of the two most active compounds, 15w and 15u, administered to female CD1 mice at 30 mg / kg of each compound in 0.5% carboxymethylcellulose at higher dosages approaching the expected therapeutic dose. The results, shown in Figures 4A and 4B, show that 15u (but not 15w) exhibited excellent PK, had a high AUC (6.9-fold higher than the AUC of 15w), and exhibited very slow clearance, with a mean serum concentration of 15u at the latest time point (8 hours) of C max This shows that the average pregnancy rate was 64.4% at 15 weeks (compared to 11.5% at 15 weeks). The PK analysis demonstrated that one of the tested thienopyridine derivatives, 15u, exhibits highly attractive PK properties, possessing very high bioavailability and stability after oral administration.

[0085] Example 4 Pharmacokinetic profiles of 15u and 15w deuterated derivatives To determine the PK of deuterated derivatives of 15u and 15w, 8-12 week old female CD-1 mice were treated with 30 mg / kg 15u or 15u-D6 or 16 or 18 mg / kg 15w, 15w-D2, 15w-D6 by oral gavage in solution. Blood samples (70-100 μL) were collected from the retro-orbital vein of anesthetized animals using heparinized microhematocrit capillaries at various time points (1, 2, 6, 8 h post-dose) into BD Microtainer blood collection tubes for serum separation. Serum samples were processed for LCMSMS to determine drug concentrations using compound-specific MRM (15u: 439-394; 15u-D6: 445-394; 15w: 453-436; 15w-D2: 455-438; 15w-D6: 459-442). PK curves were generated by plotting drug concentrations against time points using GraphPad software, and the AUC (area under the curve) within the first 8 hours after dosing was calculated using Excel software to compare the PK profiles of non-deuterated and deuterated compounds. These two PK studies show that replacement of hydrogen in the dimethylamine group with deuterium (D6 derivative) slightly improved the PK for 15u (Figure 5A) and greatly improved the PK for 15w (Figure 5B). In contrast to D6, the D2 derivative did not improve the PK of 15w (Figure 5B).

[0086] Example 5 In vivo efficacy of 15u in castration-resistant prostate cancer CDK8 / 19 inhibition reduces the expression of certain androgen receptor (AR)-induced genes, including PSA, the most common marker of prostate cancer, and the growth of castration-resistant prostate cancer (CRPC). Figures 6A-6C show the effect of various concentrations of three CDK8 / 19 inhibitors, thienopyridine derivatives 15u and 15w, and Senexin B, on PSA expression in cell culture supernatants of the CRPC cell line C4-2 after 4 days of treatment in conventional medium supplemented with FBS. All three inhibitors suppressed PSA expression, with IC50 values ​​of 27.6 nM for 15u, 15.7 nM for 15w, and 255 nM for Senexin B. The in vivo effect of a mixture of 15u and 15w (the same mixture used for the PK study in Example 3) on PSA expression by C4-2 cells was analyzed after treatment of male NSG mice bearing C4-2 xenografts (grouped based on initial serum PSA levels) with 30 mg / kg per day orally for 4 days. Both serum PSA protein levels and intratumoral PSA mRNA levels were strongly reduced by treatment with a mixture of 15u and 15w (Figures 6D-6F). Given the significantly different PK of 15u and 15w (Example 3), it appears that the effect on PSA was mediated by 15u.

[0087] In another in vivo study, the CRPC cell line 22rv1 was grown as a xenograft in castrated male nude mice. 3 When tumors reached an average size of 100 μg / kg, mice were randomized into two groups (n=13) and treated with either vehicle (0.5% carboxymethylcellulose) control or 50 mg / kg of 15u taken orally daily. As shown in Figure 7A and also by tumor mass at the end of the study (Figure 7B), 15u treatment strongly inhibited tumor growth. Notably, 15u treatment showed no obvious adverse effects or loss of mouse body weight (Figure 7C).

[0088] Example 6 Effect of 15u on breast cancer metastasis. 4T1 is a murine triple-negative breast cancer (TNBC) cell line that is highly metastatic to the lungs. The effect of CDK8 on lung metastasis in this model was demonstrated in the study shown in Figures 8A-8C. CDK8-targeting shRNA was used to almost completely knock down CDK8 expression in 4T1 cells (Figure 8A; these cells do not express detectable CDK19 protein). Parental and CDK8-knockdown 4T1 cells (n=10) were orthotopically injected into the mammary fat pad and primary tumors were resected 17 days later. After surgery, all mice eventually died of lung metastases. Primary tumor mass showed no significant effect of CDK8 knockdown on tumor growth (Figure 8B). However, loss of CDK8 was associated with a strong increase in mouse survival (Figure 8C).

[0089] In a similar study, after resection of primary tumors, mice were divided into three groups (Figure 8D, n=8) and then treated with vehicle (5% dextrose) or 15u (25 mg / kg in 5% carboxymethylcellulose, orally, bid). 15u significantly increased mouse survival of metastatic disease (Figure 8E), an effect similar to that of CDK8 knockdown (Figure 8C). In another study with this model, tumors formed by parental 4T1 cells were resected and mice were randomized into two groups (Figure 8F, n=8) and treated with Senexin B (administered in medicated food (350 ppm) in combination with a single oral dose of 50 mg / kg as described in (Liang, 2018)) or given control food and vehicle. Senexin B treatment provided a statistically significant but modest increase in survival (Figure 8G).

[0090] In summary, the favorable PK of 15u (Example 3) and its in vivo activity (Examples 4, 5), together with its favorable kinome profile (Example 2), indicate that 15u is an effective CDK8 / 19 inhibitor and composition for use in the treatment of cancers associated with CDK8 / 19 activity.

