Creatine kinase inhibitors

Abstract

The present invention relates to compounds of Formula (I): or a pharmaceutically acceptable salt thereof useful for treatment of a disease or disorder associated with CK and methods of inhibiting CK.

Description

CREATINE KINASE INHIBITORSGOVERNMENT LICENSE RIGHTS

[0001] This invention was made with government support under R21 CA259739 awarded by the National Institutes of Health. The government has certain rights in the invention.RELATED APPLICATIONS

[0002] This application claims priority to US Provisional Application No. 63 / 647,222, filed on May 14, 2024, the contents of which are hereby incorporated by reference in their entirety.FIELD

[0003] The present invention relates to inhibitors of creatine kinase useful for the treatment of cancers.BACKGROUND

[0004] Creatine kinase (CK), also known as creatine phosphokinase (CPK) or phosphocreatine kinase is known to play a key role in cellular energy buffering and transport and provide local ATP production in periods of elevated energetic demand, such as during rapid anabolism and growth. CK is essential for the growth and metastasis of aggressive acute myeloid leukemias, liver and breast cancer metastases, and pancreatic cancers. Because somatic tissues do not rely on CK for viability, the essentiality of CK for growth and metastasis of these cancers suggests CK inhibitors will be attractive targets for cancer therapy. Covalent CK inhibitors are described in WO2022087433 and Narek Darabedian, et al., “Depletion of creatine phosphagen energetics with a covalent creatine kinase inhibitor, Nature Chemical Biology. 19, 815-824 (July 2023). New CK inhibitors are required to address an ongoing unmet need for new cancer therapies.SUMMARY OF THE INVENTION

[0005] An aspect of the invention is to provide compounds of Formula I, or a pharmaceutically acceptable salt thereof:where:X is -C(O)NH- or -CH2O-; R1is H or methyl;R2is -CN, -OCH3, or -SCH3.

[0006] Another aspect of the invention is to provide compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein X is -C(O)NH-.

[0007] Another aspect of the invention is to provide compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein

[0008] Another aspect of the invention is to provide compounds of Formula I, or a pharmaceuticallyor -SCH3.

[0009] Another aspect of the invention is to provide compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein X is -C(O)NH-, R1is H,

[0010] Another aspect of the invention is to provide compounds of Formula I, or a pharmaceutically acceptable salt thereof, wherein

[0011] Another aspect of the invention is to provide compounds of Formula I, or a pharmaceutically2acceptable salt thereof, whereinandR is -SCH

[0012] Another aspect of the invention is to provide compounds of Formula I, or a pharmaceutically acceptable salt thereof, selected from the group consisting of:

[0013] Another aspect of the present invention provides a pharmaceutical composition comprising a compound of Formula I or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

[0014] A further aspect of the present invention provides a compound of Formula I or a pharmaceutically acceptable salt thereof for use in therapy.

[0015] Another aspect of the present invention provides a method for treating cancer susceptible to inhibition of CK comprising administering to a patient in need thereof a therapeutically effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof.

[0016] A further aspect of the present invention provides the use of a compound of Formula I or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment of disorders susceptible to inhibition of CK.

[0017] Another aspect of the invention is the use of a compound of Formula I or a pharmaceutically acceptable salt thereof for the treatment of a cancer associated with creatine kinase overexpression.DETAILED DESCRIPTION

[0018] Terms used herein but not separately defined are taken to have their normal and customary meaning as understood by one of ordinary skill in the art.

[0019] The term “patient” means mammal and “mammal” includes, but is not limited to, a human.

[0020] “Therapeutically effective amount” means the dosage of a compound of Formula I or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing a compound of Formula I, or a pharmaceutically acceptable salt thereof, necessary to inhibit CK in a patient in need thereof. Anticipated dosages of a compound of Formula I, or a pharmaceutically acceptable salt thereof, are in the range of 1 mg / patient / day to 2000 mg / patient / day. The exact dosage required to treat a patient and the duration of treatment will be determined by a physician in view of the stage and severity of the disease as well as the specific needs and response of the individual patient. Dosage administration may be adjusted to provide an optimal therapeutic benefit to an individual patient and to manage or avoid drug-related toxicities. For example, in addition to single daily dosing, multiple smaller daily doses or administration on a staggered daily, weekly, or monthly schedule may be appropriate.

[0021] The terms “treatment”, “treat”, and “treating” are meant to include the full spectrum of pharmaceutical intervention for a patient in need of CK inhibition, such as administration of a CK inhibitor of the present invention to alleviate, slow, or reverse one or more of a patient’s symptoms or to delay progression of the disorder even if the disorder is not actually eliminated.

