Pyridopyrimidine derivatives useful as WEE1 kinase inhibitors

Abstract

The present disclosure provides compounds or pharma- ceutically acceptable salts thereof, as well as methods of using these compounds to inhibit Wee1 kinase, e.g., to treat cancer in a subject. The claimed compounds include Compound (I). [Formula 1] TIFF2024522304000130.tif38128

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Patent Application No. 63 / 196,744, filed June 4, 2021, the disclosure of which is incorporated herein by reference.

[0002] The present invention relates to compounds that are receptor protein tyrosine kinase inhibitors, and methods of using these compounds. In particular, the present invention relates to the use of such compounds in the treatment or prevention of one or more conditions in which the functional effect of one or more kinases is increased, such as cancer. [Background technology]

[0003] Wee1 belongs to a family of protein kinases involved in the terminal phosphorylation and inactivation of cyclin-dependent kinase 1-bound cyclin B, which leads to G cell cycle arrest in response to DNA damage. Wee1 was first identified in fission yeast, where Wee1 deficiency resulted in premature mitotic entry and replication of yeast cells with reduced size. Wee1 is the major kinase involved in inhibitory phosphorylation of tyrosines. Wee1 is a tyrosine kinase that phosphorylates and inactivates Cdc2 and is involved in G checkpoint signaling. More specifically, Wee1 inhibits the G 2 p53 is involved in G-M checkpoint signaling. p53 is a key regulator in the G checkpoint, so p53-deficient tumors are solely dependent on the G checkpoint after DNA damage. More specifically, p53 is involved in the G 1 p53-deficient tumors are at increased risk of G-terminal DNA damage, as they are a key regulator of the G-S checkpoint. 2 Wee1 inhibition selectively sensitizes such tumors to DNA-damaging agents, as they are solely dependent on the Wee1-M checkpoint.

[0004] Before a cell undergoes mitosis, the cell undergoes a G 1 -S, Intra-S, and G 2The Wee1 kinase is a key G kinase that mediates the progression of the inflammatory process through a tightly regulated cascade of G-M checkpoints. 2 This tyrosine kinase has emerged as a G-M checkpoint regulator. This tyrosine kinase negatively regulates entry into mitosis by catalyzing the inhibitory phosphorylation of Cdc2 (the human homolog of cyclin-dependent kinase 1 (CDK1)) on tyrosine-15 (Y15). This leads to inactivation of the Cdc2 / cyclin B complex, resulting in the G 2 Wee1 arrests cells in G-M and allows DNA repair. Such inhibition also occurs through Chk1-mediated inhibition of Cdc25 phosphatase, removing the inhibitory phosphorylation on Cdc2. Thus, entry into mitosis depends on the balance between the opposing activities of Wee1 and Chk1 / Cdc25. Wee1 inhibition therefore arrests cells in G-M and allows DNA repair. 2 It is predicted that Wee1 abrogates the -M arrest and drives cells into premature mitosis, a hypothesis that was confirmed by studies demonstrating that inhibition of Wee1 by either small molecule inhibitors or small interfering RNA leads to premature entry into mitosis and consequent cell death by mitotic death or apoptosis (S. Muller, J. Clinical. Oncology, 2015).

[0005] Wee1 is highly expressed in several cancer types, including hepatocellular carcinoma, breast cancer, cervical cancer, lung cancer, squamous cell carcinoma, diffuse intrinsic pontine glioma (DIPG), glioblastoma, medulloblastoma, leukemia, melanoma, and ovarian cancer (P. Reigan et al., Trends in Pharmacol. Sci., 2016).

[0006] Since there are few Wee1 inhibitors in clinical development, it is necessary to improve the selectivity of Wee1 inhibitors and the properties of inhibitors that allow targeting of specific cancer types, because inhibition of Wee1 activity can selectively promote the death of cancer cells with defective cell cycle checkpoints. At the same time, it has little effect on normal cells with normal cell cycle checkpoints. Therefore, Wee1 inhibitors can be used as targeted drugs for the treatment of cancer and other cell proliferation disorders.

[0007] The present disclosure provides compounds for use in treating, preventing, or managing one or more conditions in which the functional effect of one or more kinases is increased, such as cancer, using a compound(s) disclosed herein, or a pharma- ceutically acceptable salt or prodrug thereof, as well as methods of using these compounds, e.g., to inhibit Wee1 kinase, e.g., to treat, prevent, or manage cancer, in a subject.

[0008] All references cited herein are incorporated by reference in their entirety. Summary of the Invention

[0009] The present disclosure provides compounds for use in treating, preventing or managing one or more conditions in which the functional effect of one or more kinases is increased, such as cancer, using a compound(s) disclosed herein, or a pharma- ceutically acceptable salt or prodrug thereof, as well as methods of using these compounds, e.g., to inhibit Wee1 kinase in a subject, e.g., to treat, prevent or manage cancer.

[0010] The present disclosure provides a compound having the structure of Formula I:

[0011] [ka] (In the formula, R 1 Halo, C 1-6 Alkyl, C 3-8 Cycloalkyl, or C 2-6 alkenyl, R 2 , H, C 1-6 Alkyl or C 3-8 cycloalkyl) or a pharma- ceutically acceptable salt thereof. 1 The present disclosure provides compounds where R 1 The present disclosure provides compounds where R 1 The present disclosure provides compounds where R 1 C 1-4The present disclosure provides compounds in which R 1 The present disclosure provides compounds where R 1 C 3-7 The present disclosure provides compounds in which R 1 The present disclosure provides compounds where R 1 C 1-4 The present disclosure provides compounds in which R 1 The present disclosure provides compounds where R 2 C 1-6 The present disclosure provides compounds in which R 2 The present disclosure provides compounds where R 2 The present disclosure provides compounds where R 2 C 3-8 The present disclosure provides compounds in which R 2 The present disclosure provides compounds where R 2 is cyclopropyl.

[0012] The present disclosure provides a compound having the structure of Formula II:

[0013] [ka] or a pharma- ceutically acceptable salt thereof.

[0014] The present disclosure relates to the following compound:

[0015] [ka] or a pharma- ceutically acceptable salt thereof.

[0016] The present disclosure provides a compound having the structure of Formula III:

[0017] [ka] (In the formula, R 2 , H, C 1-6 Alkyl or C 3-8 The present disclosure provides a method for the preparation of a cycloalkyl group, or a pharma- ceutically acceptable salt thereof. 2 is H. The present disclosure provides compounds in which R 2 C 1-6 The present disclosure provides compounds in which R 2 is methyl.

[0018] The present disclosure provides a compound having the structure of Formula IV:

[0019] [ka] (wherein R1, R2 are H, methyl, ethyl, or propyl; optionally, R1 and R2 are joined to form a 3-6 membered ring; R3 is a C1-4 small alkyl, or a C3-6 cycloalkyl ring; and R4 is H, C1-3 alkyl, CF3, Omethyl, OCF3, OCF2H, CN, or halo), or a pharma- ceutically acceptable salt thereof.

[0020] The present disclosure provides a compound having the structure of Formula V

[0021] [ka] wherein R2 is H, methyl, ethyl, or propyl, R3 is a C1-4 small alkyl, or a C3-6 cycloalkyl ring, and R4 is H, C1-3 alkyl, CF3, Omethyl, OCF3, OCF2H, CN, or halo, or a pharma- ceutically acceptable salt thereof.

[0022] The present disclosure provides a compound having the structure of Formula VI:

[0023] [ka] wherein R2 is H, methyl, ethyl, or propyl, R3 is a C1-4 small alkyl, or a C3-6 cycloalkyl ring, and R4 is H, C1-3 alkyl, CF3, Omethyl, OCF3, OCF2H, CN, or halo, or a pharma- ceutically acceptable salt thereof.

[0024] The present disclosure also provides a compound of formula VIa: or a pharma- ceutically acceptable salt thereof:

[0025] [ka] (In the formula, R 2 ~R 6 is defined herein).

[0026] The present disclosure further provides a compound of formula VII:

[0027] [ka] In the formula, R 10 is defined herein.

[0028] The present disclosure provides pharmaceutical compositions comprising one or more compounds disclosed herein and one or more pharma- ceutically acceptable excipients. The present disclosure provides methods for inhibiting Wee1 in a patient in need of such treatment, comprising administering to the patient a compound or composition disclosed herein. The present disclosure provides methods for treating cancer in a patient in need of such treatment, comprising administering to the patient a compound disclosed herein. The present disclosure provides methods for treating cancer in a patient in need of such treatment, comprising administering to the patient a compound disclosed herein. The present disclosure provides methods for treating cancer, including cancers such as adrenal cortical tumors, AIDS-related cancers (e.g., AIDS-related lymphomas), anal cancer, appendix cancer, astrocytoma, atypical teratoma / rhabdoid tumors, basal cell tumors, cholangiocarcinoma, bladder cancer, bone cancer, brain cancer (e.g., glioblastoma), breast cancer, bronchial cancer, cancer of unknown primary site, carcinoid tumors, castration-resistant prostate cancer, central nervous system cancers (e.g., central nervous system atypical teratoma / rhabdoid tumors, central nervous system embryonal tumors, central nervous system lymphomas, primary central nervous system lymphomas, and the like. lymphoma), cervical cancer, chordoma, chondrosarcoma, chronic myeloproliferative disorder, colon cancer, colorectal cancer, craniopharyngioma, desmoplastic round cell tumor, diffuse large B-cell lymphoma, endometrial cancer, ependymoma, esophageal cancer, Ewing's sarcoma family of tumors, extracranial germ cell tumor, extragonadal cell tumor, extrahepatic bile duct cancer, gallbladder cancer, gastric (stomach) cancer, gastrointestinal carcinoid tumor, gastrointestinal stromal tumor, germ cell tumor, gestational trophoblastic tumor, glioma, head cancer, hepatocellular (liver) carcinoma, high-grade prostate cancer, tissue Myeloproliferation, hypopharyngeal cancer, Kaposi's sarcoma, renal (kidney) cancer, Langerhans cell histiocytosis, laryngeal cancer, leptomeningeal disease, lip cancer, low-grade prostate cancer, leukemia (e.g., chronic lymphocytic leukemia, chronic myeloid leukemia, hairy cell leukemia), lung cancer, lymphoma (Burkitt's lymphoma, central nervous system lymphoma, T-cell lymphoma (e.g., cutaneous T-cell lymphoma, Hodgkin's lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, lymphoplasmacytic lymphoma), intermediate-grade prostate cancer, Medulloblastoma, medulloepithelioma, melanoma, Merkel cell carcinoma, mesothelioma, metastatic squamous cell carcinoma of unknown primary, oral cavity cancer, multiple endocrine neoplasia syndrome, multiple myeloma, multiple myeloma / plasma cell neoplasm, mycosis fungoides, myelodysplastic syndrome, myelodysplastic / myeloproliferative neoplasm, myeloproliferative disorder, nasal cavity or paranasal sinus cancer, neck cancer, nasopharyngeal cancer, neck cancer, neuroblastoma, eye cancer, ocular melanoma, oral cavity cancer, oropharyngeal cancer, oral cavity cancer, osteosarcoma or malignant fibrous histiocytoma of bone, osteosarcoma or malignant fibrous histiocytoma of bone,Ovarian cancer, ovarian germ cell tumor, ovarian epithelial cancer, ovarian low malignant potential tumor, pancreatic cancer, papillomatosis, paranasal sinus or nasal cavity cancer, parathyroid cancer, penile cancer, pharyngeal cancer, moderately differentiated pineal parenchymal tumor, pineoblastoma or supratentorial primitive neuroectodermal tumor, pituitary tumor, pleuropulmonary blastoma, pregnancy cancer, prostate cancer, rectal cancer, renal pelvis cancer, airway cancer involving the NUT gene on chromosome 15, retinoblastoma, rhabdomyoblastoma, salivary gland cancer, sarcoma, Sézary syndrome, skin cancer, skin cancer , small intestine cancer, soft tissue sarcoma, spinal cord tumor, squamous cell carcinoma, squamous cell cervical carcinoma of unknown primary, occult primary supratentorial primitive neuroectodermal tumor, testicular cancer, laryngeal cancer, thymoma or thymic carcinoma, thyroid cancer, transitional cell carcinoma of the renal pelvis or ureter, rare cancers of childhood, ureteral cancer, urethral cancer, uterine sarcoma, vaginal cancer, vulvar cancer, Waldenstrom's macroglobulinemia (lymphoplasmacytic lymphoma), Wilms' tumor, or female cancer.

[0029] The present disclosure provides a method, wherein the cancer is breast cancer, prostate cancer, pancreatic cancer, lung cancer, colorectal cancer, ovarian cancer, liver cancer, melanoma, kidney cancer, central nervous system cancer, brain cancer such as glioblastoma, leukemia, or lymphoma. The present disclosure provides a method, wherein the compound is administered in combination with at least one additional therapeutic agent. The present disclosure provides a method, wherein the at least one additional therapeutic agent is a chemotherapeutic agent.The disclosure further provides that the chemotherapeutic agent is selected from the group consisting of busulfan, melphalan, chlorambucil, cyclophosphamide, ifosfamide, temozolomide, bendamustine, cisplatin, mitomycin C, bleomycin, carboplatin, camptothecin, irinotecan, topotecan, doxorubicin, epirubicin, aclarubicin, mitoxantrone, elliptinium, etoposide, 5-azacytidine, gemcitabine, 5-fluorouracil, methotrexate, 5-fluoro-2'-deoxy-uridine, fludarabinose, cyclophosphamide ... Vin, nelarabine, ara-C, pralatrexate, pemetrexed, hydroxyurea, thioguanine, colchicine, vinblastine, vincristine, vinorelbine, paclitaxel, ixabepilone, cabazitaxel, docetaxel, campath, panitumumab, metazotuzumab, navtuzumab, pimtuzumab, remortuzumab, bevacizumab, paltuzumab, trastuzumab, cetuximab, obinutuzumab, orfamuzumab, rituximab, alemtuzumab, thiemtuzumab, toximab, benzimidazole, benzo[b]azetidine ... Mab, dalemtuzumab, erlotuzumab, T-DM1, ofatumumab, dinutuximab, blinatumomab, ipilimumab, avastin, trastuzumab, rituximab, imatinib, gefitinib, erlotinib, osimertinib, afatinib, ceritinib, aretinib, crizotinib, erlotinib, lapatinib, sorafenib, nitinib, nilotinib, dasatinib, pazopanib, temsirolimus, everolimus, vorinostat, romidepsin, panobinostat, belinostat, tamoxifen, wherein the therapeutic agent is selected from the group consisting of letrozole, fulvestrant, mitoguazone, octreotide, retinoic acid, arsenic trioxide, zoledronic acid, bortezomib, carfilzomib, ixazomib, vismodegib, sonidegib, denosumab, thalidomide, lenalidomide, venetoclax, aldesleukin (recombinant human interleukin-2), sipuleucel-T (prostate cancer therapeutic vaccine), palbociclib, olaparib, niraparib, rucaparib, tazoparib, and combinations thereof.

[0030] The present disclosure provides a method for reducing the activity of a kinase encoded by the gene WEE1, comprising contacting the kinase with an inhibitory amount of a compound disclosed herein. The present disclosure provides a method carried out in vitro. The present disclosure provides a method carried out in a subject. The present disclosure provides a method for treating or preventing a Wee1 mediated disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound disclosed herein, or a pharma- ceutically acceptable salt or prodrug thereof, or a pharmaceutical composition thereof. The present disclosure relates to a method for treating a disease, the method comprising administering to a patient ... , primary central nervous system lymphoma), cervical cancer, chordoma, chondrosarcoma, chronic myeloproliferative disorder, colon cancer, colorectal cancer, craniopharyngioma, desmoplastic round cell tumor, diffuse large B-cell lymphoma, endometrial cancer, ependymoma, esophageal cancer, Ewing's sarcoma family of tumors, extracranial germ cell tumor, extragonadal cell tumor, extrahepatic bile duct cancer, gallbladder cancer, gastric (stomach) cancer, gastrointestinal carcinoid tumor, gastrointestinal stromal tumor, germ cell tumor, gestational trophoblastic tumor, glioma, head Neck cancer, hepatocellular (liver) cancer, high-grade prostate cancer, histiocytic proliferation, hypopharyngeal cancer, Kaposi's sarcoma, renal (kidney) cancer, Langerhans cell histiocytosis, laryngeal cancer, leptomeningeal disease, lip cancer, low-grade prostate cancer, leukemia (e.g., chronic lymphocytic leukemia, chronic myeloid leukemia, hairy cell leukemia), lung cancer, lymphoma (Burkitt's lymphoma, central nervous system lymphoma), T-cell lymphoma (e.g., cutaneous T-cell lymphoma, Hodgkin's lymphoma, , non-Hodgkin's lymphoma, lymphoplasmacytic lymphoma), intermediate-grade prostate cancer, medulloblastoma, medulloepithelioma, melanoma, Merkel cell carcinoma, mesothelioma, metastatic squamous cell carcinoma of unknown primary site, oral cancer, multiple endocrine neoplasia syndrome, multiple myeloma, multiple myeloma / plasma cell neoplasm, mycosis fungoides, myelodysplastic syndrome, myelodysplastic / myeloproliferative neoplasm, myeloproliferative disorder, nasal or paranasal sinus cancer, neck cancer, nasopharyngeal cancer, cervical cancer, neuroblastoma, eye cancer, ocular melanoma,Oral cavity cancer, oropharyngeal cancer, oral cavity cancer, osteosarcoma or malignant fibrous histiocytoma of bone, osteosarcoma or malignant fibrous histiocytoma, ovarian cancer, ovarian germ cell tumor, ovarian epithelial cancer, ovarian low malignant potential tumor, pancreatic cancer, papillomatosis, paranasal sinus or nasal cavity cancer, parathyroid cancer, penile cancer, pharyngeal cancer, moderately differentiated pineal parenchymal tumor, pineoblastoma or supratentorial primitive neuroectodermal tumor, pituitary tumor, pleuropulmonary blastoma, pregnancy cancer, prostate cancer, rectal cancer, renal pelvis cancer, airway cancer involving the NUT gene on chromosome 15, retinoblastoma, rhabdomyoblastoma, salivary adenocarcinoma, sarcoma, Sezary syndrome, skin cancer, skin cancer, small intestine cancer, soft tissue sarcoma, spinal cord tumor, squamous cell carcinoma, squamous cell cervical carcinoma of unknown primary, occult primary supratentorial primitive neuroectodermal tumor, testicular cancer, throat cancer, thymoma or thymic carcinoma, thyroid cancer, transitional cell carcinoma of the renal pelvis or ureter, rare cancers of childhood, ureteral cancer, urethral cancer, uterine sarcoma, vaginal cancer, vulvar cancer, Waldenstrom's macroglobulinemia (lymphoplasmacytic lymphoma), Wilms' tumor, or female cancer.

[0031] The present disclosure provides the use of the compounds and compositions of the present disclosure for the manufacture of a medicament for the prevention and / or treatment of the indications described herein.

[0032] According to further embodiments, the present disclosure provides for the use of the compounds and pharmaceutical compositions described herein in a pharmaceutical amount, most preferably for use as a pharmaceutical for treating, for example, a disease or disorder described herein in a subject.

[0033] According to yet another embodiment, the present disclosure provides for the use of the above pharmaceutical composition and at least one additional therapeutic agent in a pharmaceutical, most preferably in an effective amount for use as a pharmaceutical for treating, e.g., a disease or disorder associated with a disease described herein, in a subject.

[0034] The present disclosure relates to a method for treating and / or preventing a disease or condition described herein in a patient, comprising selecting a patient in need of treatment and / or prevention of said disease or condition described herein, and administering to the patient a therapeutically effective amount of a composition of the present disclosure, thereby treating and / or preventing the disease in the patient. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0035] As used herein, the term "active pharmaceutical ingredient" ("API") or "pharmaceutical active agent" may be used as disclosed herein and is a drug or agent intended for use in the human or animal body to cure, alleviate, prevent, or diagnose a disease, illness, physical injury, or pathological condition, to enable identification of a state, condition, or function of the physical or mental condition, to replace active substances produced by the human or animal body or bodily fluids, to defend against, eliminate, or render harmless pathogens, parasites, or exogenous substances, or to affect a state, condition, or function of the physical or mental condition. Drugs in use can be found in reference books such as, for example, the Rote Liste or the Merck Index.

[0036] "Pharmaceutically acceptable salts" refers to derivatives of the disclosed compounds, where the parent compound is modified by making acid or base salts thereof. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines. Pharmaceutically acceptable salts include conventional non-toxic salts formed, for example, from non-toxic inorganic or organic acids. For example, such conventional non-toxic salts include those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, sulfonic acid, sulfamic acid, phosphoric acid, nitric acid, and the like, as well as those prepared from organic acids such as amino acids, acetic acid, propionic acid, succinic acid, glycolic acid, stearic acid, lactic acid, malic acid, tartaric acid, citric acid, ascorbic acid, palmitic acid, maleic acid, hydroxymaleic acid, phenylacetic acid, glutamic acid, benzoic acid, salicylic acid, sulfanilic acid, 2-acetoxybenzoic acid, fumaric acid, toluenesulfonic acid, methanesulfonic acid, ethanedisulfonic acid, oxalic acid, isothioic acid, and the like, as well as other salts known to those skilled in the pharmaceutical arts. Lists of suitable salts can be found in Remington's Pharmaceutical Sciences, 18th Edition (ed. Alfonso R. Gennaro; Mack Publishing Company, Easton, PA, 1990); Remington: the Science and Practice of Pharmacy, 19th Edition (Lippincott, Williams & Wilkins, 1995); Handbook of Pharmaceutical Excipients, 3rd Edition (ed. Arthur H. Kibbe, Amer. Pharmaceutical Assoc., 1999); Pharmaceutical Codex: Principles and Practice of Pharmaceutics, 12th Edition (Walter Lund ed. Pharmaceutical Press, London, 1994).and Goodman and Gilman's: the Pharmacological Basis of Therapeutics (eds. Louis S. Goodman and Lee E. Limbird, McGraw Hill, 1992), the disclosures of which are incorporated herein by reference.

[0037] An amount, as used herein, is "effective" if it produces an effect in a subject. As used herein, the term "effective amount" refers to an amount of a compound or composition that is sufficient to significantly induce positive benefits, including those disclosed herein, either individually or in combination, but low enough to avoid serious side effects, i.e., to provide a reasonable benefit-to-risk ratio within the scope of the sound judgment of a person skilled in the art. For those skilled in the art, effective amounts, as well as dosages and frequency of administration, can be determined according to standard methodologies of mere routine experimentation based on their knowledge and the present disclosure.

[0038] As used herein, the terms "subject" and "patient" are used interchangeably. As used herein, the term "patient" refers to an animal, preferably a mammal (e.g., non-primates (e.g., cows, pigs, horses, cats, dogs, rats, etc.) and primates (e.g., monkeys and humans), most preferably humans). In some embodiments, the subject is a non-human animal, such as a farm animal (e.g., horse, pig, or cow) or a pet (e.g., dog or cat). In a preferred embodiment, the subject is an elderly human. In another embodiment, the subject is a human. In another embodiment, the subject is an adult. In yet another embodiment, the subject is an infant.

[0039] As used herein, the phrase "pharmaceutical acceptable" means approved by a regulatory agency of the Federal or state government, or listed in the United States Pharmacopoeia, the European Pharmacopoeia, or other generally recognized pharmacopoeias for use in animals, and more particularly in humans.

[0040] As used herein, the terms "prevent," "preventing," and "prevention" in the context of administration of a therapy to a subject refer to prevention or inhibition of the recurrence, onset, and / or progression of a disease or condition, or combination of therapies (e.g., a combination of prophylactic or therapeutic agents).

[0041] As used herein, the terms "therapy" and "therapy" can refer to any method(s), composition(s), and / or agent(s) that can be used in the prevention, treatment and / or management of a disease or condition, or one or more symptoms thereof.

[0042] As used herein, the terms "treat," "treatment," and "treating" in the context of administration of a therapy to a subject refer to the reduction or inhibition of the progression and / or duration of a disease or condition, the reduction or amelioration of the severity of a disease or condition, and / or the amelioration of one or more symptoms thereof that results from the administration of one or more therapies.

[0043] As used herein, the term "about," when used in conjunction with a recited numerical value or range, has the meaning reasonably ascribed to it by one of ordinary skill in the art, i.e., indicating some more or some less than the recited value or range.

[0044] Active Agent In one aspect, a compound having the chemical structure of Formula I:

[0045] [ka] (In the formula, R 1 Halo, C 1-6 Alkyl, C 3-8 Cycloalkyl or C 2-6 alkenyl, R 2 , H, C 1-6 Alkyl or C 3-8cycloalkyl) or a pharma- ceutically acceptable salt thereof; and methods of using these compounds to inhibit WEE1 kinase and treat cancer in a subject. 1 In a further embodiment, R 1 is chloro, bromo, fluoro, or iodo. In a further embodiment, R 1 is chloro or bromo. In a further embodiment, R 1 is C 1-4 In a further embodiment, R 1 is methyl, ethyl, n-propyl, or i-propyl. 1 is methyl. In a further embodiment, R 1 is C 3-7 In a further embodiment, R 1 is cyclopropyl. In a further embodiment, R 1 is C 1-4 In a further embodiment, R 1 is vinyl or isopropenyl. In yet other embodiments, R 1 is vinyl. In a further embodiment, R 1 In another embodiment, R 2 is H. In a further embodiment, R 2 is C 1-6 In a further embodiment, R 2 is methyl, ethyl, propyl, butyl, pentyl, or hexyl. In a further embodiment, R 2 is methyl, ethyl, n-propyl, or i-propyl. 2 is C 3-8 In a further embodiment, R 2 is cyclobutyl, cyclopentyl, or cyclohexyl. In a further embodiment, R 2 is cyclopropyl.

[0046] In certain embodiments, the present disclosure provides a compound having the structure of formula II:

[0047] [ka] or a pharma- ceutically acceptable salt thereof. 1 is halo, C 1-6 Alkyl, C 3-8 Cycloalkyl, or C 2-6 alkenyl, R 2 is H, C 1-6 Alkyl or C 3-8 In some embodiments, R 1 In a further embodiment, R 1 is chloro, bromo, fluoro, or iodo. 1 is chloro or bromo. In yet other embodiments, R 1 is C 1-4 In yet another embodiment, R 1 is methyl, ethyl, n-propyl, or i-propyl. 1 is methyl. In other embodiments, R 1 is C 3-7 In yet another embodiment, R 1 is cyclopropyl. In yet other embodiments, R 1 is C 1-4 In a further embodiment, R 1 is vinyl or isopropenyl. In other embodiments, R 1 is vinyl. In a further embodiment, R 1 is isopropenyl.