[0091] Example 7 In vivo efficacy of combined 15u and enzalutamide treatment in castration-resistant prostate cancer The combined effect of 15u and the antiandrogen enzalutamide in CRPC was analyzed in the murine MYC-Cap-CR model. MYC-CaP-CR cells (Ellis L. et al., 2012. Prostate 72(6):587-591) were selected for castration resistance from engineered MYC-CaP cells expressing MYC from an AR-responsive promoter (Watson PA, et al., 2005. Cancer Res 65(24):11565-11571). Castration resistance in these cells is associated with overexpression of full-length AR but not AR variants, such as AR-V7 in 22rv1 (Olson BM, et al., 2017. Cancer immunology research 5(12):1074-1085). In short-term cell proliferation assays, the CDK8 / 19 inhibitors Senexin B and 15u had little effect on MYC-CAP-CR cell growth in androgen-containing medium, whereas enzalutamide paradoxically stimulated the proliferation of these cells (Figure 9A). However, when enzalutamide was combined with either CDK8 / 19 inhibitor, MYC-CAP-CR cell growth was strongly inhibited (Figure 9A), indicating that CDK8 / 19 inhibition can overcome enzalutamide resistance. In long-term clonogenic assays, both enzalutamide and CDK8 / 19 inhibitors reduced MYC-CaP-CR colony formation, and their combination produced a clearly synergistic effect (Figure 9B). The in vivo effect of 15u in combination with enzalutamide was tested on MYC-CaP-CR tumors growing subcutaneously in intact (non-castrated) FVB male mice. While both enzalutamide and 15u alone had modest effects on tumor volume (Figure 9C) and mass (Figure 9D) when used alone, their combination produced significant (p=0.02) tumor inhibition.

[0092] These results suggest that 15u can be advantageously combined with enzalutamide (or other antiandrogens) in the treatment of CRPC. The strongest in vivo activity of 15u as a single agent in CRPC was observed in 22rv1 cells expressing AR-V7, suggesting that prostate cancers expressing AR-V7 and possibly other androgen-independent AR variants may be particularly sensitive to CDK8 / 19 inhibition in vivo.

[0093] Example 8 Anti-leukemic effects of thienopyridine derivatives The anti-leukemic properties of 15u were examined in the acute myeloid leukemia (AML) cell line MV4-11, previously shown to be sensitive to CDK8 / 19 inhibition in vitro and in vivo (Pelish HE, et al., 2015. Nature 526(7572):273-276). The population of MV4-11 cells used for the in vivo studies was lentivirally infected to express luciferase and ZsGreen with pHIV-Luc-ZsGreen to allow leukemic proliferation analysis by bioluminescence imaging (BLI). The initial luciferase-ZsGreen transduced cell population was sorted for ZsGreen positivity by fluorescence activated cell sorting. This MV4-11 cell population was tested for sensitivity to 15u. 15u potently inhibited MV4-11 proliferation and appeared anti-proliferative with an IC50 value of 25 nM (Figure 10A).

[0094] Assay protocol. For in vivo studies, 7-week-old female NSG mice (Jackson Laboratories) were inoculated with 2 × 10 6MV4-11 cells expressing luciferase were injected into the tail vein. Following engraftment, BLI was performed on inoculated mice 5 days after cell inoculation. After BLI, mice were sorted into two matched cohorts of 10 mice and one cohort of 5 mice. BLI detection was performed on IVIS Lumina II Series Hardware for In-Vivo Imaging with optional XFOV lens and Living Image software. The IVIS environment for sorting mice into cohorts was set to Bin 8, F1.2, 180 seconds for high sensitivity. Subsequent exposures (weeks 1–5) were set to Bin 4, F1.2, 120 seconds for increased resolution.

[0095] Treatment began 6 days after cell inoculation and continued for 23 days. Ten mice received vehicle alone (5% carboxymethylcellulose) by gavage (200 μl). Ten mice received 15u at 30 mg / kg suspended in vehicle by gavage (200 μl) twice daily. Five mice were treated with medicated food containing 15u at 1 g / kg as custom Teklad diet prepared by Envigo (Madison, WI). This diet is comparable to the diet used to feed normal mice, except for the addition of dye and 15u. Control MV4-11 xenografted mice (vehicle) developed vigorous tumor masses as detected by BLI (Figures 10B-10C). The 15u gavage treatment group showed a striking response, with a 94% growth inhibition of leukemia growth, p=0.001. The 15u food treatment group showed an even more striking leukemia suppression, with a 99.7% inhibition of leukemia growth, p=0.002. Mice were followed for survival after treatment. Results. As shown in Figure 10D, mice treated with 15u by oral gavage showed superior survival rates.

[0096] Example 9 Effect of 15u on in vivo growth of MDA-MB-468 triple-negative breast cancer (TNBC) xenografts Human MDA-MB-468 triple-negative breast cancer (TNBC) cells were found to be responsive to 15u and other CDK8 / 19 inhibitors during long-term treatment in vitro. To evaluate the effect of CDK8 / 19 inhibition on the in vivo growth of MDA-MB-468 xenografts, one million cells were injected sc into the right flank of immunodeficient NSG female mice (9 weeks old) with 40% Matrigel (100 ml total volume). Eleven days after inoculation, mice were randomized into two groups (n=9) according to tumor size. The mean tumor volume in each group was 115 mm 3 The first group (control) of mice received a regular diet, while the second group (treated) of mice received a medicated diet containing 250 ppm of 15u. Thirteen days after the start of treatment, the medicated diet was supplemented with daily oral gavage providing 5 mg / kg of 15u solution in the treatment group or vehicle alone (control group). Thirty-seven days after the start of treatment, the gavage dose was increased to 8 mg / kg in the treatment group, and treatment was continued for a total of 66 days. Tumor volumes were measured twice weekly with calipers (Figure 11A), showing a significant reduction in tumor volume in the 15u treatment group. At the end of the study, mice were euthanized and tumors were dissected and weighed. Tumor mass was significantly lower in the 15u treatment group (Figure 11B). Mouse body weights (Figure 11C) showed no deleterious effects of long-term 15u treatment.

