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WO2025240240A1 - Nouveaux dérivés de spirocycle en tant qu'inhibiteurs de hck - Google Patents

Nouveaux dérivés de spirocycle en tant qu'inhibiteurs de hck

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Publication number
WO2025240240A1
WO2025240240A1 PCT/US2025/028562 US2025028562W WO2025240240A1 WO 2025240240 A1 WO2025240240 A1 WO 2025240240A1 US 2025028562 W US2025028562 W US 2025028562W WO 2025240240 A1 WO2025240240 A1 WO 2025240240A1
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WO
WIPO (PCT)
Prior art keywords
mmol
pyrazolo
pyrimidin
compound
phenoxyphenyl
Prior art date
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Pending
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PCT/US2025/028562
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English (en)
Inventor
John Michael HATCHER
Steven TREON
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Dana Farber Cancer Institute Inc
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Dana Farber Cancer Institute Inc
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Publication of WO2025240240A1 publication Critical patent/WO2025240240A1/fr
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/10—Spiro-condensed systems

Definitions

  • the present disclosure relates to compounds having activity as against one or more, including two or more, target kinases, as well as methods of using the compounds for inhibiting kinase activity as well as methods for use in the treatment and/or prevention of a disease (e.g., a proliferative disease, such as IgM gammopathy, mastocytosis, and cancer).
  • a disease e.g., a proliferative disease, such as IgM gammopathy, mastocytosis, and cancer.
  • MYD88 Activating mutations in MYD88 are prevalent in Waldenström macroglobulinemia (WM; 95% to 97%), primary central nervous system lymphoma (PCNSL; 60% to 80%), activated B-cell diffuse large B-cell lymphoma (ABC DLBCL; 30% to 40%), chronic lymphocytic leukemia (CLL; 5% to 10%), and marginal zone lymphoma (MZL; 5% to 10%).
  • WM Waldenström macroglobulinemia
  • PCNSL primary central nervous system lymphoma
  • ABSL chronic lymphocytic leukemia
  • MZL marginal zone lymphoma
  • Mutated MYD88 triggers auto-assembly of a myddosome complex that includes Bruton tyrosine kinase (BTK), which triggers downstream NF-kB pro-survival signaling.
  • BTK Bruton tyrosine kinase
  • HCK hematopoietic cell kinase
  • IL-6 interleukin-6
  • One embodiment of the present disclosure includes a compound of Formula (I): or one or more of an enantiomer, tautomer, or pharmaceutically acceptable salt thereof, wherein each of m and n is independently 0 or 1; Ring D 1 is absent (and m is 0) or Ring D 1 selected from the group consisting of cyclohexyl, a 6-membered heterocycle, an 8-10 membered heterobicycle, a 7-11 membered heterospirocycle, and a C 7 -C 11 spirocycle; Ring D 2 is selected from the group consisting of a 4-6 membered heterocycle and a 7-11 membered heterospirocycle; R D1 is oxo, –CH 3 , or –C(O)CH 3 ; and R D2 is oxo, –CH 3 , –C(O)CH 3 , or oxetane.
  • D 1 is a cyclohexane. In one aspect, D 1 is a piperidinyl. In one aspect, D 1 is an 8-10 membered heterobicycle. In one aspect, D 1 is spirocycle. In one aspect, D 1 is 7-11 membered heterospirocycle. In one aspect, D 1 is C 7 -C 11 membered spirocycle. In one aspect, D 1 is In one aspect, R D2 is oxo. In one aspect, D 1 is absent. In one aspect, D 2 is 4-6 membered heterocycle. In one aspect, D 2 is piperazine.
  • D 2 -R D2 is In one aspect, D 2 is a 7-11 membered heterospirocycle. In one aspect, D 2 -R D2 is [0007] In one aspect, R D2 is –CH 3 . In one aspect, R D2 is oxo. [0008] One embodiment of the present disclosure includes a pharmaceutical composition comprising the compound of the present disclosure, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers or vehicles. [0009] One embodiment of the present disclosure includes a method of inhibiting the activity of one or more kinase in a subject comprising administering to the subject the compound of the present disclosure, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising such compound or salt.
  • the one or more kinase is selected from the group consisting an SRC family of cytoplasmic tyrosine kinases (SFKs), a hematopoietic cell kinase (HCK), a LYN proto- oncogene tyrosine kinase (LYN), a Tec family of cytoplasmic tyrosine kinases, and a Bruton’s tyrosine kinase (BTK).
  • the one or more kinase is a mutated kinase.
  • the mutated kinase is one or more of: (i) a mutated BTK, (ii) the BTK is mutated at Cys481, and (iii) the BTK is a C481S mutated BTK.
  • the one or more kinase is resistant to treatment.
  • the BTK is resistant to inhibition by ibrutinib.
  • the subject is resistant to treatment with one or more of ibrutinib, CC-292, ONO-4059, evobrutinib, spebrutinib, BGB-3111, HM71224, and ACP-196, or a pharmaceutically acceptable salt thereof.
  • One embodiment of the present disclosure provides a method of treating a proliferative disease associated with a mutation in a MYD88 protein in a subject in need thereof comprising administering to the subject the compound of the present disclosure, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition comprising such compound or salt.
  • the disease is associated with aberrant activity of one or more of a hematopoietic cell kinase (HCK), of a LYN proto-oncogene tyrosine kinase (LYN), of Bruton’s tyrosine kinase (BTK), or a mutation in a BTK protein.
  • HCK hematopoietic cell kinase
  • LYN LYN proto-oncogene tyrosine kinase
  • BTK Bruton’s tyrosine kinase
  • the disease is associated with a mutation in a BTK protein, wherein the mutated BTK protein is a C481S mutated BTK.
  • the proliferative disease is cancer, IgM gammopathy or mastocytosis.
  • the proliferative disease is cancer, and wherein the cancer is breast cancer, colon cancer, stomach cancer, testicular cancer, cancer of the central nervous system, lymphoma, leukemia, myeloma or myeloproliferative disease.
  • the cancer is lymphoma, and wherein the lymphoma is a B-cell lymphoma.
  • the B-cell lymphoma is lymphoplasmacytic lymphoma, diffuse large B-cell lymphoma (DLBCL), follicular lymphoma, marginal zone B-cell lymphoma or small lymphocytic lymphoma.
  • the lymphoplasmacytic lymphoma is IgM secreting lymphoplasmacytic lymphoma, Waldenström’s macroglobulinemia, or non-IgM secreting lymphoplasmacytic lymphoma.
  • the diffuse large B-cell lymphoma is activated B- cell-like (ABC-DLBCL), or germinal center B-cell-like (GBC-DLBCL).
  • the small lymphocytic lymphoma is mantle cell lymphoma.
  • the cancer is leukemia, and wherein the leukemia is chronic lymphocytic leukemia, or myelogenous leukemia.
  • the myelogenous leukemia is chronic myelogenous leukemia, or acute myelogenous leukemia.
  • the acute myelogenous leukemia is mast cell leukemia.
  • the cancer is a myeloma, and wherein the myeloma is an IgM myeloma. In one aspect, the IgM myeloma is IgM multiple myeloma.
  • the cancer is a myeloproliferative disease, and wherein the myeloproliferative disease is myelodysplastic syndrome.
  • the proliferative disease is an IgM gammopathy and wherein the IgM gammopathy is an IgM Monoclonal gammopathy of undetermined significance (MGUS) or amyloid light chain (AL) amyloidosis.
  • the proliferative disease is mastocytosis and wherein the mastocytosis is systemic mastocytosis.
  • the subject is treated with one or more additional therapeutic agents, or oncologic treatments administered concurrently with, prior to, or subsequent to treatment with the compound, pharmaceutically acceptable salt, or pharmaceutical composition.
  • One embodiment of the present disclosure provides a compound according to the present disclosure, for use in inhibiting the activity of one or more kinase in a subject in need thereof.
  • One embodiment of the present disclosure provides a compound according to the present disclosure, for the manufacture of a medicament for the inhibition of the activity of one or more kinase in a subject in need thereof.
  • One embodiment of the present disclosure provides the compound of the present disclosure or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition thereof, for use in a method of treatment, e.g., using the methods described herein.
  • One embodiment of the present disclosure includes a process for preparing a compound of the present disclosure.
  • One embodiment of the present disclosure includes a compound obtained by a process of the present disclosure.
  • One or more aspects and embodiments may be incorporated in a different embodiment although not specifically described. That is, all aspects and embodiments may be combined in any way or combination to form a new embodiment or aspect.
  • DETAILED DESCRIPTION OF THE DISCLOSURE [0020] The present disclosure highlights the development and characterization of novel and selective kinase inhibitors of HCK and BTK.
  • a compound as herein disclosed and methods of using the compounds for inhibiting kinase activity as well as methods for use in the treatment and/or prevention of a disease (e.g., a proliferative disease, such as IgM gammopathy, mastocytosis, cancer) in a subject in need thereof as described herein.
  • a disease e.g., a proliferative disease, such as IgM gammopathy, mastocytosis, cancer
  • a disease e.g., a proliferative disease, such as IgM gammopathy, mastocytosis, cancer
  • Compounds described herein can comprise one or more asymmetric centers, and thus can exist in various stereoisomeric forms, e.g., enantiomers and/or diastereomers.
  • the compounds described herein can be in the form of an individual enantiomer, diastereomer or geometric isomer, or can be in the form of a mixture of stereoisomers, including racemic mixtures and mixtures enriched in one or more stereoisomer.
  • Isomers can be isolated from mixtures by methods known to those skilled in the art, including chiral high pressure liquid chromatography (HPLC) and the formation and crystallization of chiral salts; or preferred isomers can be prepared by asymmetric syntheses.
  • HPLC high pressure liquid chromatography
  • alkyl refers to a straight–chain or branched saturated hydrocarbon group having from 1 to 20 carbon atoms (“C 1–20 alkyl”). Unless otherwise specified, each instance of an alkyl group is independently unsubstituted (an “unsubstituted alkyl”) or substituted (a “substituted alkyl”) with one or more substituents (e.g., halogen, such as F).
  • cycloalkyl refers to a non–aromatic cyclic hydrocarbon group having from 3 to 10 ring carbon atoms (“C 3–10 cycloalkyl”).
  • the cycloalkyl group is either monocyclic (“monocyclic cycloalkyl”) or contain a fused, bridged or spiro ring system such as a bicyclic system (“bicyclic cycloalkyl”) and can be saturated or can be partially unsaturated.
  • heterocyclyl refers to a 3– to 10–membered non–aromatic ring system having ring carbon atoms and 1 to 4 ring heteroatoms, wherein each heteroatom is independently selected from the group consisting of nitrogen, oxygen, sulfur, boron, phosphorus, and silicon (“3–10 membered heterocyclyl”).
  • the point of attachment can be a carbon or nitrogen atom, as valency permits.
  • a heterocyclyl group can either be monocyclic (“monocyclic heterocyclyl”) or a fused, bridged or spiro ring system such as a bicyclic system (“bicyclic heterocyclyl”), and can be saturated or can be partially unsaturated.
  • Heterocyclyl bicyclic ring systems can include one or more heteroatoms in one or both rings.
  • the term “unsaturated” or “partially unsaturated” refers to a moiety that includes at least one double or triple bond.
  • a “partially unsaturated” ring system is further intended to encompass rings having multiple sites of unsaturation but is not intended to include aromatic groups (e.g., aryl or heteroaryl groups) as defined herein.
  • saturated refers to a group that does not contain a double or triple bond, i.e., contains all single bonds.
  • Halo or “halogen” refers to fluorine (fluoro, –F), chlorine (chloro, –Cl), bromine (bromo, –Br), or iodine (iodo, –I).
  • Alkoxy or “alkoxyl” refers to a functional group of the formula: –O–alkyl.
  • pharmaceutically acceptable salt refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.
  • Pharmaceutically acceptable salts are well known in the art. For example, Berge et al., describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66, 1–19, incorporated herein by reference.
  • solvate refers to forms of the compound, or a salt thereof, which are associated with a solvent, usually by a solvolysis reaction. This physical association may include hydrogen bonding.
  • Compound of the disclosure may be prepared, e.g., in crystalline form, and may be solvated. Suitable solvates include pharmaceutically acceptable solvates and further include both stoichiometric solvates and non-stoichiometric solvates.
  • hydrate refers to a compound that is associated with water. Typically, the number of the water molecules contained in a hydrate of a compound is in a definite ratio to the number of the compound molecules in the hydrate. Therefore, a hydrate of a compound may be represented, for example, by the general formula R ⁇ x H 2 O, wherein R is the compound and wherein x is a number greater than 0.
  • tautomers refer to compounds that are interchangeable forms of a particular compound structure, and that vary in the displacement of hydrogen atoms and electrons. Thus, two structures may be in equilibrium through the movement of ⁇ electrons and an atom (usually H). For example, enols and ketones are tautomers because they are rapidly interconverted by treatment with either acid or base. Another example of tautomerism is the aci- and nitro- forms of phenylnitromethane, which are likewise formed by treatment with acid or base.
  • An enantiomer can be characterized by the absolute configuration of its asymmetric center and is described by the R- and S-sequencing rules of Cahn and Prelog, or by the manner in which the molecule rotates the plane of polarized light and designated as dextrorotatory or levorotatory (i.e., as (+) or (-)-isomers respectively).
  • a chiral compound can exist as either individual enantiomer or as a mixture thereof.
  • a mixture containing equal proportions of the enantiomers is called a “racemic mixture.”