[0048] In certain embodiments, the present disclosure provides a compound having the structure of formula IIa:

[0049] [ka] or a pharma- ceutically acceptable salt thereof. 1 is halo, C 1-6Alkyl, C 3-8 Cycloalkyl, or C 2-6 alkenyl, R 2 is H, C 1-6 Alkyl or C 3-8 In some embodiments, R 1 In a further embodiment, R 1 is chloro, bromo, fluoro, or iodo. 1 is chloro or bromo. In yet other embodiments, R 1 is C 1-4 In yet another embodiment, R 1 is methyl, ethyl, n-propyl, or i-propyl. 1 is methyl. In other embodiments, R 1 is C 3-7 In a further embodiment, R 1 is cyclopropyl. In yet other embodiments, R 1 is C 1-4 In a further embodiment, R 1 is vinyl or isopropenyl. In other embodiments, R 1 is vinyl. In a further embodiment, R 1 is isopropenyl.

[0050] In certain embodiments, the present disclosure provides a compound having the structure of formula III

[0051] [ka] or a pharma- ceutically acceptable salt thereof, 2 , H, C 1-6 Alkyl or C 3-8 In certain embodiments, R 2 is H. In certain embodiments, R 2 is C 1-6 In certain embodiments, R is alkyl, or a pharma- ceutically acceptable salt thereof. 2 is methyl.

[0052] In certain embodiments, the present disclosure provides a compound having the structure of formula IV:

[0053] [ka] or a pharma- ceutically acceptable salt thereof (wherein R1, R2 are H, Me, Et, Pr, and optionally R1 and R2 are joined to form a 3-6 membered ring, R3 is a C1-4 small alkyl, C3-6 cycloalkyl ring, and R4 is H, C1-3 alkyl, CF3, OMe, OCF3, OCF2H, CN, halo).

[0054] In certain embodiments, the present disclosure provides a compound of formula IVa:

[0055] [ka] or a pharma- ceutically acceptable salt thereof. 1 and R 2 are independently H, methyl, ethyl, or propyl, or R 1 and R 2 are joined together to form a 3-6 membered ring, R 3 is C 1-4 Alkyl or C 3-6 cycloalkyl, R 4 , H, C 1-3 Alkyl, CF 3 , methoxy, OCF 3 , OCF 2 H, CN, or halo. In some embodiments, R 1 and / or R 2 is H. In other embodiments, R 1 and / or R 2 is methyl. In a further embodiment, R 1 and / or R 2 is ethyl. In yet another embodiment, R 1 and / or R 2 is propyl. In a further embodiment, R1 and R 2 are joined together to form a 3-6 membered ring. In another embodiment, R 1 and R 2 are joined together to form a cyclopropyl group. In a further embodiment, R 3 is methyl, ethyl, propyl, or butyl, etc. 1-4 In yet another embodiment, R 3 is cyclopropyl, etc. 3-6 In a further embodiment, R 4 is H. In other embodiments, R 4 is C such as methyl 1-3 In a further embodiment, R 4 CF 3 In yet another embodiment, R 4 In a further embodiment, R 4 is OCF 3 In another embodiment, R 4 is OCF 2 H. In a further embodiment, R 4 is CN. In yet another embodiment, R 4 is halo, such as fluoro, chloro, or bromo.

[0056] In certain embodiments, the present disclosure provides a compound having the structure of Formula V:

[0057] [ka] or a pharma- ceutically acceptable salt thereof, wherein R2 is H, Me, Et, Pr, R3 is C1-4 small alkyl, C3-6 cycloalkyl ring, and R4 is H, C1-3 alkyl, CF3, OMe, OCF3, OCF2H, CN, halo.

[0058] In certain embodiments, the present disclosure provides a compound of formula Va:

[0059] [ka] or a pharma- ceutically acceptable salt thereof, 2 is H, methyl, ethyl, or propyl; R 3 is C 1-4 Alkyl or C 3-6 cycloalkyl, R 4 , H, C 1-3 Alkyl, CF 3 , methoxy, OCF 3 , OCF 2 H, CN, or halo. 2 is H. In other embodiments, R 2 is methyl. In a further embodiment, R 2 is ethyl. In another embodiment, R 2 is propyl. In a further embodiment, R 3 is methyl, ethyl, propyl, or butyl, etc. 1-4 In yet another embodiment, R 3 is cyclopropyl, etc. 3-6 In a further embodiment, R 4 is H. In other embodiments, R 4 is C such as methyl 1-3 In a further embodiment, R 4 CF 3 In yet another embodiment, R 4 In a further embodiment, R 4 is OCF 3 In another embodiment, R 4 is OCF 2 H. In a further embodiment, R 4 is CN. In yet another embodiment, R 4 is halo, such as fluoro, chloro, or bromo.

[0060] In certain embodiments, the present disclosure provides a compound having the structure of formula VI:

[0061] [ka] or a pharma- ceutically acceptable salt thereof, wherein R2 is H, Me, Et, Pr, R3 is C1-4 small alkyl, C3-6 cycloalkyl ring, and R4 is H, C1-3 alkyl, CF3, OMe, OCF3, OCF2H, CN, halo.

[0062] In other embodiments, the present disclosure provides a compound of formula VIb:

[0063] [ka] or a pharma- ceutically acceptable salt thereof, 2 is H, methyl, ethyl, or propyl; R 3 is C 1-4 Alkyl or C 3-6 cycloalkyl, R 4 , H, C 1-3 Alkyl, CF 3 , methoxy, OCF 3 , OCF 2 H, CN, or halo. 2 is H. In other embodiments, R 2 is methyl. In a further embodiment, R 2 is ethyl. In another embodiment, R 2 is propyl. In a further embodiment, R 3 is methyl, ethyl, propyl, or butyl, etc. 1-4 In yet another embodiment, R 3 is cyclopropyl, etc. 3-6 In a further embodiment, R 4 is H. In other embodiments, R 4 is C such as methyl 1-3 In a further embodiment, R 4 CF 3 In yet another embodiment, R 4 In a further embodiment, R 4 is OCF 3 In another embodiment, R 4 is OCF2 H. In a further embodiment, R 4 is CN. In yet another embodiment, R 4 is halo, such as fluoro, chloro, or bromo.

[0064] In a further embodiment, the present disclosure provides a compound of formula VIa:

[0065] [ka] or a pharma- ceutically acceptable salt thereof, 2 is H, methyl, ethyl, or propyl; R 3 , H, C 1-4 Alkyl or C 3-6 cycloalkyl, R 4 , H, C 1-3 Alkyl, CF 3 , methoxy, OCF 3 , OCF 2 H, CN, or halo; R 5 and R 6 are independently H, halo, or C 1-6 In some embodiments, R 2 is H. In other embodiments, R 2 is methyl. In a further embodiment, R 2 is ethyl. In another embodiment, R 2 is propyl. In a further embodiment, R 3 is C 1-4 alkyl, such as methyl, ethyl, propyl, or butyl. In other embodiments, R 3 is C 3-6 In a further embodiment, R 4 is H. In other embodiments, R 4 is C such as methyl 1-3 In a further embodiment, R 4 CF 3 In yet another embodiment, R 4 In a further embodiment, R 4 is OCF3 In another embodiment, R 4 is OCF 2 H. In a further embodiment, R 4 is CN. In yet another embodiment, R 4 is halo, such as fluoro, chloro, or bromo. In a further embodiment, R 5 and / or R 6 is H. In other embodiments, R 5 and / or R 6 is halo, such as fluoro, chloro, or bromo, or halo, such as fluoro. 5 and / or R 6 is C 1-6 Alkyl, for example methyl, ethyl, propyl, or butyl, or for example methyl.

[0066] In other embodiments, the present disclosure provides a compound of formula IVII:

[0067] [ka] or a pharma- ceutically acceptable salt thereof, 10 are H, OH, and NH. 2 , NH(C 1-6 alkyl), or N(C 1-6 Alkyl)(C 1-6 In some embodiments, R 10 is H. In other embodiments, R 10 is OH. In a further embodiment, R 10 NH 2 In yet another embodiment, R 10 is NH(C 1-6 In still further embodiments, R 10 is N(C 1-6 Alkyl)(C 1-6 alkyl), e.g. N(CH 3 ) 2 It is.

[0068] Further embodiments of the present disclosure provide compounds shown in Table 1, or a pharma- ceutically acceptable salt thereof.

[0069] [Table 1-1]

[0070] [Table 1-2]

[0071] [Table 1-3]

[0072] [Table 1-4]

[0073] [Table 1-5]

[0074] [ka]

[0075] [ka]

[0076] [ka]

[0077] [ka]

[0078] [ka]

[0079] [ka]

[0080] [ka]

[0081] [ka]

[0082] [ka]

[0083] In exemplary embodiments, the formulations disclosed herein contain an active agent disclosed herein at about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 5 In exemplary embodiments, the formulations disclosed herein may contain an active agent in a concentration of about 1 to about 20%, about 5% to about 25%, about 10% to about 20%, or about 15% to about 18%, about 30% to about 70%, about 35% to about 65%, about 63.13%, and about 40% to about 64% w / w.

[0084] In the exemplary formulations disclosed herein, the active agents disclosed herein represent about 1% to 75% by weight, preferably 2% to 30% by weight, and more preferably 5% to 20% by weight of the total weight of the formulation.

[0085] In other embodiments, the pharmaceutical compositions disclosed herein further comprise one or more additional ingredients, such as a pharma- ceutically compatible carrier, binder, viscosity modifier, filler, suspending agent, flavoring agent, sweetening agent, disintegrating agent, surfactant, preservative, lubricant, coloring agent, excipient, solubilizing agent, wetting agent, stabilizer, humectant, anti-adhesive agent, paracellular activator, anti-foaming agent, antioxidant, chelating agent, anti-fungal agent, anti-bacterial agent, or one or more combinations thereof.

[0086] Pharmaceutical dosage form The compounds and compositions disclosed herein may be provided in the form of pharmaceutical compositions, e.g., pharmaceutical dosage forms, such as minicapsules, capsules, tablets, implants, troches, lozenges (minitablets), temporary or permanent suspensions, ovules, suppositories, wafers, chewable tablets, quick or fast dissolving tablets, effervescent tablets, granules, films, sprinkles, pellets, beads, pills, powders, powders, platelets, strips, or sachets. The compositions may also be administered after being mixed with, for example, yogurt or fruit juice and swallowed, or may be administered with a drink or beverage. These forms are well known in the art and are appropriately packaged. The compositions may be formulated for oral or rectal delivery.

[0087] Tablets prepared for oral administration according to the present invention and manufactured using direct compression generally contain other inert additives (e.g., binders, lubricants, disintegrants, fillers, stabilizers, surfactants, colorants, etc.). Binders are used to impart cohesiveness to the tablet, thus ensuring that the tablet remains intact after compression. Suitable binder materials include, but are not limited to, starches (including corn starch and pregelatinized starch), gelatin, sugars (including sucrose, glucose, dextrose and lactose), polyethylene glycol, waxes, and natural and synthetic gums, such as sodium acacia alginate, polyvinylpyrrolidone, cellulosic polymers (including hydroxypropylcellulose, hydroxypropylmethylcellulose, methylcellulose, microcrystalline cellulose, ethylcellulose, hydroxyethylcellulose, etc.), and Veegum. Lubricants are used to facilitate tablet manufacture, promote powder flow, and prevent particle capping (i.e., particle breakage) when pressure is released. Useful lubricants are, for example, magnesium stearate, calcium stearate, stearic acid, and hydrogenated vegetable oils (preferably consisting of about 1% to 5% by weight, most preferably less than about 2% by weight, of hydrogenated and purified triglycerides of stearic and palmitic acids). The lubricant may be present, for example, in a concentration of about 0.25% to about 3% by weight, 0.5% to about 2.0% by weight, or about 0.75% to about 1.5%.

[0088] Disintegrants are used to facilitate the disintegration of tablets, thereby increasing the erosion rate relative to the dissolution rate, and are generally starches, clays, celluloses, algins, gums, or crosslinked polymers (e.g., crosslinked polyvinylpyrrolidone). Fillers include, for example, materials such as silicon dioxide, titanium dioxide, alumina, talc, kaolin, powdered cellulose, and microcrystalline cellulose, as well as soluble materials such as mannitol, urea, sucrose, lactose, lactose monohydrate, dextrose, sodium chloride, and sorbitol. Dissolution enhancers, including solubilizers themselves, emulsifiers, and complexing agents (e.g., cyclodextrins), may also be advantageously included in the formulation. Stabilizers well known in the art are used to inhibit or retard drug decomposition reactions, including, for example, oxidative reactions. Disintegrants may be present, for example, in concentrations of about 0.25% to about 3% by weight, 0.5% to about 2.0% by weight, or about 0.75% to about 1.5% by weight.

[0089] Shellac, also called purified lac, is a refined product obtained from the resinous secretions of insects. This coating dissolves in media with a pH > 7.

[0090] Colorants, antiblocking agents, surfactants, defoamers, lubricants, stabilizers such as hydroxypropyl cellulose, acids / bases may be added to the coating in addition to plasticizers to solubilize or disperse the coating materials and improve coating performance and the coated product.

[0091] In practicing the methods disclosed herein, the compositions of the present disclosure may be administered to mammalian species, such as dogs, cats, humans, etc., and therefore may be incorporated into conventional systemic administration dosage forms, such as tablets, capsules, or elixirs, which may also contain necessary carrier materials, excipients, viscosity adjusters, lubricants, buffers, antibacterial agents, bulking agents (such as mannitol), antioxidants (ascorbic acid, sodium bisulfate), and the like.

[0092] The dose administered may be carefully adjusted according to the age, weight and condition of the patient, as well as the route of administration, dosage form and regimen and the desired result.

[0093] The compositions of the present disclosure may be administered in single or divided dose form one to four times daily, or may be administered multiple times daily. It may be advisable to start the patient on a low dose combination and gradually work up to a higher dose combination.

[0094] Tablets of various sizes can be prepared (e.g., tablets weighing from about 2 to 2000 mg total), which contain one or more active ingredients, the remainder being physiologically acceptable carriers of other substances in accordance with accepted practice. Gelatin capsules can be similarly formulated.

[0095] Liquid formulations can also be prepared by dissolving or suspending one or a combination of active agents in a conventional liquid vehicle acceptable for administration to provide the desired dosage, for example, 1 to 4 teaspoons.

[0096] Dosage forms can be administered to the patient, for example, 1, 2, 3, 4, 5, 6 times per day, or other multiple dose regimens.

[0097] To allow for more precise control of the dosing schedule, the active agents may be administered separately in individual dosage units, simultaneously, or at carefully coordinated times. Each agent may be individually formulated in separate unit dosage forms in a manner similar to that described herein.

[0098] In formulating the compositions, the active agents in the amounts described herein may be combined in accordance with accepted practices with physiologically acceptable vehicles, carriers, excipients, binders, viscosity modifiers, preservatives, stabilizers, flavorings, and the like in a particular type of unit dosage form.

[0099] When formulated as a capsule, the capsule can be a hard or soft gelatin capsule, a starch capsule, a cellulose capsule, or a multiparticulate system. A multiparticulate system consists of a dosage form based on a plurality of drug-loaded spheres, which can be prepared by layering the drug on a core, usually a sugar-starch mixture sphere with a diameter of about 0.8 mm, until a sufficient level is reached, and then providing a drug release barrier around the drug-loaded sphere. Drug-loaded spheres can also be made by wet massing a mixture of drug and excipients, forcing the wet mass through a perforated screen to form short strands, which are rolled in a spheronizer before drying, and applying a drug release barrier. The drug release barrier can be a wax, such as carnauba wax or glyceryl fatty acid esters, or a polymeric barrier, such as a mixture of ethylcellulose and hydroxypropylmethylcellulose. These work well for moderately soluble drugs with doses in the milligram to less than a few hundred milligram units per day. Multiparticulate systems are usually filled into capsules to provide a unit dosage form due to the damage caused to such particles when attempting to compress them into tablets. The total dose contained in a single unit is constrained by the amount of filling possible in a hard gelatin capsule of easily swallowable size, usually not more than a few hundred milligrams. Although not limited to capsules, such dosage forms can be further coated, for example, with a seal coating, an enteric coating, a sustained release coating, or a targeted delayed release coating. These various coatings are known in the art, but for clarity, a brief description of a seal coating, or a coating with an isolating layer is provided. A thin layer up to 20 microns thick can be applied for a variety of reasons, including reducing particle porosity, reducing dust, chemical protection, taste masking, reducing odor, minimizing gastrointestinal irritation, etc. The isolating effect is proportional to the thickness of the coating. Water-soluble cellulose ethers are preferred for this application. A combination of HPMC and ethylcellulose, or EUDRAGIT® E100, may be particularly suitable for taste masking applications. Conventional enteric coating materials listed elsewhere may also be applied to form the isolating layer.

[0100] The oral dosage form of the controlled release active agent formulation of the present disclosure can be in the form of a multiparticulate formulation or a tablet. The term "multiparticulate" as used herein includes discrete particles, pellets, mini-tablets and mixtures or combinations thereof. The multiparticulate oral dosage form according to the present disclosure can include a blend of two or more populations of particles, pellets, or mini-tablets having different in vivo and / or in vitro release characteristics. For example, the multiparticulate oral dosage form can include a blend of an immediate release component and a controlled release component contained in a suitable capsule, such as a hard or soft gelatin capsule. When the multiparticulate formulation is filled into a capsule, it can be administered by swallowing the capsule or by opening the capsule and sprinkling the contents on food. Alternatively, the multiparticulate can be present in a sachet.

[0101] The particles and one or more auxiliary excipient materials can be compressed into tablet form, such as a multi-layer tablet.Typically, a multi-layer tablet can contain the same or different levels of the same active ingredient with the same or different release characteristics, or can contain two layers that contain different active ingredients in each layer.Such multi-layer tablets can be optionally coated with a controlled release polymer to provide additional controlled release characteristics.

[0102] As noted above, the active agent formulations and oral dosage forms of the present disclosure may include auxiliary excipients, such as, for example, diluents, lubricants, surfactants, disintegrants, plasticizers, anti-adherents, opacifiers, pigments, flavors, etc. As will be appreciated by one of skill in the art, the exact selection of excipients and their relative amounts will depend in part on the final oral dosage form into which the controlled release active agent formulation is to be incorporated.

[0103] Suitable diluents include, for example, pharma- ceutically acceptable inert fillers (e.g., microcrystalline cellulose, lactose, dibasic calcium phosphate, sugar, and / or any mixture of the above).Examples of diluents include microcrystalline cellulose, such as those sold under the trademark Avicel, for example, Avicel pH101, Avicel pH102, Avicel pH112, Avicel pH200, Avicel pH301, Avicel pH302; lactose, for example, lactose monohydrate, lactose anhydrous, and Pharmatose DCL21 (Pharmatose is a trademark) (including anhydrous, monohydrate, and spray-dried forms); dibasic calcium phosphate, for example, Emcompress (Emcompress is a trademark); mannitol; starch; sorbitol; sucrose; and glucose. Suitable lubricants, including agents that affect the flowability of the powder to be compressed, are, for example, colloidal silicon dioxide, such as Aerosil 200; talc; stearic acid, magnesium stearate, calcium stearate, and sodium stearyl fumarate. Suitable disintegrants include, for example, lightly cross-linked polyvinylpyrrolidone, corn starch, potato starch, maize starch and modified starches, croscarmellose sodium, crospovidone, sodium starch glycolate, and combinations and mixtures thereof.

[0104] According to a further aspect of the present disclosure, there is provided a controlled release active agent formulation for oral administration comprising a blend of particles. According to a further aspect of the present disclosure, there is provided a controlled release active agent formulation for oral administration comprising a blend of particles as defined above, mixed with an immediate release form of the active agent or a pharma- ceutically acceptable salt thereof to ensure rapid achievement of effective therapeutic blood levels. Preferably, the immediate release form of the active agent comprises pellets as defined above, without the rate-controlling membrane.

[0105] The sustained or delayed release coating is designed to provide delivery over a long period of time. The sustained or delayed release coating is, for example, a pH-independent coating formed from ethyl cellulose, hydroxypropyl cellulose, methyl cellulose, hydroxymethyl cellulose, hydroxyethyl cellulose, acrylic acid esters, or sodium carboxymethyl cellulose. Depending on the selection of coating material and / or coating thickness, various sustained or delayed release dosage forms can be easily designed by those skilled in the art to achieve delivery to both small intestine and large intestine, delivery only to small intestine, or delivery only to large intestine.

[0106] An enteric coating is a mixture of acceptable excipients that is applied to, combined with, mixed with, or otherwise added to a carrier or composition. The coating may be applied to compressed or molded or extruded tablets, gelatin capsules, and / or pellets, beads, granules, or particles of the carrier or composition. The coating may be applied via aqueous dispersion or after dissolving in a suitable solvent.

[0107] The dosage form of the composition disclosed herein can also be formulated as an enteric-coated delayed release oral dosage form, i.e., an oral dosage form of the composition described herein that utilizes an enteric coating that affects the release in the lower gastrointestinal tract.The enteric-coated dosage form can be a compressed or molded or extruded tablet / form (coated or uncoated) that contains granules, pellets, beads, or particles of the active ingredient and / or other composition ingredients, which are themselves coated or uncoated.The enteric-coated oral dosage form can also be a capsule (coated or uncoated) that contains pellets, beads, or granules of the solid carrier or composition, which are themselves coated or uncoated.

[0108] The delayed release coating composition includes polymeric materials such as cellulose butyrate phthalate, cellulose hydrogen phthalate, cellulose propionate phthalate, polyvinyl acetate phthalate, cellulose acetate phthalate, cellulose acetate trimellitate, hydroxypropyl methylcellulose phthalate, hydroxypropyl methylcellulose acetate, dioxypropyl methylcellulose succinate, carboxymethylethylcellulose, hydroxypropyl methylcellulose acetate succinate, acrylic acid, methacrylic acid, and / or esters thereof, and copolymers formed therefrom. A preferred enteric coating herein is comprised of Type A, B, or C methacrylic acid copolymers available from Rohm Tech, Inc. (Malden, Massachusetts), and an aqueous dispersion of cellulose acetate phthalate latex available from Eastman Fine Chemicals (Kingsport, Tennessee).

[0109] neoplasia The present disclosure provides a method for treating and / or preventing neoplasms in a patient in need of such treatment or prevention, comprising administering to the patient a therapeutically effective amount of the compound(s) or composition(s) disclosed herein, combinations thereof, and pharma- ceutically acceptable salts thereof, optionally in combination with, for example, a neoplasm therapeutic agent. The term "neoplasia" as used herein also refers to tumors, proliferative diseases, malignant tumors, and their metastases. Examples of cancer diseases include, for example, carcinomas, sarcomas, lymphomas or leukemia cells, germ cell tumors, blastomas, or other cancers. Cancers include, but are not limited to, epithelial neoplasms, squamous cell neoplasms, squamous cell carcinomas, basal cell neoplasms, basal cell carcinomas, transitional cell carcinomas and carcinomas, adenomas and adenocarcinomas (glandular), adenomas, adenocarcinomas, scirrhous insulinomas, glucagonomas, gastrinomas, vipomas, cholangiocarcinomas, hepatocellular carcinomas, adenoid cystic carcinomas, adnexal carcinomas, prolactinomas, oncocytomas, Hurthle cell adenomas, renal cell carcinomas, Grawitz tumors, multiple endocrine adenomas, endometrioid adenomas, adnexal and cutaneous adnexal neoplasms, mucoepidermoid neoplasms, cysts, Includes mucinous and serous neoplasms, cystadenoma, pseudomyxoma peritonei, ductal, lobular and medullary neoplasms, acinar cell neoplasms, composite epithelial neoplasms, Warth's tumor, thymoma, special gonadal neoplasms, sex cord-stromal tumors, thecuma, granulosa cell tumor, ovarian male germinoma, Sertoli-Leydic cell tumor, acinar tumor, paraganglioma, pheochromocytoma, glomus tumor, nevi and melanoma, melanocytic nevi, malignant melanoma, melanoma, nodular melanoma, dysplastic nevi, malignant lentigo melanoma, superficial spreading melanoma, and malignant acral lentigo melanoma. Sarcomas include, but are not limited to, Askin tumor, botryoid sarcoma, chondrosarcoma, Ewing sarcoma, malignant hemangioma, malignant schwannoma, osteosarcoma, soft tissue sarcomas (including alveolar soft part sarcoma, angiosarcoma, phyllodes cyst sarcoma, dermatofibrosarcoma, desmoid tumor, desmoplastic small round cell tumor, epithelioid sarcoma, extraskeletal chondrosarcoma, extraskeletal osteosarcoma, fibrosarcoma, hemangiopericytoma, angiosarcoma, Kaposi's sarcoma, leiomyosarcoma, liposarcoma, lymphangiosarcoma, lymphosarcoma, malignant fibrous histocytoma, neurofibrosarcoma, rhabdomyosarcoma, and synovial sarcoma).Prolymphocytic leukemias and leukemias include, but are not limited to, chronic lymphocytic leukemia / small lymphocytic lymphoma, B-cell lymphoma, lymphoplasmacytic lymphoma (such as Waldenstrom's macroglobulinemia), splenic marginal zone lymphoma, plasma cell myeloma, plasmacytoma, M protein deposition disease, heavy chain disease, extranodal marginal zone B-cell lymphoma (also called MALT lymphoma), nodal marginal zone B-cell lymphoma (nmzl), follicular lymphoma, mantle cell lymphoma, diffuse large B-cell lymphoma, mediastinal (thymic) large B-cell lymphoma, intravascular large B-cell lymphoma, primary effusion lymphoma, Burkitt's lymphoma / leukemia, Includes: T-cell prolymphocytic leukemia, T-cell large granular lymphocytic leukemia, aggressive NK-cell leukemia, adult T-cell leukemia, extranodal NK / T-cell lymphoma, nasal type, enteropathy-type T-cell lymphoma, hepatosplenic T-cell lymphoma, blastic NK-cell lymphoma, mycosis fungoides / Sezary syndrome, primary cutaneous CD30-positive T-lymphoproliferative disorder, primary cutaneous anaplastic large cell lymphoma, lymphomatoid papulosis, angioimmunoblastic T-cell lymphoma, peripheral T-cell lymphoma, anaplastic large cell lymphoma not otherwise specified, classical Hodgkin lymphoma (nodular sclerosis, mixed cytology, lymphocyte-rich, lymphocyte-depleted or non-depleted), and nodular lymphocyte-predominant Hodgkin lymphoma. Germ cell tumors include, but are not limited to, germinoma, dysgerminoma, seminoma, nongerminomatous germ cell tumor, embryonal carcinoma, endodermal sinus tumor, choriocarcinoma, teratoma, polygerminoma, and gonadoblastoma. Blastomas include, but are not limited to, nephroblastoma, medulloblastoma, and retinoblastoma. Other cancers include, but are not limited to, lip cancer, laryngeal cancer, hypopharyngeal cancer, tongue cancer, salivary gland cancer, stomach cancer, adenocarcinoma, thyroid cancer (medullary and papillary thyroid cancer), kidney cancer, renal parenchymal cancer, cervical cancer, uterine cancer, endometrial cancer, choriocarcinoma, testicular cancer, bladder cancer, melanoma, brain tumors such as glioblastoma, astrocytoma, meningioma, medulloblastoma and peripheral neuroectodermal tumors, gallbladder cancer, bronchial carcinoma, multiple myeloma, basal cell carcinoma, teratoma, retinoblastoma, choroidal melanoma, seminoma, rhabdomyosarcoma, craniopharyngioma, osteosarcoma, chondrosarcoma, myosarcoma, liposarcoma, fibrosarcoma, Ewing's sarcoma, and plasmacytoma.