[0097] Example 10 Determination of the maximum tolerated dose (MTD) of 15u in CD-1 mice To determine the maximum tolerated dose (MTD), 8-week-old male or female CD-1 mice were randomly assigned to various dose groups and treated with increasing doses of 15u either by oral gavage of the solution or by drugged food. In one MTD in vivo study, female CD-1 mice were treated by gavage (bid) twice daily to provide 5, 10, 15, 30, 60 or 120 mg / kg 15u, and male CD-1 mice were treated by gavage bid to provide 60 or 120 mg / kg for 14 days. No adverse effects were observed in male mice in any treatment group (60 and 120 mg / kg bid) or in female mice in groups treated with 15u bid at doses up to 60 mg / kg (Figure 12A). The highest dose (120 mg / kg bid) caused approximately 10% weight loss in female mice after 7-10 days of treatment, but no further deterioration was observed throughout the remainder of the treatment period (Figure 12A).

[0098] In another long-term MTD in vivo assay, groups of male and female CD-1 mice were fed a regular diet (control) or a diet supplemented with 15u (500 ppm or 1000 ppm) for 4 or 5 weeks (Figure 12B). The daily doses for the 500 ppm and 1000 ppm groups were estimated to be approximately 50-100 mg / kg and 100-200 mg / kg, respectively, based on daily dietary consumption. Only the highest dose (1000 ppm) produced significant mass loss (5-10%) in female mice during the first week, but no further adverse effects were observed during the remaining treatment period. Considering that maximal therapeutic effect could be obtained at a daily dose of 30 mg / kg in various mouse xenograft models, these two MTD assays suggested a high therapeutic index for 15u.

[0099] Example 11 In silico modeling of the binding of thienopyridines and pyrrolopyridines to CDK8 Schrodinger Induced Fit docking was used to generate docking models for compounds 15u, 15w, 15w_APP, 15_PP, and 15u_CN that bind to CDK8. The Induced Fit protocol docks the active ligands with Glide, then generates a diverse ensemble of ligand poses, a procedure that uses a reduced van der Waals radius and an increased Coulomb-vdW cutoff to temporarily remove highly flexible side chains during the docking process. For each pose, the ligand is then adapted by reorienting neighboring side chains using Prime structure prediction. These residues and the ligand are then minimized. Finally, each ligand is redocked to its corresponding low-energy protein structure, and the resulting complexes are ranked according to GlideScore. This model was used to guide the design of the next predicted novel structures.

[0100] Figure 13 shows an overlay of the binding modes of 15u, 15w, 15w_APP, 15w_PP, and 15w_CN with CDK8. The results show the similarity of binding between various thienopyridines and pyrrolopyridines. Comparison of 15u and 15w with 15w_APP indicates that the -NH- group replacing the sulfur does not change the binding mode. In addition, 15w_App provides an extra H-bond donor to the hinge region of CDK8. The PP compounds show similar binding interactions with APP analogs. Docking models predict that -NH- provides an extra H-bond donor to the hinge region of CDK8 to increase potency while compensating for the loss of the 3-amino group. Comparison of 15u_CN with 15u and 15w suggests that the carbonitrile interacts similarly with amides.

[0101] Example 12 Structure-activity relationships Table 6 summarizes the structure activity relationships for the compositions described herein. To determine inhibitory potency, NFκB activity assays described in Example 1 (HEK238-NFκB-Luc assay) and MV4-11 assays described in Example 8 (MV4-11-Luc assay) were used. To determine PK, 8-12 week old female CD-1 mice were treated with the indicated doses (15-30 mg / kg) of test inhibitors as solution formulations (10% N-methyl-2-pyrrolidone (NMP), 27% propylene glycol (PG), 63% polyethylene glycol 400 (PEG-400)) by oral gavage. Blood samples (70-100 μL) were collected from the retro-orbital vein of anesthetized animals at various time points (1, 2, 6, 8 hours post-dose) by heparinized microhematocrit capillary tubes into BD Microtainer blood collection tubes for serum separation. Serum samples were processed for LCMSMS to determine drug concentrations using compound-specific MRM (15u: 439-394; 15u-D6: 445-394; 15w: 453-436; 15w-D2: 455-438; 15w-D6: 459-442; 6264: 483-394; 6300: 480-380; 6304: 453-408). Drug concentrations were plotted against time points to generate PK curves with GraphPad software, and AUC (area under the curve) within the first 8 hours post-dose was calculated with Excel software to compare the PK profiles of various compounds.

[0102] [Table 6-1] [Table 6-2] [Table 6-3] [Table 6-4] Example 13 Synthesis scheme Scheme 1 shows a general synthetic procedure for preparing the compounds disclosed herein. The preparation of specific compounds is shown below. Scheme 1. Small scale synthesis of thienopyridine derivatives

[0103] [ka]

[0104] Another scheme for preparing thienopyridine compounds is disclosed in Saito, K. et al., Bioorg Med Chem 2013, 21, 1628-42. Scheme 2 shows a synthetic scheme for the preparation of pyrrolopyridines such as 15u_PP. One skilled in the art can modify Schemes 1 and 2 to prepare furopyridines. Scheme 2. Synthesis of 15u_PP

[0105] [ka]

[0106] Synthesis of 3-amino-4-(4-(4-(2-(dimethylamino)-2-oxoethyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxamide (15w)

[0107] [ka]