[0022] The compounds of Formula I are useful for the treatment of cancers associated with CK overexpression. These cancers include liver cancer, pancreatic cancer, lung cancer, colon cancer, and hematological cancers, including myelogenous leukemia, myeloid leukemia (including acute myeloid leukemia), myelodysplastic syndrome, lymphoblastic leukemia (including acute lymphoblastic leukemia), chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), high risk CLL, follicular lymphoma, diffuse large B-cell lymphoma (including DLBCL or ABC-DLBCL), mantle cell lymphoma (MCL), Waldenstrom’s macroglobulinemia (WM), multiple myeloma, marginal zone lymphoma (MZL), Burkitt’s lymphoma, non-Burkitt high grade B-cell lymphoma,extranodal marginal zone B-cell lymphoma, transformed high grade B-cell lymphoma (HGBL), lymphoplasmacytic lymphoma (LPL), central nervous system lymphoma (CNSL), or MALT lymphoma.

[0023] A further aspect of the invention is the use of a compound of Formula I for the treatment of acute myeloid leukemia (AML).

[0024] The compounds of Formula I are inhibitors of CK. The CK susceptible to inhibition of a compound of Formula I may be an isoform of creatine kinase such as CK-MM, CK-BB, or CK-MB.

[0025] Compounds of Formula I or a pharmaceutically acceptable salt thereof are preferably formulated as a pharmaceutical composition using a pharmaceutically acceptable carrier and administered by a variety of routes. Preferably, such compositions are for oral administration. Such pharmaceutical compositions and methods for preparing them are well known in the art. See, for example, REMINGTON: THE SCIENCE AND PRACTICE OF PHARMACY (L.V. Allen ed., Pharmaceutical Press, 22ndEdition, 2012).

[0026] A compound of F ormula I or a pharmaceutically acceptable salt thereof may be administered either simultaneously with, or before, or after, one or more other therapeutic agents. When said compounds or pharmaceutically acceptable salts thereof are administered with one or more therapeutic agents, they may be administered separately, by the same or different route of administration, or together in the same pharmaceutical composition as the other therapeutic agent or agents. Where one or more additional therapeutic agents are administered, the administration of each therapeutic agent may be simultaneous, separate, or sequential.

[0027] Compounds of Formula I are capable of reacting with a number of inorganic and organic acids to form pharmaceutically acceptable acid addition salts. Such pharmaceutically acceptable salts and common methodology for preparing them are well known in the art. See, for example, P. Stahl, etal., HANDBOOK OF PHARMACEUTICAL SCIENCES, (VCHA / Wiley - VCH, 2002); S.M. Berge, etal., Pharmaceutical Salts, 66 JOURNAL OF PHARMACEUTICAL SCIENCES 1 (1977).

[0028] It will be understood that compounds of Formula I have a chiral center and may be depicted as single stereoisomers. For example, a compound of Formula I is illustrated below:

[0029] All stereoisomers of the compounds of Formula I are contemplated within the scope of the present invention. As used herein, references to a single stereoisomer are meant to also include stereoisomeric mixtures including the named or depicted compound of Formula I. Herein, the Cahn-Ingold-Prelog designations of (R)- and (S)- may be used to refer to specific stereoisomers. Specific stereoisomers can be prepared by stereospecific synthesis using enantiomerically pure or enriched starting materials. The specific stereoisomers of either starting materials, intermediates, or racemic mixtures including compounds of Formula I and II can be resolved by techniques well known in the art, such as those found in Stereochemistry of Organic Compounds, E. I. Eliel and S. H. Wilen (Wiley 1994) and Enantiomers, Racemates, and Resolutions, J., Jacques, A Collet, and S. H. Wilen (Wiley 1991), including chromatography on chiral stationary phases, enzymatic resolutions, or fractional crystallization or chromatography of diastereomers formed for that purpose, such as diastereomeric salts.

[0030] It should be understood that the compounds described herein, unless otherwise specified, encompass and include all isomers and stable isotopic variants of the compounds of the present disclosure, such as deuterated compounds. All isotopic variants of the compounds provided herein, whether radioactive or not, should be included within the scope of the present disclosure.

[0031] Compounds of the present invention are named according to IUPAC, and may also be named according to CAS, and other naming conventions may be used to unambiguously identify a compound of Formula I or a pharmaceutically acceptable salt thereof.