[0110] In further embodiments, the cancer may be lung cancer, including non-small cell lung cancer and small cell lung cancer (including small cell carcinoma (oat cell carcinoma), mixed small cell / large cell carcinoma, and mixed small cell carcinoma), colon cancer, breast cancer, prostate cancer, liver cancer, pancreatic cancer, brain cancer, kidney cancer, ovarian cancer, stomach cancer, skin cancer, bone cancer, stomach cancer, breast cancer, pancreatic cancer, glioma, glioblastoma, hepatocellular carcinoma, papillary renal carcinoma, squamous cell carcinoma of the head and neck, leukemia, lymphoma, myeloma, or a solid tumor.

[0111] In embodiments, the cancer is acute lymphoblastic leukemia, acute myeloid leukemia; adrenal cortical tumor; AIDS-related cancer; AIDS-related lymphoma; anal cancer; appendiceal cancer; astrocytoma; atypical teratoma / rhabdoid tumor; basal cell carcinoma; bladder cancer; brain stem glioma; brain tumor (including brain stem glioma, central nervous system atypical teratoma / rhabdoid tumor, central nervous system embryonal tumor, astrocytoma, craniopharyngioma, ependymoma, ependymoma, medulloblastoma, medulloepithelioma, intermediate differentiated pineal parenchymal tumor, supratentorial primitive neuroectodermal tumor, and pineoblastoma); breast cancer; bronchial tumor; Burkitt's lymphoma; cancer of unknown primary site; carcinoma of unknown origin; noid tumor;carcinoma of unknown primary site;central nervous system atypical teratoma / rhabdoid tumor;central nervous system embryonal tumor;cervical cancer;childhood cancer;chordoma;chronic lymphocytic leukemia;chronic myeloid leukemia;chronic myeloproliferative disorder;colon cancer;colorectal cancer;craniopharyngioma;cutaneous T-cell lymphoma;endocrine islet cell tumor;endometrial cancer;ependymomas;ependymoma;esophageal cancer;neuroblastoma;Ewing's sarcoma;extracranial germ cell tumor;extragonadal germ cell tumor;extrahepatic bile duct cancer;gallbladder cancer;gastric (stomach) cancer;gastrointestinal carcinoid tumor;gastrointestinal stromal tumor;gastrointestinal stromal tumor (GIST);gestational trophoblastic tumor;glioma;hairy cell leukemia;head and neck cancer;cardiac cancer;Hodgkin's lymphoma;hypopharyngeal cancer;intraocular melanoma;island cell tumor;Kaposi's sarcoma;kidney cancer;Langerhans cell histiocytosis;laryngeal cancer;lip cancer;liver cancer;malignant fibrous histiocytoma bone cancer;medulloblastoma;medulloepithelioma;melanoma Merkel cell carcinoma;Merkel cell skin cancer;mesothelioma;metastatic squamous neck cancer with occult primary;oral cancer;multiple endocrine neoplasia syndrome;multiple myeloma;multiple myeloma / plasma cell neoplasm;mycosis fungoides;myelodysplastic syndrome;myeloproliferative neoplasm;nasal cancer;nasopharyngeal carcinoma;neuroblastoma;non-Hodgkin's lymphoma;non-melanoma skin cancer;non-small cell lung cancer;oral cancer;oral oral cavity cancer;oropharyngeal cancer;osteosarcoma;other brain and spinal tumors;ovarian cancer;ovarian epithelial cancer;ovarian germ cell tumors;ovarian low malignant potential tumors;pancreatic cancer;papillomatosis;paranasal sinus cancer;parathyroid carcinoma;pelvic cancer;penile cancer;pharyngeal cancer;pineal parenchymal tumor of intermediate differentiation;pineoblastoma;pituitary tumors;plasma cell neoplasm / multiple myeloma;pleuropulmonary blastoma;primary central nervous system (CNS) lymphoma;primary hepatocellular liver cancer;prostate cancer;rectal cancer;renal cancer;renal cell (kidney) cancer;renal cell carcinoma;airway cancer;retinoblastoma;rhabdomyosarcoma;salivary gland cancer;Sezary syndrome;small cell lung cancer;small intestine cancer;soft tissue sarcoma;squamous cell carcinoma;squamous neck cancer;gastric (stomach) cancer;The cancers include supratentorial primitive neuroectodermal tumors; T-cell lymphoma; testicular cancer; laryngeal cancer; thymic cancer; thymoma; thyroid cancer; transitional cell carcinoma; transitional cell carcinoma of the renal pelvis and ureter; trophoblastic tumor; ureteral cancer; urethral cancer; uterine cancer; uterine sarcoma; vaginal cancer; vulvar cancer; Waldenstrom's macroglobulinemia; or Wilms' tumor. The methods of the present invention can be used to characterize these and other cancers. Characterizing the phenotype can thus provide a diagnosis, prognosis, or theranosis of one of the cancers disclosed herein.;

[0112] In some embodiments, the cancer is acute myeloid leukemia (AML), breast cancer, cholangiocarcinoma, colorectal adenocarcinoma, extrahepatic bile duct adenocarcinoma, female reproductive tract malignancies, gastric adenocarcinoma, gastroesophageal carcinoma, gastrointestinal stromal tumor (GIST), glioblastoma, head and neck squamous cell carcinoma, leukemia, hepatocellular carcinoma, low-grade glioma, lung bronchioloalveolar carcinoma (BAC), non-small cell lung cancer (NSCLC), small cell lung cancer (SCLC), lymphoma, male reproductive tract malignancies. and malignant solitary fibrous tumor of the pleura (MSFT), melanoma, multiple myeloma, neuroendocrine tumors, nodular diffuse large B-cell lymphoma, non-epithelial ovarian carcinoma (non-EOC), ovarian surface epithelial carcinoma, pancreatic adenocarcinoma, pituitary carcinoma, oligodendroglioma, prostate adenocarcinoma, retroperitoneal or peritoneal carcinoma, retroperitoneal or peritoneal sarcoma, small intestine malignancies, soft tissue tumors, thymic carcinoma, thyroid carcinoma, or uveal melanoma.

[0113] As described herein, the present invention provides methods and compositions useful for the analysis, detection, characterization, imaging, prevention, and treatment of various diseases and disorders. In various embodiments, the disease or disorder includes cancer, precancerous conditions, inflammatory diseases, immune diseases, autoimmune diseases or disorders, cardiovascular diseases or disorders, neurological diseases or disorders, infectious diseases, or pain. Cancer includes, but is not limited to, acute lymphoblastic leukemia, acute myeloid leukemia; adrenal cortical tumors; AIDS-related cancers; AIDS-related lymphomas; anal cancer; appendix cancer; astrocytoma; atypical teratoma / rhabdoid tumors; basal cell carcinoma; bladder cancer; brain stem glioma; brain tumors (brain stem glioma, central nervous system atypical teratoma / rhabdoid tumor, central nervous system embryonal tumor, astrocytoma, craniopharyngioma, ependymoma, ependymoma, medulloblastoma, medulloepithelioma, intermediate differentiation pineal parenchymal tumor, supratentorial primitive neuroectoder ... tumors, including esophageal and pineoblastomas;breast cancer;bronchial tumors;Burkitt's lymphoma;carcinoma of unknown primary site;carcinoid tumors;carcinoma of unknown primary site;central nervous system atypical teratoma / rhabdoid tumor;central nervous system embryonal tumors;cervical cancer;childhood cancer;chordoma;chronic lymphocytic leukemia;chronic myelogenous leukemia;chronic myeloproliferative disorder;colon cancer;colorectal cancer;craniopharyngioma;cutaneous T-cell lymphoma;endocrine islet cell tumors;endometrial cancer;ependymomas;ependymoma;esophageal cancer;neuroblastoma;Ewing's sarcoma;cranial Extragonadal germ cell tumor;Extrahepatic bile duct cancer;Gallbladder cancer;Gastric (stomach) cancer;Gastrointestinal carcinoid tumor;Gastrointestinal stromal tumor;Gastrointestinal stromal tumor (GIST);Gestational trophoblastic tumor;Glioma;Hairy cell leukemia;Head and neck cancer;Heart cancer;Hodgkin's lymphoma;Hypopharyngeal cancer;Intraocular melanoma;Islet cell tumor;Kaposi's sarcoma;Kidney cancer;Langerhans cell histiocytosis;Laryngeal cancer;Lip cancer;Liver cancer;Lung cancer;Malignant fibrous histiocytomaBone cancer;Medulloblastoma;Meloduloepithelioma;MelanomaMerkel cell carcinoma;Merkel cell skin cancer;Mesothelioma ;Metastatic squamous neck cancer with occult primary;Oral cancer;Multiple endocrine neoplasia syndrome;Multiple myeloma;Multiple myeloma / plasma cell neoplasm;Mycosis fungoides;Myelodysplastic syndrome;Myeloproliferative neoplasm;Nasal cancer;Nasopharyngeal carcinoma;Neuroblastoma;Non-Hodgkin's lymphoma;Non-melanoma skin cancer;Non-small cell lung cancer;Oral cancer;Oral cancer;Oral pharyngeal cancer;Osteosarcoma;Other brain and spinal tumors;Ovarian cancer;Ovarian epithelial cancer;Ovarian germ cell tumors;Ovarian low malignant potential tumors;Pancreatic cancer;Papillomatosis;Sino-nasal cancer;Paratoyroid carcinoma;Pelvic cancer;Penile cancer;Laryngeal cancer;Pineal parenchymal tumor of intermediate differentiation;Pineoblastoma;Pituitary tumor;Plasma cell neoplasm / multiple myeloma;Pleuropulmonary blastoma;Primary central nervous system (CNS) lymphoma;Primary hepatocellular carcinoma;Prostate cancer;Rectal cancer;Kidney cancer;Renal cell (kidney) cancer;Renal cell carcinoma;Airway cancer;Retinoblastoma;Rhabdomyosarcoma;Salivary gland cancer;Sezary syndrome;Small cell lung cancer;Small intestine cancer;Soft tissue sarcoma;Squamous cell carcinoma;Squamous neck cancer;Gastric (stomach) cancer;Supratentorial primitive neuroectodermal tumor;T-cell lymphoma;Testicular cancer;Laryngeal cancer;Thymic carcinoma;Thymoma;Thyroid cancer;Transitional cell carcinoma;Transitional cell carcinoma of the renal pelvis and ureter;Troblastic tumor;Ureteral cancer;Urethral cancer;Uterine cancer;Uterine sarcoma;Vaginal cancer;Vulvar cancer;Waldenström's macroglobulinemia;or Wilms' tumor. Precancerous conditions include, but are not limited to, Barrett's esophagus. Autoimmune diseases can include, but are not limited to, inflammatory bowel disease (IBD), Crohn's disease (CD), ulcerative colitis (UC), pelvic inflammation, vasculitis, psoriasis, diabetes, autoimmune hepatitis, multiple sclerosis, myasthenia gravis, type 1 diabetes, rheumatoid arthritis, psoriasis, systemic lupus erythematosus (SLE), Hashimoto's thyroiditis, Graves' disease, ankylosing spondylitis, Sjogren's disease, CREST syndrome, scleroderma, rheumatic disease, organ rejection, primary sclerosing cholangitis, or sepsis. Cardiovascular diseases can include, but are not limited to, atherosclerosis, congestive heart failure, unstable plaque, stroke, ischemia, hypertension, stenosis, vascular occlusion, or thrombotic events. The neurological disease may include, but is not limited to, one of multiple sclerosis (MS), Parkinson's (PD), Alzheimer's (AD), schizophrenia, bipolar disorder, depression, autism, prion disease, Pick's disease, dementia, Huntington's disease (HD), Down's syndrome, cerebrovascular disease, Rasmussen's encephalitis, viral meningitis, neuropsychiatric systemic lupus erythematosus (NPSLE), amyotrophic lateral sclerosis, Creutzfeldt-Jakob disease, Gerstmann-Sträussler-Scheinker disease, transmissible spongiform encephalopathy, ischemic reperfusion injury (e.g., stroke), brain trauma, microbial infection, or chronic fatigue syndrome. The pain may include, but is not limited to, one of fibromyalgia, chronic neuropathic pain, or peripheral neuropathic pain. The infectious disease may include, but is not limited to, one of a bacterial infection, a viral infection, a yeast infection, Whipple's disease, prion disease, cirrhosis, methicillin-resistant Staphylococcus aureus, HIV, Hepatitis C virus (HCV), Epstein-Barr virus, Helicobacter pylori, hepatitis, syphilis, meningitis, malaria, tuberculosis, or influenza;

[0114] treatment Any therapy (e.g., therapeutic or prophylactic agent) that is useful, has been used, is currently being used, or can be used for the prevention, treatment, and / or management of a neoplasm or cancer can be used to prevent, treat, and / or manage a patient using the compositions and methods disclosed herein, e.g., using compositions and methods for administration of compounds disclosed herein, administered simultaneously, e.g., in separate dosage forms or sequentially in the same dosage form, for the treatment and / or prevention of a neoplasm or cancer. Therapies (e.g., therapeutic or prophylactic agents) include, but are not limited to, peptides, polypeptides, fusion proteins, nucleic acid molecules, small molecules, mimetics, synthetic drugs, inorganic molecules, and organic molecules. Non-limiting examples of cancer therapies include biological therapies, including chemotherapy, radiation therapy, hormonal therapy, antiangiogenic therapy, targeted therapy, and / or immunotherapy, and surgery. In certain embodiments, a prophylactically and / or therapeutically effective regimen includes the administration of a combination of therapies. In certain embodiments, the compositions disclosed herein, combinations thereof, and pharmacologic acceptable salts thereof can be administered as medicaments for treating or preventing a neoplasm.

[0115] Examples of cancer therapies that may be used in conjunction with the compositions and methods disclosed herein include, but are not limited to, Rituxan (rituximab), Herceptin (trastuzumab), Erbitux (cetuximab), Vectibix (panitumumab), Arzerra (ofatumumab), Benlysta (belimumab), Yervoy (ipilimumab), Perjeta (pertuzumab), tremelimumab, Opdivo (nivolumab), Keytruda (pembrolizumab), dacetuzumab, urelumab, MPDL3280A, lambrolizumab, blinatumomab, Humira (adalimumab), Campath (alemtuzumab), CEA-Scan Arcitmomab (fab fragment), Erbitux (cetuximab), Myoscint (imicilomab pentetate), ProstAScint (capromab pendetide), Remicade (infliximab), ReoPro (abciximab), Simulect (basiliximab), Synagis (palivizumab), Verluma (nofetumomab), Xolair (omalizumab), Zenapax (daclizumab) (ibritumomab), Zevalin (ibritumomab tiuxetan), Orthoclone OKT3 (Muromonab-CD3), Panorex (edrecolomab), Mylotarg (gemtuzumab ozogamicin), golimumab (Centocor), Cimzia (certolizumab pegol), Soliris (eculizumab), CNTO1275 (ustekinumab), Vectibix (panitumumab), Bexxar (tositumomab and 131 and biologics such as IFN-γ-phosphate phosphate sorbitol, IFN-γ-phosphate sorbitol ...

[0116] Examples of cancer therapies that may be used in conjunction with the compositions and methods disclosed herein include, but are not limited to, acivicin; aclarubicin; acodazole hydrochloride; acronine; adzelesin; aldesleukin; altretamine; ambomycin; amethanthrone acetate; aminoglutethimide; amsacrine; anastrozole; anthracyclines; anthramycin; asparaginase; asparin; azacitidine (Vidaza); azetepa; azotomycin; batimastat; benzodepa; bicalutamide; bisantrene hydrochloride; bismafied dimesylate; Bisphosphonates (e.g., pamidronate (Aredria), clodronate sodium (Bonefos), zoledronic acid (Zometa), alendronate (Fosamax), etidronate, ifandronate, cimadronate, risedronate, and tiludronate); bizelesin; bleomycin sulfate; brequinar sodium; bropirimine; busulfan; cactinomycin; calsterone; caracemide; carbetimer; carboplatin; carmustine; carrubicin hydrochloride; carzelesin; cedefingol; chlorambucil ;Cilolemycin;Cisplatin;Cladribine;Crisnatol mesylate;Cyclophosphamide;Cytarabine (Ara-C);Dacarbazine;Dactinomycin;Daunorubicin hydrochloride;Decitabine (Dacogen);Demethylating agents, dexorumaplatin;Desaguanine;Desaguanine mesylate;Diaziquone;Docetaxel;Doxorubicin;Doxorubicin hydrochloride;Droloxifene;Droloxifene citrate;Dromostanolone propionate;Duazomycin;Edatrexate;Eflornithine hydrochloride;EphA2 inhibitors;Elsami Torsin;Enloplatin;Enpromate;Epipropizine;Epirubicin hydrochloride;Elbrozole;Erubicin hydrochloride;Estramustine;Estramustine sodium phosphate;Etanidazole;Etoposide;Etoposide phosphate;Etoprine;Fadrozole hydrochloride;Fazarabine;Fenretinide;Floxuridine;Fludarabine phosphate;Fluorouracil;Fluorocitabine;Fosquidone;Fostriecin sodium;Gemcitabine;Gemcitabine hydrochloride;Histone deacetylase inhibitors (HDAC-Is) Hydroxyurea;Idarubicin hydrochloride;Ifosfamide; Irmofosine; Imatinib mesylate (Gleevec, Gleevec); Interleukin II (including recombinant interleukin II or rIL2), interferon alpha-2a; interferon alpha-2b; interferon alpha-n1; interferon alpha-n3; interferon beta-Ia; interferon gamma-Ib; Iproplatin; Irinotecan hydrochloride; Lanreotide acetate; Lenalidomide (Revlimid); Letrozole; Leuprolide acetate; Liarozole hydrochloride; Lometrexol sodium; Lomustine; Losoxantrone hydrochloride; Masoprocol; Maytansine; Mechlorethamine hydrochloride; Anti-CD2 antibodies (e.g., siplizumab (MedImmune Inc.; WO 02 / 098370 (incorporated herein by reference in its entirety); megestrol acetate; melengestrol acetate; melphalan; menogaril; mercaptopurine; methotrexate; methotrexate sodium; metoprine; metholedepa; mifepristone; myindomide; mitocalcin; mitochromine; mitogillin; mitomarcine; mitomycin; mitospar; mitotane; mitoxantrone hydrochloride; mycophenolic acid; nocodazole; nogalamycin; ORG34517; ormaplatin; oxaliplatin; oxisuran; paclitaxel; pegaspargase; periomycin; pentamustine; peplomycin sulfate; perfosfamide; pipobroman; piposulfan; piroxantrone hydrochloride; plicamycin; Pumestane;Porfimer sodium;Porfiromycin;Prednimustine;Procarbazine hydrochloride;Puromycin;Puromycin hydrochloride;Pyrazofurin;Riboprin;Logretimide;RU486, Safingol;Safingol hydrochloride;Semustine;Cintrazene;Sparfosate sodium;Sparsomycin;Spirogermanium hydrochloride;Spiromustine;Spiroplatin;Streptonigrin;Streptozocin;Sulofenur;Tallysomycin;Tecogalan sodium;Tegafur;Teroxantrone hydrochloride;Temoporfin;Teniposide;Teroxiron;Testolactone;Thiamiprine;Thioguanine;Thiotepa;Tiazofurin;Tirapazamine;Toremifene citrate;Tresorone acetate;Triciribine phosphate;Trimetrexate;Trimetrexate glucuronide; Triptorelin; Tubrozole hydrochloride; Uracil mustard; Uredep; Vapreotide; Verteporfin; Vinblastine sulfate; Vincristine sulfate; Vindesine; Vindesine sulfate; Vinglisine sulfate; Vinulosine sulfate; Vinorelbine tartrate; Virocidin sulfate; Bizolidine sulfate; Vorozole; Zeniplatin; Zinostatin; Zorubicin hydrochloride.;

[0117] Other examples of cancer therapies that may be used in conjunction with the compositions and methods disclosed herein include, but are not limited to, 20-epi-1,25 dihydroxyvitamin D3; 5-ethynyluracil 1; abiraterone; aclarubicin; acylfulvene; adecypenol; adozelesin; aldesleukin; ALL-TK antagonists; altretamine; ambamustine; amidox; amifostine; aminolevulinic acid; amrubicin; amsacrine; anagrelide; anastrozole; andrographolide; angiogenesis inhibitors; antagonist D; antagonist G;antarelix;anti-dorsalizing morphogenetic protein-I;antiandrogens, prostate cancer;antiestrogens;antitumor drugs;antisense oligonucleotides;aphidicolin glycinate;apoptotic gene modulators;apoptotic regulators;apurinic acid;ara-CDP-DL-PTBA;arginine deaminase;asulaculin;atamestane;atrimustine;axastatin 1;axastatin 2;axastatin 3;azasetron;azatoxin;azatyrosine;baccatin III derivatives;balanol;batimastat;BCR / ABL an agonists;benzochlorins;benzoylstaurosporines;β-lactam derivatives;β-arretin;betaclamycin B;betulinic acid;bFGF inhibitors;bicalutamide;bisantrene;bisaziridinylspermine;visnafide;bistraten A;bizelesin;brefrait;bropirimine;budotitanium;buthionine sulfoximine;calcipotriol;calphostin C;camptothecin derivatives;canarypox IL-2;capecitabine;carboxamide-amino-triazoles;carboxyamidotriazoles;CaRestM3;CARN700; Cartilage-derived inhibitors;Carzelesin;Casein kinase inhibitor (ICOS);Castanospermine;Cecropin B;Cetrorelix;Chlorin;Chloroquinoxaline sulfonamides;Cicaprost;Cis-porphyrins;Cladribine;Clomiphene analogs;Clotrimazole;Colismycin A;Colismycin B;Combretastatin A4;Combretastatin analogs;Conagenin;Crambesidin 816;Crysnatol;Cryptophycin 8;Cryptophycin A derivatives;Cracin A;Cyclopentane thraquinone;Cycloplatam;Sipemycin;Cytarabine ocfosfate;Cytolytic factors;Cytostatin;Daclizumab;Decitabine;Dehydrodidemnin B;Deslorelin;Dexamethasone dexfosphamide;Dexrazoxane;Dexverapamil;Diaziquone;Didemnin B;Didox;Diethylnorspermine;Dihydro-5-azacytidine;Dihydrotaxol, dioxamycin;Diphenylspiromustine;Docetaxel;Docosanol;Dolasetron;Doxifluridine;Droloxifene;Dronabinol;Duocarmycin SA;Ebselen;Ecomustine;Edelfosine;Edrecolomab;Eflornithine;Elecromustine Menthol;Emitefur;Epirubicin;Epristeride;Estramustine analogs;Estrogen agonists;Estrogen antagonists;Etanidazole;Etoposide phosphate;Exemestane;Fadrozole;Fazarabine;Fenretinide;Filgrastim;Finasteride;Flavopiridol;Flezelastine;Fluasterone;Fludarabine;Fluorodaunornithine hydrochloride;Forfenimex;Formestane;Fostriecin;Fotemustine;Gadolinium texaphyrin;Gallium nitrate;Galocitabine;Ganirelix;Gelatinase inhibitors;Gemcitabine;Glutathione inhibitors;HMG CoA reductase inhibitors (e.g., atorvastatin, cerivastatin, fluvastatin, lescol, lupitor, lovastatin, rosuvastatin, and simvastatin); hepsulfame; heregulin; hexamethylene bisacetamide; hypericin; ibandronate; idarubicin; idoxifene; idramantone; ilmofosine; ilomastat; imidazoacridone; imiquimod; immunostimulating peptides; insulin-like growth factor-1 receptor inhibitors; interferon agonists; interferon ;Interleukins;Iobenguane;Iododoxorubicin;Ipomenol, 4-Ilopraact;Irsogladine;Isobengazole;Isohomohalochondrin B;Itasetron;Jasplakinolide;Kahalalide F;Lamellarin-N triacetate;Lanreotide;Leinamycin;Lenograstim;Lentinan sulfate;Leptolstatin;Letrozole;Leukemia inhibitory factor;Leukocyte alpha interferon;Leuprolide + estrogen + progesterone;Leuprorelin;Levamisole;LFA-3TIP, liarozole; linear polyamine analogs; lipophilic disaccharide peptides; lipophilic platinum compounds; lysoclinamide 7; lobaplatin; robirisine; lometrexol; lonidamine; losoxantrone; lovastatin; loxoribine; lurtotecan; lutetium texaphyrin; lysofylline; lytic peptides; matansine; mannostatin A; marimastat; masoprocol; maspin; matrilysin inhibitors; matrix metalloproteinase inhibitors; menogaril; merbarone; meterulline; methioninase; metoclopramide; MIF inhibitors agents;mifepristone;miltefosine;mirimostim;mismatched double-stranded RNA;mitoguazone;mitolactol;mitomycin analogues;mitonafide;mitotoxin fibroblast growth factor-saporin;mitoxantrone;mofalotene;molgramostim;human chorionic gonadotropin;monophosphoryl lipid A+myobacterium cell wall sk;mopidamol;multidrug resistance gene inhibitors;multiple tumor suppressor 1-based therapy;mustard anticancer drugs;mysaperoxide B;mycobacterial cell wall extract;myriaporone;N-acetyldinaline;N-substituted beta Nsamides;Nafarelin;Nagressip;Naloxone + Pentazocine;Navabine;Naphthoterpin;Nartograstim;Nedaplatin;Nemorubicin;Neridronic acid;Neutral endopeptidase;Nilutamide;Nisamycin;Nitric oxide regulators;Nitroxide antioxidants;Nitrulline;06-benzylguanine;Octreotide;Oxenone;Oligonucleotides;Onapristone;Oracin;Oral cytokine inducers;Ormaplatin;Osateron;Oxaliplatin;Oxanomycin;Paclitaxel;Paclitaxel analogues;Paclitaxel inducers Body;Palaumin;Palmitoyl rhizoxin;Pamidronic acid;Panaxytriol;Panomyphen;Parabactin;Pazoliptin;Pegaspargase;Perdecin;Pentosan polysulfate sodium;Pentostatin;Pentrozole;Perflubron;Perfosfamide;Perillyl alcohol;Phenazinomycin;Phenylacetate;Phosphatase inhibitors;Picibanil;Pilocarpine hydrochloride;Pirarubicin;Piritrexim;Pracetin A;Pracetin B;Plasminogen activator inhibitors;Platinum complexes;Platinum compounds;Platinum-triamine complexes;Porfimer sodium;Porfiromycin;Prednisone;Propyl bis-acridone;Prostaglandin J2;Proteasome inhibitors;Protein A-based immunomodulators;Protein kinase C inhibitors;Protein kinase C inhibitors, microalgae;Protein tyrosine phosphatase inhibitors;Purine nucleoside phosphorylase inhibitors;Purpurins;Pyrazoloacridines;Pyridoxylated hemoglobin polyoxyethylene;raf antagonists;Raltitrexed;Ramosetron;Ras farnesyl protein transferase inhibitors;R as inhibitors;ras-GAP inhibitors;demethylated rete elliptin;rhenium Re186 etidronate;rhizoxin;ribozymes;RII retinamide;logretimide;rohitukin;romurtide;roquinimex;rubidinone B1;ruboxil;safingol;cytopin;SarCNU;sarcophytol A;sargramostim;Sdi1 mimics;semustine;senescence-derived inhibitor 1;sense oligonucleotides;signal transduction inhibitors;signal transduction modulators;gamma secretase inhibitors, single-chain antigen-binding protein;sizofiran;sobuzoxane;sodium borate Lithium;Sodium phenylacetate;Sorberol;Somatomedin binding proteins;Sonermin;Sparfosic acid;Spicamycin D;Spiromustine;Splenopentin;Spongistatin 1;Squalamine;Stem cell inhibitors;Stem cell division inhibitors;Stypiamide;Stromelysin inhibitors;Sulfinosine;Superactive vasoactive intestinal peptide antagonists;Sladista;Suramin;Swainsonine;Synthetic glycosaminoglycans;Tiamustine;5-fluorouracil;Leucovorin;Tamoxifen methiodide;Tauromustine;Tazarotene;Tecogalanna Thorium;Tegafur;Terlapyrylium;Telomerase inhibitors;Temoporfin;Temozolomide;Teniposide;Tetrachlorodecaoxide;Tetrazomine;Taliblastine;Thiocoraline;Thrombopoietin;Thrombopoietin mimetics;Thymalfasin;Thymopoietin receptor agonists;Thymotrin;Thyroid-stimulating hormone;Tin ethyl etiopurpurin;Tirapazamine;Titanocene dichloride;Topsentin;Toremifene;Pluripotent stem cell factor;Translation inhibitors;Tretinoin;Triacetyluridine;Triciribine;Trimetrexate;Triptorelin;Tropisetron; turosteride; tyrosine kinase inhibitors; tyrphostins; UBC inhibitors; ubenimex; urogenital sinus-derived growth inhibitory factor; urokinase receptor antagonists; vapreotide; variolin B; vector systems, red blood cell gene therapy; thalidomide; veraresol; veramine; verudin; verteporfin; vinorelbine; bixalin; anti-integrin antibodies (e.g., anti-integrin αvβ3 antibodies); vorozole; zanoteron; zeniplatin; dilascorub, and zinostatin stimalamer.;

