How to treat splenomegaly

Abstract

Therapeutic methods and pharmaceutical compositions for treating splenomegaly in a human subject are described. In certain embodiments, the present invention includes therapeutic methods for treating splenomegaly using a BTK inhibitor.

Description

[Technical Field]

[0001]

[0001] Disclosed herein are methods for treating splenomegaly using Bruton's tyrosine kinase (BTK) inhibitors. [Background technology]

[0002] Myelofibrosis (MF) is a chronic leukemia that is a cancer affecting the blood-forming tissues in the body. It belongs to a group of diseases called myeloproliferative disorders and is a rare type of bone marrow cancer that disrupts the normal production of blood cells. Myelofibrosis causes extensive scarring in the bone marrow, leading to severe anemia that can cause weakness and fatigue; myelofibrosis can also cause a decrease in platelet count, which increases the risk of bleeding. Myelofibrosis often leads to an enlarged spleen and lymph nodes due to the accumulation of CD34+ malignant myeloid cells in the spleen.

[0003]

[0003] The clinical spectrum of MF includes primary myelofibrosis and MF occurring during essential thrombocythemia or polycythemia vera. Myelofibrosis is a chronic hematologic malignancy characterized by splenomegaly, leukoerythroblastosis, cytopenias, teardrop poikilocytosis, bone marrow fibrosis, extramedullary hematopoiesis, increased bone marrow microvascular density, and constitutive mobilization of CD34-expressing hematopoietic stem cells (HSCs) and progenitor cells (HPCs).

[0004] Myelofibrosis is also characterized by abnormal trafficking and homing of HSCs and HPCs in the bone marrow and peripheral blood, leading to constitutive mobilization of HSCs and HPCs and the establishment of splenomegaly. CXCR4-CXCL12 (CXCL12, also known as SDF-1) signaling plays a pivotal role in various processes underlying proper lymphoid and myeloid cell development and function, including the development and maintenance of progenitor cells in the bone marrow, the homing of immature and mature cells to secondary lymphoid organs, and the trafficking and homing of plasma cells to the bone marrow. In MF, constitutive mobilization of HSCs and HPCs is associated with profound alterations in the CXCR4-CXCL12 axis, resulting from downregulation of CXCR4 expression by myelofibrotic CD34+ cells due to hypermethylation of the CXCR4 promoter and proteolytic degradation of CXCL12. In the spleens of MF patients, CXCL12 and integrins such as very late antigen-4 (VLA-4) are highly expressed, and CXCL12 acts as a chemoattractant for recruited CD34+ cells. This contrasts with bone marrow and peripheral blood, where CXCL12 expression levels are abnormally low. Once attracted to the spleen via CXCL12, adhesion molecules such as VLA-4 and its ligand VCAM-1 (vascular cell adhesion molecule 1) sequester CD34+ cells, leading to the formation of splenomegaly. In addition, this aberrant stem cell behavior can be influenced not only by intrinsic properties of stem cells but also by regulatory signals provided by the MF microenvironment (Wang (2015) Experimental Hematology 43, pp. 100–109). Therefore, the ability to manipulate cell trafficking, homing, and sequestration via these pathways offers an opportunity to treat MF patients.

[0005]

[0005] Bruton's tyrosine kinase is a non-receptor tyrosine kinase belonging to the Tec family and has important functions in several types of benign and malignant cells of the hematopoietic system. Furthermore, recent clinical studies using the irreversible oral BTK inhibitors, acalabrutinib and ibrutinib, have demonstrated excellent clinical activity and tolerability in a variety of B-cell malignancies, including chronic lymphocytic leukemia (CLL), mantle cell lymphoma (MCL), Waldenström's macroglobulinemia, and diffuse large B-cell lymphoma. Furthermore, it is now clear that the mechanism of action of BTK inhibitors is multifactorial, and a key component of their function is disruption of tumor cells and their protective microenvironment. Inhibition of BTK has been shown to regulate malignant myeloid cell migration in CLL, MCL, and acute myeloid leukemia by downregulating the expression of numerous vascular adhesion molecules and inhibiting CXCR4-CXCL12-induced cell trafficking, homing, and integrin adhesion (Zaitseva (2014) Oncotarget 5, pp. 9930-9938). CXCL12 plays a central role in CLL pathogenesis and progression by regulating CLL cell interactions with the stromal microenvironment, leading to cell survival and proliferation. BTK plays a role in signal transduction activated by the CXCR4-CXCL12 signaling axis and is involved in rapid integrin activation. BTK inhibition blocks CXCL12-induced induction of lymphocyte function-associated antigen-1 (LFA-1) and VLA-4 integrins. Furthermore, BTK inhibition blocks activation of the small GTP-binding protein RhoA, which regulates integrin affinity. Importantly, BTK tyrosine phosphorylation and activation by CXCL12 is dependent on upstream activation of JAK2 (Janus kinase 2). Thus, BTK and JAK protein tyrosine kinases exhibit hierarchical activity in both chemokine and integrin activation and dependent cell adhesion (Montresor (2018) Oncotarget, 9, 35123-35140). Finally, BTK is highly expressed in both mature and primitive myeloid cells, including HSCs and HPCs.The CXCR4-CXCL12 signaling axis is a key means of recruitment and homing for CD34+ cells.