[0108] To a solution of 2-(4-bromophenyl)-N,N-dimethyl-acetamide (1 eq) and tert-butyl 1,4-diazepane-1-carboxylate (1.2 eq) in t-BuOH and 1,4-dioxane was added 2-dicyclohexylphosphino-2'-(N,N-dimethylamino)biphenyl (0.15 eq), t-BuONa (1.4 eq) and tris(dibenzylideneacetone)dipalladium (0.05 eq). The mixture was degassed, protected with nitrogen and then refluxed for 1 h. Then the mixture was cooled to rt, water was added, the mixture was extracted with EA, the organic layer was washed with brine, dried over Na2SO4, condensed and purified by flash column to give tert-butyl 4-[4-[2-(dimethylamino)-2-oxo-ethyl]phenyl]-1,4-diazepane-1-carboxylate (92% yield), ESI-MS m / z: 362 ([M+H] + ; A solution of tert-butyl 4-[4-[2-(dimethylamino)-2-oxo-ethyl]phenyl]-1,4-diazepane-1-carboxylate (1 eq) in DCM, then TFA (5 eq) was added and the mixture was stirred at rt for 3 h, after which the mixture was condensed to remove TFA to give 2-(4-(1,4-diazepan-1-yl)phenyl)-N,N-dimethylacetamide, which was used without further purification, ESI-MS m / z: 262 ([M+H] + ). To a solution of 2-(4-(1,4-diazepan-1-yl)phenyl)-N,N-dimethylacetamide (1 eq) in acetonitrile was added 2,4-dichloronicotinonitrile (1 eq) and DIPEA (2 eq). The mixture was then stirred at 80° C. overnight. After that, the mixture was cooled to rt, condensed, the mixture was then dissolved in DCM, water was added, the mixture was extracted with DCM, the organic layer was collected, washed with brine, dried over Na2SO4, condensed, and purified by flash column to give 2-(4-(4-(2-chloro-3-cyanopyridin-4-yl)-1,4-diazepan-1-yl)phenyl)-N,N-dimethylacetamide (55% yield), ESI-MS m / z: 398 ([M+H] +);To a solution of 2-(4-(4-(2-chloro-3-cyanopyridin-4-yl)-1,4-diazepan-1-yl)phenyl)-N,N-dimethylacetamide (1 eq) in MeOH, MeONa (2 eq) and methyl thioglycolate (2 eq) were added and the mixture was then stirred at 100° C. overnight. Afterwards, the mixture was cooled to rt, condensed and purified by flash column to give methyl 3-amino-4-(4-(4-(2-(dimethylamino)-2-oxoethyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxylate (72% yield), ESI-MS m / z: 468 ([M+H] + );Methyl 3-amino-4-(4-(4-(2-(dimethylamino)-2-oxoethyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxylate (1 eq) in THF and water, then LiOH (2 eq) was added and the mixture was stirred at 60° C. overnight. Afterwards, the mixture was cooled to rt, condensed and dissolved in DMF, then HATU (1.5 eq) and DIPEA (2 eq) were added and the mixture was stirred at rt for 15 min, then NH4OH (6 eq) was added to the above mixture and stirred at rt for another 2 h. Then water was added and the mixture was extracted with DCM, the organic layers were combined, dried over Na2SO4, concentrated and purified by flash column to give 3-amino-4-(4-(4-(2-(dimethylamino)-2-oxoethyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxamide as a pale yellow solid (45% yield), ESI-MS m / z: 453 ([M+H] + ).

[0109] Synthesis of 3-amino-4-(4-(4-(2-(bis(methyl-d3)amino)-2-oxoethyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxamide (15w_D6)

[0110] [ka]

[0111] For experimental procedures, see 15w above. ESI-MS m / z: 459 ([M+H] + ). Synthesis of 3-amino-4-(4-(4-(2-(dimethylamino)-2-oxoethyl-1,1-d2)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxamide (15w_D2)

[0112] [ka] For experimental procedures, see 15w above. ESI-MS m / z: 455 ([M+H] + ). Synthesis of 3-amino-4-(4-(4-(dimethylcarbamoyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxamide (15u)

[0113] [ka]

[0114] See 15w above for experimental procedure. A pale yellow solid was obtained. ESI-MS m / z: 439 ([M+H] + ). Synthesis of 3-amino-4-(4-(4-(bis(methyl-d3)carbamoyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxamide (15u_D6)

[0115] [ka]

[0116] See 15w above for experimental procedure. The synthesis of 15u_D6 was confirmed by analysis with Waters HPLC-MS (LCA-232 SQ MS detector). The retention time was 21.40 min (5-95% TFA, 0.1% formic acid) and the parent ion (M+1) was observed at 445.1919. Figure 14A shows the UV chromatograph of compound 15u_D6 eluting at approximately 21 min. Figure 14B shows the ESI chromatograph of compound 15u_D6 eluting at approximately 21 min. Figure 14C confirms the synthesis of 15u_D6, ESI-MS m / z: 445 ([M+H] + ).

[0117] Synthesis of 3-amino-4-(4-(4-((4-methylpiperazin-1-yl)methyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxamide (6300)

[0118] [ka]