[0118] According to the present invention, any suitable mode of administration may be used, including, but not limited to, parenteral administration, such as intravenous, subcutaneous, intramuscular and intrathecal administration; oral, intranasal, rectal or vaginal administration; directly to the tumor; transdermal patch; implant device (especially for sustained release); and finally, topical administration. The mode of administration varies according to the disease to be treated. In one embodiment, the arsenic trioxide of the present invention is dissolved in an aqueous sodium hydroxide solution with the pH adjusted to a physiologically acceptable range, for example, about pH 6-8.

[0119] A non-limiting list of compounds that may be used in conjunction with the compositions and methods disclosed herein includes inhibitors of interleukin-3 receptor (IL-3R) and CD123 (including peptides, peptide-conjugates, antibodies, antibody-conjugates, antibody fragments, and antibody fragment-conjugates that target IL-3R or CD123); cantharidin; norcantharidin and its analogs and derivatives; Notch pathway inhibitors, including gamma secretase inhibitors; Sonic hedgehog / smoothened pathway inhibitors, including cyclopamine and its analogs; antibodies against CD96; Certain NF-κB / proteasome inhibitors, including ruthenolides and their analogues;certain triterpenes, including celastrol;certain mTOR inhibitors;compounds and antibodies targeting the urokinase receptor;sinefungin;certain inosine monophosphate dehydrogenase (IMPDH) inhibitors;PPAR-α and PPAR-γ agonists and antagonists (pioglitazone, tesaslitazar, muraglitazar, peliglitazar, lobeglitazone, balaglitazone, ragaglitazar, rosiglitazone, farglitazar, sodeglitazar, reglitazar, etc.) , naveglitazar, oxeglitazar, metaglidasen, netoglitazone, darglitazone, englitazone, thiazolidinediones, aleglitazar, edaglitazone, rivoglitazone, troglitazone, imiglitazar, and sipoglitazar; telomerase inhibitors; antibodies against EpCAM (ESA); GSK-3β agonists and antagonists, including lithium, 6-bromoinirubin-3'-oxime (BIO), TDZD8; Wnt pathway inhibitors, such as antibodies or small molecules against Frizzled, including dishev Antibodies that inhibit eled / Frizzled or beta-catenin;anti-CD20 antibodies and conjugates for novel use in multiple myeloma or melanoma (e.g. Rituxan, Beksar, Zevalin);anti-CD133 antibodies;anti-CD44 antibodies;antibodies against IL-4;specific differentiation agents such as versnarinone;compounds targeting CD33 such as antibodies or betulinic acid;compounds targeting lactadherin such as antibodies;small molecules or antibodies targeting CXCR4 or SDF-1;small molecules or antibodies targeting multidrug resistance pumps;inhibitors of survivin;inhibitors of XIAP;These include small molecules that target Bcl-2; antibodies against CLL-1; and furin inhibitors (e.g., cucurbitacin).

[0120] A further non-limiting list of compounds that may be used in conjunction with the compositions and methods disclosed herein to target cancer include i) antibodies, antibody fragments, and proteins that are naked or conjugated to therapeutic moieties that target specific cell surface targets on cancer stem cells, or ii) small molecules known in the art, including those that can be further optimized (e.g., by chemistry) or identified by cancer stem cell-based screening (such as determining whether a compound impairs cancer stem cell proliferation or viability by standard methods), and target cell surface and intracellular targets (inclusive). The proteins involved in the expression of IL-1, IL-2, and IL-3 (not meant to imply a binding affinity to IL-1) are Rex1 (Zfp42), CTGF, activin A, Wnt, FGF-2, HIF-1, AP-2γ, Bmi-1, nucleostemin, hiwi, Moz-TIF2, Nanog, β-arrestin-2, Oct-4, Sox2, stella, GDF3, RUNX3, EBAF, TDGF-1, nodal, ZFPY, PTNE, Evi-1, Pax3, Mcl-1, c-kit, Lex-1, Zfx, lactadherin, aldehyde dehydrogenase, BCRP, telomerase, CD133, Bcl-2, CD26, Gremlin, and FoxC2.

[0121] In some embodiments, the therapy(ies) is an immunomodulatory agent that may be used in conjunction with the compositions and methods disclosed herein. Non-limiting examples of immunomodulatory agents include proteinaceous agents such as cytokines, peptidomimetics, and antibodies (e.g., human, humanized, chimeric, monoclonal, polyclonal, Fvs, scFvs, Fab or F(ab)2 fragments or epitope-binding fragments), nucleic acid molecules (e.g., antisense nucleic acid molecules and triple helices), small molecules, organic compounds, and inorganic compounds. In particular, immunomodulatory agents include, but are not limited to, methotrexate, leflunomide, cyclophosphamide, cytoxan, imuran, cyclosporine A, minocycline, azathioprine, antibacterial agents (e.g., FK506 (tacrolimus)), methylprednisolone (MP), corticosteroids, steroids, mycophenolate mofetil, rapamycin (sirolimus), mizoribine, deoxyspergualin, brequinar, malononitriloamide (e.g., leflunamide), T receptor modulators, cytokine receptor modulators, and mast cell modulators. In one embodiment, the immunomodulatory agent is a chemotherapeutic agent. In another embodiment, the immunomodulatory agent is an immunomodulatory agent other than a chemotherapeutic agent. In some embodiments, the therapy(ies) used in accordance with the present invention are not immunomodulatory agents. In some embodiments, the therapy(ies) are antiangiogenic agents. Non-limiting examples of anti-angiogenic agents include proteins, polypeptides, peptides, fusion proteins, antibodies, such as antibodies that specifically bind to TNF-α (e.g., human, humanized, chimeric, monoclonal, polyclonal, Fvs, scFvs, Fab fragments, F(ab) 2 Antibody molecules that reduce or inhibit angiogenesis include, for example, antibodies, mAbs, and antigen-binding fragments thereof, nucleic acid molecules (eg, antisense molecules or triple helices), organic molecules, inorganic molecules, and small molecules that reduce or inhibit angiogenesis.

[0122] In certain embodiments, the therapy(ies) are alkylating agents, nitrosoureas, antimetabolites, and anthracyclines, topoisomerase II inhibitors, or mitotic inhibitors. Alkylating agents include, but are not limited to, busulfan, cisplatin, carboplatin, chlorambucil, cyclophosphamide, ifosfamide, decarbazine, mechlorethamine, mephalen, and temozolomide. Nitrosoureas include, but are not limited to, carmustine (BCNU) and lomustine (CCNU). Antimetabolites include, but are not limited to, 5-fluorouracil, capecitabine, methotrexate, gemcitabine, cytarabine, and fludarabine. Anthracyclines include, but are not limited to, daunorubicin, doxorubicin, epirubicin, idarubicin, and mitoxantrone. Topoisomerase II inhibitors include, but are not limited to, topotecan, irinotecan, etoposide (VP-16), and teniposide. Mitotic inhibitors include, but are not limited to, taxanes (paclitaxel, docetaxel), and vinca alkaloids (vinblastine, vincristine, and vinorelbine). In some embodiments of the invention, the therapy(ies) include administration of cantharidin or an analog thereof. The invention includes the use of agents that target cancer stem cells. In certain embodiments, the agent acts alone. In other embodiments, the agent is attached directly or indirectly to another therapeutic moiety.Non-limiting examples of therapeutic moieties include, but are not limited to, alkylating agents, antimetabolites, plant alkaloids, cytotoxic agents, chemotherapeutic agents (e.g., steroids, cytosine arabinoside, fluorouracil, methotrexate, aminopterin, mitomycin C, demecolcine, etoposide, mithramycin, calicheamicin, CC-1065, chlorambucil, or melphalan), radionuclides, therapeutic enzymes, cytokines, toxins (including plant-derived toxins, fungal-derived toxins, bacterial-derived toxins (e.g., deglycosylated ricin A chain, ribosome-inactivating protein, alpha-sarcin, aspergillin, restylosin, ribonuclease, diphtheria toxin, pseudomonas exotoxin, bacterial endotoxin, or lipid A portion of bacterial endotoxin)), growth regulators, and RNases. In some embodiments, the agent used is an agent that binds to a marker, e.g., an antigen on a cancer stem cell. In certain embodiments, the agent binds to an antigen that is expressed at a higher level on cancer stem cells than on normal stem cells. In certain embodiments, the agent specifically binds to a cancer stem cell antigen that is not on normal stem cells. In other embodiments, the therapy(ies) is an agent that binds to a marker on cancer stem cells. In one embodiment, the agent that binds to a marker on cancer stem cells is an antibody, or an antibody conjugated to a therapeutic moiety, or an antibody fragment conjugated to a therapeutic moiety.

[0123] In certain embodiments, the antibody or fragment thereof that binds to marker on cancer stem cells is substantially non-immunogenic in the subject that is treated.Methods for obtaining non-immunogenic antibody include, but are not limited to, chimerizing antibody, humanizing antibody, and isolating antibody from the same species as the subject that is treated.Antibody or fragment thereof that binds to marker in cancer stem cells can be produced using techniques known in the art.

[0124] In some embodiments, the therapy includes the use of X-rays, gamma rays, and other radiation sources to destroy cancer stem cells and / or cancer cells. In certain embodiments, radiation therapy is administered as external beam radiation or teletherapy, where radiation is directed from a remote source. In other embodiments, radiation therapy is administered as internal therapy or brachytherapy, where a radiation source is placed inside the body near the cancer stem cells, cancer cells, and / or tumor mass.

[0125] In some embodiments, the treatment used is a proliferation-based treatment, non-limiting examples of which include chemotherapy and radiation therapy, as described above.

[0126] Currently available therapies and their dosages, routes of administration and recommended usage are known in the art and described in such references as the Physician's Desk Reference (60th ed., 2006).

[0127] In certain embodiments, cycling therapy involves administration of a first cancer therapeutic for a period of time, followed by administration of a second cancer therapeutic for a period of time, optionally followed by administration of a third cancer therapeutic for a period of time, and so on, repeating this sequential administration, i.e., cycle, to reduce the development of resistance to one of the cancer therapeutics, to avoid or reduce side effects of one of the cancer therapeutics, and / or to improve the effectiveness of the cancer therapeutics.

[0128] When two prophylactically and / or therapeutically effective regimens are administered to a subject simultaneously, the term "simultaneously" does not mean to be limited to administration of the cancer therapeutics at exactly the same time, but rather that they are administered to a subject in an order and within a time interval such that they can act together (e.g., synergistically to provide increased benefit than if they were administered otherwise). For example, the cancer therapeutics may be administered simultaneously or sequentially in any order at different times. However, if not administered simultaneously, they should be administered close enough in time to provide the desired therapeutic effect, preferably in a synergistic manner. The combined cancer therapeutics may be administered separately in any suitable form and by any suitable route. It is understood that when the components of the combined cancer therapeutics are not administered in the same pharmaceutical composition, they may be administered to a subject in need thereof in any order. For example, the first prophylactically and / or therapeutically effective regimen can be administered prior to (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks before), concomitantly with, or subsequent to (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks after) administration of the second cancer therapeutic to a subject in need thereof. In various embodiments, the cancer therapeutics are administered 1 minute apart, 10 minutes apart, 30 minutes apart, less than 1 hour apart, 1 hour apart, 1-2 hours apart, 2-3 hours apart, 3-4 hours apart, 4-5 hours apart, 5-6 hours apart, 6-7 hours apart, 7-8 hours apart, 8-9 hours apart, 9-10 hours apart, 10-11 hours apart, 11-12 hours apart, 24 hours or less apart, or 48 hours or less apart. In one embodiment, the cancer therapeutics are administered within the same office visit. In another embodiment, the combination cancer therapeutics are administered 1 minute to 24 hours apart.

[0129] Packaging / treatment kit The present disclosure provides a kit for carrying out the method according to the present disclosure in a simple and effective manner. Such a kit may be suitable for delivery of solid oral forms such as tablets or capsules. Such a kit may include several unit doses. Such a kit may include a means for containing the doses oriented in the order of their intended use. An example of a means for containing the doses in the order of their intended use is a card. An example of such a kit is a "blister pack". Blister packs are well known in the packaging industry and are widely used to package unit dosage forms. If desired, the blister can be in the form of a child-resistant blister, i.e., a blister that is difficult for a child to open, but can be easily opened by an adult. Optionally, a memory aid can be provided, for example, in the form of numbers, letters, or other markings, or with a calendar function and / or calendar insert, designating the days and the divisions of the day in the treatment schedule in which the doses can be administered, such as, for example, the morning dose is packaged with the "day" and the afternoon dose. Or the morning dose is packaged with the afternoon dose. Alternatively, placebo doses, or vitamin or nutritional supplements can be included, either in a similar form to the active dose or in a different form.

[0130] The present disclosure provides compositions, including preparations, formulations, and / or kits, comprising the above-described combinations of ingredients (including multi-ingredient combinations of drugs of the present invention), which are useful as therapies for treating, preventing, or ameliorating the conditions, conditions, and diseases provided herein. In one embodiment, each member of the combination of ingredients is manufactured in a separate package, kit, or container, or all or a subset of the combination of ingredients is manufactured in a separate package or container. In another embodiment, the package, kit, or container comprises a blister package, clamshell, tray, shrink wrap, or the like.

[0131] In one embodiment, the package, kit, or container comprises a "blister package" (also called a blister pack or bubble pack). In one embodiment, the blister package consists of two or more separate compartments. The blister package is composed of two separate material elements: a clear plastic cavity molded to the product and its blister board backing. These two elements are then joined together by a heat sealing process that allows the product to be hung or displayed. Exemplary types of "blister packages" include face seal blister packages, gang run blister packages, mock blister packages, interactive blister packages, and slide blister packages.

[0132] A blister pack, clamshell, or tray is a form of packaging used for commercial products. Thus, the present invention provides a blister pack, clamshell, or tray containing the composition of the present invention (e.g., a (multi-component combination of drugs of the present invention) combination of active ingredients). Blister packs, clamshells, or trays can be designed to be non-reclosable, so that the consumer can know if the package has already been opened. They are used to package commercial products where product tampering is a consideration, such as the medicaments of the present invention. In one aspect, the blister pack of the present invention includes the combination of the present invention, which includes a molded PVC base with a raised area ("blister") for containing tablets, pills, etc., covered with a foil laminate. The tablets, pills, etc. are removed from the pack by peeling the foil or by pressing the blister to cause the tablets to break the foil. In one aspect, a special form of blister pack is a strip pack.

[0133] In one embodiment, the blister pack also includes a packaging method in which the composition comprising the combination of ingredients of the present invention is contained between the card and the transparent PVC. The PVC can be transparent so that the article (pills, tablets, gel tabs, etc.) can be easily seen and inspected, and in one embodiment, it can be vacuum-formed around a mold so that it can snugly accommodate the article and have room to open at the time of purchase. In one embodiment, the card is light-colored and designed according to the article inside (pills, tablets, gel tabs, etc.), and the PVC is attached to the card using a pre-formed tab where the adhesive is placed. The adhesive can be strong enough to allow the pack to be hung on a nail, but weak enough that the joint can be torn open in this way to access the article. Sometimes, for large articles or multiple enclosed pills, tablets, gel tabs, etc., the card has a perforated window for access. In one embodiment, for example, a more secure blister pack for articles such as pills, tablets, gel tabs, etc. of the present invention is used, which can include two vacuum-formed PVC sheets interlocked together at the edges with an information card inside.

[0134] In one embodiment, the blister package includes at least two components (e.g., a multi-component combination of drugs of the present invention): a thermoformed "blister" that contains the product (e.g., a combination of the present invention), and then a "blister card" that is a printed card with an adhesive coating on the front side. During the assembly process, the blister components, most commonly made from PVC, are attached to the blister card using a blister machine. Conventional blister packs can also be sealed.

[0135] As discussed herein, the articles of manufacture of the invention may include packages of the therapeutic combinations of the invention, either alone or in combination, as a "blister package" or as a blister package with a lid, a blister with a lid or a blister card or packet, or multiple packets including shrink wrap.

[0136] In one aspect, any of the articles of manufacture of the invention, including the kit or blister pack, includes a memory aid to help remind the patient how and when to take the medication of the invention.

[0137] The treatment kit can be constructed in various forms well known to those skilled in the art. The kit includes at least one unit dose of an active agent for administration according to a daily regimen and a means for containing the unit dose. The treatment kit can be constructed for, for example, once daily, twice daily, three times daily, four times daily, multiple daily administration, or other dosing regimens. The kit includes a means for daily administration of the agent of the invention. In one embodiment, the kit includes about 1 to about 4 unit doses.

[0138] In one embodiment, the means for containing the unit doses is a card, including, for example, a card that can be folded. This card is referred to herein as the main card, or the principal card or the first card, and is distinguished from any additional cards, circulars, or other such materials that may be associated with the kit. This main card can be folded with a simple crease, or with a double crease, to present a spine similar to the spine of a closed book. The main card can include a printable surface, i.e., a surface on which the product name, appropriate administration instructions, product information, drawings, logos, memory aids, calendar functions, etc. can be printed. The main card can include a means for containing the unit dose or different doses designated for different times of the day, and a memory aid for administering the unit dose. The main card, especially when it is made from two or more laminated paperboard surfaces, can include, for example, a slit or pocket in one of the inner paperboard surfaces of the folded card. The slit or pocket can be used to contain a removable secondary card, i.e., a second or insert card that is not permanently attached or fixed to the main card.

[0139] The memory aid may include a list of days of the week, i.e., Sunday, Monday, Tuesday, Wednesday, Thursday, Friday, and Saturday, with appropriate space for the patient to select and indicate on the card the preferred day for administering the treatment. The memory aid may include a list of times, with appropriate space for the patient to select and indicate on the card the preferred time of day for administering the treatment (e.g., morning, afternoon, noon). The memory aid may also include a removable sticker with a suitable pressure sensitive adhesive to facilitate easy removal and reapplication to a desired surface, such as a calendar or day reminder. The removable sticker may be placed on the main card, or may be placed on a secondary card configured to be easily inserted into and removed from any slit in the main card. Additionally, any slit may include additional patient information and other circulars.

[0140] Other means for containing the unit doses can include bottles and vials, where the bottle or vial contains a memory aid, such as a printed label for administering the unit dose(s). The label can also contain a removable reminder sticker for placement on a calendar or daily reminder to further help the patient remember when to take or when the dose has been taken.

[0141] The present invention will be described in more detail below by way of examples, but the present invention is not limited to these examples.

[0142] Aspects Aspect 1: A compound having the structure of Formula I:

[0143] [ka] (In the formula, R 1 Halo, C 1-6 Alkyl, C 3-8 Cycloalkyl, or C 2-6 alkenyl, R 2 , H, C 1-6Alkyl or C 3-8 cycloalkyl) or a pharma- ceutically acceptable salt thereof.

[0144] Aspect 2: R 1 The compound of embodiment 1, wherein is halo.

[0145] Aspect 3: R 1 The compound of embodiment 1 or 2, wherein is chloro, bromo, fluoro, or iodo.

[0146] Aspect 4: R 1 The compound of embodiment 1 or 2, wherein is chloro or bromo.

[0147] Aspect 5: R 1 is C 1-4 The compound of embodiment 1, wherein the is alkyl.

[0148] Aspect 6: R 1 The compound of embodiment 1 or 5, wherein is methyl, ethyl, n-propyl, or i-propyl.

[0149] Aspect 7: R 1 is C 3-7 The compound of embodiment 1, wherein the is cycloalkyl.

[0150] Aspect 8: R 1 The compound of embodiment 1 or 7, wherein is cyclopropyl.

[0151] Aspect 9: R 1 is C 1-4 The compound of embodiment 1, wherein the compound is alkenyl.

[0152] Aspect 10: R 1 is vinyl or isopropenyl.

[0153] Aspect 11: R 2 is C 1-6 The compound of any one of the preceding embodiments, wherein is alkyl.

[0154] Aspect 12: R 2

[0036] The compound of any one of the preceding embodiments, wherein is methyl, ethyl, propyl, butyl, pentyl, or hexyl.

[0155] Aspect 13: R 2

[0026] The compound of any one of the preceding aspects, wherein is methyl, ethyl, n-propyl, or i-propyl.

[0156] Aspect 14: R 2 is C 3-8 The compound according to any one of the preceding aspects, wherein the compound is cycloalkyl.

[0157] Aspect 15: R 2 is cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl.

[0158] Aspect 16: R 2 is cyclopropyl.

[0159] Aspect 17: A compound according to aspect 1, having the structure of formula II:

[0160] [ka] or a pharma- ceutically acceptable salt thereof. A compound according to embodiment 1.

[0161] Aspect 18:

[0162] [ka] The compound of embodiment 1, wherein or a pharma- ceutically acceptable salt thereof. A compound according to embodiment 1.

[0163] Aspect 19: A compound having the structure of formula III:

[0164] [ka] (In the formula, R 2 , H, C 1-6 Alkyl or C 3-8 cycloalkyl), or a pharma- ceutically acceptable salt thereof.

[0165] Aspect 20: R 2 is H.

[0166] Aspect 21: R 2 is C 1-6 The compound of embodiment 19, wherein the aryl group is alkyl.

[0167] Aspect 22: R 2 is methyl.

[0168] Aspect 23: A compound having the structure of formula IV,

[0169] [ka] (wherein R1, R2 are H, methyl, ethyl, or propyl; optionally, R1 and R2 are joined to form a 3-6 membered ring; R3 is a C1-4 small alkyl, or a C3-6 cycloalkyl ring; and R4 is H, C1-3 alkyl, CF3, Omethyl, OCF3, OCF2H, CN, or halo), or a pharma- ceutically acceptable salt thereof.

[0170] Aspect 24: A compound having the structure of formula V:

[0171] [ka] (wherein R2 is H, methyl, ethyl, or propyl, R3 is a C1-4 small alkyl, or a C3-6 cycloalkyl ring, and R4 is H, C1-3 alkyl, CF3, Omethyl, OCF3, OCF2H, CN, or halo), or a pharma- ceutically acceptable salt thereof.

[0172] Aspect 25: A compound having the structure of formula VI,

[0173] [ka] (wherein R2 is H, methyl, ethyl, or propyl, R3 is a C1-4 small alkyl, or a C3-6 cycloalkyl ring, and R4 is H, C1-3 alkyl, CF3, Omethyl, OCF3, OCF2H, CN, or halo), or a pharma- ceutically acceptable salt thereof.

[0174] Embodiment 26: A pharmaceutical composition comprising one or more compounds of any one of the preceding embodiments and one or more pharma- ceutically acceptable excipients.

[0175] Embodiment 27: A method for inhibiting Wee1 in a patient in need of such treatment, comprising administering to the patient a compound or composition according to any one of embodiments 1 to 26.

[0176] Embodiment 28: A method of treating cancer in a patient in need thereof, comprising administering to the patient a compound of any one of embodiments 1 to 26.