[0006]

[0006] Currently, ruxolitinib, fedratinib, and allogeneic stem cell transplantation are the main means of treating patients with MF. Ruxolitinib, a drug developed to inhibit JAK2 mutations, is often the first treatment used. Ruxolitinib is also effective in people with CALR (calreticulin, located on chromosome 19p13.2) or MPL (myeloproliferative leukemia viral oncogene; located on chromosome 1p34) mutations because CALR and MPL mutations also activate JAK2. Ruxolitinib is effective in reducing spleen size and controlling symptoms, which can increase overall survival; however, ruxolitinib does not reverse fibrosis in most cases and can lead to anemia and low platelet counts. Surprisingly, ruxolitinib works equally well to reduce splenomegaly and control symptoms in MF patients lacking the JAK2V617 driver mutation, confounding early treatment predictions. However, ruxolitinib is not disease-modifying because it has no effect on reducing the number of malignant CD34+ cells or Jak allele burden. Nevertheless, with a better understanding of the cellular and molecular events leading to the development of MF, the potential exists for safer and more effective targeted therapies, such as BTK inhibitors, to treat myeloproliferative neoplasms with splenomegaly through modulation of cell trafficking, homing, and adhesion. Summary of the Invention

[0007]

[0007] The present invention relates to a method of treating splenomegaly in a human subject in need thereof, comprising administering a Bruton's tyrosine kinase inhibitor to the human subject. In some embodiments, the human subject has an accumulation of malignant CD34+ myeloid cells in the spleen. In some embodiments, the malignant CD34+ myeloid cells have decreased expression of CXCR4 compared to normal myeloid cells. In some embodiments, the human subject is suffering from myelofibrosis. In some embodiments, the myelofibrosis is selected from the group consisting of primary myelofibrosis (PMF), post-polycythemia vera myelofibrosis (post-PV-MF), and post-essential thrombocythemia myelofibrosis (post-ET-MF). In some embodiments, the human subject has failed to respond to ruxolitinib therapy. In some embodiments, the human subject has acute myeloid leukemia (AML) harboring the JAK2V617F mutation, optionally secondary to a myeloproliferative neoplasm (MPN). In some embodiments, the human subject does not have a JAK2V617F mutation and optionally has acute myeloid leukemia secondary to a myeloproliferative neoplasm.

[0008] In some embodiments, the BTK inhibitor is administered in an amount sufficient to stimulate migration of malignant CD34+ myeloid cells into the peripheral blood of a human subject. In some embodiments, the BTK inhibitor is administered in an amount sufficient to stimulate apoptosis of malignant CD34+ myeloid cells in the spleen of a human subject. In some embodiments, the BTK inhibitor is administered in an amount sufficient to reduce VLA-4 activity in malignant CD34+ myeloid cells. In some embodiments, the BTK inhibitor is administered in an amount sufficient to reduce VLA-4 expression in malignant CD34+ myeloid cells.

[0009] The present invention also relates to a method for stimulating migration of malignant CD34+ myeloid cells from the spleen to peripheral blood in a human subject suffering from splenomegaly, comprising administering a BTK inhibitor to the human subject. In some embodiments, the human subject has an accumulation of malignant CD34+ myeloid cells in the spleen. In some embodiments, the method comprises stimulating apoptosis of malignant CD34+ myeloid cells in the spleen by administering a BTK inhibitor. In some embodiments, the malignant CD34+ myeloid cells have decreased expression of CXCR4 compared to normal myeloid cells. In some embodiments, the human subject has myelofibrosis. In some embodiments, the myelofibrosis is selected from the group consisting of primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In some embodiments, the human subject has failed to respond to ruxolitinib therapy. In some embodiments, the human subject has acute myeloid leukemia harboring the JAK2V617F mutation and optionally secondary to a myeloproliferative neoplasm. In some embodiments, the human subject does not have a JAK2V617F mutation and optionally has acute myeloid leukemia secondary to a myeloproliferative neoplasm.

[0010] The methods of the invention include treating human subjects who have not been treated with a JAK2 inhibitor. The methods of the invention also include treating human subjects who are intolerant to a JAK2 inhibitor. The methods of the invention include treating human subjects who are ineligible for treatment with a JAK2 inhibitor. The methods of the invention include treating human subjects who relapse after JAK2 inhibitor treatment or who are refractory to JAK2 inhibitor treatment.

[0011] In the methods of the present invention, the BTK inhibitor is administered once daily at a dose selected from the group consisting of 15 mg, 25 mg, 30 mg, 50 mg, 60 mg, 75 mg, 90 mg, 100 mg, 120 mg, 150 mg, 175 mg, 180 mg, 200 mg, 225 mg, 240 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 360 mg, 375 mg, 480 mg, and 560 mg.

[0012] In the methods of the present invention, the BTK inhibitor is administered twice daily at a dose selected from the group consisting of 15 mg, 25 mg, 30 mg, 50 mg, 60 mg, 75 mg, 90 mg, 100 mg, 120 mg, 150 mg, 175 mg, 180 mg, 200 mg, 225 mg, 240 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 360 mg, 375 mg, 480 mg, and 560 mg.

[0013]

[0013] In the methods of the present invention, the BTK inhibitor is administered orally.

[0014]

[0014] In the methods of the present invention, the BTK inhibitor is a covalent BTK inhibitor.

[0015]

[0015] In the methods of the present invention, the BTK inhibitor is a non-covalent BTK inhibitor. [Brief explanation of the drawings]

[0016] [Figure 1A] 1 is a graph showing inhibition of cell migration toward SDF-1. [Figure 1B] 1 is a graph showing inhibition of cell migration toward SDF-1. [Figure 2] 1 is a graph showing cell release from fibronectin. [Figure 3] 1 is a graph showing the levels of cell surface molecules. [Figure 4] Graph showing cytokine and chemokine levels. DETAILED DESCRIPTION OF THE INVENTION

[0017]

[0020] While preferred embodiments of the present invention have been shown and described herein, such embodiments are provided by way of example only and are not intended to otherwise limit the scope of the invention. Various alternatives to the described embodiments of the invention may be employed in practicing the invention.