[0032] The compounds employed as initial starting materials in the synthesis of compounds of Formula I are well known and, to the extent not commercially available, are readily synthesized using specific references provided, by standard procedures commonly employed by those of ordinary skill in the art or are found in general reference texts. Examples of known procedures and methods include those in general reference texts such as: COMPREHENSIVE ORGANIC TRANSFORMATIONS (VCH Publishers Inc., 1989); COMPENDIUM OF ORGANIC SYNTHETIC METHODS (Wiley Interscience, Volumes 1 - 10, 1974 - 2002); Michael B. Smith and Jerry March, ADVANCED ORGANIC CHEMISTRY, REACTIONS, MECHANISMS, AND STRUCTURE (Wiley Interscience, 5thed. 2001); Francis A. Carey and Richard J. Sundberg, ADVANCED ORGANIC CHEMISTRY, PARTB, REACTIONS AND SYNTHESIS (Kluwer Academic / Plenum Publishers, 4thed. 2000), and references cited therein.

[0033] Certain intermediates described in the following preparations may contain one or more nitrogen protecting groups. It is understood that protecting groups may be varied as appreciated by one of skill in the art depending on the actual reaction conditions and the particular transformations to be performed. The protection and deprotection conditions are well known to the skilled artisan and are described in the literature (See for example "Greene's Protective Groups in Organic Synthesis", Fifth Edition, by Peter GM. Wuts and Theodora W. Greene, John Wiley and Sons, Inc. 2014).

[0034] Compounds of Formula I or pharmaceutically acceptable salts thereof may be prepared by a variety of procedures known in the art, some of which are illustrated in the Schemes, Preparations, and Examples below which are provided to further illustrate the invention without limiting the scope of the invention in any way. The specific steps and methodology for each of the synthetic routes described may be combined in different ways, or in conjunction with steps from different schemes, to prepare compounds of the present invention or pharmaceutically acceptable salts thereof. The products of each step in the schemes below may be isolated by conventional methods well known in the art, including extraction, evaporation, precipitation, chromatography, filtration, trituration, and crystallization. In the schemes below, all substituents are as previously defined unless otherwise indicated.

[0035] The abbreviations used herein are defined according to Al dri chimica Acta, vol. 17, No. 1, 1984. Other abbreviations are defined as follows: “ACN” refers to acetonitrile; “BF3.THF” refers to boron trifluoride tetrahydrofuran; “DCM” refers to dichloromethane; “DIPEA” refers to diisopropylethylamine; “DMF” refers to dimethylformamide; “DMSO” refers to dimethyl sulfoxide; “EtOAc” refers to ethyl acetate; “ESI-MS” refers to electrospray ionization mass spectrometry; “HATU” refers to l-[bis(dimethylamino)methylene]-17 / -l,2,3-triazolo[4,5- / >]pyridinium 3-oxid hexafluorophosphate; “h” refers to hour(s); “MeOH” refers to methanol; “PG” refers to protecting group; “PPhi” refers to triphenylphosphine; “prep-HPLC” refers to preparative high performance liquid chromatography; “TFA” refers to trifluoroacetic acid; “TMAD” refers to tetramethylazodicarboxamide.

[0036] In the schemes below, all substituents unless otherwise indicated, are as previously defined. The reagents and starting materials are either commercially available or may be prepared by methods well known to one of ordinary skill in the art, some of which are presented in the preparations below. Without limiting the scope of the invention, the following schemes, preparations, and examples are provided to further illustrate the invention.

[0037] In the schemes below, all substituents unless otherwise indicated, are as previously defined. The reagents and starting materials are either commercially available or may be prepared by methods well known to one of ordinary skill in the art, some of which are presented in the preparations below. Without limiting the scope of the invention, the following schemes, preparations, and examples are provided to further illustrate the invention.Scheme 1

[0038] Scheme 1 depicts the preparation of compounds of the present invention where X is - C(O)NH- beginning with a suitable acid (i) and amine (ii). Amide coupling conditions are well known to the skilled person and include reacting a solution of the carboxylic acid (i) and a desired amine (ii) in a suitable solvent, such as DMF or DCM, with an appropriate coupling reagent, such as HATU, in the presence of a suitable organic base, such as DIPEA, with stirring at ambient temperature for at least 12 h.Scheme 2

[0039] Scheme 2 illustrates the preparation of compounds of the present invention where X is - CH2O- under standard Mitsunobu conditions. The suitable alcohols (iii) and (iv) are reacted in the presence of an appropriate azodicarboxylate, such as TMAD, and PPh i, in a suitable solvent, such as toluene, under N2 atmosphere, at 90 °C for at least 12 h.