  • the term “prodrugs” refer to compounds, including derivatives of compounds of the disclosure, which have cleavable groups and become by solvolysis or under physiological conditions are pharmaceutically active in vivo.
  • Such examples include, but are not limited to, ester derivatives and the like.
  • Other derivatives of the compounds of this disclosure have activity in both their acid and acid derivative forms, but in the acid sensitive form often offers advantages of solubility, tissue compatibility, or delayed release in the mammalian organism (see, Bundgard, H., Design of Prodrugs, pp.7-9, 21-24, Elsevier, Amsterdam 1985).
  • a “subject” to which administration is contemplated includes, but is not limited to, humans and/or other non–human animals, for example, mammals (e.g., primates (e.g., cynomolgus monkeys, rhesus monkeys); commercially relevant mammals such as cattle, pigs, horses, sheep, goats, cats, and/or dogs) and birds (e.g., commercially relevant birds such as chickens, ducks, geese, and/or turkeys).
  • the animal is a mammal.
  • the animal may be a male or female and at any stage of development.
  • a non–human animal may be a transgenic animal.
  • a subject who is resistant to treatment with a BTK inhibitor is one who shows no or minimal response to the treatment.
  • response to a treatment is measured by reduction in tumor cells or tumor cell killing.
  • response to a treatment is measured by changes in symptoms of the disease, condition or malignancy (e.g., a proliferative disease). It has been discovered that the compounds that block ATP binding to HCK as described herein are able to cause tumor cell killing even in cells that are derived from subjects who are resistant to a BTK inhibitor treatment.
  • the terms “administer,” “administering,” or “administration,” refers to implanting, absorbing, ingesting, injecting, inhaling, or otherwise introducing a compound, or a pharmaceutical composition thereof.
  • treatment refers to reversing, alleviating, delaying the onset of, or inhibiting the progress of a “pathological condition” (e.g., a disease, disorder, or condition, or one or more signs or symptoms thereof) described herein.
  • pathological condition e.g., a disease, disorder, or condition, or one or more signs or symptoms thereof
  • treatment may be administered after one or more signs or symptoms have developed or have been observed.
  • treatment may be administered in the absence of signs or symptoms of the disease or condition.
  • treatment may be administered to a susceptible individual prior to the onset of symptoms (e.g., in light of a history of symptoms and/or in light of genetic or other susceptibility factors). Treatment may also be continued after symptoms have resolved, for example, to delay or prevent recurrence.
  • the terms “condition,” “disease,” and “disorder” are used interchangeably.
  • the treatment may be therapeutic treatment (not including prevention or prophylactic treatment).
  • An “effective amount” of Compounds of the disclosure refers to an amount sufficient to elicit the desired biological response, i.e., treating the condition.
  • the effective amount of Compounds of the disclosure may vary depending on such factors as the desired biological endpoint, the pharmacokinetics of the compound, the condition being treated, the mode of administration, and the age and health of the subject.
  • An effective amount encompasses therapeutic and prophylactic treatment.
  • an effective amount of a compound may reduce the tumor burden or stop the growth or spread of a tumor.
  • a “therapeutically effective amount” of Compounds of the disclosure is an amount sufficient to provide a therapeutic benefit in the treatment of a condition or to delay or minimize one or more symptoms associated with the condition.
  • a therapeutically effective amount of a compound means an amount of therapeutic agent, alone or in combination with other therapies, which provides a therapeutic benefit in the treatment of the condition.
  • the term “therapeutically effective amount” can encompass an amount that improves overall therapy, reduces, or avoids symptoms or causes of the condition, or enhances the therapeutic efficacy of another therapeutic agent.
  • a “proliferative disease” refers to a disease that occurs due to abnormal growth or extension by the multiplication of cells (Walker, Cambridge Dictionary of Biology; Cambridge University Press: Cambridge, UK, 1990).
  • a proliferative disease may be associated with: 1) the pathological proliferation of normally quiescent cells; 2) the pathological migration of cells from their normal location (e.g., metastasis of neoplastic cells); 3) the pathological expression of proteolytic enzymes such as the matrix metalloproteinases (e.g., collagenases, gelatinases, and elastases); or 4) the pathological angiogenesis as in proliferative retinopathy and tumor metastasis.
  • proteolytic enzymes such as the matrix metalloproteinases (e.g., collagenases, gelatinases, and elastases)
  • the pathological angiogenesis as in proliferative retinopathy and tumor metastasis.
  • Exemplary proliferative diseases include cancers (e.g., breast cancer, colon cancer, testicular cancer, CNS cancer, stomach cancer, lymphoma (e.g., B-cell Lymphoma (e.g., lymphoplasmacytic lymphoma (e.g., IgM secreting (i.e., Waldenström’s Macroglobulinemia), non-IgM secreting)), Diffuse Large B-Cell Lymphoma (e.g., activated B-cell-like (ABC)-DLBCL, germinal center B- cell-like (GBC)-DLBCL)), Follicular Lymphoma, Marginal zone B-cell lymphoma, Small lymphocytic lymphoma (e.g.
  • B-cell Lymphoma e.g., lymphoplasmacytic lymphoma (e.g., IgM secreting (i.e., Waldenström’s Macroglobulinemia), non-IgM secreting)
  • Chronic lymphocytic leukemia CLL
  • Mantle cell lymphoma Leukemia
  • Leukemia e.g., myelogenous leukemia (e.g., chronic myelogenous leukemia, acute myelogenous leukemia)
  • benign neoplasms angiogenesis, inflammatory diseases, auto inflammatory diseases, and autoimmune diseases.
  • neoplasm and “tumor” are used interchangeably and refer to an abnormal mass of tissue wherein the growth of the mass surpasses and is not coordinated with the growth of a normal tissue.
  • a neoplasm or tumor may be “benign” or “malignant,” depending on the following characteristics: degree of cellular differentiation (including morphology and functionality), rate of growth, local invasion, and metastasis.
  • a “benign neoplasm” is generally well differentiated, has characteristically slower growth than a malignant neoplasm, and remains localized to the site of origin.
  • a benign neoplasm does not have the capacity to infiltrate, invade, or metastasize to distant sites.
  • Exemplary benign neoplasms include, but are not limited to, lipoma, chondroma, adenomas, acrochordon, senile angiomas, seborrheic keratoses, lentigos, and sebaceous hyperplasias.
  • certain “benign” tumors may later give rise to malignant neoplasms, which may result from additional genetic changes in a subpopulation of the tumor’s neoplastic cells, and these tumors are referred to as “pre-malignant neoplasms.”
  • An exemplary pre-malignant neoplasm is a teratoma.
  • a malignant neoplasm is generally poorly differentiated (anaplasia) and has characteristically rapid growth accompanied by progressive infiltration, invasion, and destruction of the surrounding tissue. Furthermore, a malignant neoplasm generally has the capacity to metastasize to distant sites.
  • the term “metastasis,” “metastatic,” or “metastasize” refers to the spread or migration of cancerous cells from a primary or original tumor to another organ or tissue and is typically identifiable by the presence of a “secondary tumor” or “secondary cell mass” of the tissue type of the primary or original tumor and not of that of the organ or tissue in which the secondary (metastatic) tumor is located.
  • a prostate cancer that has migrated to bone is said to be metastasized prostate cancer and includes cancerous prostate cancer cells growing in bone tissue.
  • cancer refers to a malignant neoplasm (Stedman’s Medical Dictionary, 25th ed.; Hensyl ed.; Williams & Wilkins: Philadelphia, 1990).
  • the cancer may be a solid tumor.
  • the cancer may be a hematological malignancy.
  • Exemplary cancers include, but are not limited to, acoustic neuroma; adenocarcinoma; adrenal gland cancer; anal cancer; angiosarcoma (e.g., lymphangiosarcoma, lymphangioendotheliosarcoma, hemangiosarcoma); appendix cancer; benign monoclonal gammopathy; biliary cancer (e.g., cholangiocarcinoma); bladder cancer; breast cancer (e.g., adenocarcinoma of the breast, papillary carcinoma of the breast, mammary cancer, medullary carcinoma of the breast); brain cancer (e.g., meningioma, glioblastomas, glioma (e.g., astrocytoma, oligodendroglioma), medulloblastoma); bronchus cancer; carcinoid tumor; cervical cancer (e.g., cervical adenocarcinoma); choriocar
  • Wilms tumor, renal cell carcinoma); liver cancer (e.g., hepatocellular cancer (HCC), malignant hepatoma); lung cancer (e.g., bronchogenic carcinoma, small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), adenocarcinoma of the lung); leiomyosarcoma (LMS); mastocytosis (e.g., systemic mastocytosis); muscle cancer; myelodysplastic syndrome (MDS); mesothelioma; myeloproliferative disorder (MPD) (e.g., polycythemia vera (PV), essential thrombocytosis (ET), agnogenic myeloid metaplasia (AMM) a.k.a.
  • HCC hepatocellular cancer
  • lung cancer e.g., bronchogenic carcinoma, small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), adenocarcinoma of the lung
  • myelofibrosis MF
  • chronic idiopathic myelofibrosis chronic myelocytic leukemia (CML), chronic neutrophilic leukemia (CNL), hypereosinophilic syndrome (HES)
  • neuroblastoma e.g., neurofibromatosis (NF) type 1 or type 2, schwannomatosis
  • neuroendocrine cancer e.g., gastroenteropancreatic neuroendocrinetumor (GEP-NET), carcinoid tumor
  • osteosarcoma e.g., bone cancer
  • ovarian cancer e.g., cystadenocarcinoma, ovarian embryonal carcinoma, ovarian adenocarcinoma
  • papillary adenocarcinoma pancreatic cancer
  • pancreatic cancer e.g., pancreatic andenocarcinoma, intraductal papillary mucinous neoplasm (IPMN), Islet cell tumors
  • angiogenesis refers to the formation and the growth of new blood vessels. Normal angiogenesis occurs in the healthy body of a subject for healing wounds and for restoring blood flow to tissues after injury.
  • the healthy body controls angiogenesis through a number of means, e.g., angiogenesis-stimulating growth factors and angiogenesis inhibitors.
  • Many disease states such as cancer, diabetic blindness, age-related macular degeneration, rheumatoid arthritis, and psoriasis, are characterized by abnormal (i.e., increased or excessive) angiogenesis.
  • Abnormal or pathological angiogenesis refers to angiogenesis greater than that in a normal body, especially angiogenesis in an adult not related to normal angiogenesis (e.g., menstruation or wound healing).
  • Abnormal angiogenesis can provide new blood vessels that feed diseased tissues and/or destroy normal tissues, and in the case of cancer, the new vessels can allow tumor cells to escape into the circulation and lodge in other organs (tumor metastases).
  • the angiogenesis is pathological angiogenesis.
  • An “autoimmune disease” refers to a disease arising from an inappropriate immune response of the body of a subject against substances and tissues normally present in the body. In other words, the immune system mistakes some part of the body as a pathogen and attacks its own cells.
  • autoimmune thyroiditis This may be restricted to certain organs (e.g., in autoimmune thyroiditis) or involve a particular tissue in different places (e.g., Goodpasture’s disease which may affect the basement membrane in both the lung and kidney).
  • the treatment of autoimmune diseases is typically with immunosuppression, e.g., medications which decrease the immune response.
  • Exemplary autoimmune diseases include, but are not limited to, glomerulonephritis, Goodpasture’s syndrome, necrotizing vasculitis, lymphadenitis, peri-arteritis nodosa, systemic lupus erythematosis, rheumatoid arthritis, psoriatic arthritis, systemic lupus erythematosis, psoriasis, ulcerative colitis, systemic sclerosis, dermatomyositis/polymyositis, anti-phospholipid antibody syndrome, scleroderma, pemphigus vulgaris, ANCA-associated vasculitis (e.g., Wegener’s granulomatosis, microscopic polyangiitis), uveitis, Sjogren’s syndrome, Crohn’s disease, Reiter’s syndrome, ankylosing spondylitis, Lyme disease, Guillain-Barré syndrome, Hashimoto’s thyroiditis, and cardio
  • inflammatory disease refers to a disease caused by, resulting from, or resulting in inflammation.
  • inflammatory disease may also refer to a dysregulated inflammatory reaction that causes an exaggerated response by macrophages, granulocytes, and/or T-lymphocytes leading to abnormal tissue damage and/or cell death.
  • An inflammatory disease can be either an acute or chronic inflammatory condition and can result from infections or non- infectious causes.
  • Inflammatory diseases include atherosclerosis, arteriosclerosis, autoimmune disorders, multiple sclerosis, systemic lupus erythematosus, polymyalgia rheumatica (PMR), gouty arthritis, degenerative arthritis, tendonitis, bursitis, psoriasis, cystic fibrosis, arthrosteitis, rheumatoid arthritis, inflammatory arthritis, Sjogren’s syndrome, giant cell arteritis, progressive systemic sclerosis (scleroderma), ankylosing spondylitis, polymyositis, dermatomyositis, pemphigus, pemphigoid, diabetes (e.g., Type I), myasthenia gravis, Hashimoto’s thyroiditis, Graves’ disease, Goodpasture’s disease, mixed connective tissue disease, sclerosing cholangitis, inflammatory bowel disease, Crohn’s disease, ulcerative colitis, pernicious anemia
  • kinase refers to any enzyme that catalyzes the addition of phosphate groups to an amino acid residue of a protein.
  • a serine kinase catalyzes the addition of a phosphate group to serine residue in a protein.
  • the kinase is a protein kinase.