[0018]

[0021] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0019]

[0022] As used herein, the terms "administered in combination with" and "co-administration" include administration of two or more active pharmaceutical ingredients to a subject such that both agents and / or metabolic products of both agents are present in the subject at the same time. Co-administration includes simultaneous administration in separate compositions, administration at different times in separate compositions, or administration in a single composition where two or more agents are present.

[0020]

[0023] The term "effective amount" or "therapeutically effective amount" or "sufficient amount" refers to an amount of an active pharmaceutical ingredient or active pharmaceutical ingredient combination described herein that is sufficient to achieve the intended application, including but not limited to, disease treatment. A therapeutically effective amount may vary depending on the intended application (in vitro or in vivo), or the subject and disease state being treated (e.g., the subject's weight, age, and sex), the severity of the disease state, the mode of administration, and other factors that can be readily determined by one skilled in the art. The term also applies to a dose that will induce a specific response in target cells (e.g., malignant CD34+ myeloid cells). The specific dose will vary depending on the particular compound selected, the dosing regimen to be followed, whether the compound is administered in combination with other compounds, the timing of administration, the tissue to which the compound is administered, and the physical delivery system by which the compound is delivered.

[0021]

[0024] "Myelofibrosis" refers to spontaneous scarring (fibrosis) of the bone marrow, which disrupts the normal production of blood cells and leads to severe anemia and enlargement of the spleen, lymph nodes, and liver. Myelofibrosis can be associated with a variety of diseases, primarily myeloproliferative (preleukemic) disorders. Myelofibrosis is also known as myeloid metaplasia of unknown etiology. As used herein, myelofibrosis includes, but is not limited to, primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. As used herein, myelofibrosis is characterized by the accumulation of malignant CD34+ myeloid cells in the bone marrow, spleen, and lymph nodes.

[0022]

[0025] A "pharmaceutically acceptable carrier" or "pharmaceutically acceptable excipient" is intended to include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic agents, and absorption delaying agents. The use of such media and agents for active pharmaceutical ingredients is known in the art. Except insofar as any conventional media or agent is incompatible with the active pharmaceutical ingredient, use of any conventional media or agent in the therapeutic compositions of the invention is contemplated. Supplementary active ingredients can also be incorporated into the described compositions.

[0023]

[0026] The term "pharmaceutically acceptable salt" refers to salts derived from a variety of organic and inorganic counterions known in the art. Pharmaceutically acceptable acid addition salts can be formed with inorganic and organic acids. Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, and phosphoric acid. Organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and salicylic acid. Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases. Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum. Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, substituted amines, including naturally occurring substituted amines, cyclic amines, and basic ion exchange resins. Specific examples include isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. In selected embodiments, pharmaceutically acceptable base addition salts are selected from ammonium, potassium, sodium, calcium, and magnesium salts. The term "cocrystal" refers to a molecular complex derived from several cocrystal formers known in the art. Unlike salts, cocrystals typically do not involve proton transfer between the cocrystal and the drug, but instead involve intermolecular interactions such as hydrogen bonding, aromatic ring stacking, or dispersion forces between the cocrystal former and the drug in the crystal structure.

[0024]

[0027] The terms "QD", "qd", or "qd" mean daily, once a day, or once a day. The terms "BID", "bid", or "bid" mean twice per day, twice a day, or twice a day. The terms "TID", "tid", or "tid" mean three times per day, three times a day, or three times a day. The terms "QID", "qid", or "qid" mean four times per day, four times a day, or four times a day.

[0025]

[0028] As used herein, the term "splenomegaly" refers to an enlargement of the spleen, as assessed by size or weight. In some embodiments, the enlargement results from the sequestration of malignant CD34+ myeloid cells and the resulting development of extramedullary hematopoiesis.

[0026]

[0029] A "therapeutic effect," as that term is used herein, encompasses the therapeutic benefits and / or prophylactic benefits described above. A prophylactic effect includes delaying or eliminating the appearance of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing, halting, or reversing the progression of a disease or condition, or any combination thereof.

[0027]

[0030] When ranges are used herein to describe physical or chemical properties, such as molecular weight or chemical formula, all combinations and subcombinations of the ranges and specific embodiments within the ranges are intended to be included. The use of the term "about" when referring to a number or numerical range means that the stated number or numerical range is approximate within experimental variation (or within statistical experimental error), and thus the number or numerical range may vary, for example, by 1% to 15% of the stated number or numerical range. The term "comprising" (and related terms such as "comprise" or "comprises" or "having" or "including") includes, for example, embodiments "consisting of" or "consisting essentially of" the described characteristic, such as any composition of matter, method, or process.

[0028]

[0031] The BTK inhibitor compounds of the present invention also include any crystalline and amorphous forms of the compounds in Table 1, including, for example, polymorphs, pseudopolymorphs, solvates, hydrates, non-solvated polymorphs (including anhydrous forms), conformational polymorphs, and amorphous forms of the compounds, and mixtures thereof. "Crystalline form" and "polymorph" are intended to include all crystalline and amorphous forms of the compounds, including, for example, polymorphs, pseudopolymorphs, solvates, hydrates, non-solvated polymorphs (including anhydrous forms), conformational polymorphs, and amorphous forms, and mixtures thereof, unless a specific crystalline or amorphous form is referenced.

[0029] Methods for treating complications of myelofibrosis

[0032] The present disclosure relates to the discovery that BTK inhibitors can be used to treat various complications of myelofibrosis, including, for example, splenomegaly, extramedullary hematopoiesis, and fibrosis. Accordingly, in certain aspects, the present disclosure relates to methods for treating splenomegaly, extramedullary hematopoiesis, and fibrosis by administering an effective amount of a BTK inhibitor to a human subject in need of treatment, optionally in combination with one or more other supportive therapies or active agents for treating splenomegaly. The disclosure herein demonstrates that desirable therapeutic agents can be selected based on BTK inhibition. Therefore, without wishing to be bound by a particular mechanism of action, it is anticipated that BTK inhibition, which alters one or more downstream signaling components (e.g., CXCR-4, CXCL12, VLA-4, VCAM-1) to recruit CD34+ cell migration to the peripheral blood, will be useful in treating complications associated with myelofibrosis, particularly in treating or preventing one or more myelofibrosis complications, including, but not limited to, splenomegaly, extramedullary hematopoiesis, and fibrosis.