[0040] The skilled artisan will appreciate that the order of the synthetic steps required to prepare compounds of Formula I may be varied as necessary or desired.PREPARATIONSPreparation 11 -Acryloylindoline-3-carboxylic acid

[0041] To a solution of indoline-3 -carboxylic acid (1.0 g, 6.1 mmol) in DCM (3 mL) was added acryloyl chloride (665.6 mg, 7.4 mmol) at 0 °C. The mixture was stirred at 25 °C for 2 h. Then the reaction mixture was concentrated under reduced pressure to give the title compound.Preparation 2 / c / V-Butyl 3-((2-cyanobenzo[<7]thiazol-6-yl)carbamoyl)indoline- l -carboxylate

[0042] To a solution of 1 -( / / 7-butoxycarbonyl)indoline-3 -carboxylic acid (90 mg, 0.3 mmol) and 6-aminobenzo[<7]thiazole-2-carbonitrile (50 mg, 0.3 mmol) in DCM (2 mL) was added HATU (220 mg, 0.6 mmol) and DIPEA (180.0 mg, 1.4 mmol) at 0 °C. The mixture was stirred at 25 °C for 12 h. The reaction was poured into water (15 mL) and extracted with EtOAc (3 x 50 mL). The reaction mixture was dried over NaiSO-i, filtered, and concentrated under reduced pressure to give a residue, and the residue was purified by silica-gel column chromatography with DCM and MeOH to give the title compound (80 mg, 67%).Preparation 35-Hydroxybenzo[< / ]thiazole-2-carbonitrile

[0043] A mixture of 5-methoxy-l,3-benzothiazole-2-carbonitrile (500 mg, 3 mmol) and pyridine; hydrochloride (10.00 g, 86.53 mmol) was stirred at 180 °C for 1.5 h. Then the reaction mixture was diluted with H2O (10 mL), and then extracted with EtOAc (3 x 10 mL). The combined organic layers were dried over anhydrous NaiSO-i, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (TFA condition; column: Phenomenex luna Cl 8 100 * 40 mm * 3 um; mobile phase: [H2O (0.1% TFA) - ACN]; gradient: 10% - 45% B over 8.0 min) to give the title compound (150 mg, 851 pmol, 32%) as a yellow solid.

[0044] ’H NMR (400 MHz, DMSO-t / s) 8 (ppm) 10.43 - 9.93 (m, 1H), 8.11 (d, J = 8.9 Hz, 1H), 7.51 (d, J = 2.3 Hz, 1H), 7.36 - 7.14 (m, 1H)3Preparation 4 tert-Butyl 3 -(hydroxymethyl)indoline- 1 -carboxylate

[0045] To a mixture of l-tert-butoxycarbonylindoline-3-carboxylic acid (2 g, 8 mmol) in THF (20 mL) was added BH3 THF (1 M, 15.19 mL) dropwise at 0 °C under N2 atmosphere, stirred at 0 °C for 0.5 h. Then the mixture was stirred at 20 °C for 3 h. The reaction mixture was quenched by added MeOH (30 mL) at 0 °C and stirred at 80 °C for 2 h. Then the reaction mixture was diluted with H2O (20 mL), and then extracted with EtOAc (3 x 20 mL). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the title compound (2 g) as a yellow oil.

[0046] ’H NMR (400 MHz, DMSO-t / s) 8 = 7.85 - 7.37 (m, 1H), 7.28 - 7.12 (m, 2H), 6.99 - 6.84 (m, 1H), 5.09 - 4.73 (m, 1H), 3.95 (br t, J = 10.3 Hz, 1H), 3.84 - 3.73 (m, 1H), 3.69 - 3.56 (m, 1H), 3.53 - 3.33 (m, 2H), 1.60 - 1.42 (m, 9H)Preparation 5 Indolin-3-ylmethanol

[0047] To a solution of tert-butyl 3-(hydroxymethyl)indoline-l-carboxylate (300 mg, 1 mmol) in DCM (5 mL) was added TFA (1 mL). The mixture was stirred at 20 °C for 1 h. The reaction mixture was concentrated under reduced pressure to remove solvent and provide the title compound (300 mg, TFA salt) as a yellow oil.

[0048] ESLMS (m / z) = 150 (M+H).

[0049] MW: 149.19Preparation 61 -(3-(Hydroxymethyl)indolin- 1 -yl)prop-2-en- 1 -one

[0050] To a solution of indolin-3-ylmethanol (300 mg, 2 mmol) in DCM (3 mL) was added TEA (610.43 mg, 6.03 mmol, 840 pL) and prop-2-enoyl chloride (182.00 mg, 2.01 mmol, 163 pL) at 0 °C. The mixture was stirred at 0 °C for 1 h. The reaction mixture was concentrated under reduced pressure to remove solvent. The residue was purified by prep-HPLC (TFA condition; column: Phenomenex luna Cl 8 100 * 40mm * 3 um; mobile phase: [H2O (0.1% TFA) - ACN]; gradient: 1% - 35% B over 8.0 min) to give the title compound (190 mg, 935 pmol, 47%) as a yellow oil.