  • kinases include, but are not limited to, cytoplasmic tyrosine kinases (e.g., SRC family kinases (e.g., HCK, LYN, BLK, FRK), Tec family kinases (e.g., BTK)), a cyclin- dependent kinase (CDK, e.g., CDK1, CDK2, CDK2, CDK4, CDK5, CDK7, CDK8, CDK9, CDK10, CDK11, CDK12, CDK13, CDK14, CDK16, CDK20)), a mitogen-activated protein kinase (MAPK, e.g., MAPK1 , MAPK3 , MAPK4 , MAPK6 , MAPK7 , MAPK8 , MAPK9 , MAPK10 , MAPK11 , MAPK12 , MAPK13 , MAPK14 , MAPK15), a glycogen synthase kinase 3
  • HCK Hematopoietic cell kinase
  • IL6 interleukin 6
  • GP130 IL6 co-receptor IL6ST
  • Bruton’s tyrosine kinase is a member of the src-related BTK/Tec family of cytoplasmic tyrosine kinases, is required for B cell receptor signaling, plays a key role in B-cell maturation, and exhibits increased activation in a number of B-cell malignancies.
  • LYN proto-oncogene is a member of the src-family of protein tyrosine kinases, plays an important role in the regulation of B-cell differentiation, proliferation, survival and apoptosis, is important for immune self-tolerance, and acts downstream of several immune receptors, including the B-cell receptor (BCR).
  • BCR signaling is thought to be involved in pro-growth and survival signaling in MYD88 mutated disease, as well as being involved in non-MYD88 mutated disease.
  • BCR signaling is thought to be active in Waldenström’s Macroglobulinemia, ABC subtype of diffuse large B-cell lymphoma, and chronic lymphocytic leukemia.
  • Proto-oncogene tyrosine-protein kinase SRC is a protein tyrosine kinase, plays a central role in the regulation of a variety of biological processes, such as cell proliferation, migration, adhesion, and survival in solid tumors, and is overexpressed in Waldenström’s Macroglobulinemia.
  • inhibitors refer to the ability of a compound to reduce, slow, halt, block, or prevent activity of a particular biological process (e.g., a kinase (e.g., SFK (e.g., HCK, LYN, BLK, FRK), Tec family kinases (e.g., BTK)) in a cell relative to vehicle.
  • a kinase e.g., SFK (e.g., HCK, LYN, BLK, FRK), Tec family kinases (e.g., BTK)
  • block or blocking refer to the ability of a compound to prevent a biological interaction (e.g., binding) in a cell relative to a negative control, e.g., vehicle.
  • a compound can block ATP from binding to the ATP binding pocket of a kinase. Such blocking may occur by direct binding of the compound to the ATP binding pocket itself, or indirect blocking.
  • the term refers to a reduction in the level of binding of ATP to a kinase, e.g., BTK and/or HCK, and/or LYN, and/or SRC, to a level that is statistically significantly lower than an initial level, which may, for example, be a baseline level of ATP binding.
  • the term refers to a reduction in the level of ATP binding to a kinase, e.g., BTK and/or HCK, and/or LYN, and/or SRC, to a level that is less than 75%, less than 50%, less than 40%, less than 30%, less than 25%, less than 20%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, less than 1%, less than 0.5%, less than 0.1%, less than 0.01%, less than 0.001%, or less than 0.0001% of an initial level, which may, for example, be a baseline level of ATP binding.
  • a kinase e.g., BTK and/or HCK, and/or LYN, and/or SRC
  • blocking ATP binding leads to a reduction in the level of enzyme activity, e.g., BTK and/or HCK, and/or LYN, and/or SRC activity, to a level that is less than 75%, less than 50%, less than 40%, less than 30%, less than 25%, less than 20%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, less than 1%, less than 0.5%, less than 0.1%, less than 0.01%, less than 0.001%, or less than 0.0001% of an initial level, which may, for example, be a baseline level of enzyme activity.
  • enzyme activity e.g., BTK and/or HCK, and/or LYN, and/or SRC activity
  • a compound or pharmaceutical composition When a compound or pharmaceutical composition is referred to as “selectively,” “specifically,” or “competitively” binding a first protein, the compound binds the first protein, e.g., BTK or HCK or LYN or SRC, with a higher binding affinity (e.g., not less than about 2-fold, not less than about 5-fold, not less than about 10-fold, not less than about 30-fold, not less than about 100-fold, not less than about 1,000-fold, or not less than about 10,000-fold) than binding a second protein that is different from the first protein, e.g., BTK.
  • a higher binding affinity e.g., not less than about 2-fold, not less than about 5-fold, not less than about 10-fold, not less than about 30-fold, not less than about 100-fold, not less than about 1,000-fold, or not less than about 10,000-fold
  • a compound blocks ATP binding to a first protein, e.g., HCK or LYN or SRC, at a lower concentration (e.g., not less than about 10-fold) than it blocks ATP binding a second protein that is different from the first protein, e.g., BTK.
  • a first protein e.g., HCK or LYN or SRC
  • a second protein that is different from the first protein, e.g., BTK.
  • Compounds which selectively block ATP binding to a kinase e.g., BTK, HCK, LYN
  • BTK kinase
  • Methods include purified enzyme and cell based biochemical and binding assays such as an HCK gatekeeper mutant rescue assay, an in vitro kinase assay, e.g., using HCK gatekeeper mutated kinase, competitive binding assays using KiNativTM technology or biotin tagged inhibitors, e.g., HCK inhibitors.
  • Suitable assays for determining selective inhibition of HCK by a compound include, but are not limited to, Life Technology Z-Lyte activity assays (e.g., including HCK gatekeeper mutants and GK + 6 mutants); DiscoverX KINOMEscan ® binding assays; MRC radioactivity assays; ACD Ba/F3 viability assays (e.g., including HCK gatekeeper mutants and GK + 6 mutants); Yeast hybrid proliferation assays; Protein thermostability assays; and cancer cells with HCK gatekeeper mutants or GK + 6 mutants proliferation-rescue assays. Such assays can also be used to determine selective inhibition of LYN and/or SRC by a compound.
  • Life Technology Z-Lyte activity assays e.g., including HCK gatekeeper mutants and GK + 6 mutants
  • DiscoverX KINOMEscan ® binding assays MRC radioactivity assays
  • ACD Ba/F3 viability assays e
  • MYD88 mutation means any change or difference in the nucleic acid or protein sequence of MYD88 as compared to the wild type sequence that results in the activation of MYD88 which leads to the activation of NF- ⁇ B. Mutations include, but are not limited to, nonsense mutations, missense mutations, frameshift mutations, rearrangement mutations, insertion mutations and deletion mutations.
  • the mutation is a somatic mutation at position 38182641 in chromosome 3p22.2 which results in a single nucleotide change from T ⁇ C in the myeloid differentiation primary response (MYD88) gene, and a predicted non-synonymous change at amino acid position 265 from leucine to proline (L265P).
  • the mutation is another activating mutation in MYD88, such as V217F, W218R, I220T, S222R, M232T, S243N, T294P.
  • MYD88 mutated disease or “disease associated with mutated MYD88” means any disease in a subject that is related to a change or difference in the nucleic acid or protein sequence of MYD88 as compared to the wild type sequence that results in the activation of MYD88 which leads to the activation of NF- ⁇ B.
  • mutated MYD88 is associated with Waldenström’s Macroglobulinemia (IgM secreting lymphoplasmacytic lymphoma), non-IgM secreting lymphoplasmacytic lymphoma, ABC subtype of diffuse large B-cell lymphoma, primary central nervous system (CNS) lymphoma, immune privileged lymphomas that include testicular lymphoma, marginal zone lymphoma, and chronic lymphocytic leukemia.
  • mutated MYD88 is associated with susceptibility to infectious disease.
  • mutated MYD88 is associated with susceptibility to autoimmune disease.
  • One embodiment of the present disclosure relates to a compound of Formula (I) or a pharmaceutically acceptable salt thereof: wherein Ring D 1 is absent, a C6 cycloalkyl, a 6 membered heterocycle, an 8-10 membered heterobicycle, a 7-11 membered heterospirocycle, or a C 7 -C 11 spirocycle; Ring D 2 is a 4-6 membered heterocycle or 7-11 membered heterospirocycle; and R D2 is absent, oxo2, –CH 3 , –C(O)CH 3 , or oxetane.
  • D 1 is a C 6 cycloalkyl. In some embodiments, D 1 is a cyclohexane. [0065] In some embodiments, D 1 is a 6 membered heterocycle. In some embodiments, D 1 is a piperidinyl. [0066] In some embodiments, D 1 is an 8-10 membered heterobicycle. [0067] In some embodiments, D 1 is: [0068] In some embodiments, D 1 is a spirocycle. In some embodiments, D 1 is 7-11 membered heterospirocycle. In some embodiments, D1 is, [0069] In some embodiments, D 1 is C 7 -C 11 membered spirocycle.
  • D 1 [0070] In some embodiments, D 1 is absent. [0071] In some embodiments, D 2 is a 4-6 membered heterocycle. In some embodiments, D 2 is piperazine. In some embodiments, D 2 -R D2 is [0072] In some embodiments, D 2 is a 7-11 membered heterospirocycle. In some embodiments, D 2 -R D2 is [0073] In some embodiments, R D2 is absent. In some embodiments, R D2 is –CH 3 . In some embodiments, R D2 is oxo. In some embodiments, R D2 is oxetane. In some embodiments, R D2 is – C(O)CH 3 .
  • D 1 when D 1 is 2 D is a 4-6 membered heterocycle. [0075] In some embodiments, when D 1 is 2 D is a 7-11 membered heterospirocycle. [0076] In some embodiments, when D 1 is 2 D is a 4-6 membered heterocycle. [0077] In some embodiments, when D 1 is D 2 is a 4-6 membered heterocycle. [0078] In some embodiments, when D1 is D2 is a 4-6 membered heterocycle. [0079] In some embodiments, when D 1 is absent, D 2 is a 7-11 membered heterospirocycle.
  • D 1 when D 1 is cyclohexane, D 2 is a 7-11 membered heterospirocycle.
  • D 1 when D 1 is piperidinyl, D 2 is a 7-11 membered heterospirocycle.
  • D 1 when D 1 is D 2 is a 7-11 membered heterospirocycle or piperazine.
  • One embodiment of the present disclosure includes a compound of Formula (I) or a pharmaceutically acceptable salt thereof.
  • the disclosure relates to a compound of Formula (I) in a non-salt form.
  • the disclosure relates to a compound of Formula (I) is a salt form.
  • the disclosure relates to a compound selected from Table A, or a pharmaceutically acceptable salt thereof. In other embodiments, the disclosure relates to a compound selected from Table A, i.e., the compound in non-salt form. [0084] Table A:
  • the disclosure relates to a compound of formula
  • the disclosure relates to the foregoing compound in non-salt form.
  • Such compound is considered to be a “compound of the disclosure,” as that term is used herein.
  • the disclosure relates to a compound of formula or a pharmaceutically acceptable salt thereof. In other embodiments, the disclosure relates to the foregoing compound in non-salt form. Such compound is considered to be a “compound of the disclosure,” as that term is used herein.
  • the disclosure relates to a compound of formula
  • the disclosure relates to the foregoing compound in non-salt form. Such compound is considered to be a “compound of the disclosure,” as that term is used herein.
  • the disclosure relates to a compound of formula or a pharmaceutically acceptable salt thereof.
  • the disclosure relates to the foregoing compound in non-salt form. Such compound is considered to be a “compound of the disclosure,” as that term is used herein.
  • the disclosure relates to a compound of formula
  • the disclosure relates to the foregoing compound in non-salt form. Such compound is considered to be a “compound of the disclosure,” as that term is used herein.
  • the disclosure relates to a compound of formula or a pharmaceutically acceptable salt thereof.
  • the disclosure relates to the foregoing compound in non-salt form. Such compound is considered to be a “compound of the disclosure,” as that term is used herein.
  • the disclosure relates to radiolabeled analogs of the compounds of the disclosure.
  • the term “radiolabeled analogs of the compounds of the disclosure” refers to compounds that are identical to the compounds of the disclosure, as described herein, including all embodiments thereof, except that one or more atoms has been replaced with a radioisotope of the atom present in the compounds of the disclosure.
  • the term “radioisotope” refers to an isotope of an element that is known to undergo spontaneous radioactive decay. Examples of radioisotopes include 3 H, 14 C, 35 S, 18 F, 36 Cl, and the like, as well as the isotopes for which a decay mode is identified in V.S. Shirley & C.M.
  • the disclosure relates to pharmaceutically acceptable salts of the radiolabeled analogs, in accordance with any of the embodiments described herein in connection with the compounds of the disclosure.
  • the disclosure relates to pharmaceutical compositions comprising the radiolabeled analogs, or pharmaceutically acceptable salts thereof, and a pharmaceutically acceptable carrier, adjuvant or vehicle, in accordance with any of the embodiments described herein in connection with the compounds of the disclosure.
  • the disclosure relates to radiolabeled analogs, pharmaceutically acceptable salts thereof, and pharmaceutical compositions thereof, for use, in accordance with any of the embodiments described herein in connection with the compounds of the disclosure.
  • Salts, Compositions, Uses, Formulations, Administration, and Additional Agents Pharmaceutically acceptable salts and compositions [0096]
  • the disclosure provides compounds, and pharmaceutically acceptable salts thereof, that inhibit the activity of one or more kinases, and thus the present compounds, and pharmaceutically acceptable salts thereof, are useful for the treatment of diseases, disorders, and conditions including those described herein.