[0030]

[0033] Thus, the present invention relates to methods of treating splenomegaly, comprising administering to a human in need thereof a BTK inhibitor compound selected from Table 1 or a pharmaceutically acceptable salt thereof. In some embodiments, the splenomegaly is secondary to myelofibrosis. In some embodiments, the MF is primary myelofibrosis, also known as chronic idiopathic myelofibrosis (cIMF). This is in contrast to myelofibrosis secondary to polycythemia vera or essential thrombocythemia. However, in some embodiments, the present invention encompasses treating splenomegaly due to myelofibrosis secondary to polycythemia vera or essential thrombocythemia. In some embodiments, the BTK inhibitor is any of the compounds in Table 1 or a pharmaceutically acceptable salt thereof.

[0031]

[0034] The present invention also relates to a method for treating splenomegaly, comprising administering a BTK inhibitor or a pharmaceutically acceptable salt thereof to a human subject in need thereof. In one embodiment, the splenomegaly is in a human subject suffering from a myelofibrosis selected from the group consisting of primary myelofibrosis, secondary myelofibrosis, myelofibrosis secondary to polycythemia vera (PV), myelofibrosis secondary to essential thrombocythemia (ET), myelofibrosis secondary to chronic myeloid leukemia (CML), and idiopathic myelofibrosis. In one embodiment, the myelofibrosis is selected from the group consisting of primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of prefibrotic / early PMF and overt fibrotic stage PMF. In one embodiment, the human is determined to be hydroxyurea (HU) intolerant (unacceptable side effect). In one embodiment, the human subject is determined to be hydroxyurea resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy dependent. In one embodiment, the human subject is phlebotomy dependent without splenomegaly.

[0032]

[0035] In one embodiment, the human subject is JAK2 inhibitor naive (i.e., has not received therapy with a JAK2 inhibitor). In one embodiment, the human subject is JAK2 inhibitor intolerant. In one embodiment, the human subject is JAK2 inhibitor ineligible due to low platelet count. In one embodiment, the human subject has relapsed after JAK2 inhibitor treatment. In one embodiment, the human subject is refractory to JAK2 inhibitor treatment. In one embodiment, the human subject has failed ruxolitinib or fedratinib therapy. Failure of ruxolitinib or fedratinib therapy includes, but is not limited to, (i) a lack of reduction in the severity or progression of any myeloproliferative neoplasm in a human subject receiving ruxolitinib or fedratinib, or (ii) a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, failure of ruxolitinib or fedratinib therapy is a failure to reduce the severity or progression of any myelofibrosis in a human subject receiving ruxolitinib or fedratinib. In one embodiment, failure of ruxolitinib or fedratinib therapy is a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, the BTK inhibitor is a compound selected from Table 1, and pharmaceutically acceptable salts thereof.

[0033]

[0036] In one embodiment, the invention relates to a method of treating splenomegaly in a human, comprising the step of administering to the human a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from Table 1.

[0034]

[0037] In one embodiment, the invention relates to a method of treating extramedullary hematopoiesis in a human, comprising the step of administering to the human a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from Table 1.

[0035]

[0038] In one embodiment, the invention relates to a method of treating fibrosis in a human, comprising the step of administering to the human a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from any of Table 1.

[0036]

[0039] In one embodiment, a human subject has an accumulation of malignant CD34+ myeloid cells in their spleen. These malignant CD34+ myeloid cells have reduced expression of CXCR4 compared to normal myeloid cells. In one embodiment, a BTK inhibitor is administered in a therapeutically effective amount sufficient to stimulate migration of the malignant CD34+ myeloid cells into the peripheral blood of the human subject. In one embodiment, the BTK inhibitor is administered in an amount sufficient to inactivate VLA-4 on the malignant CD34+ myeloid cells.

[0037]

[0040] In one embodiment, the present invention relates to a method for stimulating the migration of malignant CD34+ myeloid cells from the spleen to peripheral blood in a human subject suffering from myelofibrosis, comprising administering a BTK inhibitor to the human subject. In one embodiment, the BTK inhibitor is administered in an amount sufficient to reduce the activity of CXCR4 and CXCL12, thereby reducing the chemoattractant effect of these molecules on malignant CD34+ myeloid cells. The reduction in CXCR4 and CXCL12 activity contributes to the sequestration of malignant myeloid CD34+ cells in the spleen. In one embodiment, malignant CD34+ myeloid cells have accumulated in the spleen of a human subject. These malignant CD34+ myeloid cells have reduced expression of CXCR4 compared to normal myeloid cells.

[0038]

[0041] In one embodiment, the method includes treating complications associated with myelofibrosis, such as splenomegaly, extramedullary hematopoiesis, and fibrosis, but not treating myelofibrosis itself (e.g., only the complications of myelofibrosis are treated in a human, and not myelofibrosis).

[0039]

[0042] In one embodiment, the BTK inhibitor is administered at a dose of 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 560 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg 200 mg BID, 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID. In one embodiment, the BTK inhibitor is administered to a human according to the Dosages and Dosing Regimen section.

[0040]

[0043] In one embodiment, the human suffering from splenomegaly, extramedullary hematopoiesis, or fibrosis has myelofibrosis characterized by the presence of a CALR mutation (calreticulin, located on chromosome 19p13.2) in a human subject, as described in Massie, New Engl. J. Med. (2013) 25, pp. 2379-2390, which is incorporated herein by reference in its entirety.