[0177] Aspect 29: The cancer is an adrenal cortical tumor, an AIDS-related cancer (e.g., AIDS-related lymphoma), anal cancer, appendix cancer, astrocytoma, atypical teratoma / rhabdoid tumor, basal cell tumor, cholangiocarcinoma, bladder cancer, bone cancer, brain cancer (e.g., glioblastoma), breast cancer, bronchial cancer, cancer of unknown primary site, carcinoid tumor, castration-resistant prostate cancer, central nervous system cancer (e.g., central nervous system atypical teratoma / rhabdoid tumor, central nervous system embryonal tumor, central nervous system lymphoma, primary central nervous system lymphoma), cervical cancer, chordoma, chondrosarcoma, chronic myeloproliferative disorder, colon cancer, colorectal cancer, craniopharyngioma, desmoplastic circle, and the like. B-cell tumors, diffuse large B-cell lymphoma, endometrial cancer, ependymoma, esophageal cancer, Ewing's sarcoma family of tumors, extracranial germ cell tumors, extragonadal cell tumors, extrahepatic bile duct cancer, gallbladder cancer, gastric (stomach) cancer, gastrointestinal carcinoid tumors, gastrointestinal stromal tumors, germ cell tumors, gestational trophoblastic tumors, gliomas, head cancer, hepatocellular (liver) cancer, high-grade prostate cancer, histiocytic proliferation, hypopharyngeal cancer, Kaposi's sarcoma, kidney (kidney) cancer, Langerhans cell histiocytosis, laryngeal cancer, leptomeningeal disease, lip cancer, low-grade prostate cancer, leukemia (e.g., chronic lymphocytic leukemia, chronic myeloid leukemia, hairy cell leukemia), lung cancer, lymphoma lymphoma (Burkitt's lymphoma, central nervous system lymphoma, T-cell lymphoma (e.g., cutaneous T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, lymphoplasmacytic lymphoma), intermediate-grade prostate cancer, medulloblastoma, medulloepithelioma, melanoma, Merkel cell carcinoma, mesothelioma, metastatic squamous cell carcinoma of unknown primary, oral cancer, multiple endocrine neoplasia syndrome, multiple myeloma, multiple myeloma / plasma cell neoplasm, mycosis fungoides, myelodysplastic syndrome, myelodysplastic / myeloproliferative neoplasm, myeloproliferative disorder, nasal cavity or paranasal sinus cancer, neck cancer, nasopharyngeal cancer, neck cancer, neuroblastoma, eye cancer, ocular melanoma , oral cancer, oropharyngeal cancer, oral cancer, osteosarcoma or malignant fibrous histiocytoma of bone, osteosarcoma or malignant fibrous histiocytoma, ovarian cancer, ovarian germ cell tumor, ovarian epithelial cancer, ovarian low malignant potential tumor, pancreatic cancer, papillomatosis, paranasal sinus or nasal cancer, parathyroid cancer, penile cancer, pharyngeal cancer, moderately differentiated pineal parenchymal tumor, pineoblastoma or supratentorial primitive neuroectodermal tumor, pituitary tumor, pleuropulmonary blastoma, pregnancy cancer, prostate cancer, rectal cancer, renal pelvis cancer, airway cancer involving the NUT gene on chromosome 15, retinoblastoma, rhabdomyoblastoma, salivary gland cancer, sarcoma, Sezary syndrome, skin cancer, skin cancer, small intestine cancer, soft tissue sarcoma, spinal tumor, squamous cell carcinoma,29. The method of embodiment 28, wherein the cancer is squamous cell cervical cancer of unknown primary, occult primary supratentorial primitive neuroectodermal tumor, testicular cancer, laryngeal cancer, thymoma or thymic carcinoma, thyroid cancer, transitional cell carcinoma of the renal pelvis or ureter, rare cancer of childhood, ureteral cancer, urethral cancer, uterine sarcoma, vaginal cancer, vulvar cancer, Waldenstrom's macroglobulinemia (lymphoplasmacytic lymphoma), Wilms' tumor, or female cancer.

[0178] Embodiment 30: The method of embodiment 29, wherein the cancer is breast cancer, prostate cancer, pancreatic cancer, lung cancer, colorectal cancer, ovarian cancer, liver cancer, melanoma, renal cancer, central nervous system cancer, brain cancer such as glioblastoma, leukemia, or lymphoma.

[0179] Embodiment 31: The method according to any one of embodiments 27 to 30, wherein the compound is administered in combination with at least one additional therapeutic agent.

[0180] Embodiment 32: The method of embodiment 31, wherein the at least one additional therapeutic agent is a chemotherapeutic agent.

[0181] Aspect 33: The chemotherapeutic agent is selected from the group consisting of busulfan, melphalan, chlorambucil, cyclophosphamide, ifosfamide, temozolomide, bendamustine, cisplatin, mitomycin C, bleomycin, carboplatin, camptothecin, irinotecan, topotecan, doxorubicin, epirubicin, aclarubicin, mitoxantrone, elliptinium, etoposide, 5-azacytidine, gemcitabine, 5-fluorouracil, methotrexate, 5-fluoro-2′-deoxy-uridine, fludara. Vin, nelarabine, ara-C, pralatrexate, pemetrexed, hydroxyurea, thioguanine, colchicine, vinblastine, vincristine, vinorelbine, paclitaxel, ixabepilone, cabazitaxel, docetaxel, campath, panitumumab, metazotuzumab, navtuzumab, pimtuzumab, remortuzumab, bevacizumab, paltuzumab, trastuzumab, cetuximab, obinutuzumab, orfamuzumab, rituximab, alemtuzumab, thiemtuzumab, toximab, benzimidazole, benzo[b]azetidine ... Mab, dalemtuzumab, erlotuzumab, T-DM1, ofatumumab, dinutuximab, blinatumomab, ipilimumab, avastin, trastuzumab, rituximab, imatinib, gefitinib, erlotinib, osimertinib, afatinib, ceritinib, aretinib, crizotinib, erlotinib, lapatinib, sorafenib, nitinib, nilotinib, dasatinib, pazopanib, temsirolimus, everolimus, vorinostat, romidepsin, panobinostat, belinostat, tamoxifen, 33. The method of embodiment 32, wherein the therapeutic agent is selected from the group consisting of letrozole, fulvestrant, mitoguazone, octreotide, retinoic acid, arsenic trioxide, zoledronic acid, bortezomib, carfilzomib, ixazomib, vismodegib, sonidegib, denosumab, thalidomide, lenalidomide, venetoclax, aldesleukin (recombinant human interleukin-2), sipuleucel-T (prostate cancer therapeutic vaccine), palbociclib, olaparib, niraparib, rucaparib, tazoparib, and combinations thereof.

[0182] Embodiment 34: A method for reducing the activity of a kinase encoded by gene WEE1, comprising contacting the kinase with an inhibitory amount of a compound according to any one of embodiments 1 to 26.

[0183] Embodiment 35: The method according to embodiment 34, wherein the method is carried out in vitro.

[0184] Embodiment 36: The method of embodiment 34, wherein the method is performed in a subject.

[0185] Embodiment 37: A method of treating or preventing a Wee1 mediated disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound according to any one of embodiments 1 to 26, or a pharma- ceutically acceptable salt or prodrug thereof, or a pharmaceutical composition thereof.

[0186] Aspect 38: The disease is a cancer such as an adrenal cortical tumor, AIDS-related cancer (e.g., AIDS-related lymphoma), anal cancer, appendiceal cancer, astrocytoma, atypical teratoma / rhabdoid tumor, basal cell tumor, cholangiocarcinoma, bladder cancer, bone cancer, brain cancer (e.g., glioblastoma), breast cancer, bronchial cancer, cancer of unknown primary site, carcinoid tumor, castration-resistant prostate cancer, central nervous system cancer (e.g., central nervous system atypical teratoma / rhabdoid tumor, central nervous system embryonal tumor, central nervous system lymphoma, primary central nervous system lymphoma), cervical cancer, chordoma, chondrosarcoma, chronic myeloproliferative disorder, colon cancer, colorectal cancer, craniopharyngioma, desmoplastic Round cell tumor, diffuse large B-cell lymphoma, endometrial cancer, ependymoma, esophageal cancer, Ewing's sarcoma family of tumors, extracranial germ cell tumors, extragonadal cell tumors, extrahepatic bile duct cancer, gallbladder cancer, gastric (stomach) cancer, gastrointestinal carcinoid tumors, gastrointestinal stromal tumors, germ cell tumors, gestational trophoblastic tumors, glioma, head cancer, hepatocellular (liver) cancer, high-grade prostate cancer, histiocytic proliferation, hypopharyngeal cancer, Kaposi's sarcoma, kidney (kidney) cancer, Langerhans cell histiocytosis, laryngeal cancer, leptomeningeal disease, lip cancer, low-grade prostate cancer, leukemia (e.g., chronic lymphocytic leukemia, chronic myeloid leukemia, hairy cell leukemia), lung cancer, lymphoma lymphoma (Burkitt's lymphoma, central nervous system lymphoma, T-cell lymphoma (e.g., cutaneous T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, lymphoplasmacytic lymphoma), intermediate-grade prostate cancer, medulloblastoma, medulloepithelioma, melanoma, Merkel cell carcinoma, mesothelioma, metastatic squamous cell carcinoma of unknown primary, oral cancer, multiple endocrine neoplasia syndrome, multiple myeloma, multiple myeloma / plasma cell neoplasm, mycosis fungoides, myelodysplastic syndrome, myelodysplastic / myeloproliferative neoplasm, myeloproliferative disorder, nasal cavity or paranasal sinus cancer, neck cancer, nasopharyngeal cancer, neck cancer, neuroblastoma, eye cancer, ocular melanoma , oral cancer, oropharyngeal cancer, oral cancer, osteosarcoma or malignant fibrous histiocytoma of bone, osteosarcoma or malignant fibrous histiocytoma, ovarian cancer, ovarian germ cell tumor, ovarian epithelial cancer, ovarian low malignant potential tumor, pancreatic cancer, papillomatosis, paranasal sinus or nasal cancer, parathyroid cancer, penile cancer, pharyngeal cancer, moderately differentiated pineal parenchymal tumor, pineoblastoma or supratentorial primitive neuroectodermal tumor, pituitary tumor, pleuropulmonary blastoma, pregnancy cancer, prostate cancer, rectal cancer, renal pelvis cancer, airway cancer involving the NUT gene on chromosome 15, retinoblastoma, rhabdomyoblastoma, salivary gland cancer, sarcoma, Sezary syndrome, skin cancer, skin cancer, small intestine cancer, soft tissue sarcoma, spinal tumor, squamous cell carcinoma,38. The method of embodiment 37, wherein the cancer is selected from squamous cell cervical cancer of unknown primary, occult primary supratentorial primitive neuroectodermal tumor, testicular cancer, laryngeal cancer, thymoma or thymic carcinoma, thyroid cancer, transitional cell carcinoma of the renal pelvis or ureter, rare cancers of childhood, ureteral cancer, urethral cancer, uterine sarcoma, vaginal cancer, vulvar cancer, Waldenstrom's macroglobulinemia (lymphoplasmacytic lymphoma), Wilms' tumor, or female cancer.

[0187] Working Example Preparation of intermediate compounds Example 1: 1-(1-Methyl-4-piperidyl)pyrazol-4-amine (IM1)

[0188] [ka] Step 1: 4-nitro-1H-pyrazole (10 g, 88.44 mmol, 1.0 equiv.), tert-butyl 4-hydroxypiperidine-1-carboxylate (1-1, 17.80 g, 88.44 mmol, 1.0 equiv.), and PPh in THF (400 mL). 3 (34.79 g, 132.66 mmol, 1.5 equiv.) was added to a solution of DIAD (26.82 g, 132.66 mmol, 1.5 equiv.) with N 2 The mixture was added dropwise at -60°C under reduced pressure. The mixture was warmed to 20°C and stirred for 16 h. The mixture was then diluted with EtOAc (200 mL), washed with water (200 mL) / brine (200 mL), dried, filtered and concentrated. The residue was purified by silica gel column chromatography to give tert-butyl 4-(4-nitropyrazol-1-yl)piperidine-1-carboxylate (1-2, 25 g) as a white solid. ESI[M+H-100]=197.3

[0189] Step 2: A solution of tert-butyl 4-(4-nitropyrazol-1-yl)piperidine-1-carboxylate (1-2, 10 g, 33.75 mmol) in HCl / EtOAc (4 M, 100 mL) was stirred at 20° C. for 1 h and then concentrated to give 4-(4-nitropyrazol-1-yl)piperidine (1-3) as a white solid. ESI [M+H]=197.3

[0190] Step 3: To a solution of 4-(4-nitropyrazol-1-yl)piperidine (1-3, 8.2 g, 35.24 mmol, HCl salt) in MeOH (100 mL) was added DIEA (9.11 g, 70.49 mmol, 2.0 equiv.) and formaldehyde (1.59 g, 52.87 mmol, 1.5 equiv.). The mixture was stirred at 20° C. for 15 min. Then, NaBH 3 CN (4.43 g, 70.49 mmol, 2.0 equiv) was added and the mixture was stirred at 20° C. for an additional 45 min. The reaction mixture was cooled to 5° C. 2 The mixture was diluted with 200 mL of EtOAc (300 mL) and extracted with EtOAc (500 mL x 2). The combined organic layers were washed with Na 2 SO 4 The mixture was dried at 40° C., filtered and concentrated. The residue was purified by silica gel column chromatography to give 1-methyl-4-(4-nitropyrazol-1-yl)piperidine (1-4, 2 g) as a white solid. ESI [M+H]=211.2.

[0191] Step 4: A mixture of 1-methyl-4-(4-nitropyrazol-1-yl)piperidine (1-4, 2 g, 9.51 mmol) and Pd / C (2 g, 10% purity) in EtOAc (20 mL) is degassed and diluted with H 2 After purging three times with H 2 (15 Psi) at 30° C. for 2 h. The reaction mixture was filtered and concentrated to give 1-(1-methyl-4-piperidyl)pyrazol-4-amine (IM1, 1.8 g, crude product) as a pink oil. ESI [M+H]=181.2.

[0192] Example 2: tert-Butyl 4-(4-amino-1H-pyrazol-1-yl)piperidine-1-carboxylate (IM2)

[0193] [ka] A mixture of tert-butyl 4-(4-nitropyrazol-1-yl)piperidine-1-carboxylate (1-2, 3 g, 10.12 mmol) and Pd / C (1 g, 10% purity) in EtOAc (30 mL) was diluted with H 2 (15 psi) at 20° C. for 12 h. The mixture was filtered and concentrated to give tert-butyl 4-(4-aminopyrazol-1-yl)piperidine-1-carboxylate (IM2, 1.4 g) as a brown oil. ESI [M+H]=267.2.

[0194] Example 3: 8-Bromo-6-(2,6-dichlorophenyl)-2-(methylthio)pyrido[4,3-d]pyrimidin-5(6H)-one (IM3).

[0195] [ka] Step 1: A mixture of 2-methylisothiourea, sulfuric acid (28.88 g, 103.74 mmol, 0.5 equiv.), and 1,1-dimethoxy-N,N-dimethyl-methanamine (3-1, 24.72 g, 207.47 mmol) was heated at 100° C. for 3 h. The mixture was cooled to 20° C., a solution of ethyl 3-oxobutanoate (27 g, 207.47 mmol, 1 equiv.) in EtOH (120 mL) was added, and the resulting mixture was heated at 100° C. for 12 h, cooled to room temperature, and concentrated. The residue was diluted with EtOAc (1.0 L), washed with water (500 mL) / brine (500 mL), dried, and concentrated to give ethyl 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylate (3-2, 41.12 g) as a yellow oil. ESI[M+H]=213.1.

[0196] Step 2: A solution of 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylate (3-2, 41.12 g, 193.72 mmol) in THF (150 mL) / MeOH (50 mL) was treated with LiOH.H 2 HO (15 g, 357.45 mmol, 1.85 equiv.) 22H2O solution (150 mL) was added. The mixture was stirred at 20° C. for 12 h and concentrated to remove MeOH and THF. The remaining aqueous layer was extracted with methyl tert-butyl ether (300 mL). The aqueous layer was acidified with 1N HCl to pH 2. The precipitate was collected and dried to give 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylic acid (3-3, 24.4 g) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ=8.88(s,1H),2.66(s,3H),2.54(s,3H).

[0197] Step 3: A mixture of 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylic acid (3-3, 18.2 g, 98.80 mmol) and 2,6-dichloroaniline (16.01 g, 98.80 mmol, 1 equiv.) in chlorobenzene (455 mL) was degassed and diluted with N 2 The mixture was purged three times with PCl 3 (13.57 g, 98.80 mmol, 1 equiv.) was added and the mixture was treated with N 2 The mixture was stirred at 135° C. for 16 hours under reduced pressure. The mixture was concentrated, and the residue was washed with saturated Na 2 CO 3 Poured into aqueous solution (500 mL) and extracted with 2-methyl-THF (1 L) / EtOAc (500 mL). The organic phase was dried and concentrated. The residue was triturated with EtOAc (100 mL) to give N-(2,6-dichlorophenyl)-4-methyl-2-methylsulfanyl-pyrimidine-5-carboxamide (3-4, 18.0 g) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ=10.54(s,1H),8.74(s,1H),7.61(d,J=8.00Hz,2H),7.40-7.45(m,1H),2.61(s,3H),2.57(s,3H). ESI[M+H]=328.0 / 330.0.

[0198] Step 4: To a solution of N-(2,6-dichlorophenyl)-4-methyl-2-methylsulfanyl-pyrimidine-5-carboxamide (3-4, 15 g, 45.70 mmol) in DMF (150 mL) was added DMF-DMA (16.34 g, 137.10 mmol, 3 equiv). The resulting mixture was stirred at 80° C. for 12 h and then concentrated. The residue was triturated with EtOH (50 mL) to give 6-(2,6-dichlorophenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (3-5, 5.3 g) as a white solid. ESI [M+H]=338.1 / 340.1.

[0199] Step 5: To a solution of 6-(2,6-dichlorophenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (3-5, 3.88 g, 11.47 mmol) in MeCN (56 mL) was added NBS (3.06 g, 17.21 mmol, 1.5 equiv.) and the mixture was stirred at 80° C. for 16 h. The mixture was concentrated and the residue was triturated with EtOH (20 mL) to give 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (IM3, 3.1 g, as a pale yellow solid). ESI [M+H]=417.9 / 415.9.

[0200] Example 4: 8-Bromo-6-(2,6-dichlorophenyl)-2-(methylsulfinyl)pyrido[4,3-d]pyrimidin-5(6H)-one (IM4).

[0201] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (IM3, 100 mg, 239.75 μmol) in DCM (5 mL) was added m-CPBA (62.06 mg, 359.62 μmol, 85% purity, 1.5 equiv.). The mixture was stirred at 20° C. for 1 h and then concentrated to give IM4, which was used for further purification. ESI [M+H]=433.9 / 431.9

[0202] Example 5: tert-Butyl 4-(4-((8-bromo-6-(2,6-dichlorophenyl)-5-oxo-5,6-dihydropyrido[4,3-d]pyrimidin-2-yl)amino)-1H-pyrazol-1-yl)piperidine-1-carboxylate (IM5)

[0203] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (IM4, 1 g, 2.31 mmol) in DCE (50 mL) / DCM (50 mL) was added tert-butyl 4-(4-aminopyrazol-1-yl)piperidine-1-carboxylate (IM2, 1.23 g, 4.62 mmol, 2 equiv.) followed by AcOH (138.65 mg, 2.31 mmol, 1 equiv.). The mixture was stirred at 50° C. for 12 h. The mixture was concentrated and the residue was purified by reverse phase preparative HPLC to give tert-butyl 4-[4-[[8-bromo-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (IM5, 1.4 g) as a yellow solid. ESI[M+H-56]=580.1 / 578.2.

[0204] Example 6: tert-Butyl 4-[4-[[6-(2,6-dichlorophenyl)-8-isopropenyl-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (IM6)

[0205] [ka] Dioxane (5 mL) and H 2tert-Butyl 4-[4-[[8-bromo-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (IM5, 200 mg, 314.79 μmol), 2-isopropenyl-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (105.80 mg, 629.58 μmol, 2 equiv.), Pd(dppf)Cl in O (1 mL). 2 DCM (128.53 mg, 157.40 μmol, 0.5 equiv.), and K 2 CO 3 A mixture of (87.01 mg, 629.58 μmol, 2 equiv.) was degassed and diluted with N 2 The mixture was stirred at 100° C. for 12 h, cooled to room temperature, and purified with H 2 The combined organic layers were poured into 2024 (5 mL) and extracted with EtOAc (20 mL x 3). 2 SO 4 The residue was purified by preparative HPLC to give tert-butyl 4-[4-[[6-(2,6-dichlorophenyl)-8-isopropenyl-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (IM6, 80 mg) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ=10.40(s,1H),9.27-9.07(m,1H),8.20-8.02(m,1H),7.77-7.66(m,3H),7.65-7.57(m,2H),5.48-5.13(m,2H),4.45-4 .28(m,1H),4.14-3.94(m,2H),3.10-2.82(m,2H),2.24-2.16(m,3H),2.11-1.97(m,2H),1.84-1.65(m,2H),1.45(s,9H). ESI[M+H]=596.2 / 598.2

[0206] Preparation of exemplary compounds of the invention Example 7: 8-Bromo-6-(2,6-dichlorophenyl)-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P1)

[0207] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (IM4, 100 mg, 230.89 μmol) in DCE (8 mL) was added 1-(1-methyl-4-piperidyl)pyrazol-4-amine (IM1, 83.24 mg, 461.78 μmol, 2 equiv.) and TFA (26.33 mg, 230.89 μmol, 1 equiv.). The mixture was stirred at 50° C. for 12 h, concentrated, and purified by preparative HPLC to give 8-bromo-6-(2,6-dichlorophenyl)-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P1, 141.97 mg) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ=10.71(s,1H),9.09(s,1H),8.41(d,J=18.39Hz,2H),7.82(s,1H),7.71-7.76(m,2H ),7.56-7.66(m,1H),4.21(brs,1H),3.00(brs,2H),2.34(brs,4H),1.91-2.15(m,5H). ESI[M+H]=550.0 / 548.0.

[0208] Example 8: 6-(2,6-dichlorophenyl)-8-methyl-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P2)

[0209] [ka] Step 1: 8-Bromo-6-(2,6-dichlorophenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (IM3, 0.1 g, 239.75 μmol), 2,4,6-trimethyl-1,3,5,2,4,6-trioxatriborinane (120.39 mg, 479.49 μmol, 50% purity, 2 equiv.), Pd(dppf)Cl in dioxane (12 mL). 2 DCM (9.79 mg, 11.99 μmol, 0.05 equiv.), and K 2 CO 3 (99.41 mg, 719.24 μmol, 3 equiv.) was added to a mixture of N 2 The mixture was stirred at 100° C. for 6 h under reduced pressure. The mixture was concentrated and the residue was purified by preparative HPLC to give 6-(2,6-dichlorophenyl)-8-methyl-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (8-1, 30 mg) as an off-white solid. ESI [M+H]=352.0 / 354.0.

[0210] Step 2: To a solution of 6-(2,6-dichlorophenyl)-8-methyl-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (8-1, 20 mg, 56.78 μmol, 1 eq.) in DCM (1 mL) was added m-CPBA (9.80 mg, 56.78 μmol, 85% purity, 1 eq.). The mixture was stirred at 20° C. for 1 h and then used in the next step without further purification. ESI [M+H]=368.0 / 370.0.

[0211] Step 3: To a solution of 6-(2,6-dichlorophenyl)-8-methyl-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (8-2, 9.58 mg, 26.03 μmol) in DCE (0.5 mL) was added a solution of 1-(1-methyl-4-piperidyl)pyrazol-4-amine (IM1, 9.38 mg, 52.05 μmol, 2 equiv.) in DCE (1 mL), followed by TFA (2.97 mg, 26.03 μmol, 1 equiv.). The mixture was stirred at 50° C. for 12 h and then concentrated. The residue was purified by preparative HPLC to give 6-(2,6-dichlorophenyl)-8-methyl-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P2, 9.39 mg, HCl) as a yellow gum. 1 H NMR (400MHz, DMSO-d 6 )δ=10.17-10.48(m,1H),9.47-9.90(m,1H),9.11(s,1H),8.12(s,1H),7.79(s,1H),7.73(d,J=8.13Hz,2H),7.67(s, 1H),7.56-7.61(m,1H),4.47(brs,1H),3.41(brs,2H),3.16(brd,J=10.88Hz,2H),2.83(brs,3H),2.07-2.33(m,7H). ESI[M+H]=484.1 / 486.1.

[0212] Example 9: 8-Cyclopropyl-6-(2,6-dichlorophenyl)-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P3)

[0213] [ka] Step 1: tert-Butyl 4-[4-[[8-bromo-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (IM5, 200 mg, 314.79 μmol), cyclopropylboronic acid (135.20 mg, 1.57 mmol, 5 equiv.), Pd(dppf)Cl2 A mixture of dioxane (4 mL) and H 2 DCM (128.54 mg, 157.40 μmol, 0.5 equiv.) and K 2 CO 3 (87.01 mg, 629.59 μmol, 2 equiv.) was degassed and diluted with N 2 The resulting mixture was stirred at 100° C. for 12 h, cooled to room temperature, concentrated, and purified with H 2 The mixture was diluted with 200 mL of ethyl acetate (10 mL × 3) and extracted with ethyl acetate (10 mL × 3). The combined organic layers were washed with Na 2 SO 4 The residue was purified by preparative HPLC to give tert-butyl 4-[4-[[8-cyclopropyl-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (9-1, 70 mg) as a yellow solid. ESI [M+H] = 596.2 / 598.2.

[0214] Step 2: To a solution of tert-butyl 4-[4-[[8-cyclopropyl-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (9-1, 70 mg, 117.35 μmol) in DCM (10 mL) was added TFA (7.70 g, 67.53 mmol), the mixture was stirred at 20 °C for 1 h, concentrated, and the residue was purified by preparative HPLC to give 8-cyclopropyl-6-(2,6-dichlorophenyl)-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P3, 13.81 mg, HCl) as a yellow gum. 1 H NMR (400MHz, DMSO-d 6)δ=10.47(s,1H),9.36(brs,2H),9.10(s,1H),8.20(s,1H),7.81(s,1H),7.70(d,J=8.00Hz,2H),7.50-7.60(m,1H),7.46(s,1H) ,4.50(brs,1H),3.36(brd,J=12.26Hz,2H),3.06(brs,2H),2.18(brs,5H),0.97(brd,J=8.13Hz,2H),0.65(brd,J=4.50Hz,2H). ESI[M+H]=486.2 / 498.2.