[0051] ESI-MS (m / z) = 204 (M+H).

[0052] MW: 203.24EXAMPLESExample 1 l-Acryloyl-A-(2-cyanobenzo[<7]thiazol-5-yl)indohne-3-carboxamide

[0053] To a solution of 1 -acryloylindoline-3 -carboxylic acid (130 mg, 1 mmol) and 5- aminobenzo[<7]thiazole-2-carbonitrile (88 mg, 1 mmol) in DMF (2 mL) was added HATU (380 mg, 1 mmol) and DIPEA (320.0 mg, 2.5 mmol) at 0 °C. The mixture was stirred at 25 °C for 12 h. The reaction is poured into H2O (15 mL) and the reaction is extracted with EtOAc (3 x 50 mL). The reaction mixture is dried over NaiSO-i, filtered, and concentrated under reduced pressure to give a residue. The residue is purified by prep-HPLC to give the title compound (100 mg, 53%) as a white solid.

[0054] ’H NMR (500 MHz, DMSO- ,) 8 10.93 (s, 1H), 8.65 (d, J = 2.0 Hz, 1H), 8.30 (d, J = 8.9 Hz, 1H), 8.21 (d, J = 8.2 Hz, 1H), 7.89 (dd, J = 8.9, 2.0 Hz, 1H), 7.48 (d, J = 7.5 Hz, 1H), 7.28 (td, J = 7.8, 1.3 Hz, 1H), 7.08 (td, J = 7.5, 1.1 Hz, 1H), 6.84 (dd, J = 16.5, 10.3 Hz, 1H), 6.36 (dd, J = 16.6, 2.3 Hz, 1H), 5.88 (dd, J = 10.3, 2.2 Hz, 1H), 4.59 (dd, J = 9.4, 4.3 Hz, 1H), 4.55 - 4.43 (m, 2H).

[0055] The compounds in the following table were prepared essentially as described in Example1.

[0056] Example 2 10.71 (s, 1H), 8.39 (d, J= 2.1 Hz, 1H),8.20 (d, J= 8.2 Hz, 1H), 7.81 (d, J= 8.8 Hz, 1H), 7.60 (dd, J= 8.8, 2.1 Hz, 1H), 7.45 (d, J= 7.5 Hz, 1H), 7.31 - 7.24 (m, 1H), 7.08 (td, J= 7.5, 1.1 Hz, 1H), 6.83 (dd, J= 16.6, 10.3 Hz, 1H), 6.36 (dd, J = 16.7, 2.2 Hz, 1H), 5.88 (dd, J= 10.3, 2.2 Hz, 1H), 4.58 - 4.52 (m, 1H), 4.51 - 4.41 (m, 2H), 2.78 (s, 3H).

[0057] Example 3: ’H NMR (500 MHz, DMSO-t / s) 8 10.62 (s, 1H), 8.21 (d, J= 8.1 Hz, 1H),8.06 (d, J= 2.0 Hz, 1H), 7.81 (d, J= 8.6 Hz, 1H), 7.51 (dd, J= 8.7, 2.1 Hz, 1H), 7.45 (d, J= 7.5 Hz, 1H), 7.28 (td, J= 7.8, 1.3 Hz, 1H), 7.08 (td, J= 7.5, 1.1 Hz, 1H), 6.84 (dd, J= 16.6, 10.3 Hz, 1H), 6.36 (dd, J= 16.6, 2.2 Hz, 1H), 5.88 (dd, J= 10.3, 2.2 Hz, 1H), 4.58 - 4.51 (m, 1H), 4.51 - 4.42 (m, 2H), 4.15 (s, 3H).Example 4 l-Acryloyl-A-(2-cyanobenzo[<7]thiazol-6-yl)indohne-3-carboxamide

[0058] To a solution of tert-butyl 3-((2-cyanobenzo[<7]thiazol-6-yl)carbamoyl)indoline-l - carboxylate (80 mg, 0.2 mmol) in DCM (2 mL) was added TFA (1 mL). The mixture was warmed to RT and stirred for 0.5 h. The solvent was then evaporated, and the residue was dissolved into DMF (3 mL), followed by a slow addition of DIPEA (120 mg, 1 mmol) and acryloyl chloride (0.57 mL, 0.57 mmol, 1.0 M in DCM) in an ice bath. The reaction was poured into water (15 mL) and the reaction is extracted with EtOAc (3 x 50 mL). The reaction mixture is dried over NaiSO-i, filtered,and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC to give the title compound (30 mg, 42%) as a white solid.