[0041]

[0044] In one embodiment, the human suffering from splenomegaly, extramedullary hematopoiesis, or fibrosis has myelofibrosis characterized by the presence of the MPL mutation (myeloproliferative leukemia viral oncogene; located on chromosome 1p34) in a human subject, as described in Pikman, Plos Med. (2006) 3, e270, which is incorporated herein by reference in its entirety.

[0042]

[0045] In one embodiment, the human suffering from splenomegaly, extramedullary hematopoiesis, or fibrosis has myelofibrosis characterized by the JAK2V617F mutation in the human subject. JAK2V617F is a functional mutation that promotes cytokine-independent proliferation of myeloid cells, as described by Nakatake (Oncogene (2012) 31, pp. 1323-1333), the entire contents of which are incorporated herein by reference.

[0043]

[0046] In one embodiment, the human suffering from splenomegaly, extramedullary hematopoiesis, or fibrosis has myelofibrosis characterized by one or more mutations selected from the group consisting of JAK2V617F, MPL, CALR, and combinations thereof.

[0044]

[0047] In one embodiment, the human suffering from splenomegaly, extramedullary hematopoiesis, or fibrosis has myelofibrosis characterized by the absence of the JAK2V617F mutation.

[0045]

[0048] In one embodiment, the invention relates to a method of treating splenomegaly in a human subject suffering from myelofibrosis secondary to essential thrombocythemia in the human, comprising the step of administering to the human a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from Table 1.

[0046]

[0049] In one embodiment, the invention relates to a method of treating myelofibrosis secondary to chronic myeloid leukemia in a human, comprising the step of administering to the human a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from Table 1.

[0047] [Table 1] JPEG0007771064000002.jpg200149 JPEG0007771064000003.jpg216149 JPEG0007771064000004.jpg200149 JPEG0007771064000005.jpg200149 JPEG0007771064000006.jpg201149 JPEG0007771064000007.jpg193149 JPEG0007771064000008.jpg200149 JPEG0007771064000009.jpg200149 JPEG0007771064000010.jpg208149 JPEG0007771064000011.jpg201149 JPEG0007771064000012.jpg162149

[0048]

[0050] In some embodiments, the BTK inhibitor is TG-1701 or Loxo-305.

[0049]

[0051] In one embodiment, the present invention provides a pharmaceutical composition comprising 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg BID, 200 mg

[0013] A method of treating myelofibrosis in a human, comprising administering to the human a therapeutically effective amount of a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof, at a dosage selected from the group consisting of 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID. In one embodiment, the MF is selected from the group consisting of myelofibrosis, primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of fibrillary / early PMF and fibrotic PMF.

[0050]

[0052] In one embodiment, the present invention provides a pharmaceutical composition comprising 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg BID, 200 mg 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID. The present invention relates to a method of treating myelofibrosis in a human, comprising the step of administering to the human a therapeutically effective amount of a BTK inhibitor compound selected from Table 1 or a pharmaceutically acceptable salt thereof, at a dosage selected from the group consisting of 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID, wherein the MF is selected from the group consisting of MF secondary to polycythemia vera, MF secondary to essential thrombocythemia, and MF secondary to CML.

[0051]

[0053] In one embodiment, the present invention provides a pharmaceutical composition comprising 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg BID, 200 mg 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID.

[0052]

[0054] In one embodiment, the present invention provides a pharmaceutical composition comprising 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg BID, 200 mg 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID.

[0053]

[0055] In one embodiment, the present invention provides a pharmaceutical composition comprising 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg BID, 200 mg 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID.

[0054]

[0056] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating myelofibrosis, comprising administering to a human one or more doses of a BTK inhibitor compound from Table 1 or a pharmaceutically acceptable salt thereof. In one embodiment, the MF is selected from the group consisting of primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of fibrillary / early PMF and fibrotic PMF.

[0055]

[0057] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating primary myelofibrosis, wherein treating comprises administering one or more doses of a BTK inhibitor compound from Table 1 or a pharmaceutically acceptable salt thereof to a human being.

[0056]

[0058] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating post-polycythemia vera myelofibrosis, wherein treating comprises administering to a human one or more doses of a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof.

[0057]

[0059] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating post-essential thrombocythemia myelofibrosis, wherein treating comprises administering to a human one or more doses of a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof.

[0058]

[0060] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating myelofibrosis secondary to polycythemia vera, wherein treating comprises administering to a human one or more doses of a BTK inhibitor compound from Table 1 or a pharmaceutically acceptable salt thereof.

[0059]

[0061] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating myelofibrosis secondary to essential thrombocythemia, wherein treating comprises administering to a human one or more doses of a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof.

[0060]

[0062] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating myelofibrosis secondary to chronic myeloid leukemia, wherein treating comprises administering to a human one or more doses of a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof.

[0061]

[0063] In one embodiment, the invention relates to the use of a composition comprising a BTK inhibitor selected from Table 1 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating myelofibrosis, comprising administering one or more doses of the composition comprising the BTK inhibitor or a pharmaceutically acceptable salt thereof to a human. In one embodiment, the myelofibrosis is selected from the group consisting of primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of fibrillary / early PMF and fibrotic PMF.

[0062]

[0064] The methods described above may be used as a first-line cancer therapy or following treatment with conventional therapy, including ruxolitinib or fedratinib.

[0063]

[0065] A BTK inhibitor or a pharmaceutically acceptable salt thereof may be used in combination with radiation therapy, hormone therapy, surgery, and immunotherapy, therapies known to those skilled in the art, to treat myelofibrosis selected from the group consisting of primary myelofibrosis, idiopathic myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of fibrillary / early PMF and fibrotic PMF.