  • compositions comprising a compound as described herein, or a pharmaceutically acceptable salt thereof, and optionally comprise a pharmaceutically acceptable carrier, adjuvant or vehicle.
  • these compositions optionally further comprise one or more additional therapeutic agents.
  • Pharmaceutical preparations and compounds are administered to a subject by any suitable route.
  • compositions can be administered orally, including sublingually, rectally, parenterally, intracisternally, intravaginally, intraperitoneally, topically and transdermally (as by powders, ointments, or drops), bucally, or nasally.
  • the pharmaceutical preparations of the present disclosure may include or be diluted into a pharmaceutically-acceptable carrier.
  • pharmaceutically-acceptable carrier means one or more compatible fillers, diluants or other such substances, which are suitable for administration to a human or other mammal such as a dog, cat, or horse.
  • carrier denotes an organic or inorganic ingredient, natural or synthetic, with which the active ingredient is combined to facilitate the application.
  • the carriers are capable of being commingled with the preparations of the present disclosure, and with each other, in a manner such that there is no interaction which would substantially impair the desired pharmaceutical efficacy or stability.
  • Carriers suitable for oral, subcutaneous, intravenous, intramuscular, etc. formulations can be found in Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, Pa.
  • the present disclosure also provides pharmaceutical compositions comprising a compound described herein and optionally a pharmaceutically acceptable excipient.
  • the disclosure features a pharmaceutical composition comprising a compound of the disclosure, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.
  • the disclosure features a pharmaceutical composition comprising a therapeutically effective amount of a compound, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers or vehicles.
  • a proliferative disease e.g., an IgM gammopathy (e.g., an IgM Monoclonal gammopathy of undetermined significance (MGUS), amyloid light chain (AL) amyloidosis), mastocytosis (e.g., systemic mastocytosis) cancer (e.g., breast cancer, colon cancer, testicular cancer, CNS cancer, stomach cancer, lymphoma (e.g., B- cell lymphoma (e.g., lymphoplasmacytic lymphoma (e.g., IgM secreting lymphoplasmacytic lymphoma (i.e., Waldenström’s Macroglobulinemia), non-IgM secreting lymphoplasmacytic lymphoma)), diffuse large B-cell lymphoma (e.g., activated B-cell-like (
  • MGUS amyloid light chain amyloidosis
  • mastocytosis e.g., systemic mastocytosis
  • the method comprises administering a compound of the disclosure, or a pharmaceutically acceptable salt thereof.
  • the disease is associated with a mutated MYD88 protein.
  • the disease is associated with a mutated BTK protein.
  • the disease is associated with a C481 mutated BTK protein.
  • the disease is associated with a C481S mutated BTK protein.
  • the diseases are associated with aberrant activity of a kinase (e.g., SRC Family kinases (e.g., HCK, LYN, BLK, FRK), Tec family kinases (e.g., BTK).
  • the disease is resistant to inhibition by a BTK inhibitor (e.g., ibrutinib, CC-292, ONO-4059, evobrutinib, spebrutinib, BGB-3111, HM71224, or ACP-196).
  • a BTK inhibitor e.g., ibrutinib, CC-292, ONO-4059, evobrutinib, spebrutinib, BGB-3111, HM71224, or ACP-196.
  • the disease is resistant to treatment with ibrutinib.
  • the disease is associated with a mutated BTK protein (e.g., a C481S mutated BTK), and the disease is resistant to treatment with ibrutinib.
  • the one or more kinase is an SRC Family kinase (e.g., HCK, LYN, BLK, FRK).
  • the one or more kinase is a Tec family kinase (e.g., BTK).
  • the BTK is resistant to inhibition by a BTK inhibitor (e.g., ibrutinib, CC-292, ONO-4059, evobrutinib, spebrutinib, BGB-3111, HM71224, or ACP-196).
  • a BTK inhibitor e.g., ibrutinib, CC-292, ONO-4059, evobrutinib, spebrutinib, BGB-3111, HM71224, or ACP-196.
  • the BTK is ibrutinib resistant.
  • a method of treating a subject comprising administering to a subject with an MYD88 mutated disease a therapeutically effective amount of a compound of the disclosure or a pharmaceutically acceptable salt, solvate, hydrate, crystal, isotopically-labeled derivative, stereoisomer, or prodrug thereof.
  • An MYD88 mutated disease can include, but is not limited to a proliferative disease (e.g., an IgM gammopathy (e.g., an IgM Monoclonal gammopathy of undetermined significance (MGUS), amyloid light chain (AL) amyloidosis), mastocytosis (e.g., systemic mastocytosis) cancer (e.g., breast cancer, colon cancer, testicular cancer, CNS cancer, stomach cancer, lymphoma (e.g., B-cell lymphoma (e.g., lymphoplasmacytic lymphoma (e.g., IgM secreting lymphoplasmacytic lymphoma (i.e., Waldenström’s Macroglobulinemia), non-IgM secreting lymphoplasmacytic lymphoma)), diffuse large B-cell lymphoma (e.g., activated B-cell- like (ABC)- DLBCL, germinal center B-cell-like (GBC)
  • the subject being treated has previously undergone treatment with ibrutinib. In certain embodiments, the subject being treated has developed ibrutinib resistance.
  • the provided methods include inhibiting LYN and/or SRC comprising the steps of administering to the subject a therapeutically effective amount of a compound of the disclosure, or a pharmaceutically acceptable salt, solvate, hydrate, crystal, isotopically-labeled derivative, stereoisomer, or prodrug thereof. In some embodiments, the method further comprises administering an agent which inhibits LYN and/or SRC.
  • a compound of the disclosure as described herein is administered to the subject in combination (e.g., concurrently or sequentially) with an agent which blocks ATP binding to SRC.
  • a compound of the disclosure as described herein is administered to the subject in combination (e.g., concurrently or sequentially) with an agent which blocks ATP binding to LYN.
  • a compound of the disclosure as described herein is administered to the subject in combination (e.g., concurrently or sequentially) with an agent which blocks ATP binding to HCK.
  • the provided methods include inhibiting Tec family kinases comprising the steps of administering to the subject a therapeutically effective amount of a compound of the disclosure, or a pharmaceutically acceptable salt, solvate, hydrate, crystal, isotopically-labeled derivative, stereoisomer, or prodrug thereof.
  • the provided methods include inhibiting BTK comprising the steps of administering to the subject a therapeutically effective amount of a compound of the disclosure, or a pharmaceutically acceptable salt, solvate, hydrate, crystal, isotopically-labeled derivative, stereoisomer, or prodrug thereof.
  • the method further comprises administering an agent which inhibits BTK and/or other Tec family kinases.
  • a compound of the disclosure as described herein is administered to the subject in combination (e.g., concurrently or sequentially) with an agent which blocks ATP binding to a Tec family kinase.
  • a compound of the disclosure as described herein is administered to the subject in combination (e.g., concurrently or sequentially) with an agent which blocks ATP binding to BTK.
  • Detection methods include, but are not limited to, direct sequencing, DNA chip technologies, mass spectroscopy, polymerase chain reaction (PCR), allele specific polymerase chain reaction, real time polymerase chain reaction, reverse transcriptase PCR, electrophoretic mobility, nucleic acid hybridization, fluorescent in situ hybridization, and denaturing high performance liquid chromatography.
  • PCR polymerase chain reaction
  • allele specific polymerase chain reaction e.g., as described in WO 2013/006443.
  • LPL LPL
  • the subject may also present one or more of the following clinical features or symptoms of other B cell neoplasms: asymptomatic localized or generalized peripheral lymphadenopathy, plasmacytic difference, bone marrow involvement, autoimmune thrombocytopenia, peripheral blood villous lymphocytes, end organ damage (hypercalcemia, renal insufficiency, bone lesions), recurrent infections, elevated creatine, hyperuricemia, and hypoalbunemia.
  • a subject suspected of having one or more of Waldenström’s Macroglobulinemia i.e., IgM secreting lymphoplasmacytic lymphoma), non-IgM secreting lymphoplasmacytic lymphoma, ABC subtype of diffuse large B-cell lymphoma, primary central nervous system (CNS) lymphoma, immune privileged lymphomas that include testicular lymphoma, marginal zone lymphoma, and chronic lymphocytic leukemia may be assessed for the presence of a mutation in the gene encoding MYD88, e.g., at position 38182641 in chromosome 3p22.2, as well as other activating mutations in MYD88 (including but not limited to V217F, W218R, I220T, S222R, M232T, S243N, and T294P).
  • MYD88 e.g., at position 38182641 in chromosome 3p22.2
  • the compounds described herein are able to bind the one or more kinase being inhibited.
  • a compound described herein is able to bind to the one or more kinase.
  • the one or more kinase is an SFK (e.g., HCK, LYN, BLK, FRK).
  • the one or more kinase is HCK.
  • the one or more kinase is LYN.
  • the one or more kinase is a Tec family kinase (e.g., BTK).
  • the one or more kinase is BTK.
  • a kinase e.g., SFK (e.g., HCK, LYN, BLK, FRK), Tec family kinase (e.g., BTK)
  • SFK e.g., HCK, LYN, BLK, FRK
  • Tec family kinase e.g., BTK
  • the activity of a kinase in a subject is decreased by at least about 1%, at least about 3%, at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, or at least about 90%.
  • the activity of a kinase in a subject is selectively inhibited by the method.
  • the activity of a kinase (e.g., HCK, LYN, BTK) in a subject is selectively decreased by a compound or pharmaceutical composition described herein.
  • a disease including proliferative disease, may be associated with aberrant or undesired activity of a kinase, and/or overexpression of the kinase.
  • Aberrant or undesired activity of a kinase may be an increased or a decreased level of activity of the kinase.
  • Proliferative diseases are sometimes associate with abnormal levels of JAK activity, frequently through increased or decreased JAK activation. Inhibition of the activity of JAK2 would be expected to inhibit phosphorylation.
  • JAK2 is not overexpressed, but the activity of JAK2 is increased.
  • JAK2 is overexpressed, and the activity of JAK2 is increased.
  • the compounds and pharmaceutical compositions described herein may inhibit the activity of JAK2 and be useful in treating and/or preventing diseases, such as diseases associated with the aberrant, increased, or undesired activity of a kinase, overactivation of the kinase, and/or overexpression of the kinase.
  • the disease e.g., the disease to be treated or prevented by a method described herein
  • a kinase e.g., SFK (e.g., HCK, LYN, BLK, FRK), Tec Family kinases (e.g., BTK)).
  • the disease is associated with overexpression of a kinase (e.g., SFK (e.g., HCK, LYN, BLK, FRK), Tec Family kinases (e.g., BTK)).
  • a kinase e.g., SFK (e.g., HCK, LYN, BLK, FRK), Tec Family kinases (e.g., BTK)
  • the disease is a proliferative disease.
  • the proliferative disease is cancer.
  • the cancer is associated with a mutation in MYD88.
  • the cancer is associated with mutated BTK.
  • the proliferative disease is mastocytosis.
  • the mastocytosis is systemic mastocytosis.
  • the proliferative disease is an IgM gammopathy.
  • the IgM gammopathy is IgM monoclonal gammopathy with undetermined significance.
  • the disease is breast cancer. In certain embodiments, the disease is colon cancer. In certain embodiments, the disease is testicular cancer. In certain embodiments, the disease is cancer of the CNS. In certain embodiments, the disease is stomach cancer. In certain embodiments, the disease is lymphoma. In certain embodiments, the lymphoma is B-cell Lymphoma. In certain embodiments, B-cell lymphoma is lymphoplasmacytic lymphoma.
  • the lymphoplasmacytic lymphoma is IgM secreting lymphoplasmacytic lymphoma (i.e., Waldenström’s Macroglobulinemia). In certain embodiments, the disease is Waldenström’s Macroglobulinemia. In certain embodiments, the lymphoplasmacytic lymphoma is non-IgM secreting lymphoplasmacytic lymphoma. In certain embodiments, the lymphoma is Diffuse Large B-Cell Lymphoma (DLBCL). In certain embodiments, the DLBCL is activated B- cell-like (ABC)-DLBCL. In certain embodiments, the DLBCL is germinal center B-cell-like (GBC)-DLBCL.
  • BCC B-cell-like
  • the lymphoma is Follicular Lymphoma. In certain embodiments, the lymphoma is marginal zone B-cell lymphoma. In certain embodiments, the lymphoma is Small lymphocytic lymphoma. In certain embodiments, the small lymphocytic lymphoma is Mantle cell lymphoma. In certain embodiments, the cancer is leukemia. In certain embodiments, the leukemia is chronic lymphocytic leukemia (CLL). In certain embodiments, the leukemia is myelogenous leukemia. In certain embodiments, the myelogenous leukemia is chronic myelogenous leukemia. In certain embodiments, the myelogenous leukemia is acute myelogenous leukemia.
  • CLL chronic lymphocytic leukemia
  • the acute myelogenous leukemia is mast cell leukemia.
  • the cancer is myeloma.
  • the myeloma is IgM myeloma.
  • the IgM myeloma is IgM multiple myeloma.
  • the cancer is a myeloproliferative disease.
  • the myeloproliferative disease is myelodysplastic syndrome.
  • the method described herein is superior (e.g., showing improved safety and/or therapeutic effects) or comparable to existing therapy (e.g., chemotherapy, treatment with a BTK inhibitor). In certain embodiments, the method described herein is associated with decreased toxicity when compared to existing therapy (e.g., chemotherapy, treatment with a BTK inhibitor).
  • the cell is a malignant cell (e.g., cancer cell).