[0059] ’H NMR (500 MHz, DMSO- ,) 8 10.99 (s, 1H), 8.75 (d, J = 2.1 Hz, 1H), 8.24 (d, J = 9.0 Hz, 1H), 8.21 (d, J = 8.1 Hz, 1H), 7.84 (dd, J = 9.0, 2.1 Hz, 1H), 7.46 (d, J = 7.5 Hz, 1H), 7.28 (td, J = 7.8, 1.3 Hz, 1H), 7.08 (td, J = 7.5, 1.1 Hz, 1H), 6.83 (dd, J = 16.6, 10.3 Hz, 1H), 6.36 (d, J = 16.8 Hz, 1H), 5.88 (dd, J = 10.3, 2.2 Hz, 1H), 4.59 - 4.44 (m, 3H).

[0060] The compounds in the following table were prepared essentially as described in Example4.

[0061] Example 5: ’H NMR (500 MHz, DMS 10.70 (s, 1H), 8.24 (d, J= 2.1 Hz, 1H),8.21 (d, J= 8.1 Hz, 1H), 7.95 (d, J= 8.7 Hz, 1H), 7.57 (dd, J= 8.7, 2.1 Hz, 1H), 7.46 (d, J= 7.5 Hz,1H), 7.28 (td, J= 7.8, 1.4 Hz, 1H), 7.08 (td, J= 7.5, 1.1 Hz, 1H), 6.84 (dd, J= 16.6, 10.3 Hz, 1H),6.36 (dd, J= 16.8, 2.3 Hz, 1H), 5.88 (dd, J= 10.3, 2.2 Hz, 1H), 4.59 - 4.54 (m, 1H), 4.47 (d, J= 9.8 Hz, 2H), 2.79 (s, 3H).

[0062] Example 6: ’H NMR (500 MHz, DMSO- ,) 8 9.91 (s, 1H), 8.57 (d, J= 2.0 Hz, 1H), 8.23 (d, J= 8.9 Hz, 1H), 8.19 (d, J= 8.2 Hz, 1H), 7.89 (dd, J= 9.0, 2.1 Hz, 1H), 7.54 (dd, J= 7.5, 1.3 Hz, 1H), 7.29 (td, J= 7.8, 1.3 Hz, 1H), 7.11 (td, J= 7.5, 1.2 Hz, 1H), 6.79 (dd, J= 16.7, 10.3 Hz, 1H), 6.34 (dd, J= 16.6, 2.1 Hz, 1H), 5.87 (dd, J= 10.4, 2.1 Hz, 1H), 4.86 (d, J= 10.8 Hz, 1H), 4.12 (d, J= 10.8 Hz, 1H), 1.71 (s, 3H).

[0063] Example 7: 1H NMR (500 MHz, DMSO-t / g) 8 10.36 (s, 1H), 8.19 (d, J= 8.1 Hz, 1H), 7.43 - 7.36 (m, 2H), 7.33 (dd, J= 8.5, 2.4 Hz, 1H), 7.26 (td, J= 7.8, 1.3 Hz, 1H), 7.07 (td, J= 7.5, 1.1 Hz, 1H), 6.99 (d, J= 8.5 Hz, 1H), 6.81 (dd, J= 16.6, 10.2 Hz, 1H), 6.40 - 6.30 (m, 1H), 5.87 (dd, J= 10.3, 2.2 Hz, 1H), 4.51 (d, J= 6.8 Hz, 1H), 4.41 (d, J= 7.6 Hz, 2H), 3.04 - 2.96 (m, 2H), 2.73 (t, J= 6.1 Hz, 2H), 1.99 (dtd, J= 8.4, 6.1, 4.8 Hz, 2H).Example 85-((l-Acryloylindolin-3-yl)methoxy)benzo[<7]thiazole-2-carbonitrile

[0064] To a solution of 5-hydroxybenzo[<7]thiazole-2-carbonitrile (85 mg, 482 pmol) and l-(3- (hydroxymethyl)indolin-l-yl)prop-2-en-l-one (98.05 mg, 482.42 pmol) in toluene (3 mL) was added PPhi (316.33 mg, 1.21 mmol) and TMAD (207.67 mg, 1.21 mmol). The mixture was stirred at 90 °C for 12 h under N2 atmosphere. The reaction mixture was concentrated under reduced pressure to remove solvent. The residue was purified by prep-HPLC (TFA condition; column: Phenomenex Luna C18 100 * 30mm * 5um; mobile phase: [H2O (0.1% TFA) - ACN]; gradient: 25% - 65% B over 9.0 min) to give the title compound (32.2 mg, 87.5 pmol, 18% yield) as a white solid.