[0215] Example 10: 6-(2,6-dichlorophenyl)-8-ethyl-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P4) and 6-(2,6-dichlorophenyl)-8-ethyl-2-((1-(1-methylpiperidin-4-yl)-1H-pyrazol-4-yl)amino)pyrido[4,3-d]pyrimidin-5(6H)-one (P5)

[0216] [ka] Step 1: 2-Methyl-2-nutanol (18 mL) and H 2 tert-Butyl 4-[4-[[8-bromo-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (IM5, 300 mg, 472.19 μmol), ethylboronic acid (139.55 mg, 1.89 mmol, 4 equiv.), CataCXiumA Pd G2 (63.14 mg, 94.44 μmol, 0.2 equiv.) and Cs in 200 (4.5 mL). 2 CO 3 (307.70 mg, 944.38 μmol, 2 equiv.) was degassed and diluted with N 2The mixture was purged three times with 500 cc of 1000 rt, and then the mixture was stirred at 90° C. for 12 h. The mixture was concentrated and the residue was purified by preparative HPLC to give tert-butyl 4-[4-[[6-(2,6-dichlorophenyl)-8-ethyl-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (10-1, 30 mg) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ=10.38(s,1H),9.11(s,1H),8.12(s,1H),7.69-7.75(m,3H),7.49-7.64(m,2H),4.38(brs,1H),4.04(brd,J=12.26Hz,2H),2 .87-3.01(m,2H),2.75(brd,J=7.38Hz,2H),2.05(brd,J=10.76Hz,2H),1.70-1.80(m,2H),1.42(s,9H),1.26(t,J=7.38Hz,3H). ESI[M+H]=584.2 / 586.2.

[0217] Step 2: A mixture of tert-butyl 4-[4-[[6-(2,6-dichlorophenyl)-8-ethyl-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (10-1, 30 mg, 51.33 μmol) in TFA / DCM (1:3, 4 mL) was stirred for 1 h at 20° C. The mixture was concentrated and the residue was purified by preparative HPLC to give 6-(2,6-dichlorophenyl)-8-ethyl-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P4, 23.78 mg, HCl) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6)δ=10.43(s,1H),9.24-9.46(m,3H),8.10(s,1H),7.78(s,1H),7.71(d,J=8.13Hz,2H),7.49-7.60(m,2H),4.45 -4.59(m,1H),3.37(brd,J=12.38Hz,2H),3.07(brs,2H),2.61-2.79(m,2H),2.19(brs,4H),1.14-1.34(m,3H). ESI[M+H]=484.1 / 486.1.

[0218] Step 3: To a solution of 6-(2,6-dichlorophenyl)-8-ethyl-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P4, 80 mg, 133.69 μmol) in MeOH (4 mL) was added HCHO (32.55 mg, 401.07 μmol, 37% purity, 3 equiv.). The mixture was stirred at 20° C. for 0.5 h followed by addition of NaBH 3 CN (8.40 mg, 133.69 μmol, 1 equiv.) was added. The mixture was stirred at 20° C. for 1 h, then concentrated and the residue was purified by preparative HPLC to give 6-(2,6-dichlorophenyl)-8-ethyl-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P5, 7.77 mg, HCl) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ=11.10(brs,1H),10.39-10.48(m,1H),9.11(s,1H),8.01-8.22(m,1H),7.65-7.86(m,3H),7.47-7.64(m,2H),4.46(b rs,1H),3.52(brd,J=11.88Hz,2H),3.16(brd,J=10.01Hz,2H),2.63-2.84(m,5H),2.18-2.44(m,4H),1.09-1.33(m,3H). ESI[M+H]=498.2 / 500.1.

[0219] Example 11: 6-(2,6-dichlorophenyl)-2-((1-(piperidin-4-yl)-1H-pyrazol-4-yl)amino)-8-(prop-1-en-2-yl)pyrido[4,3-d]pyrimidin-5(6H)-one (P6)

[0220] [ka] A mixture of tert-butyl 4-[4-[[6-(2,6-dichlorophenyl)-8-isopropenyl-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (IM6, 10 mg, 16.76 μmol) in DCM (1 mL) / TFA (0.2 mL) was stirred for 0.5 h at 20° C. The mixture was concentrated and the residue was purified by preparative HPLC to give 6-(2,6-dichlorophenyl)-8-isopropenyl-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P6, 8.32 mg, HCl) as a yellow gum. 1 H NMR (400MHz, DMSO-d 6 )δ=10.56-10.15(m,1H),9.45-8.86(m,3H),8.18-7.98(m,1H),7.83-7.47(m,5H),5.52-5.40(m,1H), 5.36-5.15(m,1H),4.58-4.43(m,1H),3.41(brd,J=11.4Hz,2H),3.20-3.00(m,2H),2.38-2.03(m,7H). ESI[M+H]=496.1 / 498.1

[0221] Example 12: 6-(2,6-dichlorophenyl)-8-isopropenyl-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P7)

[0222] [ka] To a solution of 6-(2,6-dichlorophenyl)-8-isopropenyl-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P6, 70 mg, 114.68 μmol, 1 equiv., TFA) in MeOH (2 mL) was added DIEA (29.64 mg, 229.35 μmol, 2 equiv.) and HCHO (27.92 mg, 344.03 μmol, 37% purity, 3 equiv.). The mixture was stirred at 20° C. for 0.5 h and then NaBH 3 CN (7.21 mg, 114.68 μmol, 1 equiv.) was added. The mixture was stirred at 20° C. for 1 h, then concentrated and the residue was purified by preparative HPLC to give 6-(2,6-dichlorophenyl)-8-isopropenyl-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P7, 13.78 mg) as a white solid. 1 H NMR (400MHz, DMSO-d 6 )δ=10.37(s,1H),9.14(s,1H),8.18-7.99(m,1H),7.74(d,J=8.1Hz,2H),7.70-7.57(m,3H),5.50 -5.15(m,2H),4.17-4.01(m,1H),2.88(brd,J=11.5Hz,2H),2.26-2.16(m,6H),2.13-1.85(m,6H). ESI[M+H]=510.1 / 512.2

[0223] Example 13: 6-(2,6-dichlorophenyl)-8-isopropyl-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P8) and 6-(2,6-dichlorophenyl)-8-isopropyl-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P9)

[0224] [ka] Step 1: tert-Butyl 4-[4-[[6-(2,6-dichlorophenyl)-8-isopropenyl-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (IM6, 10 mg, 16.76 μmol) in EtOAc (1 mL) and PtO 2 A mixture of (10 mg) was degassed and H 2 The mixture was then purged three times with H 2 The mixture was stirred under reduced pressure (15 Psi) for 0.5 h at 0° C. The mixture was filtered, concentrated, and the residue was purified by preparative HPLC to give tert-butyl 4-[4-[[6-(2,6-dichlorophenyl)-8-isopropyl-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (13-1, 2.28 mg) as a white solid. 1 H NMR (400MHz, METHANOL-d 4 )δ=9.38-9.17(m,1H),8.30-8.10(m,1H),7.85-7.77(m,1H),7.70-7.62(m,2H),7.59-7.51(m,1H),7.33-7.19(m,1H),4.47-4.33(m,1H) ,4.25(brd,J=13.6Hz,2H),3.70-3.52(m,1H),3.12-2.90(m,2H),2.22-2.08(m,2H),2.00-1.86(m,2H),1.51(s,9H),1.44-1.28(m,6H). ESI[M+H]=598.2 / 600.2

[0225] Step 2: A mixture of tert-butyl 4-[4-[[6-(2,6-dichlorophenyl)-8-isopropyl-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (13-1, 95 mg, 158.72 μmol) in DCM (2 mL) / TFA (0.4 mL) was stirred for 0.5 h at 20° C. The mixture was concentrated and the residue was purified by preparative HPLC to give 6-(2,6-dichlorophenyl)-8-isopropyl-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P8, 50 mg, 81.64 μmol, TFA) as a grey solid. 1 H NMR (400MHz, DMSO-d 6 )δ=10.47-10.15(m,1H),9.26-9.09(m,1H),8.85-8.37(m,2H),8.15-8.01(m,1H),7.82(s,1H),7.74(d,J=8.2Hz,2H),7.71-7 .57(m,1H),7.52-7.45(m,1H),4.59-4.46(m,1H),3.54-3.44(m,3H),3.23-3.02(m,2H),2.27-2.05(m,4H),1.43-0.92(m,6H). ESI[M+H]=498.1 / 500.2

[0226] Step 3: To a solution of 6-(2,6-dichlorophenyl)-8-isopropyl-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P8, 35 mg, 57.15 μmol, TFA) in MeOH (2 mL) was added DIEA (14.77 mg, 114.30 μmol, 2 equiv.) and HCHO (13.91 mg, 171.45 μmol, 37% purity, 3 equiv.). The mixture was stirred at 20° C. for 0.5 h and then NaBH 3CN (3.59 mg, 57.15 μmol, 1 equiv.) was added. The mixture was stirred at 20° C. for 1 h, then concentrated and the residue was purified by preparative HPLC to give 6-(2,6-dichlorophenyl)-8-isopropyl-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (P9, 11.8 mg) as an off-white solid. 1 H NMR (400MHz, DMSO-d 6 )δ=10.46-10.09(m,1H),9.27-9.05(m,1H),8.17-7.92(m,1H),7.73(brd,J=8.4Hz,2H),7.59(brd,J=7.1Hz,1H),7.52-7.4 1(m,1H),4.24-4.06(m,1H),3.30-3.11(m,3H),3.02-2.86(m,2H),2.31-2.13(m,4H),2.09-1.92(m,3H),1.51-0.83(m,6H). ESI[M+H]=512.2 / 514.2

[0227] Example 14: 8-Bromo-6-(2,6-dichlorophenyl)-2-(1,2,3,4-tetrahydroisoquinolin-7-ylamino)pyrido[4,3-d]pyrimidin-5-one (P10) and 8-Bromo-6-(2,6-dichlorophenyl)-2-[(2-methyl-3,4-dihydro-1H-isoquinolin-7-yl)amino]pyrido[4,3-d]pyrimidin-5-one (P11)

[0228] [ka] Step 1: To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (IM5, 180 mg, 415.60 μmol) in DCE (2 mL) was added tert-butyl 7-amino-3,4-dihydro-1H-isoquinoline-2-carboxylate (206.40 mg, 831.20 μmol, 2.0 equiv) followed by AcOH (124.78 mg, 2.08 mmol, 5.0 equiv). The mixture was stirred at 50° C. for 1 h and then concentrated to give tert-butyl 7-[8-bromo-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]-3,4-dihydro-1H-isoquinoline-2-carboxylate (14-1, 250 mg, crude) as a yellow solid. ESI [M+H]=618.2 / 616.2.

[0229] Step 2: A mixture of tert-butyl 7-[[8-bromo-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]-3,4-dihydro-1H-isoquinoline-2-carboxylate (14-1, 250 mg, 404.98 μmol) in HCl / MeOH (3 mL, 4 M) was stirred for 1 h at 20° C. The mixture was concentrated and the residue was purified by preparative HPLC to give 8-bromo-6-(2,6-dichlorophenyl)-2-(1,2,3,4-tetrahydroisoquinolin-7-ylamino)pyrido[4,3-d]pyrimidin-5-one (P10, 148.05 mg, HCl) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ=10.71(brs,1H),9.37(brs,2H),9.20(s,1H),8.47(s,1H),8.17(brs,1H),7.76(d,J=8.00Hz,3H) ,7.58-7.66(m,1H),7.24(d,J=8.38Hz,1H),4.61(brs,2H),4.30(brs,2H),3.00(brt,J=5.88Hz,2H). ESI[M+H]=518.0 / 516.0

[0230] Step 3: To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-(1,2,3,4-tetrahydroisoquinolin-7-yl)amino)pyrido[4,3-d]pyrimidin-5-one (P10, 60 mg, 108.37 μmol, HCl) in MeOH (2 mL) was added DIEA (28.01 mg, 216.74 μmol, 2.0 equiv.) and HCHO (26.39 mg, 325.11 μmol, 37% pure, 3.0 equiv.). The mixture was stirred at 20° C. for 0.5 h and then NaBH 3 CN (6.81 mg, 108.37 μmol, 1.0 equiv.) was added. The mixture was stirred at 20° C. for 0.5 h, concentrated, and the residue was purified by preparative HPLC to give 8-bromo-6-(2,6-dichlorophenyl)-2-[(2-methyl-3,4-dihydro-1H-isoquinolin-7-yl)amino]pyrido[4,3-d]pyrimidin-5-one (P11, 26.22 mg, HCl) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ=11.58(brs,1H),10.78(brs,1H),9.24(s,1H),8.22-8.56(m,2H),7.79(d,J=8.13Hz,3H),7.61-7.70(m,1H) ),7.28(d,J=8.50Hz,1H),4.31-4.58(m,2H),3.67(brd,J=8.76Hz,1H),3.24-3.44(m,2H),2.90-3.08(m,4H). ESI[M+H]=532.0 / 530.0

[0231] Example 15: Preparation of compound P30

[0232] [ka]

[0233] (i) Preparation of Compound 2 A mixture of 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylic acid (3 g, 16.29 mmol, 1.0 equiv.), 2,6-dimethylaniline (1.97 g, 16.29 mmol, 1.0 equiv.) in chlorobenzene (50 mL) was degassed and purified with N 2purged three times with 3 (2.24 g, 16.29 mmol, 1.0 equiv.) was added. The mixture was diluted with N 2 The mixture was stirred at 135° C. for 3 hours under atmospheric pressure. The mixture was concentrated, and the residue was washed with saturated Na 2 CO 3 The organic layer was poured into aqueous solution (50 mL) and extracted with 2-methyltetrahydrofuran (50 mL x 3). 2 SO 4 The residue was triturated with EtOH (20 mL) to give N-(2,6-dimethylphenyl)-4-methyl-2-methylsulfanyl-pyrimidine-5-carboxamide (2.45 g, 8.53 mmol, 52.35% yield) as a yellow solid. ESI [M+H] = 288.1.

[0234] (ii) Preparation of Compound 3

[0235] [ka] To a solution of N-(2,6-dimethylphenyl)-4-methyl-2-methylsulfanyl-pyrimidine-5-carboxamide (2.45 g, 8.53 mmol, 1.0 equiv.) in DMF (50 mL) was added DMF-DMA (6.10 g, 51.15 mmol, 6.0 equiv.) and the mixture was stirred at 80° C. for 12 h. The mixture was concentrated and the residue was triturated with EtOH (15 mL) to give 6-(2,6-dimethylphenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (1.76 g, 5.92 mmol, 69.42% yield) as a pale yellow solid. ESI [M+H]=298.1.

[0236] (iii) Preparation of Compound 4

[0237] [ka] To a solution of 6-(2,6-dimethylphenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (1.76 g, 5.92 mmol, 1.0 equiv.) in ACN (30 mL) was added NBS (1.58 g, 8.88 mmol, 1.5 equiv.), the mixture was degassed and purified with N 2 The mixture was purged with N 2 The mixture was stirred under atmosphere at 80° C. for 12 h. The mixture was concentrated and the residue was triturated with EtOH (10 mL) to give 8-bromo-6-(2,6-dimethylphenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (2 g, 5.32 mmol, 89.81% yield) as a pale yellow solid. ESI [M+H]=376.2 / 378.2.

[0238] (iv) Preparation of Compound 5

[0239] [ka] To a solution of 8-bromo-6-(2,6-dimethylphenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (500 mg, 1.33 mmol, 1.0 equiv.) in DCM (10 mL) was added m-CPBA (404.67 mg, 1.99 mmol, 85% purity, 1.5 equiv.) at 0° C. The mixture was stirred at 20° C. for 1 h and then used in the next step without further purification. ESI [M+H]=391.9 / 393.9.

[0240] (v) Preparation of compound 6

[0241] [ka] To a solution of 8-bromo-6-(2,6-dimethylphenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (520 mg, 1.33 mmol, 1.0 equiv) in DCE (12 mL) was added tert-butyl 4-(4-aminopyrazol-1-yl)piperidine-1-carboxylate (708.46 mg, 2.66 mmol, 2.0 equiv) and AcOH (2.00 g, 33.25 mmol, 25.0 equiv). The mixture was stirred at 50° C. for 1 h and then concentrated to give tert-butyl 4-[4-[[8-bromo-6-(2,6-dimethylphenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (710 mg, crude) as a brown gum. ESI[M+H-56]=538.3 / 540.3.

[0242] (vi) Preparation of compound P30

[0243] [ka] A mixture of tert-butyl 4-[4-[[8-bromo-6-(2,6-dimethylphenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (700 mg, 1.18 mmol, 1.0 equiv) in DCM (6 mL) and TFA (2 mL) was stirred for 12 h at 20° C. The mixture was concentrated and purified by preparative HPLC (column: Phenomenex luna C18 250 * 50mm * 10 μm; mobile phase: [water (0.1% TFA)-ACN]; B%: 15% to 45%, 10 min) to give 8-bromo-6-(2,6-dimethylphenyl)-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (400 mg, 655.60 μmol, yield 55.68%, purity 99.718%, TFA) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6)δ=10.68(s,1H),9.13(s,1H),8.88-8.70(m,1H),8.62-8.44(m,2H),8.29-7.98(m,1H),7.89-7.66(m,1H),7.3 7-7.23(m,3H),4.62-4.48(m,1H),3.52-3.39(m,2H),3.20-3.04(m,2H),2.30-2.20(m,2H),2.20-2.03(m,8H). ESI[M+H]=494.1 / 496.1.

[0244] Example 16: Preparation of compound P27

[0245] [ka] To a solution of 8-bromo-6-(2,6-dimethylphenyl)-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (250 mg, 505.68 μmol, 1.0 equiv., TFA) in MeOH (4 mL) was added DIEA (130.71 mg, 1.01 mmol, 2.0 equiv.) and HCHO (123.13 mg, 1.52 mmol, 37% purity, 3.0 equiv.). The mixture was stirred at 20° C. for 0.5 h followed by addition of NaBH 3 CN (63.56 mg, 1.01 mmol, 2.0 equiv) was added. The mixture was stirred at 20° C. for 1 h, then concentrated and purified by preparative HPLC (column: Welch Xtimate C18 100 * 25mm * 3um; mobile phase: [water (0.05% HCl)-ACN]; B%: 10% to 30%, 8 min) to give 8-bromo-6-(2,6-dimethylphenyl)-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (144.15 mg, 258.29 μmol, yield 51.08%, purity 97.630%, HCl) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6)δ11.06-10.78(m,1H),10.72-10.43(m,1H),9.10(s,1H),8.60-8.42(m,1H),8.22-7.91(m,1H),7.63(d,J=1.5Hz,1H),7.34-7.2 0(m,3H),4.54-4.40(m,1H),3.58-3.33(m,2H),3.23-3.07(m,2H),2.75(brd,J=4.6Hz,3H),2.40-2.19(m,4H),2.08-1.99(m,6H). ESI[M+H]=508.1 / 510.1.

[0246] Example 17: Preparation of compound P24 (i) Preparation of Compound 2

[0247] [ka] To a solution of 6-aminoindan-1-one (250 mg, 1.70 mmol, 1.0 equiv) in MeOH (4 mL) was added NaBH 4 (0.02 g, 528.65 μmol, 0.3 equiv.) was added and stirred at 20° C. for 2 h. The mixture was quenched with 1 N HCl (5 mL) followed by saturated Na 2 CO 3 Add aqueous solution (10 mL), extract with 2-methyltetrahydrofuran (20 mL x 3), and add Na 2 SO 4 The organic layer was filtered and concentrated to give 6-aminoindan-1-ol (240 mg, crude) as a pink solid. 1 H NMR (400MHz, DMSO-d 6 )δ=6.90-6.83(m,1H),6.58(d,J=1.7Hz,1H),6.44(dd,J=2.1,7.9Hz,1H),5.04(d,J=6.4Hz,1H),4.94(b rs,2H),4.91-4.87(m,1H),2.79-2.68(m,1H),2.59-2.53(m,1H),2.31-2.19(m,1H),1.77-1.65(m,1H). ESI[M+H]=150.3.

[0248] (ii) Preparation of Compound 4

[0249] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (100 mg, 239.75 μmol, 1.0 equiv.) in DCM (2 mL) was added m-CPBA (63.28 mg, 311.67 μmol, 85% purity, 1.3 equiv.) at 0° C. The mixture was stirred at 20° C. for 1 h and then used in the next step without further purification. ESI [M+H]=431.8 / 433.8.

[0250] (iii) Preparation of compound P24

[0251] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (100 mg, 230.89 μmol, 1.0 equiv.) in DCE (3 mL) was added 6-aminoindan-1-ol (68.89 mg, 461.78 μmol, 2.0 equiv.) and AcOH (525.00 mg, 8.74 mmol, 0.5 mL, 37.87 equiv.). The mixture was stirred at 50° C. for 1 h. The mixture was concentrated and the residue was purified by preparative HPLC (column: Phenomenex luna C18 100 * 40mm * 3 μm; mobile phase: [water (0.04% HCl)-ACN]; B%: 30%-60%, 7 min) to give 8-bromo-6-(2,6-dichlorophenyl)-2-[(3-hydroxyindan-5-yl)amino]pyrido[4,3-d]pyrimidin-5-one (90 mg, 166.17 μmol, yield 71.97%, purity 95.673%) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6)δ10.73-10.49(m,1H),9.18(d,J=1.0Hz,1H),8.43(s,1H),8.16-7.87(m,2H),7.76(d,J=8.3Hz,2H),7.67-7.58(m,1H),7.22(brd,J=8. 3Hz,1H), 5.24(brd,J=5.9Hz,1H),5.07(brd,J=5.9Hz,1H),2.98-2.83(m,1H),2.76-2.63(m,1H),2.42-2.32(m,1H),1.89-1.75(m,1H). ESI[M+H]=517.0 / 518.9.

[0252] Example 18: Preparation of compound P23 (i) Preparation of Compound 2

[0253] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-[(3-hydroxyindan-5-yl)amino]pyrido[4,3-d]pyrimidin-5-one (50 mg, 96.49 μmol, 1.0 equiv.) in THF (4 mL) was added DPPA (53.11 mg, 192.98 μmol, 2.0 equiv.), followed by DBU (44.07 mg, 289.47 μmol, 3.0 equiv.) in N 2 The mixture was added under atmospheric pressure at 0°C. 2 The mixture was stirred at 60° C. for 3 hours under atmospheric conditions. 2 O (15 mL) was added, extracted with ethyl acetate (10 mL x 3), and then diluted with Na 2 SO 4 The organic layer was filtered and concentrated to give 2-[(3-azidoindan-5-yl)amino]-8-bromo-6-(2,6-dichlorophenyl)pyrido[4,3-d]pyrimidin-5-one (60 mg, crude) as a yellow gum. ESI [M+H]=542.1 / 544.1.

[0254] (ii) Preparation of compound P23

[0255] [ka] A solution of 2-[(3-azidoindan-5-yl)amino]-8-bromo-6-(2,6-dichlorophenyl)pyrido[4,3-d]pyrimidin-5-one (30 mg, 55.23 μmol, 1.0 equiv.) in ACN (5 mL) was added with CeCl 3 .7H 2 O (41.15 mg, 110.46 μmol, 2.0 equiv.) and NaI (74.51 mg, 497.05 μmol, 9.0 equiv.) were added. The reaction mixture was stirred at 100° C. for 12 h and then concentrated. The residue was purified by preparative HPLC (column: mobile phase: [water (0.04% HCl)-ACN]; B%: 25%-45%, 7 min) to give 2-[(3-aminoindan-5-yl)amino]-8-bromo-6-(2,6-dichlorophenyl)pyrido[4,3-d]pyrimidin-5-one (11.65 mg, 20.72 μmol, 37.52% yield, 98.485% purity, HCl) as a dark yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ10.83-10.60(m,1H),9.19(s,1H),8.57(brs,3H),8.43(s,1H),8.03(s,1H),7.76(d,J=8.1Hz,2H),7.68-7.57(m, 1H),7.38-7.20(m,2H),4.71(brs,1H),3.16-3.02(m,1H),2.94-2.80(m,1H),2.50-2.44(m,1H),2.17-2.02(m,1H). ESI[M+H]=516 / 517.9.

[0256] Example 19: Preparation of compound P22

[0257] [ka] To a solution of 2-[(3-aminoindan-5-yl)amino]-8-bromo-6-(2,6-dichlorophenyl)pyrido[4,3-d]pyrimidin-5-one (290 mg, 560.71 μmol, 1.0 equiv.) in MeOH (5 mL) was added DIEA (144.93 mg, 1.12 mmol, 2.0 equiv.) and HCHO (136.52 mg, 1.68 mmol, 37% purity, 3.0 equiv.). The mixture was stirred at 20° C. for 0.5 h followed by addition of NaBH 3 CN (70.4 mg, 1.12 mmol, 2.0 equiv) was added. The mixture was stirred at 20° C. for 1 h. The mixture was concentrated and the residue was purified by preparative HPLC (column: Welch Xtimate C18 100 * 25mm * 3 μm; mobile phase: [water (0.04% HCl)-ACN]; B%: 20% to 40%, 8 min) to give 8-bromo-6-(2,6-dichlorophenyl)-2-[[3-(dimethylamino)indan-5-yl]amino]pyrido[4,3-d]pyrimidin-5-one (13.01 mg, 21.69 μmol, yield 3.87%, purity 96.970%, HCl) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ10.92-10.62(m,2H),9.20(s,1H),8.49-8.42(m,1H),8.06-7.85(m,1H),7.76(d,J=8.1Hz,2H),7.65-7.58(m,1H),7.41-7.32(m,1H), 7.21-7.10(m,1H),4.99-4.90(m,1H),3.18-3.06(m,1H),2.93-2.83(m,1H),2.75(brs,3H),2.63(brd,J=3.4Hz,3H),2.47-2.40(m,2H). ESI[M+H]=544.0 / 546.0.

[0258] Example 20: Preparation of compound P21 (i) Preparation of Compound 2

[0259] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (200 mg, 479.49 μmol, 1.0 equiv.) in DCM (5 mL) was added m-CPBA (146.02 mg, 719.24 μmol, 85% purity, 1.5 equiv.) at 0° C. The mixture was stirred at 20° C. for 1 h and then used in the next step without further purification. ESI [M+H]=431.8 / 433.8.