  • the cell is a malignant blood cell.
  • the cell is a malignant bone marrow cell.
  • the cell is an adenocarcinoma cell, blastoma cell, carcinoma cell, or sarcoma cell.
  • the cell is a pre-malignant cell (e.g., pre- cancerous cell).
  • the present disclosure provides compounds and pharmaceutical compositions described herein for use in the treatment of a disease (e.g., a proliferative disease, such as an IgM gammopathy, mastocytosis, or cancer) in a subject in need thereof.
  • a disease e.g., a proliferative disease, such as an IgM gammopathy, mastocytosis, or cancer
  • the present disclosure provides compounds and pharmaceutical compositions described herein for use in the prevention of a disease (e.g., a proliferative disease, such as an IgM gammopathy, mastocytosis, or cancer) in a subject in need thereof.
  • a disease e.g., a proliferative disease, such as an IgM gammopathy, mastocytosis, or cancer
  • the present disclosure provides compounds and pharmaceutical compositions described herein for use in therapy, e.g., using the methods described herein.
  • the present disclosure provides uses of compounds and pharmaceutical compositions described herein in the manufacture of a medicament for treating a disease in a subject in need thereof, e.g., using the methods described herein.
  • the present disclosure provides uses of compounds and pharmaceutical compositions described herein in the manufacture of a medicament for preventing a disease in a subject in need thereof, e.g., using the methods described herein.
  • the compounds, pharmaceutical compositions, and kits described herein may synergistically augment inhibition of a kinase (e.g., SFK (e.g., HCK, LYN, BLK, FRK), a TEC family kinase (e.g., BTK)) induced by the additional pharmaceutical agent(s) in the subject.
  • a kinase e.g., SFK (e.g., HCK, LYN, BLK, FRK)
  • a TEC family kinase e.g., BTK
  • an “effective amount” of a compound of the disclosure, a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof is that amount effective for treating or lessening the severity of one or more of the conditions recited above.
  • effective amounts of the therapeutic agent will depend on the particular disease being treated; the severity of the disease; individual patient parameters including age, physical condition, size and weight, concurrent treatment, frequency of treatment, and the mode of administration.
  • an effective amount of a compound typically will vary from about 0.001 mg/kg to about 1000 mg/kg in one or more dose administrations, for one or several days (depending of course of the mode of administration and the factors discussed above). It will be understood that an effective amount of a compound is preferably provided as a unit dose with 0.1 to 1000 mg/dose of the compound to be administered, preferably one or two times per day.
  • Actual dosage levels of the therapeutic agent can be varied to obtain an amount that is effective to achieve the desired therapeutic response for a particular patient, compositions, and mode of administration.
  • the selected dosage level depends upon the activity of the particular compound, the route of administration, the tissue being treated, and prior medical history of the patient being treated.
  • an effective amount of a selective HCK inhibitor is that amount which slows the progression of the disease, halts the progression of the disease, or reverses the progression of the disease.
  • An effective amount includes, but is not limited to, that amount necessary to slow, reduce, inhibit, ameliorate or reverse one or more symptoms associated with the MYD88 mutated disease.
  • such terms refer to a reduction in the levels of IgM serum paraprotein, anemia, hyper-viscosity, neuropathy, coagulopathies, splenomegaly, hepatomegaly, and adenopathy.
  • the compound described herein is provided in an effective amount (e.g., effective for inhibiting kinase activity, such as a SRC family kinases (e.g., HCK, LYN, BLK, FRK), or Tec family kinases (e.g., BTK)) in the pharmaceutical composition.
  • the effective amount is a therapeutically effective amount.
  • a therapeutically effective amount is an amount effective for inhibiting a kinase.
  • a therapeutically effective amount is an amount effective for treating a disease (e.g., a disease associated with aberrant activity of a kinase (e.g., proliferative disease)).
  • a therapeutically effective amount is an amount effective for inhibiting the activity of a kinase and treating a disease (e.g., a disease associated with aberrant activity of a kinase (e.g., proliferative disease)).
  • a therapeutically effective amount is an amount effective for inducing apoptosis in a cell (e.g., malignant cell, premalignant cell).
  • the effective amount is an amount effective for inhibiting the activity of a kinase by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 98%.
  • the effective amount is an amount effective for inhibiting the activity of a kinase by not more than 10%, not more than 20%, not more than 30%, not more than 40%, not more than 50%, not more than 60%, not more than 70%, not more than 80%, not more than 90%, not more than 95%, or not more than 98%.
  • the subject is an animal. The animal may be of either sex and may be at any stage of development.
  • the subject described herein is a human (e.g., an adult, juvenile, or child).
  • the subject is a non-human animal.
  • the subject is a mammal.
  • the subject is a non-human mammal.
  • the subject is a domesticated animal, such as a dog, cat, cow, pig, horse, sheep, or goat.
  • the subject is a dog.
  • the subject is a companion animal, such as a dog or cat.
  • the subject is a livestock animal, such as a cow, pig, horse, sheep, or goat.
  • the subject is a zoo animal.
  • the subject is a research animal, such as a rodent (e.g., mouse, rat), dog, pig, or non-human primate.
  • the subject is a genetically engineered animal.
  • the subject is a transgenic animal (e.g., transgenic mice, transgenic pigs). In certain embodiments, the subject is a fish or reptile.
  • Pharmaceutical compositions described herein can be prepared by any method known in the art of pharmacology. In general, such preparatory methods include bringing the compound described herein (i.e., the “active ingredient”) into association with a carrier or excipient, and/or one or more other accessory ingredients, and then, if necessary and/or desirable, shaping, and/or packaging the product into a desired single- or multi-dose unit.
  • Pharmaceutical compositions can be prepared, packaged, and/or sold in bulk, as a single unit dose, and/or as a plurality of single unit doses.
  • a “unit dose” is a discrete amount of the pharmaceutical composition comprising a predetermined amount of the active ingredient.
  • the amount of the active ingredient is generally equal to the dosage of the active ingredient which would be administered to a subject and/or a convenient fraction of such a dosage, such as one-half or one-third of such a dosage.
  • Relative amounts of the active ingredient, the pharmaceutically acceptable excipient, and/or any additional ingredients in a pharmaceutical composition described herein will vary, depending upon the identity, size, and/or condition of the subject treated and further depending upon the route by which the composition is to be administered.
  • the composition may comprise between 0.1% and 100% (w/w) active ingredient.
  • compositions include inert diluents, dispersing and/or granulating agents, surface active agents and/or emulsifiers, disintegrating agents, binding agents, preservatives, buffering agents, lubricating agents, and/or oils.
  • Liquid dosage forms for oral and parenteral administration include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active ingredients, the liquid dosage forms may comprise inert diluents commonly used in the art.
  • Injectable preparations for example, sterile injectable aqueous or oleaginous suspensions can be formulated according to the known art using suitable dispersing or wetting agents and suspending agents.
  • the sterile injectable preparation can be a sterile injectable solution, suspension, or emulsion in a nontoxic parenterally acceptable diluent or solvent
  • Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules.
  • the active ingredient is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or (a) fillers or extenders, (b) binders, (c) humectants, (d) disintegrating agents, (e) solution retarding agents, (g) wetting agents, (h) absorbents, and (I) lubricants.
  • the dosage form may include a buffering agent.
  • Solid compositions of a similar type can be employed as fillers in soft and hard-filled gelatin capsules.
  • the solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the art of pharmacology.
  • Dosage forms for topical and/or transdermal administration of a compound described herein may include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants, and/or patches.
  • a pharmaceutical composition described herein can be prepared, packaged, and/or sold in a formulation suitable for pulmonary administration via the buccal cavity.
  • Pharmaceutical compositions described herein formulated for pulmonary delivery may provide the active ingredient in the form of droplets of a solution and/or suspension.
  • Formulations for nasal administration may, for example, comprise from about as little as 0.1% (w/w) to as much as 100% (w/w) of the active ingredient, and may comprise one or more of the additional ingredients described herein.
  • a pharmaceutical composition described herein can be prepared, packaged, and/or sold in a formulation for ophthalmic administration. Such formulations may, for example, be in the form of eye drops including, for example, a 0.1-1.0% (w/w) solution and/or suspension of the active ingredient in an aqueous or oily liquid carrier or excipient.
  • compositions provided herein are principally directed to pharmaceutical compositions which are suitable for administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to animals of all sorts. Modification of pharmaceutical compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals is well understood, and the ordinarily skilled veterinary pharmacologist can design and/or perform such modification with ordinary experimentation.
  • Compounds provided herein are typically formulated in dosage unit form for ease of administration and uniformity of dosage. It will be understood, however, that the total daily usage of the compositions described herein will be decided by a physician within the scope of sound medical judgment.
  • the specific therapeutically effective dose level for any particular subject or organism will depend upon a variety of factors including the disease being treated and the severity of the disorder; the activity of the specific active ingredient employed; the specific composition employed; the age, body weight, general health, sex, and diet of the subject; the time of administration, route of administration, and rate of excretion of the specific active ingredient employed; the duration of the treatment; drugs used in combination or coincidental with the specific active ingredient employed; and like factors well known in the medical arts.
  • the compounds and compositions provided herein can be administered by any route, including enteral (e.g., oral), parenteral, intravenous, intramuscular, intra-arterial, intramedullary, intrathecal, subcutaneous, intraventricular, transdermal, interdermal, rectal, intravaginal, intraperitoneal, topical (as by powders, ointments, creams, and/or drops), mucosal, nasal, bucal, sublingual; by intratracheal instillation, bronchial instillation, and/or inhalation; and/or as an oral spray, nasal spray, and/or aerosol.
  • enteral e.g., oral
  • parenteral intravenous, intramuscular, intra-arterial, intramedullary
  • intrathecal subcutaneous, intraventricular, transdermal, interdermal, rectal, intravaginal, intraperitoneal
  • topical as by powders, ointments, creams, and/or drops
  • mucosal nasal,
  • Specifically contemplated routes are oral administration, intravenous administration (e.g., systemic intravenous injection), regional administration via blood and/or lymph supply, and/or direct administration to an affected site.
  • intravenous administration e.g., systemic intravenous injection
  • regional administration via blood and/or lymph supply e.g., systemic intravenous injection
  • direct administration e.g., direct administration to an affected site.
  • the exact amount of a compound required to achieve an effective amount will vary from subject to subject, depending, for example, on species, age, and general condition of a subject, severity of the side effects or disorder, identity of the particular compound, mode of administration, and the like.
  • An effective amount may be included in a single dose (e.g., single oral dose) or multiple doses (e.g., multiple oral doses).
  • the frequency of administering the multiple doses to the subject is three doses a day, two doses a day, one dose a day, one dose every other day, one dose every third day, one dose every week, one dose every two weeks, one dose every three weeks, or one dose every four weeks.
  • a dose (e.g., a single dose, or any dose of multiple doses) described herein includes independently between 0.1 ⁇ g and 10 mg, inclusive, of a compound described herein. In certain embodiments, a dose described herein includes independently between 10 mg and 100 mg, inclusive, of a compound described herein.
  • the present disclosure provides methods of treating a disease in a subject in need thereof, the method comprising administering to the subject in need thereof an effective amount (e.g., therapeutically effective amount) of a compound of the disclosure described herein or a pharmaceutical composition described herein.
  • an effective amount e.g., therapeutically effective amount
  • the present disclosure provides methods of preventing a disease in a subject in need thereof, the method comprising administering to the subject in need thereof an effective amount (e.g., prophylactically effective amount) of a compound described herein or a pharmaceutical composition described herein.
  • the present disclosure provides methods of inhibiting the activity of a kinase in a subject in need thereof, the method comprising administering to the subject in need thereof an effective amount of a compound described herein or a pharmaceutical composition described herein.
  • Additional Therapeutic Agents [0148] It will also be appreciated that the compounds, salts, and pharmaceutically acceptable compositions of the disclosure can be employed in combination therapies, that is, the compounds, salts, and pharmaceutically acceptable compositions can be administered concurrently with, prior to, or subsequent to, one or more other desired therapeutics or medical procedures. [0149] A compound or composition, as described herein, can be administered in combination with one or more additional pharmaceutical agents (e.g., therapeutically and/or prophylactically active agents).
  • additional pharmaceutical agents e.g., therapeutically and/or prophylactically active agents.
  • the compounds or compositions can be administered in combination with additional pharmaceutical agents that improve their activity (e.g., activity (e.g., potency and/or efficacy) in treating a disease in a subject in need thereof, in preventing a disease in a subject in need thereof, in inhibiting the activity of a kinase (e.g., SFK (e.g., LYN, HCK), Tec family kinases (e.g., BTK)) in a subject, improve bioavailability, improve safety, reduce drug resistance, reduce and/or modify metabolism, inhibit excretion, and/or modify distribution in a subject.
  • a kinase e.g., SFK (e.g., LYN, HCK), Tec family kinases (e.g., BTK)
  • a kinase e.g., SFK (e.g., LYN, HCK), Tec family kinases (e.g., BTK)
  • a pharmaceutical composition described herein including a compound described herein and an additional pharmaceutical agent shows a synergistic effect that is absent in a pharmaceutical composition including one of the compound and the additional pharmaceutical agent, but not both.
  • treatment further includes administering to the subject an agent, e.g., an anti-cancer agent, in combination with a compound described herein.
  • treatment further includes administering to the subject one or more of bendamustine, fludarabine, bortezomib, or idelalisib.