[0064] Mechanism of action

[0066] Myelofibrosis is characterized by constitutive mobilization of hematopoietic stem and progenitor cells and the establishment of extramedullary hematopoiesis (EMH). Spleen and peripheral blood myelofibrotic CD34+ cells share a defective ability to home to the bone marrow but not to the spleen. This trafficking pattern cannot be attributed to discrepant expression of integrin or chemokine receptors, other than downregulation of CXCR4 by both peripheral blood and spleen CD34+ cells. The concentration of the intact chemoattractant CXCL12 (the ligand for CXCR4) is greater in spleen myelofibrotic plasma than in peripheral blood myelofibrotic plasma. A functionally inactive truncated product of CXCL12, a product of proteolytic degradation by serine proteases, was detected at similar levels in both spleen and peripheral blood myelofibrotic plasma. The myelofibrotic splenic microenvironment is characterized by increased levels of intact and functional CXCL12, which contributes to the localization of malignant CD34+ myeloid cells to the spleen (Wang (2015) Exp. Hematol. 43, pp. 100-109).

[0065]

[0067] Abnormal CD34+ cell trafficking and extramedullary hematopoiesis are essential components of the pathobiology of myelofibrosis. Constitutive recruitment of malignant bone marrow CD34+ cells is explained by downregulation of CXCR4 on these cells and reduced amounts of intact CXCL12, which acts as a chemoattractant for CD34+ cells. Increased concentrations of intact, fully functional CXCL12 in the myelofibrotic spleen, but not in the peripheral blood and possibly the bone marrow, contribute to the homing of malignant bone marrow CD34+ cells to the spleen rather than the bone marrow in myelofibrosis patients, ultimately leading to extramedullary hematopoiesis in the spleen. The initial establishment of extramedullary hematopoiesis in myelofibrosis patients is due in part to the presence of intact CXCL12 in the normal spleen, whose production is localized to cells lining blood vessels. As myelofibrosis progresses, the bone marrow is gradually depleted of CD34+ cells, but CD34+ cells are present in the peripheral blood, resulting from CD34+ cell trafficking between the myelofibrotic spleen, peripheral blood, and bone marrow (Wang (2015) Exp. Hematol. 43, pp. 100-109).

[0066]

[0068] Disease progression in myelofibrosis is frequently accompanied by greater splenomegaly due to increased extramedullary hematopoiesis. For EMH to occur, a permissive microenvironment must be established that provides the signals necessary for myelofibrotic hematopoietic stem cells to enter the spleen and then initiate and sustain hematopoiesis. Alterations within the splenic microenvironment contribute to abnormal trafficking of myelofibrotic stem cells (MF-SCs), leading to the development of EMH. Splenic myelofibrotic plasma is characterized by increased concentrations of CXCL12, which leads to the initiation and development of EMH in myelofibrosis patients. The spleen cells responsible for excessive production of CXCL12 appear to be endothelial cells. The microenvironments within the bone marrow and spleen differ in myelofibrosis, in part due to increased levels of intact CXCL12 present in the spleen. These distinct microenvironments within the bone marrow and spleen contribute to the sequestration of malignant CD34+ myeloid cells in the spleen and subsequent disease progression (Wang (2015) Exp. Hematol. 43, 100-109).

[0067]

[0069] Reduction of CXCR4 in peripheral blood and bone marrow releases malignant CD34+ cells from the bone marrow, while high CXCL12 levels attract malignant CD34+ myeloid cells to the spleen, while low levels of CXCL12 in peripheral blood promote their sequestration in the spleen. This leads to the accumulation of malignant CD34+ myeloid cells in the spleen and depletion of CD34+ myeloid cells from the bone marrow. Inhibition of BTK reduces the expression and activity of CXCL12, thus reducing the chemoattractant effect of CXCL12, which leads to the accumulation of malignant CD34+ myeloid cells in the spleen. This inhibition of CXCL12 in the spleen leads to the migration of malignant CD34+ myeloid cells from the spleen to peripheral blood, effectively reducing the formation of myelofibrous tissue in the spleen.

[0068]

[0070] The concentration of soluble VCAM-1, a degradation product of VCAM-1, is elevated in the plasma of patients with primary myelofibrosis and correlates with the absolute number of CD34+ cells in the peripheral blood of these patients. Furthermore, CXCR-4 expression by CD34+ cells is downregulated, and plasma CXCL12 levels are elevated, explaining the altered CXCR12 / CXCR-4 interaction that leads to CD34+ cell recruitment. Constitutive recruitment of myelofibrotic HSCs and HPCs is associated with profound alterations in the CXCR4 / CXCL12 axis, resulting from downregulation of CXCR4 expression by myelofibrotic CD34+ cells due to hypermethylation of the CXCR4 promoter and proteolytic degradation of CXCL12 and vascular adhesion molecule-1 (VCAM-1). Drugs targeting proteases involved in constitutive CD34+ cell recruitment offer an interesting strategy for preventing or eliminating the establishment of extramedullary sites of hematopoiesis in patients with primary myelofibrosis.

[0069]

[0071] The migration of leukocytes and progenitor cells is crucial for the process of extramedullary hematopoiesis. As mentioned above, this process is regulated by chemokines such as CXCL12 and is also mediated by integrins LFA1 and VLA4. JAKV617F, but not CALR, stimulates integrin signaling through activation of the small GTPase Rap1, leading to increased granulocyte binding to ICAM-1 and VCAM-1 (abundantly expressed in the spleen). The differential chemotaxis of JAKV617F, coupled with differential integrin binding to CALR-mutant leukocytes, may contribute to extramedullary hematopoiesis. In primary myelofibrosis, the risk of splenomegaly is less pronounced in CALR-mutant patients than in JAKV617F-positive individuals. JAK2V617F kinase mediates JAK2V617F signaling through BTK, PI3K / AKT, PLCγ1, and RhoA, and cooperates with the chemokine CXCL12 to regulate cell migration. This mechanism provides a theoretical basis for the contribution of these downstream molecules to the abnormal cell motility of JAKV617F-positive myeloid progenitor and stem cells that migrate from the bone marrow to peripheral blood and extramedullary organs. Therefore, the use of BTK inhibitors to inhibit the abnormal migration and homing of JAKV617F-positive clones in MPNs is encompassed by the present invention. However, in some embodiments, the presence of JAKV617F-positive clones does not affect treatment with BTK inhibitors. Therefore, BTK inhibitor treatment of human subjects with and without JAKV617F mutations is encompassed by the present invention.