[0119] To a solution of 4-fluorobenzaldehyde (1 eq) and benzyl 1,4-diazepane-1-carboxylate (1.1 eq) in DMF was added K2CO3 (3 eq). The mixture was stirred at 90 °C overnight. Afterwards, the mixture was cooled to rt, water was added, the mixture was extracted with DCM, the organic layers were combined, washed with brine, dried over Na2SO4, condensed and purified by flash column to give benzyl 4-(4-formylphenyl)-1,4-diazepane-1-carboxylate (32% yield), ESI-MS m / z: 339 ([M+H] +), benzyl 4-(4-formylphenyl)-1,4-diazepane-1-carboxylate (1 eq) and 1-methylpiperazine (2 eq) in DCM, the solution was adjusted to pH 5 with acetic acid, then NaBH3CN (1.5 eq) was added and the mixture was stirred at rt overnight. After that, saturated aqueous NaHCO3 was added and the mixture was extracted with DCM, the organic layers were combined, washed with brine, dried over Na2SO4, condensed and purified by flash column to give benzyl 4-[4-[(4-methylpiperazin-1-yl)methyl]phenyl]-1,4-diazepane-1-carboxylate (66% yield), ESI-MS m / z: 423 ([M+H] + ), a solution of benzyl 4-[4-[(4-methylpiperazin-1-yl)methyl]phenyl]-1,4-diazepane-1-carboxylate (1 eq) was dissolved in concentrated HCl and stirred at rt for 2 h, then condensed to give 1-(4-((4-methylpiperazin-1-yl)methyl)phenyl)-1,4-diazepane, which was used without further purification, ESI-MS m / z: 289 ([M+H] + ), 1-(4-((4-methylpiperazin-1-yl)methyl)phenyl)-1,4-diazepane (1 eq) in acetonitrile was added with 2,4-dichloronicotinonitrile (1 eq) and DIPEA (2 eq). The mixture was then stirred at 80° C. overnight. Afterwards, the mixture was cooled to rt, condensed, the mixture was then dissolved in DCM, water was added, the mixture was extracted with DCM, the organic layer was collected, washed with brine, dried over Na2SO4, condensed and purified by flash column to give 2-chloro-4-(4-(4-((4-methylpiperazin-1-yl)methyl)phenyl)-1,4-diazepan-1-yl)nicotinonitrile (44% yield), ESI-MS m / z: 425 ([M+H] +To a solution of 2-chloro-4-(4-(4-((4-methylpiperazin-1-yl)methyl)phenyl)-1,4-diazepan-1-yl)nicotinonitrile (1 eq) in MeOH was added MeONa (2 eq) and methyl thioglycolate (2 eq), then the mixture was stirred at 90° C. overnight. Afterwards, the mixture was cooled to rt, condensed and purified by flash column to give methyl 3-amino-4-(4-(4-((4-methylpiperazin-1-yl)methyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxylate (79% yield), ESI-MS m / z: 495 ([M+H] + ), a solution of methyl 3-amino-4-(4-(4-((4-methylpiperazin-1-yl)methyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxylate (1 eq) in THF and water, then LiOH (2 eq) was added and the mixture was stirred at 60° C. overnight. Afterwards, the mixture was cooled to rt, condensed and dissolved in DMF, then HATU (1.5 eq) and DIPEA (2 eq) were added and the mixture was stirred at rt for 15 min, then NH4OH (6 eq) was added to the above mixture and stirred at rt for another 2 h. Water was then added and the mixture was extracted with DCM, the organic layers were combined, dried over Na2SO4, concentrated and purified by flash column to give 3-amino-4-(4-(4-((4-methylpiperazin-1-yl)methyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxamide (30% yield). 1 H NMR (300 MHz, DMSO-d6) δ: 8.39 (d, J=5.2 Hz, 1H), 7.08 (d, J=9.6 Hz, 3H), 7.07 (s, 2H), 6.97 (s, 2H), 6.71 (d, J=8.4 Hz, 2H), 3.75 (m, 2H), 3.52 (t, J=6.1 Hz, 2H), 3.32 (s, 2H), 3.29 (m, 2H), 2.33 (m, 8H), 2.16 (s, 3H), 2.13 (m, 2H); 13C NMR (300 MHz, DMSO-d6) δ: 167.07, 160.35, 159.35, 150.53, 147.59, 146.38, 130.06, 130.06, 124.87, 119.21, 111.74, 111.34, 111.34, 94.99, 61.68, 55.83, 54.76, 54.66, 54.66, 52.30, 52.30, 47.99, 47.99, 45.63, 27.39; + ).

[0120] Synthesis of 3-amino-4-(4-(4-(dimethylcarbamoyl)phenyl)-1,4-diazepan-1-yl)-6-methylthieno[2,3-b]pyridine-2-carboxamide (6304)

[0121] [ka]