  • treatment further includes administering to the subject one or more of a BCL-2 inhibitor (e.g., venetoclax, navitoclax, obatoclax), a BCL-2/BCL-xL inhibitor (e.g., APG-1252, BM-1197), a proteasome inhibitor (e.g., bortezomib, carfilzomib, ixazomib or oprozomib), a monoclonal antibody (e.g., rituximab, daratumumab, ofatumumab or obinituzumab), an alkylator drug (e.g., bendamustine, cyclophosphamide), a nucleoside analogue (e.g., fludarabine or cladribine), an MTOR inhibitor (e.g., everolimus), a BTK inhibitor (e.g., ibrutinib, acalabrutini
  • the anti-cancer agent is a monoclonal antibody, e.g., rituximab.
  • the anti-cancer agent is a chemotherapeutic drug such as chlorambucil, cyclophosphamide, or vincristine or thalidomide.
  • Corticosteroids such as Prednisone, may also be used in combination.
  • Plasmapheresis can be used to treat the hyperviscosity syndrome by removing the paraprotein from the blood.
  • Autologous bone marrow transplantation may be used in combination with compounds described herein.
  • treatment further includes administering to the subject an agent that inhibits LYN and/or SRC.
  • the compound or composition can be administered concurrently with, prior to, or subsequent to one or more additional pharmaceutical agents, which may be useful as, e.g., combination therapies.
  • Pharmaceutical agents include therapeutically active agents.
  • Pharmaceutical agents also include prophylactically active agents.
  • Pharmaceutical agents include small organic molecules such as drug compounds (e.g., compounds approved for human or veterinary use by the U.S.
  • CFR Code of Federal Regulations
  • proteins proteins, carbohydrates, monosaccharides, oligosaccharides, polysaccharides, nucleoproteins, mucoproteins, lipoproteins, synthetic polypeptides or proteins, small molecules linked to proteins, glycoproteins, steroids, nucleic acids, DNAs, RNAs, nucleotides, nucleosides, oligonucleotides, antisense oligonucleotides, lipids, hormones, vitamins, and cells.
  • CFR Code of Federal Regulations
  • the additional pharmaceutical agent is a pharmaceutical agent useful for treating and/or preventing a disease (e.g., proliferative disease, cancer, inflammatory disease, autoimmune disease, genetic disease, hematological disease, neurological disease, painful condition, psychiatric disorder, or metabolic disorder) or premalignant condition.
  • a disease e.g., proliferative disease, cancer, inflammatory disease, autoimmune disease, genetic disease, hematological disease, neurological disease, painful condition, psychiatric disorder, or metabolic disorder
  • Each additional pharmaceutical agent may be administered at a dose and/or on a time schedule determined for that pharmaceutical agent.
  • the additional pharmaceutical agents may also be administered together with each other and/or with the compound or composition described herein in a single dose or administered separately in different doses.
  • the particular combination to employ in a regimen will take into account compatibility of the compound described herein with the additional pharmaceutical agent(s) and/or the desired therapeutic and/or prophylactic effect to be achieved.
  • the additional pharmaceutical agents include, but are not limited to, cytotoxic chemotherapeutic agents, epigenetic modifiers, glucocorticoids, immunotherapeutic agents, anti- proliferative agents, anti-cancer agents, anti-angiogenesis agents, anti-inflammatory agents, immunosuppressants, anti-bacterial agents, anti-viral agents, cardiovascular agents, cholesterol- lowering agents, anti-diabetic agents, anti-allergic agents, contraceptive agents, pain-relieving agents, and a combination thereof.
  • the additional pharmaceutical agent is an anti-proliferative agent (e.g., anti-cancer agent). In certain embodiments, the additional pharmaceutical agent is an anti-leukemia agent. In certain embodiments, the compounds described herein or pharmaceutical compositions can be administered in combination with an anti-cancer therapy including, but not limited to, surgery, radiation therapy, and transplantation (e.g., stem cell transplantation, bone marrow transplantation).
  • the additional pharmaceutical agent is a BCL-2 inhibitor (e.g., venetoclax, navitoclax, obatoclax), or a BCL-2/BCL-xL inhibitor (e.g., APG-1252, BM-1197).
  • the additional pharmaceutical agent is venetoclax.
  • the compounds of the disclosure are used in combination with one or more additional immunotherapies. Synthesis of the Compounds of the Disclosure [0156]
  • the compounds of the disclosure can be prepared from known materials by the methods described in the Examples, other similar methods, and other methods known to one skilled in the art.
  • the functional groups of the intermediate compounds in the methods described below may need to be protected by suitable protecting groups.
  • Protecting groups may be added or removed in accordance with standard techniques, which are well-known to those skilled in the art. The use of protecting groups is described in detail in T.G.M.
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0158] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of 8-hydroxy-2-azaspiro[4.5]decan-1-one [0159] To a mixture of 2-azaspiro[4.5]decane-1,8-dione (200 mg, 1.19 mmol) in MeOH (5 mL) was added NaBH 4 (90 mg, 2.38 mmol) at 0oC. The mixture was stirred at room temperature overnight. The mixture was quenched with water (20 mL), and the aqueous phase was extracted with EtOAc (20 mL*3). The combined organic phase was concentrated, and the residue was purified by Prep-HPLC to give 8-hydroxy-2-azaspiro[4.5]decan-1-one (150 mg, 75% yield) as a white solid.
  • Step 3 Synthesis of 1-oxo-2-azaspiro[4.5]decan-8-yl 4-methylbenzenesulfonate [0160] To a mixture of 8-hydroxy-2-azaspiro[4.5]decan-1-one (150 mg, 0.88 mmol) and DIPEA (230 mg, 1.77 mmol) in DCM (5 mL) was added Methanesulfonyl chloride (151 mg, 1.33 mmol) at 0oC. The mixture was stirred at 0 oC for 30 minutes, diluted with DCM (30 mL), and the organic phase was washed with aq. sat.
  • Step 4 Synthesis of 8-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2- azaspiro[4.5]decan-1-one [0161] To a solution of 1-oxo-2-azaspiro[4.5]decan-8-yl 4-methylbenzenesulfonate (80 mg, 0.25 mmol) in DMF (3 mL) was added 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4- amine (100 mg, 0.32 mmol) and Cs 2 CO 3 (163 mg, 0.50 mmol).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0168] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of ethyl 2-cyano-2-(1,4-dioxaspiro[4.5]decan-8-ylidene)acetate
  • 1,4-dioxaspiro[4.5]decan-8-one 10 g, 64.1 mmol
  • toluene 100 mL
  • ethyl 2-cyanoacetate 9.42 g, 83.3 mmol
  • NH 4 OAc 8.88 g, 115.4 mmol
  • Step 3 Synthesis ofethyl 2-cyano-2-(8-(2-ethoxy-2-oxoethyl)-1,4-dioxaspiro[4.5]decan-8- yl)acetate
  • ethyl acetate 13.6 g, 54.2 mmol
  • THF 100 mL
  • LDA 54.2 mL, 2.0M, 108.4 mmol
  • Step 4 Synthesis of ethyl 2-(8-(cyanomethyl)-1,4-dioxaspiro[4.5]decan-8-yl)acetate
  • ethyl 2-cyano-2-(8-(2-ethoxy-2-oxoethyl)-1,4-dioxaspiro[4.5]decan- 8-yl)acetate 7 g, 20.65 mmol
  • LiI 5.53 g, 41.30 mmol
  • Step 5 Synthesis of 1,4-dioxa-11-azadispiro[4.2.5 8 .2 5 ]pentadecan-10-one [0172] To a solution of ethyl 2-(8-(cyanomethyl)-1,4-dioxaspiro[4.5]decan-8-yl)acetate (3.69 g, 13.82 mmol) in ethanol (30 mL) was added NH 3 .H 2 O (6 mL) and Raney-Ni (5 mL).
  • Step 6 Synthesis of 3-azaspiro[5.5]undecane-2,9-dione [0173] To a solution of 1,4-dioxa-11-azadispiro[4.2.5 8 .2 5 ]pentadecan-10-one (1 g, 4.44 mmol) in THF (20 mL) was added HCl (7M in water, 6 mL), and the resulting mixture was stirred at room temperature overnight. The reaction solution was concentrated to give 3-azaspiro[5.5]undecane- 2,9-dione (1 g, crude), which was used directly for the next step. LCMS (m/z): 182.4[M+H] + .
  • Step 7 Synthesis of 9-hydroxy-3-azaspiro[5.5]undecan-2-one [0174] To a solution of 3-azaspiro[5.5]undecane-2,9-dione (1 g, crude) in CH 3 OH (20 mL) was added NaBH 4 (420 mg, 11.05 mmol). The resulting mixture was stirred at room temperature overnight, quenched with water (3 mL) and concentrated. The residue was purified by Prep-HPLC (ACN and H 2 O with 0.01% TFA as mobile phase) to give 9-hydroxy-3-azaspiro[5.5]undecan-2- one (325mg, 40.0% yield of 2 steps) as white solid. LCMS (m/z): 184.3[M+H] + .
  • Step 8 Synthesis of 2-oxo-3-azaspiro[5.5]undecan-9-yl 4-methylbenzenesulfonate [0175] To a solution of 9-hydroxy-3-azaspiro[5.5]undecan-2-one (275 mg, 1.50 mmol), DMAP (36.7 mg, 0.3 mmol) and Et3N (455.3 mg, 4.51 mmol) in DCM (20 mL) was add TsCl (572.8 mg, 3.00 mmol). The mixture was refluxed overnight, diluted with DCM (30 mL), the organic phase was washed with aq. sat.
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0178] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Example 5 Synthesis of 1-((1r,4r)-4-(6-methyl-2,6-diazaspiro[3.3]heptan-2-yl)cyclohexyl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (5).
  • Step 1 Synthesis of 3-iodo-1-(1,4-dioxaspiro[4.5]decan-8-yl)-1H-pyrazolo[3,4-d]pyrimidin-4- amine.
  • Step 4 Synthesis of tert-butyl 6-(4-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin- 1-yl)cyclohexyl)-2,6-diazaspiro[3.3]heptane-2-carboxylate.
  • Step 5 Synthesis of 1-((1r,4r)-4-(2,6-diazaspiro[3.3]heptan-2-yl)cyclohexyl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 6 Synthesis of 1-((1r,4r)-4-(6-methyl-2,6-diazaspiro[3.3]heptan-2-yl)cyclohexyl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0194] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of 6-oxospiro[3.3]heptan-2-yl 4-methylbenzenesulfonate [0195] To a mixture of 6-hydroxyspiro[3.3]heptan-2-one (500 mg, 3.97 mmol) and TEA (800 mg, 7.93 mmol) in DCM (20 mL) was added TsCl (1.13 g, 5.95 mmol) and DMAP (50 mg, 0.416 mmol). The resulting mixture was stirred at room temperature overnight, quenched with water (50 mL), and the aqueous phase was extracted with DCM (50 mL x 3).
  • Step 3 Synthesis of 6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)spiro[3.3]heptan-2-one [0196] To a solution of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (691 mg, 2.28 mmol) and 6-oxospiro[3.3]heptan-2-yl 4-methylbenzenesulfonate (639 mg, 2.28 mmol) in DMF (10 mL) was added Cs 2 CO 3 (1.12 g, 3.42 mmol).
  • Step 4 Synthesis of tert-butyl 6-methyl-2,6-diazaspiro[3.3]heptane-2-carboxylate [0197] To a mixture of tert-butyl 2,6-diazaspiro[3.3]heptane-2-carboxylate (400 mg, 2.02 mmol) and CH 3 COOH (2 drops) in DCM (10 mL) was added HCHO (400 mg, 37% in water, 4.03 mmol). The resulting mixture was stirred at room temperature for 1 hour, and then added NaBH(OAc) 3 (859 mg, 4.04 mmol).
  • Step 5 Synthesis of 2-methyl-2,6-diazaspiro[3.3]heptane [0198] To a mixture of tert-butyl 6-methyl-2,6-diazaspiro[3.3]heptane-2-carboxylate (400 mg, crude) in DCM (10 mL) was added TFA (2 mL). The resulting mixture was stirred at room temperature overnight and concentrated to give 2-methyl-2,6-diazaspiro[3.3]heptane (210 mg, crude) as a pale solid, which was used directly for the next step. LCMS: m/z 113.1 [M+H] + .
  • Step 6 Synthesis of 1-(6-(6-methyl-2,6-diazaspiro[3.3]heptan-2-yl)spiro[3.3]heptan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0199] To a mixture of 6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)spiro[3.3]heptan-2-one (150 mg, 0.36 mmol) and CH 3 COOH (3 drops) in DCM (5 mL) was added 2-methyl-2,6-diazaspiro[3.3]heptane (210 mg, crude).
  • Example 7 Synthesis of 1-(6-(4-methylpiperazin-1-yl)spiro[3.3]heptan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (7).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0201] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of 6-oxospiro[3.3]heptan-2-yl 4-methylbenzenesulfonate [0202] To a mixture of 6-hydroxyspiro[3.3]heptan-2-one (500 mg, 3.97 mmol) and TEA (800 mg, 7.93 mmol) in DCM (20 mL) was added TsCl (1.13 g, 5.95 mmol) and DMAP (50 mg, 0.416 mmol). The resulting mixture was stirred at room temperature for 12 hours, quenched with water (50 mL), and the aqueous phase was extracted with DCM (50 mL x 3).
  • Step 3 Synthesis of 6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)spiro[3.3]heptan-2-one [0203] To a solution of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (691 mg, 2.28 mmol) and 6-oxospiro[3.3]heptan-2-yl 4-methylbenzenesulfonate (639 mg, 2.28 mmol) in DMF (10 mL) was added Cs 2 CO 3 (1.12 g, 3.42 mmol).