[0070] Methods for treating myelofibrosis

[0072] The present invention relates to a method of treating myelofibrosis, comprising administering to a human in need thereof a BTK inhibitor compound selected from Table 1 or a pharmaceutically acceptable salt thereof. In some embodiments, the MF is primary myelofibrosis, also known as chronic idiopathic myelofibrosis. Primary myelofibrosis is in contrast to myelofibrosis occurring secondary to polycythemia vera or essential thrombocythemia. However, in some embodiments, the present invention encompasses treating myelofibrosis occurring secondary to polycythemia vera or essential thrombocythemia. In some embodiments, the BTK inhibitor is any of the compounds in Table 1 or a pharmaceutically acceptable salt thereof.

[0071]

[0073] The present invention also relates to a method for treating myelofibrosis, comprising administering a BTK inhibitor or a pharmaceutically acceptable salt thereof to a human in need of treatment. In one embodiment, the myelofibrosis is selected from the group consisting of primary myelofibrosis, secondary myelofibrosis, myelofibrosis secondary to polycythemia vera, myelofibrosis secondary to essential thrombocythemia, myelofibrosis secondary to chronic myeloid leukemia, and idiopathic myelofibrosis. In one embodiment, the myelofibrosis is selected from the group consisting of primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of profibrillary / early PMF and fibrotic PMF. In one embodiment, the human is assessed as hydroxyurea intolerant (unacceptable side effects). In one embodiment, the human subject is assessed as hydroxyurea resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy-dependent. In one embodiment, the human subject is phlebotomy-dependent without splenomegaly.

[0072]

[0074] In one embodiment, the human subject is JAK2 inhibitor naive (i.e., has not received therapy with a JAK2 inhibitor). In one embodiment, the human subject is JAK2 inhibitor intolerant. In one embodiment, the human subject is JAK2 inhibitor ineligible due to low platelet count. In one embodiment, the human subject has relapsed after JAK2 inhibitor treatment. In one embodiment, the human subject is refractory to JAK2 inhibitor treatment. In one embodiment, the human subject has failed ruxolitinib or fedratinib therapy. Failure of ruxolitinib or fedratinib therapy includes, but is not limited to, (i) a lack of reduction in the severity or progression of any MPN in a human subject receiving ruxolitinib or fedratinib, or (ii) a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, failure of ruxolitinib or fedratinib therapy is a failure to reduce the severity or progression of any myelofibrosis in a human subject receiving ruxolitinib or fedratinib. In one embodiment, failure of ruxolitinib or fedratinib therapy is a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, the BTK inhibitor is a compound selected from Table 1, and pharmaceutically acceptable salts thereof.

[0073]

[0075] In one embodiment, the invention relates to a method of treating primary myelofibrosis in a human, comprising administering to the human a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from Table 1. In some embodiments, the BTK inhibitor is a covalent or irreversible BTK inhibitor. In some embodiments, the BTK inhibitor is a non-covalent or reversible BTK inhibitor.

[0074]

[0076] In one embodiment, the invention relates to a method of treating post-polycythemia vera myelofibrosis in a human, comprising the step of administering to the human a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from Table 1.

[0075]

[0077] In one embodiment, the invention relates to a method of treating post-essential thrombocythemia myelofibrosis in a human being, comprising the step of administering to the human being a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from any of Table 1.

[0076]

[0078] In one embodiment, a human subject has an accumulation of malignant CD34+ myeloid cells in their spleen. These malignant CD34+ myeloid cells have decreased expression of CXCR4 compared to normal myeloid cells. In one embodiment, a BTK inhibitor is administered in a therapeutically effective amount sufficient to stimulate migration of malignant CD34+ myeloid cells from the human subject's bone marrow or spleen to the peripheral blood. In one embodiment, the BTK inhibitor is administered in an amount sufficient to inactivate VLA-4 on the malignant CD34+ myeloid cells.

[0077]

[0079] In one embodiment, the present invention relates to a method of stimulating migration of malignant CD34+ myeloid cells from the spleen to peripheral blood in a human subject suffering from myelofibrosis, comprising administering a BTK inhibitor to the human subject. In one embodiment, the human subject has an accumulation of malignant CD34+ myeloid cells in their spleen. These malignant CD34+ myeloid cells have decreased expression of CXCR4 compared to normal myeloid cells.

[0078]

[0080] In one embodiment, the present invention relates to a method of treating secondary myelofibrosis, comprising administering a BTK inhibitor to a human in need thereof, wherein the BTK inhibitor is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof, and the secondary myelofibrosis is selected from the group consisting of myelofibrosis secondary to polycythemia vera and myelofibrosis secondary to essential thrombocythemia. In one embodiment, the polycythemia vera is phlebotomy-dependent polycythemia vera. In one embodiment, the human subject is assessed as hydroxyurea-intolerant (unacceptable side effects). In one embodiment, the human subject is assessed as hydroxyurea-resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy-dependent. In one embodiment, the human subject is phlebotomy-dependent without splenomegaly. In one embodiment, the human subject has failed previous MF therapy with ruxolitinib or fedratinib.