[0065] ESI-MS (m / z) = 362 (M+H)

[0066] MW: 361.42Biological AssaysCK Target Engagement AssayCKB Protein Expression and purification

[0067] The N-terminal His tag construct of human CKB (residues 1 - 381) was overexpressed in Escherichia coli BL21 (DE3) and purified using affinity chromatography and size-exclusion chromatography. Briefly, cells were grown at 37 °C in TB medium in the presence of 50 pg / mL kanamycin to an optical density of 0.8, cooled to 17 °C, collected by centrifugation and stored at -80 °C. Cell pellets were lysed in buffer A (25 mM HEPES, pH 7.5, 500 mM NaCl, 7 mM mercaptoethanol and 20 mM imidazole) using a Microfluidizer (Microfluidics) and the resulting lysate was centrifuged at 30,000 g for 40 min. Ni-NTA beads (Qiagen) were mixed with cleared lysate for 30 min and washed with buffer A. Beads were transferred to a fast protein liquid chromatography-compatible column, and the bound protein was washed further with buffer A for 10 column volumes and eluted with buffer B (25 mM HEPES, ph 7.5, 500 mM NaCl, 7 mM mercaptoethanol, and 400 mM imidazole). The eluted samples were concentrated and purified further using a Superdex 200 16 / 600 column (Cytvia) in buffer C (20 mM HEPES, ph 7.5, 200 mM Nacl, 5% glycerol, 1 mm dithiothreitol, and 0.5 mM TCEP). Fractions containing CKB were concentrated to 50 mg / mL and stored at -80 °C.Intact protein mass spectrometry

[0068] 1 pL of test compound (lOOx stock) was added to 50 pL of 800 nM human recombinant CKB and incubated for 2 h at 37 °C. The sample was then subjected to LC-MS analysis using a PLRP- S 1000 A, 2.1 x 50 mm, 5 pm (Agilent) column on a Q-Exactive HF-X. The buffers used for separation were 2% formic Acid in water (Buffer A) and 2% Formic Acid in acetonitrile (Buffer B) which was run at 0.3 mL / min at 60 °C. The gradient was 15% B for 0.5 mins, followed up by ramping up to 95% over 4.5 mins. Positive ion modes were collected with full scan analysis over m / z 900- 2600 m / z at 7,500 resolution, le5 AGC, 25 ms maximum ion accumulation time and 60 eV in-source CID. Data processing was conducted using Thermo Biopharma Finder software version 4.1 (Thermo Fisher Scientific). Masses were identified demonstrating one (single), two (double), or three (triple) molecules of test compound were bound to CKB protein.

[0069] Exemplified compounds were tested essentially as described above and exhibited the following binding data.

[0070] These data demonstrate that the exemplified compounds label CKB.CK Inhibition Assay

[0071] Recombinant CKB was diluted to 250 nM using 50 mM TRIS, 100 pM TCEP, pH 9, and 19 pL was added to each well of a 384 well plate, lul of compound dissolved in DMSO was added to each well, resulting in 12 different concentrations ranging between 125uM to 0.03nM [each concentration was serially diluted by 1 / 4] then incubated for 2 h at 37 °C. Next, samples were diluted into assay buffer with the following final concentrations: 100 nM CKB, 40 pM TCEP, 3 mM ATP, 1.2 mM PEP, 12 mM MgCh, 210 pM NADH, 60 mM KC1, 50 mM TRIS, 9 umts / mL Pyruvate Kinase, 4.2 units / mL L-Lactate Dehydrogenase, and 6 mM creatine at pH 9. The absorbance was measured at a wavelength of 340 nm over 30 mins. Measured absorbance over time in each well was then subjected to analyze the linear regression. And the slope measured from each well based on the linear regression analysis were used for calculating IC50 by PRISM software based on the following equation:Y=Bottom + (Top-Bottom) / (l+10A((LogIC50-X)*HillSlope))• X: Log of dose or concentration• Y: Response• Top and Bottom: Plateaus in same units as Y, Bottom-Y min, Top- Y max.• LogIC50: same log units as X- Valus of X at Ymid• HillSlope: Slope factor or Hill slope- SIGN(YatMAX-YatMin)

[0072] Exemplified compounds were tested essentially as described above and exhibited the following IC50 for CK:n.d. = not determined

[0073] These data demonstrate that the compounds of Formula I are inhibitors of CK.AML and A549 Inhibition Assay