  • Step 4 Synthesis of 1-(6-(4-methylpiperazin-1-yl)spiro[3.3]heptan-2-yl)-3-(4-phenoxyphenyl)- 1H-pyrazolo[3,4-d]pyrimidin-4-amine
  • Example 8 Synthesis of 1-(1-(2-azaspiro[3.3]heptan-6-yl)piperidin-4-yl)-3-(4-phenoxyphenyl)- 1H-pyrazolo[3,4-d]pyrimidin-4-amine (8) and 1-(1-(2-methyl-2-azaspiro[3.3]heptan-6- yl)piperidin-4-yl)-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (9)
  • Step 1 Synthesis of tert-butyl 4-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidine- 1-carboxylate.
  • 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine 860 mg, 3.3 mmol
  • tert-butyl 4-hydroxypiperidine-1-carboxylate (1.32 g, 6.6 mmol
  • PPh 3 (1.73 g, 6.6 mmol
  • DIAD 1.33 g, 6.6 mmol
  • Step 3 Synthesis of tert-butyl 6-(4-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1- yl)piperidin-1-yl)-2-azaspiro[3.3]heptane-2-carboxylate.
  • Step 4 Synthesis of tert-butyl 6-(4-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin- 1-yl)piperidin-1-yl)-2-azaspiro[3.3]heptane-2-carboxylate.
  • Step 5 Synthesis of 1-(1-(2-azaspiro[3.3]heptan-6-yl)piperidin-4-yl)-3-(4-phenoxyphenyl)-1H- pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 6 Synthesis of 1-(1-(2-methyl-2-azaspiro[3.3]heptan-6-yl)piperidin-4-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 1 Synthesis of tert-butyl 6-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2- azaspiro[3.3]heptane-2-carboxylate [0214] To a suspension of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol), tert-butyl 6-hydroxy-2-azaspiro[3.3]heptane-2-carboxylate (783 mg, 3.68 mmol) and PPh 3 (964 mg, 3.68 mmol) in THF (30 mL) was added dropwisely DIAD (743 mg, 3.68 mmol) at 0 oC over a period of 10 mins.
  • Step 2 Synthesis of 3-iodo-1-(2-azaspiro[3.3]heptan-6-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0215] To a mixture of tert-butyl 6-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2- azaspiro[3.3]heptane-2-carboxylate (620 mg, 1.36 mmol) in DCM (10 mL) was added HCl (4 M in 1,4-dioxane, 2 mL).
  • Step 3 Synthesis of tert-butyl 6-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2'- diaza[2,6'-bispiro[3.3]heptane]-2'-carboxylate [0216] To a mixture of 3-iodo-1-(2-azaspiro[3.3]heptan-6-yl)-1H-pyrazolo[3,4-d]pyrimidin- 4-amine (400 mg, 1.12 mmol) in DCM (10 mL) was added tert-butyl 6-oxo-2- azaspiro[3.3]heptane-2-carboxylate (356 mg, 1.69 mmol) and CH 3 COOH (2 drops).
  • Step 4 Synthesis of tert-butyl 6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)-2,2'-diaza[2,6'-bispiro[3.3]heptane]-2'-carboxylate [0217] To a mixture of tert-butyl 6-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2,2'- diaza[2,6'-bispiro[3.3]heptane]-2'-carboxylate (300 mg, 0.54 mmol) in 1,4-dioxane/H 2 O (10 mL/3 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2-dioxaborolane (242 mg, 0.82 mmol), PdCl 2 (dppf) (40 mg,
  • Step 5 Synthesis of 1-(2,2'-diaza[2,6'-bispiro[3.3]heptan]-6-yl)-3-(4-phenoxyphenyl)-1H- pyrazolo[3,4-d]pyrimidin-4-amine [0218] To a mixture of tert-butyl 6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)-2,2'-diaza[2,6'-bispiro[3.3]heptane]-2'-carboxylate (210 mg, 0.24 mmol) in DCM (10 mL) was added TFA (2 mL).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0221] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of tert-butyl (3aR,5s,6aS)-5-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)hexahydrocyclopenta[c]pyrrole-2(1H)-carboxylate.
  • Step 3 Synthesis of 1-((3aR,5s,6aS)-octahydrocyclopenta[c]pyrrol-5-yl)-3-(4-phenoxyphenyl)- 1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 4 Synthesis of tert-butyl 3-((3aR,5s,6aS)-5-(4-amino-3-(4-phenoxyphenyl)-1H- pyrazolo[3,4-d]pyrimidin-1-yl)hexahydrocyclopenta[c]pyrrol-2(1H)-yl)azetidine-1-carboxylate.
  • Step 5 Synthesis of 1-((3aR,5s,6aS)-2-(azetidin-3-yl)octahydrocyclopenta[c]pyrrol-5-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 6 Synthesis of 1-((3aR,5s,6aS)-2-(1-methylazetidin-3-yl)octahydrocyclopenta[c]pyrrol-5- yl)-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine
  • 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine 800 mg, 3.06 mmol
  • 1,4-dioxane/H 2 O 10 mL/2 mL
  • 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol)
  • XPhos Pd G2 120 mg, 0.16 mmol
  • K 3 PO 4 1.3 g, 6.13 mmol
  • Step 2 Synthesis of tert-butyl (3aR,5r,6aS)-5-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)hexahydrocyclopenta[c]pyrrole-2(1H)-carboxylate [0229] To a suspension of tert-butyl (3aR,5s,6aS)-5-hydroxyhexahydrocyclopenta[c]pyrrole- 2(1H)-carboxylate (681 mg, 3 mmol), 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4- amine (1.09 g, 3.6 mmol) and PPh 3 (1.57 g, 6 mmol) in THF (50 mL) was added dropwisely DIAD (1.21 g, 6 mmol) at 0oC over a period of 5 mins.
  • the reaction was stirred at room temperature for 12 hours under nitrogen atmosphere.
  • the mixture was diluted with water (30 mL), and the aqueous phase was extracted with EtOAc (30 mL x 3).
  • the combined organic phase was washed with brine (20 mL x 3), dried over Na 2 SO 4 , filtered and concentrated.
  • Step 3 Synthesis of 1-((3aR,5r,6aS)-octahydrocyclopenta[c]pyrrol-5-yl)-3-(4-phenoxyphenyl)- 1H-pyrazolo[3,4-d]pyrimidin-4-amine [0230] To a mixture of tert-butyl (3aR,5r,6aS)-5-(4-amino-3-(4-phenoxyphenyl)-1H- pyrazolo[3,4-d]pyrimidin-1-yl)hexahydrocyclopenta[c]pyrrole-2(1H)-carboxylate (600 mg, 1.17 mmol) in DCM (10 mL) was added HCl (4 M in 1,4-dioxane, 5 mL).
  • Step 4 Synthesis of tert-butyl 3-((3aR,5r,6aS)-5-(4-amino-3-(4-phenoxyphenyl)-1H- pyrazolo[3,4-d]pyrimidin-1-yl)hexahydrocyclopenta[c]pyrrol-2(1H)-yl)azetidine-1-carboxylate
  • Step 5 Synthesis of 1-((3aR,5r,6aS)-2-(azetidin-3-yl)octahydrocyclopenta[c]pyrrol-5-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine
  • Step 6 Synthesis of 1-((3aR,5r,6aS)-2-(1-methylazetidin-3-yl)octahydrocyclopenta[c]pyrrol-5- yl)-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0235] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of tert-butyl (3aR,5s,6aS)-5-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)hexahydrocyclopenta[c]pyrrole-2(1H)-carboxylate.
  • Step 3 Synthesis of 1-((3aR,5s,6aS)-octahydrocyclopenta[c]pyrrol-5-yl)-3-(4-phenoxyphenyl)- 1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 4 Synthesis of 1-((3aR,5s,6aS)-2-(1-methylpiperidin-4-yl)octahydrocyclopenta[c]pyrrol-5- yl)-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Example 13 Synthesis of 1-((3aR,5r,6aS)-2-(1-methylpiperidin-4- yl)octahydrocyclopenta[c]pyrrol-5-yl)-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4- amine (14).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0240] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of tert-butyl (3aR,5r,6aS)-5-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)hexahydrocyclopenta[c]pyrrole-2(1H)-carboxylate.
  • Step 3 Synthesis of 1-((3aR,5r,6aS)-octahydrocyclopenta[c]pyrrol-5-yl)-3-(4-phenoxyphenyl)- 1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 4 Synthesis of 1-((3aR,5r,6aS)-2-(1-methylpiperidin-4-yl)octahydrocyclopenta[c]pyrrol-5- yl)-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0245] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • aqueous phase was extracted with EtOAc (40 mL x 3), the combined organic phase was washed with brine (40 mL x 2), dried over Na 2 SO 4 , filtered and concentrated to give tert-butyl 9-hydroxy-3- azaspiro[5.5]undecane-3-carboxylate (560 mg, 93% yield) as a white solid, which was used directly for the next step.
  • LCMS m/z 214.2 [M-55] + .
  • Step 3 Synthesis of tert-butyl 9-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)-3-azaspiro[5.5]undecane-3-carboxylate.
  • Step 4 Synthesis of 3-(4-phenoxyphenyl)-1-(3-azaspiro[5.5]undecan-9-yl)-1H-pyrazolo[3,4- d]pyrimidin-4-amine.
  • tert-butyl 9-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)-3-azaspiro[5.5]undecane-3-carboxylate 150 mg, 0.27 mmol
  • DCM 10 mL
  • HCl 4 M in 1,4-dioxane, 2 mL
  • Step 5 Synthesis of tert-butyl 3-(9-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)-3-azaspiro[5.5]undecan-3-yl)azetidine-1-carboxylate.
  • Step 6 Synthesis of 1-(3-(azetidin-3-yl)-3-azaspiro[5.5]undecan-9-yl)-3-(4-phenoxyphenyl)-1H- pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 7 Synthesis of 1-((3aR,5r,6aS)-2-(1-methylazetidin-3-yl)octahydrocyclopenta[c]pyrrol-5- yl)-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Example 15 Synthesis of 1-(7-(1-methylazetidin-3-yl)-7-azaspiro[3.5]nonan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (16).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0253] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 3 Synthesis of 3-(4-phenoxyphenyl)-1-(7-azaspiro[3.5]nonan-2-yl)-1H-pyrazolo[3,4- d]pyrimidin-4-amine [0255] To a solution of tert-butyl 2-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)-7-azaspiro[3.5]nonane-7-carboxylate (360 mg, 0.68 mmol) in DCM (10 mL) was added HCl (4 M in 1,4-dioxane, 2 mL).
  • Step 4 Synthesis of tert-butyl 3-(2-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)-7-azaspiro[3.5]nonan-7-yl)azetidine-1-carboxylate [0256] To a mixture of 3-(4-phenoxyphenyl)-1-(7-azaspiro[3.5]nonan-2-yl)-1H-pyrazolo[3,4- d]pyrimidin-4-amine (300 mg, 0.70 mmol) in DCE (10 mL) was added tert-butyl 3-oxoazetidine- 1-carboxylate (181 mg, 1.06 mmol).
  • Step 5 Synthesis of 1-(7-(azetidin-3-yl)-7-azaspiro[3.5]nonan-2-yl)-3-(4-phenoxyphenyl)-1H- pyrazolo[3,4-d]pyrimidin-4-amine
  • Step 6 Synthesis of 1-(7-(1-methylazetidin-3-yl)-7-azaspiro[3.5]nonan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0258] To a mixture of 1-(7-(azetidin-3-yl)-7-azaspiro[3.5]nonan-2-yl)-3-(4-phenoxyphenyl)- 1H-pyrazolo[3,4-d]pyrimidin-4-amine (200 mg, 0.41 mmol) in DCE (10 mL) was added HCHO (71 mg, 37% in water, 0.83 mmol).
  • Example 16 Synthesis of 1-(7-(1-methylpiperidin-4-yl)-7-azaspiro[3.5]nonan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (17)
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0260] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of tert-butyl 2-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)-7-azaspiro[3.5]nonane-7-carboxylate [0261] To a suspension of tert-butyl 2-hydroxy-7-azaspiro[3.5]nonane-7-carboxylate (1.19 g, 4.95 mmol), 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (750 mg, 2.48 mmol) and PPh 3 (1.30 g, 4.95 mmol)in THF (50 mL) was added dropwisely DIAD (1.0 g, 4.95 mmol) at 0oC over a period of 15 mins.
  • DIAD 1.0 g, 4.95 mmol
  • the reaction was stirred at room temperature overnight under nitrogen atmosphere.
  • the mixture was diluted with water (50 mL), and the aqueous phase was extracted with EtOAc (30 mL x 3).
  • the combined organic phase was washed with brine (20 mL x 3), dried over Na 2 SO 4 , filtered and concentrated.
  • Step 3 Synthesis of 3-(4-phenoxyphenyl)-1-(7-azaspiro[3.5]nonan-2-yl)-1H-pyrazolo[3,4- d]pyrimidin-4-amine [0262] To a solution of tert-butyl 2-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)-7-azaspiro[3.5]nonane-7-carboxylate (360 mg, 0.68 mmol) in DCM (10 mL) was added HCl (4 M in 1,4-dioxane, 2 mL).