[0122] To a solution of 4-bromo-N,N-dimethylbenzamide (1 eq) and tert-butyl 1,4-diazepane-1-carboxylate (1.2 eq) in t-BuOH and 1,4-dioxane, 2-dicyclohexylphosphino-2'-(N,N-dimethylamino)biphenyl (0.15 eq), t-BuONa (1.4 eq) and tris(dibenzylideneacetone)dipalladium (0.05 eq) were added. The mixture was degassed, protected with nitrogen and then refluxed for 1 h. Afterwards, the mixture was cooled to rt, water was added, the mixture was extracted with EA, the organic layer was washed with brine, dried over Na2SO4, condensed and purified by flash column to give tert-butyl 4-(4-(dimethylcarbamoyl)phenyl)-1,4-diazepane-1-carboxylate (94% yield), ESI-MS m / z: 348 ([M+H] +), a solution of tert-butyl 4-(4-(dimethylcarbamoyl)phenyl)-1,4-diazepane-1-carboxylate (1 eq) in DCM followed by TFA (5 eq) was added and the mixture was stirred at rt for 3 h, after which the mixture was condensed to remove the TFA to give 4-(1,4-diazepan-1-yl)-N,N-dimethylbenzamide which was used without further purification, ESI-MS m / z: 248 ([M+H] + ), 2,4-dichloro-6-methylnicotinonitrile (1 eq) and DIPEA (2 eq) were added to a solution of 4-(1,4-diazepan-1-yl)-N,N-dimethylbenzamide (1 eq) in acetonitrile. The mixture was then stirred at 80° C. overnight. After that, the mixture was cooled to rt, condensed, the mixture was then dissolved in DCM, water was added, the mixture was extracted with DCM, the organic layer was collected, washed with brine, dried over Na2SO4, condensed, and purified by flash column to give 4-(4-(2-chloro-3-cyano-6-methylpyridin-4-yl)-1,4-diazepan-1-yl)-N,N-dimethylbenzamide (66% yield), ESI-MS m / z: 398 ([M+H] + ), 4-(4-(2-chloro-3-cyano-6-methylpyridin-4-yl)-1,4-diazepan-1-yl)-N,N-dimethylbenzamide (1 eq) in MeOH was added with MeONa (2 eq) and methyl thioglycolate (2 eq), then the mixture was stirred at 100 °C overnight. Afterwards, the mixture was cooled to rt, condensed and purified by flash column to give methyl 3-amino-4-(4-(4-(dimethylcarbamoyl)phenyl)-1,4-diazepan-1-yl)-6-methylthieno[2,3-b]pyridine-2-carboxylate (77% yield), ESI-MS m / z: 468 ([M+H] +), a solution of methyl 3-amino-4-(4-(4-(dimethylcarbamoyl)phenyl)-1,4-diazepan-1-yl)-6-methylthieno[2,3-b]pyridine-2-carboxylate (1 eq) in THF and water, then LiOH (2 eq) was added and the mixture was stirred at 60° C. overnight. Afterwards, the mixture was cooled to rt, condensed and dissolved in DMF, then HATU (1.5 eq) and DIPEA (2 eq) were added and the mixture was stirred at rt for 15 min, then NH4OH (6 eq) was added to the above mixture and stirred at rt for another 2 h. Then water was added and the mixture was extracted with DCM, the organic layers were combined, dried over Na2SO4, concentrated and purified by flash column to give 3-amino-4-(4-(4-(dimethylcarbamoyl)phenyl)-1,4-diazepan-1-yl)-6-methylthieno[2,3-b]pyridine-2-carboxamide as a pale yellow solid (yield 33%). 1 H NMR (300 MHz, DMSO-d6) δ: 7.30 (d, J=8.9 Hz, 2H), 7.02 (s, 2H), 6.96 (s, 3H), 6.77 (d, J=8.9 Hz, 2H), 3.81 (m, 2H) 3.58 (m, 2H), 3.27 (m, 2H), 3.16 (m, 2H), 2.97 (s, 6H), 2.45 (s, 3H), 2.14 (m, 2H); 13 C NMR (300 MHz, DMSO-d6) δ: 170.50, 167.18, 160.03, 159.94, 159.39, 149.42, 146.46, 129.35, 129.35, 122.56, 117.13, 111.78, 110.45, 110.45, 94.25, 55.52, 54.93, 47.88, 47.75, 39.52, 39.52, 27.23, 24.22. + ).

[0123] Synthesis of 3-amino-4-(4-(4-((3-hydroxypropyl)(methyl)carbamoyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxamide (6264)

[0124] [ka]

[0125] A solution of ethyl 4-fluorobenzoate (1 eq) and 1,4-diazepane (2 eq) in DMSO was added and stirred at 120° C. overnight. Afterwards, the mixture was cooled to rt, water was added, the mixture was extracted with DCM, the organic layers were combined, washed with brine, dried over Na2SO4, condensed and purified by flash column to give ethyl 4-(1,4-diazepan-1-yl)benzoate (83% yield), ESI-MS m / z: 249 ([M+H] + To a solution of benzyl 4-(4-(ethoxycarbonyl)phenyl)-1,4-diazepane-1-carboxylate (80% yield), ESI-MS m / z: 383 ([M+H] 2H2O 2 , ethyl 4-(1,4-diazepan-1-yl)benzoate (1 eq) and benzyl carbochloridate (1.5 eq) in DCM was added DIPEA (2 eq) and the mixture was then stirred at rt overnight. Afterwards, saturated aqueous NaHCO3 was added and the mixture was extracted with DCM, the organic layers were combined, washed with brine, dried over Na2SO4, condensed and purified by flash column to give benzyl 4-(4-(ethoxycarbonyl)phenyl)-1,4-diazepane-1-carboxylate (80% yield), ESI-MS m / z: 383 ([M+H] 2H2O 2 , ethyl 4-(1,4-diazepan-1-yl)benzoate (1 eq) and benzyl carbochloridate (1.5 eq) in DCM. + ), a solution of benzyl 4-(4-(ethoxycarbonyl)phenyl)-1,4-diazepan-1-carboxylate (1 eq) was dissolved in MeOH and water, then NaOH (2.5 eq) was added and the mixture was refluxed for 2 h, the mixture was then cooled to rt, condensed, water was added, the mixture was acidified to pH=4 with 1N HCl and extracted 3 times with DCM, the organic layers were combined, dried over Na2SO4 and condensed to give 4-(4-((benzyloxy)carbonyl)-1,4-diazepan-1-yl)benzoic acid which was used without further purification, ESI-MS m / z: 355 ([M+H] +). HATU (1.5 eq) and DIPEA (3 eq) were added to a solution of 4-(4-((benzyloxy)carbonyl)-1,4-diazepan-1-yl)benzoic acid in DCM, then the mixture was stirred at rt for 20 min, after which 3-(methylamino)propan-1-ol (1.5 eq) was added and the mixture was stirred at rt for another 4 h. Then water was added, the mixture was extracted with DCM, the organic layers were combined, dried over Na2SO4, condensed and purified by flash column to give benzyl 4-(4-((3-hydroxypropyl)(methyl)carbamoyl)phenyl)-1,4-diazepan-1-carboxylate (85% yield), ESI-MS m / z: 426 ([M+H] + ). To a solution of benzyl 4-(4-((3-hydroxypropyl)(methyl)carbamoyl)phenyl)-1,4-diazepane-1-carboxylate (1 eq) in EtOH and ethyl acetate was added Pd / C, the solution was then saturated with hydrogen and stirred at rt overnight. Afterwards, the mixture was filtered, the residue was washed with methanol, and the solutions were collected, combined and condensed to give 4-(1,4-diazepan-1-yl)-N-(3-hydroxypropyl)-N-methylbenzamide (90% yield), ESI-MS m / z: 292 ([M+H] + ). To a solution of 4-(1,4-diazepan-1-yl)-N-(3-hydroxypropyl)-N-methylbenzamide (1 eq) in acetonitrile was added 2,4-dichloronicotinonitrile (1 eq) and DIPEA (2 eq). The mixture was then stirred at 80° C. overnight. Afterwards, the mixture was cooled to rt, condensed, the mixture was then dissolved in DCM, water was added, the mixture was extracted with DCM, the organic layer was collected, washed with brine, dried over Na2SO4, condensed and purified by flash column to give 4-(4-(2-chloro-3-cyanopyridin-4-yl)-1,4-diazepan-1-yl)-N-(3-hydroxypropyl)-N-methylbenzamide (47% yield), ESI-MS m / z: 428 ([M+H] +). To a solution of 4-(4-(2-chloro-3-cyanopyridin-4-yl)-1,4-diazepan-1-yl)-N-(3-hydroxypropyl)-N-methylbenzamide (1 eq) in MeOH was added MeONa (2 eq) and methyl thioglycolate (2 eq), then the mixture was stirred at 90° C. overnight. Afterwards, the mixture was cooled to rt, condensed and purified by flash column to give methyl 3-amino-4-(4-(4-((3-hydroxypropyl)(methyl)carbamoyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxylate (78% yield), ESI-MS m / z: 498 ([M+H] + ). A solution of methyl 3-amino-4-(4-(4-((3-hydroxypropyl)(methyl)carbamoyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxylate (1 eq) in THF and water, then LiOH (2 eq) was added and the mixture was stirred at 60° C. overnight. Afterwards, the mixture was cooled to rt, condensed, dissolved in DMF, then HATU (1.5 eq) and DIPEA (2 eq) were added and the mixture was stirred at rt for 15 min, then NH4OH (6 eq) was added to the above mixture and stirred at rt for another 2 h. Then water was added and the mixture was extracted with DCM, the organic layers were combined, dried over Na2SO4, concentrated and purified by flash column to give 3-amino-4-(4-(4-((3-hydroxypropyl)(methyl)carbamoyl)phenyl)-1,4-diazepan-1-yl)thieno[2,3-b]pyridine-2-carboxamide as a yellow solid (yield 51%). 11H NMR (300 MHz, DMSO-d6) δ: 8.40 (d, J=5.6 Hz, 1H), 7.27 (d, J=8.7 Hz, 2H), 7.10 (s, 2H), 7.08 (d, J=5.6 Hz, 1H), 7.00 (s, 2H), 6.78 (d, J=8.7 Hz, 2H), 4.47 (t, J=5.4 Hz, 1H), 3.81 (m, 2H), 3.59 (t, J=6.5 Hz, 2H), 3.41 (m, 4H), 3.30 (m, 2H), 3.20 (m, 2H), 2.95 (s, 3H), 2.16 (m, 2H), 1.70 (m, 2H); 13 13C NMR (300 MHz, DMSO-d6) δ: 170.69, 167.08, 160.35, 159.23, 150.55, 149.36, 146.38, 129.03, 129.03, 123.02, 119.19, 111.70, 110.50, 110.50, 95.13, 58.36, 58.36, 55.72, 54.82, 47.82, 47.82, 39.53, 39.53, 27.21; ESI-MS m / z: 483 ([M + H] + ). Yet another aspect of the present invention may be as follows. [1] A compound of formula 1.