[0260] (ii) Preparation of compound P21

[0261] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (200 mg, 461.78 μmol, 1.0 equiv.) in DCE (5 mL) was added 4-(4-methylpiperazin-1-yl)aniline (176.65 mg, 923.56 μmol, 2.0 equiv.) and AcOH (138.65 mg, 2.31 mmol, 5.0 equiv.). The mixture was stirred at 50° C. for 1 h. The mixture was concentrated and the residue was purified by preparative HPLC (HCl conditions, column: Phenomenex Luna C18 75 * 30mm * 3 μm; mobile phase: [water (0.04% HCl)-ACN]; B%: 20% to 50%, 8 min) to give 8-bromo-6-(2,6-dichlorophenyl)-2-[4-(4-methylpiperazin-1-yl)anilino]pyrido[4,3-d]pyrimidin-5-one (194.04 mg, 323.22 μmol, 69.99% yield, 99.40% purity, HCl) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6)δ10.43-10.58(m,2H),9.13-9.14(m,1H),9.14(s,1H),8.42(s,1H),8.01(brs,2H),7.75(d,J=8.13Hz,2H),7.61(dd,J=7.69,8.69Hz,1H),7 .05(brd,J=8.75Hz,2H),3.81(brd,J=12.76Hz,2H),3.50(brd,J=11.63Hz,2H),3.12-3.22(m,2H),3.01-3.10(m,2H),2.83(d,J=4.50Hz,3H). ESI[M+H]=559.0 / 560.9.

[0262] Example 21: Preparation of compound P19 (i) Preparation of Compound 2

[0263] [ka] To a solution of NaH (913.33 mg, 22.83 mmol, 60% purity, 2.0 equiv) in DMF (30 mL) was added methyl 2-(2-cyanophenyl)acetate (2 g, 11.42 mmol, 1.0 equiv) dropwise at 0° C. The mixture was stirred at 0° C. for 0.5 h, and then 1,2-methoxybenzylamine (4.29 g, 22.83 mmol, 2.0 equiv) was added dropwise at 0° C. The resulting mixture was stirred at 20° C. for 0.5 h, and then saturated NH 4 The reaction mixture was quenched with aqueous Cl (80 mL). The reaction mixture was concentrated to remove the organic phase and then diluted with H 2 The mixture was diluted with 200 mL of ethyl acetate (30 mL), extracted with ethyl acetate (40 mL x 3), and then washed with brine (30 mL). The combined organic layers were washed with Na 2 SO 4 Drying at 40° C., filtering and concentrating gave methyl 1-(2-cyanophenyl)cyclopropanecarboxylate (2.1 g, crude) as a dark yellow oil. 1 H NMR (400MHz, DMSO-d 6)δ7.79-7.90(m,1H),7.62-7.76(m,1H),7.45-7.57(m,2H),3.54-3.64(m,3H),1.57-1.71(m,2H),1.27-1.43(m,2H). ESI[M+H]=202.1.

[0264] (ii) Preparation of Compound 3

[0265] [ka] To a solution of methyl 1-(2-cyanophenyl)cyclopropanecarboxylate (2 g, 9.94 mmol, 1.0 equiv) in MeOH (60 mL) / TEA (6 mL) / HO (7 mL) was added H 2 Raney Ni (2.4 g, 9.94 mmol, 1.0 equiv.) was added under atmospheric pressure. The suspension was degassed and diluted with H 2 The mixture was purged with H 2 The mixture was stirred under (15 Psi) at 20° C. for 12 hours. The reaction mixture was filtered and the filtrate was concentrated to give spiro[1,2-dihydroisoquinoline-4,1′-cyclopropane]-3-one (1.6 g, crude product) as a yellow solid. ESI [M+H]=174.2.

[0266] (iii) Preparation of Compound 4

[0267] [ka] H 2 SO 4 To a solution of spiro[1,2-dihydroisoquinolin-4,1'-cyclopropan]-3-one (600 mg, 3.46 mmol, 1.0 equiv) in 10 mL of KNO 3 (420.25 mg, 4.16 mmol, 1.2 equiv) was added at 0° C. The mixture was stirred at 0° C. for 15 min, then at 20° C. for 45 min. The reaction solution was poured into ice water (20 mL) and filtered to give 7-nitrospiro[1,2-dihydroisoquinolin-4,1′-cyclopropane]-3-one (670 mg, crude) as a pale yellow solid. ESI[M+H]=219.1.

[0268] (iv) Preparation of Compound 5

[0269] [ka] 7-Nitrospiro[1,2-dihydroisoquinoline-4,1'-cyclopropane]-3-one (670 mg, 3.07 mmol, 1 equiv.) was treated with BH 3 THF (1M, 20 mL) was added at 0° C. The reaction mixture was stirred at 50° C. for 12 h. The reaction mixture was quenched with 4N HCl (5 mL) at 0° C. and stirred at 50° C. for 1 h. The reaction mixture was diluted with saturated Na 2 CO 3 Adjustment to pH 8 with aqueous solution gave 7-nitrospiro[2,3-dihydro-1H-isoquinoline-4,1′-cyclopropane] (600 mg, crude) as a yellow oil. ESI [M+H]=205.2.

[0270] (v) Preparation of compound 6

[0271] [ka] 7-Nitrospiro[2,3-dihydro-1H-isoquinoline-4,1'-cyclopropane] (600 mg, 2.94 mmol, 1.0 equiv.) in THF (8 mL) and H 2 To the solution of 2O (8 mL) was added tert-butoxycarbonyl tert-butyl carbonate (769.43 mg, 3.53 mmol, 1.2 equiv). The mixture was stirred at 20° C. for 1 h. The reaction mixture was poured into water (10 mL) and extracted with ethyl acetate (20 mL×3). The combined organic layers were extracted with Na 2 SO 4 The residue was purified by silica gel column chromatography (SiO 2 , petroleum ether:ethyl acetate=1:0-5:1) to give tert-butyl 7-nitrospiro[1,3-dihydroisoquinoline-4,1'-cyclopropane]-2-carboxylate as a yellow solid. ESI[M+H-tBu]=249.1.

[0272] (vi) Preparation of compound 7

[0273] [ka] tert-Butyl 7-nitrospiro[1,3-dihydroisoquinoline-4,1'-cyclopropane]-2-carboxylate (470 mg, 1.54 mmol, 1.0 equiv) in THF (3 mL), EtOH (3 mL) and H 2 To a solution of Fe (431.25 mg, 7.72 mmol, 5.0 equiv) and NH 4 Cl (413.03 mg, 7.72 mmol, 5.0 equiv) was added. The mixture was stirred at 80° C. for 1 h. The mixture was filtered and concentrated to give tert-butyl 7-aminospiro[1,3-dihydroisoquinoline-4,1′-cyclopropane]-2-carboxylate (400 mg, crude) as a yellow gum. 1 H NMR (400MHz, DMSO-d 6 )δ7.40(brs,1H),7.28(brs,1H),7.15(brs,1H),6.35-6.52(m,2H),4.46(brs,2H),3.35-3.39(m,2H),1.40(brs,9H),0.79(s,4H). ESI[M+H-tBu]=219.3.

[0274] (vii) Preparation of compound 9

[0275] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (50 mg, 119.87 μmol, 1.0 equiv.) in DCM (2 mL) was added m-CPBA (36.50 mg, 179.81 μmol, 85% purity, 1.5 equiv.) at 0° C. The mixture was stirred at 20° C. for 1 h and then used in the next step without further purification. ESI [M+H]=432.0 / 434.0.

[0276] (viii) Preparation of compound 10

[0277] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (50 mg, 115.44 μmol, 1.0 equiv) in DCE (5 mL) was added tert-butyl 7-aminospiro[1,3-dihydroisoquinoline-4,1′-cyclopropane]-2-carboxylate (63.35 mg, 230.89 μmol, 2.0 equiv) and AcOH (41.60 mg, 692.67 μmol, 6.0 equiv). The mixture was stirred at 50° C. for 1 h. The mixture was concentrated and diluted with Na 2 CO 3 (3 mL) and then extracted with ethyl acetate (5 mL x 3). The combined organic layers were washed with Na 2 SO 4 The residue was purified by preparative TLC ((SiO 2 , petroleum ether:ethyl acetate=5:1) to give tert-butyl 7-[[8-bromo-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]spiro[1,3-dihydroisoquinoline-4,1'-cyclopropane]-2-carboxylate (50 mg, 77.72 μmol, 67.32% yield) as a yellow oil. ESI[M+H]=641.9 / 643.9.

[0278] (ix) Preparation of compound P19

[0279] [ka] A mixture of tert-butyl 7-[[8-bromo-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]spiro[1,3-dihydroisoquinoline-4,1'-cyclopropane]-2-carboxylate (20 mg, 31.09 μmol, 1.0 equiv.) in TFA / DCM (2 mL, v / v=3 / 1) was stirred for 1 h at 20° C. The mixture was concentrated and purified by preparative HPLC (column: Phenomenex Luna C18 150 * 30mm * 5 μm; mobile phase: [water (0.1% TFA)-ACN]; B%: 5% to 45%, 8 min) to give 8-bromo-6-(2,6-dichlorophenyl)-2-(spiro[2,3-dihydro-1H-isoquinoline-4,1'-cyclopropane]-7-ylamino)pyrido[4,3-d]pyrimidin-5-one (8.59 mg, 13.07 μmol, 42.95% yield, 98.795% purity, TFA) as a pale yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ10.69(brs,1H),9.15-9.29(m,3H),8.46(s,1H),8.14(brs,1H),7.75(d,J=8.13Hz,3H),7 .55-7.66(m,1H),6.89(d,J=8.63Hz,1H),4.43(brs,2H),3.28(brs,2H),1.06-1.13(m,4H). ESI[M+H]=541.9 / 544.

[0280] Example 22: Preparation of compound P20

[0281] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-(spiro[2,3-dihydro-1H-isoquinoline-4,1'-cyclopropane]-7-ylamino)pyrido[4,3-d]pyrimidin-5-one (20 mg, 30.43 μmol, 1.0 equiv., TFA) in MeOH (2 mL) was added HCHO (7.41 mg, 91.29 μmol, 37% purity, 3.0 equiv.) and DIEA (3.93 mg, 30.43 μmol, 1.0 equiv.). The mixture was stirred at 20° C. for 0.5 h followed by addition of NaBH 3 CN (4 mg, 60.86 μmol, 2.0 equiv.) was added. The mixture was stirred at 20° C. for 1 h. The reaction mixture was diluted with saturated Na 2 CO 3 The mixture was diluted with aqueous solution (3 mL) and extracted with DCM (2 mL×3). The combined organic layers were washed with Na 2 SO 4 The residue was purified by preparative HPLC (column: Phenomenex Luna C18 150 * 30mm * 5 μm; mobile phase: [water (0.1% TFA)-ACN]; B%: 10%-50%, 8 min) to give 8-bromo-6-(2,6-dichlorophenyl)-2-[(2-methylspiro[1,3-dihydroisoquinoline-4,1'-cyclopropane]-7-yl)amino]pyrido[4,3-d]pyrimidin-5-one (4.42 mg, 6.58 μmol, yield 21.64%, purity 94.528%, TFA) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ10.72(brs,1H),10.24(brs,1H),9.19(s,1H),8.46(s,1H),8.22(brs,1H),7.75(d,J=8.13Hz,3H),7.57-7.66(m,1H) ),6.90(d,J=8.75Hz,1H),4.39-4.66(m,2H),3.23(brd,J=12.13Hz,2H),2.96(brd,J=2.63Hz,3H),0.96-1.40(m,4H). ESI[M+H]=555.9 / 558.0.

[0282] Example 23: Preparation of compound P25 (i) Preparation of Compound 2

[0283] [ka] 4-Nitro-1H-pyrazole (10 g, 88.44 mmol, 1.0 equiv.), tert-butyl 4-hydroxypiperidine-1-carboxylate (17.80 g, 88.44 mmol, 1.0 equiv.) and PPh 3 (34.79 g, 132.66 mmol, 1.5 equiv.) in THF (400 mL) was treated with DIAD (26.82 g, 132.66 mmol, 1.5 equiv.) 2 The mixture was added dropwise at −60° C. The mixture was warmed to 20° C. and stirred for 16 h. The mixture was then diluted with EtOAc (200 mL), washed with water (200 mL) / brine (200 mL), dried, filtered and concentrated. The residue was purified by silica gel chromatography (PE:EA=10:1 to 1:1) to give tert-butyl 4-(4-dichloropyridazin-1-yl)-3-(1-hydroxyethyl)piperazine-1-carboxylate (24 g, 80.99 mmol, 91.58% yield) as a yellow solid. LCMS: ESI[M+H-Boc]=197.3

[0284] (ii) Preparation of Compound 3

[0285] [ka] A mixture of tert-butyl 4-(4-nitropyrazol-1-yl)piperidine-1-carboxylate (8 g, 27.00 mmol, 1 equiv.), Pd / C (2 g, 10% purity) in EtOAc (80 mL) was degassed and diluted with H 2 The mixture was then purged three times with H 2 The mixture was stirred under atmospheric pressure (15 Psi) at 20° C. for 2 hours. The reaction mixture was filtered and concentrated. The residue was purified by column chromatography (SiO 2, petroleum ether / ethyl acetate=10 / 1 to 0 / 1) to give tert-butyl 4-(4-aminopyrazol-1-yl)piperidine-1-carboxylate (4 g, 14.40 mmol, 53.34% yield, 95.88% purity) as a red oil. LCMS: ESI[M+H-tBu]=211.3.

[0286] (iii) Preparation of Compound 5

[0287] [ka] A mixture of ethyl 3-oxobutanoate (61.80 g, 474.87 mmol, 1.0 equiv) and 1,1-dimethoxy-N,N-dimethyl-methanamine (68.17 g, 572.10 mmol, 1.20 equiv) was heated to 80° C. for 10 min. The mixture was cooled to 20° C., followed by the addition of 2-methylisothiourea, sulfuric acid (66.10 g, 237.43 mmol, 0.5 equiv). The mixture was heated at 85° C. for 12 h, then concentrated and the residue was dissolved in EtOAc (800 mL), water (800 mL), saturated NaHCO 3 Washing with aqueous solution (400 mL), brine (400 mL), drying and concentration gave 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylate (76.8 g, crude) as a yellow oil. ESI [M+H]=213.2.

[0288] (iv) Preparation of Compound 6

[0289] [ka] LiOH.H 2 HO (27.72 g, 660.63 mmol, 1.85 equiv.) 2The 278 mL of 0 solution was added to a solution of 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylate (75.8 g, 357.10 mmol, 1.0 equiv) in THF (278 mL) / MeOH (92 mL). The mixture was stirred at 20° C. for 1 h and then concentrated to remove the solvent. The remaining aqueous layer was extracted with methyl tert-butyl ether (500 mL) and then acidified to pH 2 with 1N HCl. The precipitate was filtered and the filter cake was dried to give 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylic acid (38 g, 201.66 mmol, 56.47% yield, 97.76% purity) as a yellow solid. ESI [M+H]=185.1.

[0290] (v) Preparation of compound 7

[0291] [ka] A mixture of 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylic acid (3 g, 16.29 mmol, 1.0 equiv.) and 2-fluoro-6-methyl-aniline (2.04 g, 16.29 mmol, 1.0 equiv.) in chlorobenzene (45 mL) was degassed and purified with N 2 3 times at 20 °C, followed by purging with PCl 3 (2.24 g, 16.29 mmol, 1.0 equiv.) was added. The mixture was then diluted with N 2 The mixture was stirred at 135° C. for 12 hours under atmospheric pressure. The mixture was concentrated, and the residue was washed with saturated Na 2 CO 3 Poured into aqueous solution (100 mL) and extracted with 2-methyl-THF (50 mL x 3). The organic phase was dried and concentrated. The residue was triturated with EtOAc (6 mL) to give N-(2-fluoro-6-methyl-phenyl)-4-methyl-2-methylsulfanyl-pyrimidine-5-carboxamie (2.3 g, 6.23 mmol, 38.23% yield, 78.87% pure) as a yellow solid. ESI [M+H] = 292.2.

[0292] (vi) Preparation of compound 8

[0293] [ka] To a solution of N-(2-fluoro-6-methyl-phenyl)-4-methyl-2-methylsulfanyl-pyrimidine-5-carboxamide (2.3 g, 7.89 mmol, 1.0 equiv.) in DMF (57 mL) was added DMF-DMA (5.64 g, 47.37 mmol, 6.29 mL, 6 equiv.). The mixture was stirred at 80° C. for 12 h. The mixture was concentrated, purified, and triturated with EtOH (8 mL) to give 6-(2-fluoro-6-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (2.13 g, 6.35 mmol, 80.42% yield, 89.82% purity) as a white solid. ESI [M+H]=302.3.

[0294] (vii) Preparation of compound 9

[0295] [ka] CH 3 To a solution of 6-(2-fluoro-6-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (2.13 g, 7.07 mmol, 1.0 equiv.) in CN (60 mL) was added NBS (1.89 g, 10.60 mmol, 1.5 equiv.). The mixture was stirred at 80° C. for 12 h. The mixture was concentrated and the crude product was triturated with EtOH (10 mL) to give 8-bromo-6-(2-fluoro-6-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (2.3 g, 5.51 mmol, 78.02% yield, 91.17% purity) as a yellow solid. ESI [M+H]=380.2 / 382.1.

[0296] (viii) Preparation of compound 10

[0297] [ka] To a solution of 8-bromo-6-(2-fluoro-6-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (1 g, 2.63 mmol, 1.0 equiv.) in DCM (43 mL) was added m-CPBA (694.11 mg, 3.42 mmol, 85% purity, 1.3 equiv.) at 0° C. The mixture was then stirred at 20° C. for 1 h. The mixture was used in the next step without further purification. ESI [M+H]=398.1 / 396.1.

[0298] (ix) Preparation of compound 11

[0299] [ka] To a solution of 8-bromo-6-(2-fluoro-6-methyl-phenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (1 g, 2.52 mmol, 1.0 equiv.) in DCE (60 mL) was added tert-butyl 4-(4-aminopyrazol-1-yl)piperidine-1-carboxylate (1.01 g, 3.79 mmol, 1.5 equiv.). Then AcOH (909.34 mg, 15.14 mmol, 6 equiv.) was added. The mixture was stirred at 50° C. for 1 h. The mixture was concentrated and the residue was purified by preparative HPLC (column: Kromasil C18 (250 * 50mm * Mobile phase: [Water (10mM NH 4 HCO 3 )-ACN]; B%: 45%-65%, 10 min) to give tert-butyl 4-[4-[[8-bromo-6-(2-fluoro-6-methyl-phenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (0.7 g, 1.17 mmol, 46.35% yield) as a yellow solid. ESI [M+H]=542.2 / 544.2.

[0300] (x) Preparation of compound P25

[0301] [ka] To a solution of tert-butyl 4-[4-[[8-bromo-6-(2-fluoro-6-methyl-phenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (150 mg, 250.64 μmol, 1.0 equiv.) in DCM (6 mL) was added TFA (2 mL). The mixture was stirred at 20° C. for 1 h. The mixture was concentrated and purified by preparative HPLC (column: Phenomenex Luna C18 150 * 30mm * 5 μm; mobile phase: [water (0.1% TFA)-ACN]; B%: 10%-40%, 9 min) to give 8-bromo-6-(2-fluoro-6-methyl-phenyl)-2-[[1-(4-piperzyl)pyrazol-4-yl]amino]pyrido[4.3-d]pyrimidin-5-one (34.37 mg, 68.97 μmol, 84.47% yield, 100% purity, TFA) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ10.69(s,1H),9.10(s,1H),8.72(brs,1H),8.27-8.49(m,2H),7.66-8.04(m,1H),7.47(dt,J=5.82,7.97Hz,1H ),7.21-7.35(m,2H),4.46-4.62(m,1H),3.44(brd,J=12.76Hz,2H),3.11(q,J=11.30Hz,2H),1.98-2.30(m,7H). ESI[M+H]=498.1 / 500.1.

[0302] Example 24: Preparation of compound P26

[0303] [ka] To a solution of 8-bromo-6-(2-fluoro-6-methyl-phenyl)-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (100 mg, 163.30 μmol, 1 equiv., TFA) in MeOH (4 mL) was added DIEA (21.11 mg, 163.30 μmol, 1.0 equiv.) and HCHO (39.76 mg, 489.90 μmol, 37% purity, 3.0 equiv.). The mixture was stirred at 20° C. for 0.5 h. Then, NaBH 3 CN (10.26 mg, 163.30 μmol, 1.0 equiv.) was added. The mixture was stirred at 20° C. for 1 h. The mixture was concentrated and the residue was purified by preparative HPLC (column: Phenomenex Luna C18 150 * 30mm * 5 μm; mobile phase: [water (0.1% TFA)-ACN]; B%: 12% to 42%, 9 min) to give 8-bromo-6-(2-fluoro-6-methyl-phenyl)-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (56.6 mg, 90.36 μmol, yield 55.33%, purity 100%, TFA) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ10.46-10.80(m,1H),9.57-9.95(m,1H),9.10(s,1H),8.27-8.50(m,2H),7.66-8.09(m,1H),7.40-7.55(m,1H),7. 21-7.35(m,2H),4.41-4.63(m,1H),3.59(brd,J=11.63Hz,2H),3.10-3.25(m,2H),2.83(brs,3H),2.06-2.36(m,7H). ESI[M+H]=512.1 / 514.1.

[0304] Example 25: Preparation of compound P14 (i) Preparation of Compound 2

[0305] [ka] To a solution of 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylic acid (3.14 g, 17.04 mmol, 1.0 equiv.) and 2,6-dichloro-3-methylaniline (3 g, 17.04 mmol, 1.0 equiv.) in chlorobenzene (30 mL) was added PCl 3 (2.34 g, 17.04 mmol, 1.0 equiv) was added. The mixture was stirred at 135° C. for 12 h. The mixture was concentrated and the residue was washed with saturated Na 2 CO 3 Poured into aqueous solution (100 mL) and extracted with 2-methyl-THF (50 mL x 3). The organic phase was dried and concentrated. The residue was triturated with EtOH (6 mL) to give N-(2,6-dichloro-3-methyl-phenyl)-4-methyl-2-methylsulfanyl-pyrimidine-5-carboxamide (2.3 g, 6.72 mmol, 39.44% yield) as a yellow solid. ESI [M+H] = 342.1 / 344.1.

[0306] (ii) Preparation of Compound 3

[0307] [ka] To a solution of N-(2,6-dichloro-3-methyl-phenyl)-4-methyl-2-methylsulfanyl-pyrimidine-5-carboxamide (2.1 g, 6.14 mmol, 1.0 equiv.) in DMF (50 mL) was added DMF-DMA (4.39 g, 36.82 mmol, 6.0 equiv.). The mixture was stirred at 80° C. for 12 h. The mixture was concentrated and triturated with EtOH (6 mL) to give 6-(2,6-dichloro-3-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (1.3 g, 3.69 mmol, 60.15% yield) as a yellow solid. ESI [M+H]=352.0 / 354.0.

[0308] (iii) Preparation of Compound 4

[0309] [ka] CH 3A mixture of 6-(2,6-dichloro-3-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (1.2 g, 3.41 mmol, 1.0 equiv.) and NBS (909.51 mg, 5.11 mmol, 1.5 equiv.) in CN (12 mL) was degassed and cooled with N 2 Purge the mixture three times with N 2 The mixture was stirred at 80° C. under atmospheric pressure for 12 h. The mixture was concentrated and then triturated with EtOH (6 mL) to give 8-bromo-6-(2,6-dichloro-3-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (1.2 g, 2.78 mmol, 81.70% yield) as a light yellow solid. ESI [M+H]=431.8 / 429.8.

[0310] (iv) Preparation of Compound 5

[0311] [ka] To a solution of 8-bromo-6-(2,6-dichloro-3-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (100 mg, 231.95 μmol, 1.0 equiv.) in DCM (2 mL) was added m-CPBA (70.63 mg, 347.92 μmol, 85% purity, 1.5 equiv.) at 0° C. The mixture was stirred at 20° C. for 1 h. The mixture was concentrated to give 8-bromo-6-(2,6-dichloro-3-methyl-phenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (100 mg, crude product) as a yellow solid, which was used in the next step without further purification. ESI [M+H]=447.9 / 445.9.

[0312] (v) Preparation of compound 6

[0313] [ka] To a solution of 8-bromo-6-(2,6-dichloro-3-methyl-phenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (100 mg, 223.65 μmol, 1.0 equiv.) in DCE (5 mL) was added tert-butyl 4-(4-aminopyrazol-1-yl)piperidine-1-carboxylate (119.13 mg, 447.29 μmol, 2.0 equiv.) and AcOH (40.29 mg, 670.94 μmol, 38.37 μL, 3.0 equiv.). The mixture was stirred at 50° C. for 1 h. The mixture was concentrated and analyzed by preparative TLC (SiO 2 , petroleum ether:ethyl acetate=1:1) to give tert-butyl 4-[4-[[8-bromo-6-(2,6-dichloro-3-methyl-phenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate acid (130 mg, 200.20 μmol, 89.51% yield) as a yellow oil. ESI[M+H]=650.1 / 648.1.

[0314] (vi) Preparation of compound P14

[0315] [ka] To a solution of tert-butyl 4-[4-[[8-bromo-6-(2,6-dichloro-3-methyl-phenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (130 mg, 200.20 μmol, 1.0 equiv.) in DCM (3 mL) was added TFA (1 mL). The mixture was stirred at 20° C. for 1 h. The mixture was concentrated and the residue was purified by preparative HPLC (column: Phenomenex Luna C18 150 * 30mm *5 μm; mobile phase: [water (TFA)-ACN]; B%: 5%-45%, 8 min) to give 8-bromo-6-(2,6-dichloro-3-methyl-phenyl)-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (21.65 mg, 32.64 μmol, yield: 43.30%, purity: TFA) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ10.74(s,1H),9.10(s,1H),8.74(brs,1H),8.46(s,1H),8.40(s,1H),7.81(s,1H),7.57-7.68(m,2H),4.47-4. 60(m,1H),3.42(brs,2H),3.11(brd,J=10.01Hz,2H),2.42(s,3H),2.24(brd,J=10.76Hz,2H),2.03-2.15(m,2H). ESI[M+H]=549.9 / 548.0.

[0316] Example 26: Preparation of compound P15

[0317] [ka] To a solution of 8-bromo-6-(2,6-dichloro-3-methyl-phenyl)-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (50 mg, 75.38 μmol, 1.0 equiv., TFA) in MeOH (2 mL) was added HCHO (18.35 mg, 226.15 μmol, 37% purity, 3.0 equiv.) and DIEA (9.74 mg, 75.38 μmol, 13.13 μL, 1.0 equiv.). The mixture was stirred at 20° C. for 0.5 h followed by addition of NaBH 3 CN (4.74 mg, 75.38 μmol, 1.0 equiv) was added. The reaction was stirred at 20° C. for 1 h. The reaction was diluted with saturated Na 2 CO 3 The mixture was quenched with aqueous solution (3 mL) and extracted with DCM (1 mL x 3). The combined organic layers were washed with Na 2 SO 4The residue was purified by preparative HPLC (column: Phenomenex luna C18 100 * 40mm * 5 μm; mobile phase: [water (TFA)-ACN]; B%: 15%-60%, 8 min) to give 8-bromo-6-(2,6-dichloro-3-methyl-phenyl)-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (9.28 mg, 13.70 μmol, 18.18% yield, 100% purity, TFA) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ10.52-10.82(m,1H),9.61(brs,1H),9.07-9.21(m,1H),8.31-8.48(m,2H),7.82(s,1H),7.54-7.71(m,2H),4.49(brs, 1H),3.59(brd,J=11.38Hz,2H),3.17(brs,2H),2.83(s,3H),2.42(s,3H),2.31(brd,J=13.01Hz,2H),2.05-2.20(m,2H). ESI[M+H]=563.9 / 562.0.