  • Step 4 Synthesis of 1-(7-(1-methylpiperidin-4-yl)-7-azaspiro[3.5]nonan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0263] To a mixture of 3-(4-phenoxyphenyl)-1-(7-azaspiro[3.5]nonan-2-yl)-1H-pyrazolo[3,4- d]pyrimidin-4-amine (100 mg, 0.23 mmol) in DCE (10 mL) was added 1-methylpiperidin-4-one (40 mg, 0.35 mmol).
  • Example 17 Synthesis of 1-(2-(1-methylazetidin-3-yl)-2-azaspiro[3.5]nonan-7-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (18).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0265] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of tert-butyl 7-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)-2-azaspiro[3.5]nonane-2-carboxylate [0266] To a suspension of tert-butyl 7-hydroxy-2-azaspiro[3.5]nonane-2-carboxylate (286 mg, 1.18 mmol) (227 mg, 1 mmol), 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (300 mg, 0.99 mmol) and PPh 3 (518 mg, 1.98 mmol)in THF (20 mL) was added dropwisely DIAD (400 mg, 1.98 mmol) at 0oC over a period of 5 mins.
  • DIAD 400 mg, 1.98 mmol
  • Step 3 Synthesis of 3-(4-phenoxyphenyl)-1-(2-azaspiro[3.5]nonan-7-yl)-1H-pyrazolo[3,4- d]pyrimidin-4-amine [0267] To a solution of 7-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)- 2-azaspiro[3.5]nonane-2-carboxylate (320 mg, 0.61 mmol) in DCM (10 mL) was added TFA (2 mL).
  • Step 4 Synthesis of tert-butyl 3-(7-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)-2-azaspiro[3.5]nonan-2-yl)azetidine-1-carboxylate [0268] To a mixture of 3-(4-phenoxyphenyl)-1-(2-azaspiro[3.5]nonan-7-yl)-1H-pyrazolo[3,4- d]pyrimidin-4-amine (200 mg, 0.46 mmol) in DCE (10 mL) was added tert-butyl 3-oxoazetidine- 1-carboxylate (162 mg, 0.93 mmol).
  • Step 5 Synthesis of 1-(2-(azetidin-3-yl)-2-azaspiro[3.5]nonan-7-yl)-3-(4-phenoxyphenyl)-1H- pyrazolo[3,4-d]pyrimidin-4-amine [0269] To a solution of tert-butyl 3-(7-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)-2-azaspiro[3.5]nonan-2-yl)azetidine-1-carboxylate (260 mg, 0.47 mmol) in DCM (10 mL) was added TFA (2 mL).
  • Step 6 Synthesis of 1-(2-(1-methylazetidin-3-yl)-2-azaspiro[3.5]nonan-7-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine
  • Example 18 Synthesis of 1-(2-(1-methylpiperidin-4-yl)-2-azaspiro[3.5]nonan-7-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (19).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0272] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of tert-butyl 7-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)-2-azaspiro[3.5]nonane-2-carboxylate [0273] To a suspension of tert-butyl 7-hydroxy-2-azaspiro[3.5]nonane-2-carboxylate (286 mg, 1.18 mmol) (227 mg, 1 mmol), 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (300 mg, 0.99 mmol) and PPh 3 (518 mg, 1.98 mmol)in THF (20 mL) was added dropwisely DIAD (400 mg, 1.98 mmol) at 0oC over a period of 5 mins.
  • DIAD 400 mg, 1.98 mmol
  • Step 3 Synthesis of 3-(4-phenoxyphenyl)-1-(2-azaspiro[3.5]nonan-7-yl)-1H-pyrazolo[3,4- d]pyrimidin-4-amine [0274] To a solution of 7-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)- 2-azaspiro[3.5]nonane-2-carboxylate (320 mg, 0.61 mmol) in DCM (10 mL) was added TFA (2 mL).
  • Step 4 Synthesis of 1-(2-(1-methylpiperidin-4-yl)-2-azaspiro[3.5]nonan-7-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0275] To a solution of 3-(4-phenoxyphenyl)-1-(2-azaspiro[3.5]nonan-7-yl)-1H-pyrazolo[3,4- d]pyrimidin-4-amine (260 mg, 0.61 mmol) in DCM (10 mL) was added 1-methylpiperidin-4-one (82 mg, 0.73 mmol).
  • Example 19 Synthesis of 1-(3-(oxetan-3-yl)-3-azaspiro[5.5]undecan-9-yl)-3-(4-phenoxyphenyl)- 1H-pyrazolo[3,4-d]pyrimidin-4-amine (20).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0277] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 6 Synthesis of 1-(3-(oxetan-3-yl)-3-azaspiro[5.5]undecan-9-yl)-3-(4-phenoxyphenyl)-1H- pyrazolo[3,4-d]pyrimidin-4-amine.
  • Example 20 Synthesis of 1-(6-(4-(oxetan-3-yl)piperazin-1-yl)spiro[3.3]heptan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (21) and 1-(4-(6-(4-amino-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)spiro[3.3]heptan-2-yl)piperazin-1-yl)ethan- 1-one (22).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine
  • 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine 800 mg, 3.06 mmol
  • 1,4-dioxane/H 2 O 10 mL/2 mL
  • 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol)
  • XPhos Pd G2 120 mg, 0.16 mmol
  • K 3 PO 4 1.3 g, 6.13 mmol
  • Step 2 Synthesis of 6-oxospiro[3.3]heptan-2-yl 4-methylbenzenesulfonate [0285] To a mixture of 6-hydroxyspiro[3.3]heptan-2-one (500 mg, 3.97 mmol) and TEA (800 mg, 7.93 mmol) in DCM (20 mL) was added TsCl (1.13 g, 5.95 mmol) and DMAP (50 mg, 0.416 mmol). The resulting mixture was stirred at room temperature for 12 hours, quenched with water (50 mL), and the aqueous phase was extracted with DCM (50 mL x 3).
  • Step 3 Synthesis of 6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)spiro[3.3]heptan-2-one [0286] To a solution of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (691 mg, 2.28 mmol) and 6-oxospiro[3.3]heptan-2-yl 4-methylbenzenesulfonate (639 mg, 2.28 mmol) in DMF (10 mL) was added Cs 2 CO 3 (1.12 g, 3.42 mmol).
  • Step 4 Synthesis of tert-butyl 4-(6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)spiro[3.3]heptan-2-yl)piperazine-1-carboxylate.
  • Step 5 Synthesis of 3-(4-phenoxyphenyl)-1-(6-(piperazin-1-yl)spiro[3.3]heptan-2-yl)-1H- pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 6 Synthesis of 1-(6-(4-(oxetan-3-yl)piperazin-1-yl)spiro[3.3]heptan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (21) [0289] To a mixture of 3-(4-phenoxyphenyl)-1-(6-(piperazin-1-yl)spiro[3.3]heptan-2-yl)-1H- pyrazolo[3,4-d]pyrimidin-4-amine (300 mg, 0.62 mmol) in DCM (20 mL) was added oxetan-3- one (89 mg, 1.24 mmol) and DIPEA (240 mg, 1.86 mmol).
  • Step 9 Synthesis of 1-(4-(6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)spiro[3.3]heptan-2-yl)piperazin-1-yl)ethan-1-one (22).
  • Example 21 Synthesis of 1-(6-(6-(oxetan-3-yl)-2,6-diazaspiro[3.3]heptan-2-yl)spiro[3.3]heptan- 2-yl)-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (23) and 1-(6-(6-(4-amino-3- (4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)spiro[3.3]heptan-2-yl)-2,6- diazaspiro[3.3]heptan-2-yl)ethan-1-one (24).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0295] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H 2 O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 2 Synthesis of 6-oxospiro[3.3]heptan-2-yl 4-methylbenzenesulfonate [0296] To a mixture of 6-hydroxyspiro[3.3]heptan-2-one (500 mg, 3.97 mmol) and TEA (800 mg, 7.93 mmol) in DCM (20 mL) was added TsCl (1.13 g, 5.95 mmol) and DMAP (50 mg, 0.416 mmol). The resulting mixture was stirred at room temperature for 12 hours, quenched with water (50 mL), and the aqueous phase was extracted with DCM (50 mL x 3).
  • Step 3 Synthesis of 6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)spiro[3.3]heptan-2-one [0297] To a solution of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (691 mg, 2.28 mmol) and 6-oxospiro[3.3]heptan-2-yl 4-methylbenzenesulfonate (639 mg, 2.28 mmol) in DMF (10 mL) was added Cs 2 CO 3 (1.12 g, 3.42 mmol).
  • Step 4 Synthesis of tert-butyl 6-(6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4- d]pyrimidin-1-yl)spiro[3.3]heptan-2-yl)-2,6-diazaspiro[3.3]heptane-2-carboxylate.
  • Step 5 Synthesis of 1-(6-(2,6-diazaspiro[3.3]heptan-2-yl)spiro[3.3]heptan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine.
  • Step 6 Synthesis of 1-(6-(6-(oxetan-3-yl)-2,6-diazaspiro[3.3]heptan-2-yl)spiro[3.3]heptan-2-yl)- 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (23) [0300] To a mixture of 1-(6-(2,6-diazaspiro[3.3]heptan-2-yl)spiro[3.3]heptan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (300 mg, 0.41mmol) in DCM (20 mL) was added oxetan-3-one (200 mg, 1.24 mmol) and DIPEA (211 mg, 1.64 mmol).
  • Step 7 Synthesis of 1-(6-(6-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)spiro[3.3]heptan-2-yl)-2,6-diazaspiro[3.3]heptan-2-yl)ethan-1-one (24) [0302] To a solution of 1-(6-(2,6-diazaspiro[3.3]heptan-2-yl)spiro[3.3]heptan-2-yl)-3-(4- phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (110 mg, 0.22 mmol) in THF (5 mL) and NaHCO 3 solution (5%, 3 mL) was added a solution of acetyl chloride (26 mg, 0.33 mmol) in THF (1 mL) drop-wisely at 0 oC.
  • Example 22 Synthesis of 1-(9-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1- yl)-3-azaspiro[5.5]undecan-3-yl)ethan-1-one. (25).
  • Step 1 Synthesis of 3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine [0304] To a mixture of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (800 mg, 3.06 mmol) in 1,4-dioxane/H2O (10 mL/2 mL) was added 4,4,5,5-tetramethyl-2-(4-phenoxyphenyl)-1,3,2- dioxaborolane (1.08 g, 3.67 mmol), XPhos Pd G2 (120 mg, 0.16 mmol) and K 3 PO 4 (1.3 g, 6.13 mmol).
  • Step 5 Synthesis of 3-(4-phenoxyphenyl)-1-(3-azaspiro[5.5]undecan-9-yl)-1H-pyrazolo[3,4- d]pyrimidin-4-amine.
  • Step 6 Synthesis of 1-(9-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-3- azaspiro[5.5]undecan-3-yl)ethan-1-one.
  • 3-(4-phenoxyphenyl)-1-(3-azaspiro [5.5] undecan-9-yl)-1H- pyrazolo [3,4-d] pyrimidin-4-amine 80 mg, 0.17 mmol
  • DCM 3 mL
  • Et3N 35mg, 0.34mmol
  • Example 23 Selective Inhibition of HCK and BTK [0311] The in vitro cellular efficacies of dual HCK/BTK inhibitors were assessed using CellTiter-Glo® Luminescent cell viability assay (Promega, Madison WI). Cells were seeded into 384 well plates with the EL406 Combination Washer Dispenser (BioTek Instruments, Inc.) and HCK/BTK inhibitors were injected into the cells culture media with the JANUS Automated Workstation (PerkinElmer Inc., Waltham MA). Cells were treated with serial diluted HCK/BTK inhibitors (20 ⁇ 0.0006 ⁇ M) for 72 hours at 37°C. Luminescent measurement was performed using the 2104 Envision® Multilabel Reader (PerkinElmer Inc.).
  • Tested compounds with activity in the tested concentration range are reported: ⁇ 0.5 nM ‘++++’, 0.5 nM to 1 nM ‘+++’, 1 nM to 10 nM ‘++’, and > 10 nM ‘+’.
  • Tested compounds with activity in the tested concentration range are reported: ⁇ 100 nM ‘A’, 100 nM to 500 nM ‘B’, 500 nM to 1000 nM ‘C’, and > 1000 nM ‘D’.
  • Compounds with values of ‘NA’ were not determined.
  • One aspect of the invention includes compounds characterized at ++++.
  • One aspect of the invention includes compounds characterized at +++.
  • One aspect of the invention includes compounds characterized at ++.
  • One aspect of the invention includes compounds characterized at +.
  • One aspect of the invention includes compounds characterized at A.
  • One aspect of the invention includes compounds characterized at B.
  • One aspect of the invention includes compounds characterized at C.
  • One aspect of the invention includes compounds characterized at D. Table B: Inhibition Data by Compound Number.
  • Test compounds for the experiments described herein were employed in free or salt form.

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Abstract

L'invention concerne des composés, et des sels pharmaceutiquement acceptables de ceux-ci, utiles en tant qu'inhibiteurs de l'activité kinase des kinases HCK et BTK. L'invention concerne également des compositions pharmaceutiques comprenant les composés ou des sels pharmaceutiquement acceptables et des procédés d'utilisation des composés, des sels pharmaceutiquement acceptables et des compositions pharmaceutiques dans le traitement de divers troubles, y compris d'une maladie proliférative telle que la gammopathie d'IgM, la mastocytose, le cancer.
PCT/US2025/028562 2024-05-14 2025-05-09 Nouveaux dérivés de spirocycle en tant qu'inhibiteurs de hck Pending WO2025240240A1 (fr)

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