[0074] UCSD-AML1 or A549 cells were diluted to 20,000cell / ml and then 45 pL of this suspension was introduced into every well of a 384-well plate (1125cell / well). Following a 24-hour incubation period, the compound was first dissolved in DMSO. Subsequently, 2.5 pL of the dissolved compound was mixed with 122.5 pL of culture media. Each concentration was serially diluted by 1 / 2. Then, 5 pL of the culture media, containing the compounds, was added to each well, resulting in 10 different concentrations ranging from 20 pM to 40 nM. After 3 days of incubation, 10 pL of CellTiter-Glo Luminescent Cell Viability Assay was added to each well incubated at RT for 10 min. Luminescence is measured using plate reader. All experiments are performed in 8 biological replicates per condition / titrated concentration. All the data analysis was performed using PRISM. Each luminescent value was normalized to present result as percentages by averaging the subcolumns and normalizing the means. Then LD50 was calculated based on the following equation:Y=100 / (l+10A((LogLD50-X)*HillSlope))• X: log of dose or concentration,• Y: Normalized response, 100% down to 0%,• LogLD50: Same log units as X -Value of X at Ymid• HillSlope: Slope factor or Hill slope- SIGN(YatMAX-YatMin)*ICso was greater than 20 pM. A value of 25 pM was arbitrarily assigned to allow the calculation of the A549 / AML ratio.**CK is essential for AML cell viability but A549 cells are a representative non-CK sensitive cell type. The ratio of A549 IC50 to AML IC50 (A549 / AML) greater than 1 for a compound of Formula I demonstrates engagement of the CK protein.

Claims

CLAIMSWe claim:

1. A compound of Formula I, or a pharmaceutically acceptable salt thereof:where:X is -C(O)NH- or -CH2O-;R1is H or methyl;R2is -CN, -OCH3, or -SCH3.

2. A compound of Claim 1 , or a pharmaceutically acceptable salt thereof, where X is - C(O)NH-.

3. A compound of Claim 1 or Claim 2, or a pharmaceutically acceptable salt thereof, where A is4. A compound of any of Claims 1 - 3, or a pharmaceutically acceptable salt thereof, where R2is -CN or -SCH3.

5. A compound of Claim 4, or a pharmaceutically acceptable salt thereof, where X is -6. A compound of Claim 4, or a pharmaceutically acceptable salt thereof, where X is -7. A compound of Claim 1 , or a pharmaceutically acceptable salt thereof, selected from the group consisting of:

8. A pharmaceutical composition comprising a compound of any of Claims 1 - 7 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

9. A compound of any of Claims 1 - 7 or a pharmaceutically acceptable salt thereof for use in therapy.

10. A method for treating a disorder susceptible to inhibition of CK comprising administering to a patient in need thereof a therapeutically effective amount of a compound of any of Claims 1 - 7 or a pharmaceutically acceptable salt thereof.

11. A method of Claim 10 where the disorder is cancer.

12. A method of Claim 11 where the disorder is liver cancer, pancreatic cancer, lung cancer, colon cancer, and hematological cancers, including myelogenous leukemia, myeloid leukemia (including acute myeloid leukemia), myelodysplastic syndrome, lymphoblastic leukemia (including acute lymphoblastic leukemia), chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), high risk CLL, follicular lymphoma, diffuse large B-cell lymphoma (including DLBCL or ABC-DLBCL), mantle cell lymphoma (MCL), Waldenstrom’s macroglobulinemia (WM), multiple myeloma, marginal zone lymphoma (MZL), Burkitt’s lymphoma, non-Burkitt high grade B-cell lymphoma, extranodal marginal zone B-cell lymphoma, transformed high grade B-cell lymphoma (HGBL), lymphoplasmacytic lymphoma (LPL), central nervous system lymphoma (CNSL), or MALT lymphoma.

13. A method of Claim 12 where the disorder is acute myeloid leukemia.

14. A compound of any of Claims 1 - 7 or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment of disorders susceptible to inhibition of CK.

15. The use of Claim 14 where the disorder is cancer.

16. The use of Claim 15 where the disorder is liver cancer, pancreatic cancer, lung cancer, colon cancer, and hematological cancers, including myelogenous leukemia, myeloid leukemia (including acute myeloid leukemia), myelodysplastic syndrome, lymphoblastic leukemia (including acute lymphoblastic leukemia), chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), high risk CLL, follicular lymphoma, diffuse large B-cell lymphoma (including DLBCL or ABC-DLBCL), mantle cell lymphoma (MCL), Waldenstrom’s macroglobulinemia (WM), multiple myeloma, marginal zone lymphoma (MZL), Burkitt’s lymphoma, non-Burkitt high grade B-cell lymphoma, extranodal marginal zone B-cell lymphoma, transformed high grade B-cell lymphoma(HGBL), lymphoplasmacytic lymphoma (LPL), central nervous system lymphoma (CNSL), or MALT lymphoma.

17. The use of Claim 16 where the disorder is acute myeloid leukemia.

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