[0318] Example 27: Preparation of compound P16 (i) Preparation of Compound 2

[0319] [ka] To a solution of 4-methyl-2-methylsulfanyl-pyrimidine-5-carboxylic acid (3.14 g, 17.04 mmol, 1.0 equiv.) and 2,6-dichloro-4-methylaniline (3 g, 17.04 mmol, 1.0 equiv.) in chlorobenzene (30 mL) was added PCl 3 (2.34 g, 17.04 mmol, 1.0 equiv) was added. The mixture was stirred at 135° C. for 12 h. The mixture was concentrated and the residue was washed with saturated Na 2 CO 3Poured into aqueous solution (100 mL) and extracted with 2-methyl-THF (50 mL x 3). The organic phase was dried and concentrated. The residue was triturated with EtOH (6 mL) to give N-(2,6-dichloro-4-methyl-phenyl)-4-methyl-2-methylsulfanyl-pyrimidine-5-carboxamide (2.5 g, 7.30 mmol, 42.86% yield) as a yellow solid. ESI [M+H] = 342.0 / 344.0.

[0320] (ii) Preparation of Compound 3

[0321] [ka] To a solution of N-(2,6-dichloro-4-methyl-phenyl)-4-methyl-2-methylsulfanyl-pyrimidine-5-carboxamide (2.24 g, 6.55 mmol, 1.0 equiv.) in DMF (50 mL) was added DMF-DMA (4.68 g, 39.27 mmol, 5.22 mL, 6.0 equiv.). The mixture was stirred at 80° C. for 12 h. The mixture was concentrated and triturated with EtOH (6 mL) to give 6-(2,6-dichloro-4-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (1.8 g, 5.11 mmol, 78.08% yield) as a pale yellow solid. ESI [M+H]=352.0 / 354.0.

[0322] (iii) Preparation of Compound 4

[0323] [ka] CH 3 A mixture of 6-(2,6-dichloro-4-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (1.7 g, 4.83 mmol, 1.0 equiv.) and NBS (1.29 g, 7.24 mmol, 1.5 equiv.) in CN (20 mL) was degassed and cooled with N 2 Purge the mixture three times with N 2The mixture was stirred at 80° C. under atmospheric pressure for 12 h. The mixture was concentrated and triturated with EtOH (6 mL) to give 8-bromo-6-(2,6-dichloro-4-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (2 g, 4.64 mmol, 96.12% yield) as a pale yellow solid. ESI [M+H]=431.9 / 429.9.

[0324] (iv) Preparation of Compound 5

[0325] [ka] To a solution of 8-bromo-6-(2,6-dichloro-4-methyl-phenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (100 mg, 231.95 μmol, 1.0 equiv.) in DCM (2 mL) was added m-CPBA (70.63 mg, 347.92 μmol, 85% purity, 1.5 equiv.) at 0° C. The mixture was stirred at 20° C. for 1 h. The mixture was concentrated to give 8-bromo-6-(2,6-dichloro-4-methyl-phenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (100 mg, crude product) as a yellow oil, which was used in the next step without further purification. ESI [M+H]=447.9 / 445.9.

[0326] (v) Preparation of compound 6

[0327] [ka] To a solution of 8-bromo-6-(2,6-dichloro-4-methyl-phenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (100 mg, 223.65 μmol, 1.0 equiv) in DCE (5 mL) was added tert-butyl 4-(4-aminopyrazol-1-yl)piperidine-1-carboxylate (119.13 mg, 447.29 μmol, 2.0 equiv) and AcOH (80.58 mg, 1.34 mmol, 76.75 μL, 6.0 equiv). The mixture was stirred at 50° C. for 1 h. The mixture was concentrated and analyzed by preparative TLC (SiO2 , petroleum ether:ethyl acetate=1:1) to give tert-butyl 4-[4-[[8-bromo-6-(2,6-dichloro-4-methyl-phenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate acid (120 mg, crude) as a yellow oil. ESI[M+H]=649.9 / 647.9.

[0328] (vi) Preparation of compound P16

[0329] [ka] To a solution of tert-butyl 4-[4-[[8-bromo-6-(2,6-dichloro-4-methyl-phenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (120 mg, 184.80 μmol, 1 equiv.) in DCM (4 mL) was added TFA (1 mL). The mixture was stirred at 20° C. for 1 h. The mixture was concentrated and purified by preparative HPLC (column: Phenomenex Luna 80 * 30mm * 3um; mobile phase: [water (TFA)-ACN]; B%: 15%-45%, 8 min) to give 8-bromo-6-(2,6-dichloro-4-methyl-phenyl)-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (27.90 mg, 42.06 μmol, yield 55.80%, purity 100%, TFA) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ10.73(s,1H),9.10(s,1H),8.78(brs,1H),8.45(s,2H),8.40(s,1H),7.81(s,1H),7.58(s,2H),4.46-4.64(m,1H) ),3.43(brd,J=12.76Hz,2H),3.11(brd,J=7.63Hz,2H),2.41(s,3H),2.24(brd,J=11.01Hz,2H),2.02-2.17(m,2H). ESI[M+H]=549.9 / 548.0.

[0330] Example 28: Preparation of compound P17

[0331] [ka] To a solution of 8-bromo-6-(2,6-dichloro-4-methyl-phenyl)-2-[[1-(4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (60 mg, 90.46 μmol, 1.0 equiv., TFA) in MeOH (2 mL) was added HCHO (22.03 mg, 271.38 μmol, 37% purity, 3.0 equiv.) and DIEA (11.69 mg, 90.46 μmol, 15.76 μL, 1 equiv.). The mixture was stirred at 20° C. for 0.5 h followed by addition of NaBH 3 CN (5.68 mg, 90.46 μmol, 1 equiv.) was added. The mixture was stirred at 20° C. for 1 h. The mixture was diluted with saturated Na 2 CO 3 The mixture was quenched with aqueous solution (3 mL) and extracted with DCM (1 mL x 3). The combined organic layers were washed with Na 2 SO 4 The residue was purified by preparative HPLC (column: Phenomenex Luna 80 * 30mm * 3um; mobile phase: [water (TFA)-ACN]; B%: 15% to 45%, 8 min) to give 8-bromo-6-(2,6-dichloro-4-methyl-phenyl)-2-[[1-(1-methyl-4-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (47.24 mg, 69.75 μmol, 77.10% yield, 100% purity, TFA) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6)δ10.51-10.83(m,1H),9.74-10.03(m,1H),9.10(s,1H),8.29-8.50(m,2H),7.83(s,1H),7.58(s,2H),4.43-4.62(m,1 H),3.59(brd,J=11.51Hz,2H),3.18(brs,2H),2.83(s,3H),2.41(s,3H),2.31(brd,J=12.88Hz,2H),2.06-2.24(m,2H). ESI[M+H]=563.9 / 561.9.

[0332] Example 29: Preparation of compound P13 (i) Preparation of Compound 2

[0333] [ka] 4-Nitro-1H-pyrazole (500 mg, 4.42 mmol, 1.0 equiv), tert-butyl 3-hydroxypiperidine-1-carboxylate (889.95 mg, 4.42 mmol, 1.0 equiv) and PPh in THF (30 mL). 3 A mixture of (1.74 g, 6.63 mmol, 1.5 equiv.) was degassed and diluted with N 2 The mixture was purged three times with 500 cc of 1000 cc, then DIAD (1.34 g, 6.63 mmol, 1.5 equiv) was added dropwise at -60 °C. The mixture was stirred at 20 °C for 12 h. The mixture was concentrated and the residue was purified by preparative HPLC (neutral conditions, column: Waters Xbridge Prep OBD C18 150 * 40mm * 10μm; Mobile phase: [water (NH 4 HCO 3 )-ACN]; B%: 30%-60%, 8 min) to give tert-butyl 3-(4-nitropyrazol-1-yl)piperidine-1-carboxylate (120 mg, 404.97 μmol, 9.16% yield) as a colorless oil. ESI [M+Ht]=241.1.

[0334] (ii) Preparation of Compound 3

[0335] [ka] A mixture of tert-butyl 3-(4-nitropyrazol-1-yl)piperidine-1-carboxylate (110 mg, 371.22 μmol, 1 equiv.), Pd / C (110 mg, 10% purity) in EtOAc (10 mL) was degassed and purified with H 2 (15 Psi) three times, and then the mixture was 2 The mixture was stirred at 30° C. under (15 Psi) for 1 h. The reaction mixture was filtered and the filtrate was concentrated to give tert-butyl 3-(4-aminopyrazol-1-yl)piperidine-1-carboxylate (80 mg, crude) as a red oil, which was used in the next step without further purification. ESI [M+H]=267.2.

[0336] (iii) Preparation of Compound 5

[0337] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfanyl-pyrido[4,3-d]pyrimidin-5-one (60 mg, 143.85 μmol, 1.0 equiv.) in DCM (2 mL) was added m-CPBA (43.81 mg, 215.77 μmol, 85% purity, 1.5 equiv.) at 0° C. The mixture was stirred at 20° C. for 1 h and used in the next step without further purification. ESI [M+H]=433.9 / 431.9.

[0338] (iv) Preparation of Compound 6

[0339] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-methylsulfinyl-pyrido[4,3-d]pyrimidin-5-one (62 mg, 143.15 μmol, 1.0 equiv) in DCE (2 mL) was added tert-butyl 3-(4-aminopyrazol-1-yl)piperidine-1-carboxylate (57.19 mg, 214.73 μmol, 1.5 equiv). Then AcOH (42.98 mg, 715.76 μmol, 40.93 μL, 5 equiv) was added. The mixture was stirred at 50° C. for 1 h. The reaction mixture was concentrated and analyzed by preparative TLC (SiO 2 , petroleum ether:ethyl acetate=1:1) to give tert-butyl 3-[4-[[8-bromo-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (90 mg, 141.66 μmol, 98.96% yield) as a yellow oil. ESI [M+H-tBu]=580.1 / 578.1.

[0340] (v) Preparation of compound P13

[0341] [ka] To a solution of tert-butyl 3-[4-[[8-bromo-6-(2,6-dichlorophenyl)-5-oxo-pyrido[4,3-d]pyrimidin-2-yl]amino]pyrazol-1-yl]piperidine-1-carboxylate (90 mg, 141.66 μmol, 1.0 equiv.) in DCM (1.5 mL) was added TFA (0.5 mL). The mixture was stirred at 25° C. for 1 h. The reaction mixture was concentrated and the residue was purified by preparative HPLC (TFA conditions, column: Phenomenex Luna 80 * 30mm * 3 μm; mobile phase: [water (TFA)-ACN]; B%: 10% to 40%, 8 min) to give 8-bromo-6-(2,6-dichlorophenyl)-2-[[1-(3-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (18.99 mg, 29.25 μmol, 54.26% yield, 100% purity, TFA) as a yellow gum.1 H NMR (400MHz, DMSO-d 6 )δ10.78(s,1H),9.13(s,1H),8.87(brs,1H),8.46(s,1H),8.37-8.43(m,1H),7.95(s,1H),7.75(d,J=8.13Hz,2H),7.59-7.65(m,1H),4 .53-4.63(m,1H),3.60(brd,J=8.76Hz,1H),3.24-3.33(m,2H),3.02(brt,J=9.76Hz,1H),2.22(brd,J=8.88Hz,1H),1.73-2.10(m,4H). ESI[M+H]=535.9 / 533.9.

[0342] Example 30: Preparation of compound P12

[0343] [ka] To a solution of 8-bromo-6-(2,6-dichlorophenyl)-2-[[1-(3-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (35 mg, 65.39 μmol, 1.0 equiv.) in MeOH (2 mL) was added DIEA (16.90 mg, 130.79 μmol, 2.0 equiv.) and formaldehyde (7.96 mg, 98.09 μmol, 37% purity, 1.5 equiv.). The mixture was stirred at 25° C. for 0.25 h followed by addition of NaBH 3 CN (8.22 mg, 130.79 μmol, 2.0 equiv.) was added. The mixture was stirred at 25° C. for 0.75 h. The reaction mixture was diluted with saturated Na 2 CO 3 The pH was adjusted to 7 with aqueous solution (80 mL) and extracted with DCM (5 mL x 2). The combined organic layers were washed with Na 2 SO 4 The residue was purified by preparative HPLC (neutral conditions, column: Waters Xbridge BEH C18 100 * 30mm * 10um; Mobile phase: [Water (NH 4 HCO 3)-ACN]; B%: 30%~60%, 10 min) to give 8-bromo-6-(2,6-dichlorophenyl)-2-[[1-(1-methyl-3-piperidyl)pyrazol-4-yl]amino]pyrido[4,3-d]pyrimidin-5-one (10.34 mg, 17.97 μmol, yield 27.49%, purity 95.48%) as a yellow solid. 1 H NMR (400MHz, DMSO-d 6 )δ10.69(s,1H),9.10(s,1H),8.44(d,J=3.75Hz,2H),7.82(s,1H),7.75(d,J=8.00Hz,2H),7.58-7.65(m,1H), 4.26-4.35(m,1H),3.31(s,1H),2.99(brd,J=8.00Hz,1H),2.22(s,3H),1.97-2.05(m,2H),1.57-1.81(m,4H). ESI[M+H]=549.9 / 547.9.

[0344] Example 31 Compounds were tested against two kinases. Compounds were tested at 10 dose IC in 3-fold serial dilutions starting at 10 μM. 50 The control compound staurosporine was tested in 10 dose IC 50 A surrogate control compound, Wee-1 inhibitor, was tested in 10 dose IC 50 The reaction was performed in the ATP-dependent mode. The reaction was carried out at 10 μM ATP. See Figures 2 and 3.

[0345] Data includes raw data, % enzyme activity (relative to DMSO control) and curve fitting.

[0346] * Curve fitting was performed where the enzyme activity at the highest concentration of compound was less than 65%.

[0347] [Table 2]

[0348] [Table 3]

[0349] Example 32 In this example, the compounds described herein were subjected to an assay to detect changes in cell cycle phase upon drug treatment, allowing the identification of drugs that affect cell proliferation in a specific manner. For example, problematic DNA replication increases the percentage of cells in S phase. Thus, quantification of cells in S phase allows the identification of drugs that cause defects in DNA replication. Similarly, drugs that selectively cause defects in progression through G2 / M can also be identified. Furthermore, synergy between the assayed drug and known cell cycle regulators can be determined, as exemplified by the effect of ATR inhibition when combined with an agent that slows DNA replication, causing replication fork collapse into DSBs and complete loss of DNA synthesis. Two novel drugs can also be combined to quantify synergistic effects on specific cell cycle phases.

[0350] The assay operates by quantification of DNA content per cell by flow cytometry. A Guava EasyCyte™ flow cytometer (Austin, TX) is used to assess cells for effects on the cell cycle. Drug-treated cells are fixed, stained with propidium iodide (PI), and resuspended in phosphate-buffered saline containing PI staining buffer. During flow cytometry, PI is excited at 528 nm and detected with a 610 / 20 bandpass. The x-axis readout is cell number. The y-axis readout is the optical density of the DNA staining dye. The assay generates a histogram showing cell number by amount of DNA, an indicator of position in the cell cycle. This rapid assay detects relative changes in cell cycle upon drug treatment compared to vehicle-treated controls and single-agent controls for detection of combination interactions. See Table 4.

[0351] [Table 4]

[0352] Although the present invention has been described in detail and with reference to specific embodiments thereof, it will be apparent to those skilled in the art that various changes and modifications can be made therein without departing from the spirit and scope of the invention.

Claims

1. A compound of formula I or III below: (i) A compound of formula I below 【Chemical 1】 (wherein, R １ is halo, C １－６ alkyl, C ３－８ cycloalkyl, or C ２－６ alkenyl, and R ２ is H, C １－６ alkyl, or C ３－８ cycloalkyl), or (ii) A compound of formula III below [Chemical Formula 2] (wherein, R2 is H, C1-6 alkyl or C3-8 cycloalkyl), or a pharmaceutically acceptable salt thereof.

2. R １ is halo, for example chloro, bromo, fluoro, or iodo, or a compound according to claim 1 which is, for example, chloro or bromo.

3. R １ and / or R2 is C １－４ alkyl, for example, methyl, ethyl, n-propyl, or i-propyl, the compound according to claim 1.

4. R １ is a C ３－７ cycloalkyl, for example, cyclopropyl, the compound according to claim 1.

5. R １ The compound according to claim 1, wherein R is C2-4 alkenyl, for example vinyl or isopropenyl.

6. R ２ is a C ３－８ cycloalkyl, for example, cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl, or for example cyclopropyl, the compound according to any one of claims 1 to 5.

7. A compound, which is of formula II below, [Chemical Formula 3] or a pharmaceutically acceptable salt thereof, the compound according to claim 1.

8. A compound, which is of the following formula, [Chemical Formula 4] or a pharmaceutically acceptable salt thereof, the compound according to claim 1.

9. A compound, which is of the following formula, [Chemical Formula 5] 【Chemical Formula 6】 or a pharmaceutically acceptable salt thereof, the compound according to claim 1. The compound of formula III according to claim 10, and R ２ is H, the compound according to claim 1.

11. A compound of formula IVa, Va, VIa, or VII below, (i) A compound of formula IVa below 【Chemical Formula 7】 (wherein, R １ and R ２ are each, independently, H, methyl, ethyl, or propyl, or R １ and R ２ are combined to form a 3- to 6-membered ring, R ３ is H, C １－４ alkyl or C ３－６ cycloalkyl, and R ４ is H, C １－３ alkyl, CF ３ , -O-methyl, OCF ３ , OCF ２ H, CN, or halo), (ii) A compound of formula Va below 【Chemical Formula 8】 (wherein, R2 is H, methyl, ethyl, or propyl, R3 is H, C1-4 alkyl or C3-6 cycloalkyl, R4 is H, C1-3 alkyl, CF3, -O methyl, OCF3, OCF2H, CN, or halo), (iii) A compound of formula VIa below 【Chemical Formula 9】 (wherein, R2 is H, methyl, ethyl, or propyl, R3 is H, C1-4 alkyl or C3-6 cycloalkyl, R4 is H, C1-3 alkyl, CF3, -O methyl, OCF3, OCF2H, CN, or halo, R5 and R6 are independently H, halo, or C1-6 alkyl), (iv) A compound of formula I below 【Chemical Formula 10】 (wherein R10 is H, OH, NH2, NH(C1-6 alkyl) or N(C1-6 alkyl)(C1-6 alkyl)), or a pharmaceutically acceptable salt thereof.

12. 【Fig. 11】 is the compound according to claim 11, or a pharmaceutically acceptable salt thereof.

13. R ２ The compound according to claim 11, wherein R, R3, and / or R4 or R10 is H.

14. R ３ , R4, R5, and / or R6 are C １－６ alkyl, such as methyl, the compound according to claim 11.

15. R ５ The compound according to claim 11, wherein R and / or R6 is a halo such as F.

16. R １０ The compound according to claim 11, wherein R is OH.

17. R １０ is NH ２ and is the compound according to claim 11.

18. R １０ is NH(C １－６ alkyl), the compound according to claim 11.

19. R １０ is N(C １－６ alkyl)(C １－６ alkyl), for example N(CH ３ ), ２ the compound according to claim 11, wherein it is such.

20. A pharmaceutical composition comprising one or more compounds according to claim 1 or 11 and one or more pharmaceutically acceptable excipients.

21. A pharmaceutical composition for inhibiting Wee1, treating cancer, reducing the activity of a kinase encoded by the gene WEE1, or treating or preventing a Wee1-mediated disease in a patient in need of such treatment, the pharmaceutical composition comprising the compound according to claim 1 or 11. **Claim 22**: The cancer is adrenal cortical tumor, AIDS-related cancer (e.g., AIDS-related lymphoma), anal cancer, appendiceal cancer, astrocytoma, atypical teratoid / rhabdoid tumor, basal cell carcinoma, bile duct cancer, bladder cancer, bone cancer, brain cancer (e.g., glioblastoma), breast cancer, bronchial cancer, cancer of unknown primary site, etc., carcinoid tumor, castration-resistant prostate cancer, central nervous system cancer (e.g., central nervous system atypical teratoid / rhabdoid tumor, central nervous system fetal tumor, central nervous system lymphoma, primary central nervous system lymphoma), cervical cancer, chordoma, chondrosarcoma, chronic myeloproliferative disorder, colon cancer, colorectal cancer, craniopharyngioma, fibromatosis colli, diffuse large B-cell lymphoma, endometrial cancer, epithelioma, esophageal cancer, Ewing sarcoma family tumor, extracranial germ cell tumor, extragonadal germ cell tumor, extrahepatic bile duct cancer, gallbladder cancer, gastric (stomach) cancer, gastrointestinal carcinoid tumor, gastrointestinal stromal tumor, germ cell tumor, gestational trophoblastic tumor, glioma, head cancer, hepatocellular (liver) cancer, high-grade prostate cancer, histiocytosis, hypopharyngeal cancer, Kaposi sarcoma, kidney (renal) cancer, Langerhans cell histiocytosis, laryngeal cancer, leptomeningeal disease, lip cancer, low-grade prostate cancer, leukemia (e.g., chronic lymphocytic leukemia, chronic myeloid leukemia, hairy cell leukemia), lung cancer, lymphoma (Burkitt lymphoma, central nervous system lymphoma, T-cell lymphoma (e.g., cutaneous T-cell lymphoma, Hodgkin lymphoma, Hodgkin lymphoma, non-Hodgkin lymphoma, lymphoplasmacytic lymphoma)), intermediate-grade prostate cancer, medulloblastoma, medulloepithelioma, melanoma, Merkel cell carcinoma, mesothelioma, metastatic squamous cell carcinoma of unknown primary, oral cancer, multiple endocrine neoplasia syndrome, multiple myeloma, multiple myeloma / plasma cell neoplasm, mycosis fungoides, myelodysplastic syndrome, myelodysplasia / myeloproliferative neoplasm, myeloproliferative disorder, nasal or paranasal sinus cancer, neck cancer, nasopharyngeal cancer, neck cancer, neuroblastoma, ocular cancer, uveal melanoma, oral cancer, oropharyngeal cancer, oral cancer, osteosarcoma or malignant fibrous histiocytoma of bone, osteosarcoma or malignant fibrous histiocytoma, ovarian cancer, ovarian germ cell tumor, ovarian epithelial cancer, ovarian low-grade tumor, pancreatic cancer, papillomatosis, paranasal sinus or nasal cavity cancer, parathyroid cancer, penile cancer, pharyngeal cancer, moderately differentiated pineal parenchymal tumor, pineoblastoma or supratentorial primitive neuroectodermal tumor, pituitary tumor, pleuropulmonary blastoma, pregnancy cancer, prostate cancer, rectal cancer, renal pelvis cancer, airway cancer involving the NUT gene on chromosome 15, retinoblastoma, rhabdomyosarcoma, salivary gland cancer, sarcoma, Sézary syndrome, skin cancer, skin cancer, small intestine cancer,The pharmaceutical composition according to claim 21, which is for soft tissue sarcoma, spinal cord tumor, squamous cell carcinoma, squamous cell carcinoma of the neck of unknown primary origin, potential primitive neuroectodermal tumor on the tentorium, testicular cancer, pharyngeal cancer, thymoma or thymic carcinoma, thyroid cancer, transitional cell carcinoma of the renal pelvis or ureter, rare cancers in children, ureteral cancer, urethral cancer, uterine sarcoma, vaginal cancer, vulvar cancer, Waldenström macroglobulinemia (lymphoplasmacytic lymphoma), Wilms tumor, or cancer in women, or, for example, the cancer is breast cancer, prostate cancer, pancreatic cancer, lung cancer, colorectal cancer, ovarian cancer, liver cancer, melanoma, kidney cancer, central nervous system cancer, brain cancer such as glioblastoma, leukemia, or lymphoma.

23. The pharmaceutical composition according to claim 21, which is administered in combination with a chemotherapeutic agent such as at least one additional therapeutic agent including a chemotherapeutic agent, busulfan, melphalan, chlorambucil, cyclophosphamide, ifosfamide, temozolomide, bendamustine, cisplatin, mitomycin C, bleomycin, carboplatin, camptothecin, irinotecan, topotecan, doxorubicin, epirubicin, aclarubicin, mitoxantrone, elliptinium, etoposide, 5-azacitidine, gemcitabine, 5-fluorouracil, methotrexate, 5-fluoro-2'-deoxy-uridine, fludarabine, nelarabine, ara-C, pralatrexate, pemetrexed, hydroxyurea, thioguanine, colchicine, vinblastine, vincristine, vinorelbine, paclitaxel, ixabepilone, cabazitaxel, docetaxel, camptosar, panitumumab, matuzumab, nabumetuzumab, pimuzumab, remolizumab, bevacizumab, pertuzumab, trastuzumab, cetuximab, obinutuzumab, ofatumumab, rituximab, alemtuzumab, gemtuzumab, tositumomab, bentsuximab, daratumumab, elotuzumab, T-DM1, ofatumumab, dinutuximab, blinatumomab, ipilimumab, avastin, trastuzumab, rituximab, imatinib, gefitinib, erlotinib, osimertinib, afatinib, ceritinib, alectinib, crizotinib, erlotinib, lapatinib, sorafenib, nintedanib, nilotinib, dasatinib, pazopanib, temsirolimus, everolimus, vorinostat, romidepsin, panobinostat, belinostat, tamoxifen, letrozole, fulvestrant, mitoguazone, octreotide, retinoic acid, arsenic trioxide, zoledronic acid, bortezomib, carfilzomib, ixazomib, besimodegib, sonidegib, denosumab, thalidomide, lenalidomide, venetoclax, aldesleukin (recombinant human interleukin-2), sipuleucel-T (prostate cancer treatment vaccine), palbociclib, olaparib, niraparib, rucaparib, talazoparib, or a combination thereof.

24. The pharmaceutical composition according to claim 21, which is used for in vitro performance.

25. The pharmaceutical composition according to claim 21, which is used for performance in a subject.

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