Chem.

change

[11] Q is sulfur, n is 0, and R 4 is methyl and R 3 -C(O)NH 2 and R 2 Ga-NH 2 The compound according to

[10] above, 〔12〕R 1 The compound according to

[11] above, wherein is N,N-dimethylformamide. 〔13〕R 1 The compound according to [2] above, wherein is N-(3-hydroxypropyl)formamide, N-(3-aminopropyl)-N-methylformamide, or tert-butyl(3-(N-methylformamido)propyl)carbamate. 〔14〕R 1 The compound according to

[13] above, wherein is N-(3-hydroxypropyl)formamide.

[15] Q is sulfur, n is 0, and R 4 is hydrogen or methyl, and R 3 -C(O)NH 2 and R 2 Ga-NH 2 The compound according to

[13] or

[14] above, 〔16〕R 4 The compound according to

[15] above, wherein is hydrogen. 〔17〕R 1 but

change

[17] above, wherein is (4-methylpiperazin-1-yl)methylene, 4-methylpiperazine-1-carbaldehyde, piperazine-1-carbaldehyde, or tert-butyl 4-formylpiperazine-1-carboxylate. 〔19〕R 1 The compound according to the above

[18] , wherein is (4-methylpiperazin-1-yl)methylene.

[20] Q is sulfur, n is 0, and R 4 is hydrogen or methyl, and R 3 -C(O)NH 2 and R 2 Ga-NH 2 The compound according to any one of the above items

[17] to

[19] , 〔21〕R 4 The compound according to

[20] above, wherein is hydrogen.

[22] A pharmaceutical composition comprising a therapeutically effective amount of the compound according to any one of [1] to

[21] above and a pharma- ceutically acceptable carrier, excipient, or diluent.

[23] A method for treating a subject having cancer, comprising administering a therapeutically effective amount of a compound of formula 1.

change

[24] The method according to

[23] above, wherein the cancer is prostate cancer, leukemia, breast cancer, colon cancer, ovarian cancer, pancreatic cancer, or melanoma.

[25] The method according to

[24] above, wherein the cancer is prostate cancer.

[26] The method according to

[25] above, wherein the prostate cancer is castration-resistant prostate cancer or is resistant to androgen deprivation therapy.

[27] The method according to

[24] or

[25] , wherein the subject is undergoing androgen deprivation therapy prior to administration of the compound to the subject.

[28] The method according to any one of

[24] to

[27] above, wherein the subject is undergoing androgen deprivation therapy simultaneously with administration of the compound to the subject.

[29] The method according to

[24] above, wherein the cancer is leukemia.

[30] The method according to

[29] above, wherein the leukemia is acute myeloid leukemia.

[31] The method according to

[24] above, wherein the cancer is breast cancer.

[32] The method according to

[31] above, wherein the breast cancer is metastatic breast cancer or triple-negative breast cancer. 〔33〕R 1 but

change

[23] to

[32] , wherein together are oxo. 〔34〕R 1 The method according to claim 33, wherein is N,N-bis(methyl-d3)formamide, N,N-bis(methyl-d3)acetamide, or N,N-dimethylacetamide-2,2-d2. 〔35〕R 1 The method according to claim 34, wherein is N,N-bis(methyl-d3)formamide.

[36] Q is sulfur, n is 0, and R 4 is hydrogen or methyl, and R 3 -C(O)NH 2 and R 2 Ga-NH 2 The method according to any one of the above

[33] to

[35] , 〔37〕R 4 The method according to claim 36, wherein is hydrogen. 〔38〕R 1 The method according to claim 33 or 34, wherein is N,N-bis(methyl-d3)propionamide.

[39] Q is sulfur, n is 0, and R 4 is hydrogen or methyl, and R 3 -C(O)NH 2 and R 2 Ga-NH 2 and R 1 The method according to claim 38, wherein is N,N-bis(methyl-d3)propionamide. 〔40〕R 4 The method according to claim 39, wherein is hydrogen. 〔41〕R 1 The method according to claim 33, wherein is N,N-dimethylformamide, N,N-dimethylacetamide, or N-methylformamide. 〔42〕R 1 The method according to claim 41, wherein is N,N-dimethylformamide.

[43] Q is sulfur, n is 0, and R 4 is hydrogen or methyl, and R 3 -C(O)NH 2 and R 2 Ga-NH 2 The method according to

[41] or

[42] , 〔44〕R 4 The method according to claim 43, wherein is methyl. 〔45〕R 4 The method according to claim 43, wherein is hydrogen. 〔46〕R 1 The method according to claim 41, wherein is N,N-dimethylacetamide.

[47] Q is sulfur, n is 0, and R 4 is hydrogen or methyl, and R 3 -C(O)NH 2 and R 2 Ga-NH 2 The method according to

[41] or

[46] , 〔48〕R 4 The method according to claim 47, wherein is methyl. 〔49〕R 4 The method according to claim 47, wherein is hydrogen. 〔50〕R 1 The method according to claim 33, wherein is N-(3-hydroxypropyl)formamide, N-(3-aminopropyl)-N-methylformamide, or tert-butyl(3-(N-methylformamido)propyl)carbamate. 〔51〕R 1 The method according to claim 50, wherein is N-(3-hydroxypropyl)formamide.

[52] Q is sulfur, n is 0, and R 4 is hydrogen or methyl, and R 3 -C(O)NH 2 and R 2 Ga-NH 2 The method according to claim 50 or 51, 〔53〕R 4 The method according to claim 52, wherein is hydrogen. 〔54〕R 1 but

change

[23] to

[32] , wherein the heterocycle is optionally deuterated. 〔55〕R 1 55. The method of claim 54, wherein is (4-methylpiperazin-1-yl)methylene, 4-methylpiperazine-1-carbaldehyde, piperazine-1-carbaldehyde, or tert-butyl 4-formylpiperazine-1-carboxylate. 〔56〕R 1 The method according to claim 55, wherein is (4-methylpiperazin-1-yl)methylene.

[57] Q is sulfur, n is 0, and R 4 is hydrogen or methyl, and R 3 -C(O)NH 2 and R 2 Ga-NH 2 The method according to any one of the above

[54] to

[56] , 〔58〕R 4 The method according to claim 57, wherein is hydrogen.

[59] The method according to any one of

[23] to

[58] above, wherein the compound is administered as a pharmaceutical composition comprising a therapeutically effective amount of the compound and a pharma- ceutically acceptable carrier, excipient, or diluent.

[60] A method for inhibiting CDK8 or CDK19, comprising contacting CDK8 or CDK19 with the compound according to any one of [1] to

[21] above.

[61] The method according to

[60] , wherein the compound contacts CDK8 and specifically inhibits CDK8, or the compound contacts CDK19 and specifically inhibits CDK19.

[62] The method according to

[61] above, wherein the compound specifically inhibits both CDK8 and CDK19.

Claims

【Request 1】 【Chemistry 1】 or a pharma- ceutically acceptable salt thereof for treating a subject having cancer.

2. 2. The pharmaceutical composition of claim 1, wherein the cancer is prostate cancer, leukemia, breast cancer, colon cancer, ovarian cancer, pancreatic cancer, or melanoma.

3. The pharmaceutical composition of claim 2 , wherein the cancer is prostate cancer.

4. The pharmaceutical composition described in claim 3, wherein the prostate cancer is castration-resistant prostate cancer.

5. The pharmaceutical composition described in claim 3, wherein the prostate cancer is resistant to androgen deprivation therapy.

6. The pharmaceutical composition of claim 2 , wherein the cancer is leukemia.

7. The pharmaceutical composition described in claim 6, wherein the leukemia is acute myeloid leukemia.

8. The pharmaceutical composition according to claim 2 , wherein the cancer is breast cancer.

9. The pharmaceutical composition described in claim 8, wherein the breast cancer is metastatic breast cancer.

10. The pharmaceutical composition described in claim 8, wherein the breast cancer is triple-negative breast cancer.

11. The compound comprising: 【Chemistry 2】 The pharmaceutical composition according to any one of claims 1 to 10,

12. The compound comprising: 【Chemistry 3】 The pharmaceutical composition according to any one of claims 1 to 10,

13. The following formula 【Chemistry 4】 or a pharma- ceutically acceptable salt thereof.

14. The compound is 【Chemistry 5】 14. The compound of claim 13,

15. A pharmaceutical composition comprising a therapeutically effective amount of a compound according to any one of claims 13-14 and a pharma- ceutically acceptable carrier, excipient, or diluent.

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