[0079]

[0081] In one embodiment, the BTK inhibitor is administered at a dose of 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 560 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg 200 mg BID, 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID. In one embodiment, the BTK inhibitor is administered to a human according to the Dosages and Dosing Regimen section.

[0080]

[0082] In one embodiment, the myelofibrosis is selected from primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the myelofibrosis is selected from primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis, and the human subject has failed ruxolitinib or fedratinib therapy for PMF, post-PV-MF, or post-ET-MF.

[0081]

[0083] In one embodiment, the myelofibrosis is characterized by the presence of a CALR mutation.

[0082]

[0084] In one embodiment, the myelofibrosis is characterized by the presence of an MPL mutation.

[0083]

[0085] In one embodiment, the myelofibrosis is characterized by the JAK2V617F mutation in a human subject.

[0084]

[0086] In one embodiment, the myelofibrosis is characterized by one or more mutations selected from the group consisting of JAK2V617F, MPL, CALR, and combinations thereof.

[0085]

[0087] In one embodiment, the invention relates to a method of treating myelofibrosis secondary to polycythemia vera in a human, comprising the step of administering to the human a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from Table 1.

[0086]

[0088] In one embodiment, the invention relates to a method of treating myelofibrosis secondary to essential thrombocythemia in a human being, comprising the step of administering to the human being a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from Table 1.

[0087]

[0089] In one embodiment, the invention relates to a method of treating myelofibrosis secondary to chronic myeloid leukemia in a human, comprising the step of administering to the human a therapeutically effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof, wherein the BTK inhibitor is a compound selected from Table 1.

[0088]

[0090] In one embodiment, the present invention provides a pharmaceutical composition comprising 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg BID, 200 mg

[0013] A method of treating myelofibrosis in a human, comprising administering to the human a therapeutically effective amount of a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof, at a dosage selected from the group consisting of 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID. In one embodiment, the MF is selected from the group consisting of myelofibrosis, primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of fibrillary / early PMF and fibrotic PMF.

[0089]

[0091] In one embodiment, the present invention provides a pharmaceutical composition comprising 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg BID, 200 mg 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID. The present invention relates to a method of treating myelofibrosis in a human, comprising the step of administering to the human a therapeutically effective amount of a BTK inhibitor compound selected from Table 1 or a pharmaceutically acceptable salt thereof, at a dosage selected from the group consisting of 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID, wherein the MF is selected from the group consisting of MF secondary to polycythemia vera, MF secondary to essential thrombocythemia, and MF secondary to CML.

[0090]

[0092] In one embodiment, the present invention provides a pharmaceutical composition comprising 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg BID, 200 mg 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID.

[0091]

[0093] In one embodiment, the present invention provides a pharmaceutical composition comprising 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg BID, 200 mg 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID.

[0092]

[0094] In one embodiment, the present invention provides a pharmaceutical composition comprising 15 mg QD, 25 mg QD, 30 mg QD, 50 mg QD, 60 mg QD, 75 mg QD, 90 mg QD, 100 mg QD, 120 mg QD, 150 mg QD, 175 mg QD, 180 mg QD, 200 mg QD, 225 mg QD, 240 mg QD, 250 mg QD, 275 mg QD, 300 mg QD, 325 mg QD, 350 mg QD, 360 mg QD, 375 mg QD, 480 mg QD, 15 mg BID, 25 mg BID, 30 mg BID, 50 mg BID, 60 mg BID, 75 mg BID, 90 mg BID, 100 mg BID, 120 mg BID, 150 mg BID, 175 mg BID, 180 mg BID, 200 mg 225 mg BID, 240 mg BID, 250 mg BID, 275 mg BID, 300 mg BID, 325 mg BID, 350 mg BID, 360 mg BID, 375 mg BID, and 480 mg BID.

[0093]

[0095] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating myelofibrosis, comprising administering to a human one or more doses of a BTK inhibitor compound or a pharmaceutically acceptable salt thereof from Table 1. In one embodiment, the MF is selected from the group consisting of primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of fibrillary / early PMF and fibrotic PMF.

[0094]

[0096] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating primary myelofibrosis, wherein treating comprises administering one or more doses of a BTK inhibitor compound from Table 1 or a pharmaceutically acceptable salt thereof to a human being.

[0095]

[0097] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating post-polycythemia vera myelofibrosis, wherein treating comprises administering to a human one or more doses of a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof.

[0096]

[0098] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating post-essential thrombocythemia myelofibrosis, wherein treating comprises administering to a human one or more doses of a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof.

[0097]

[0099] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating myelofibrosis secondary to polycythemia vera, wherein treating comprises administering to a human one or more doses of a BTK inhibitor compound from Table 1 or a pharmaceutically acceptable salt thereof.

[0098]

[0100] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating myelofibrosis secondary to essential thrombocythemia, wherein treating comprises administering to a human one or more doses of a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof.

[0099]

[0101] In one embodiment, the invention relates to the use of a BTK inhibitor or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating myelofibrosis secondary to chronic myeloid leukemia, wherein treating comprises administering to a human one or more doses of a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof.

[0100]

[0102] In one embodiment, the invention relates to the use of a composition comprising a BTK inhibitor selected from Table 1 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating myelofibrosis, comprising administering one or more doses of the composition comprising the BTK inhibitor or a pharmaceutically acceptable salt thereof to a human. In one embodiment, the myelofibrosis is selected from the group consisting of primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of fibrillary / early PMF and fibrotic PMF.

[0101]

[0103] The methods described above may be used as a first-line cancer therapy or following treatment with conventional therapy, including ruxolitinib or fedratinib.

[0102]

[0104] A BTK inhibitor or a pharmaceutically acceptable salt thereof may be used in combination with radiation therapy, hormone therapy, surgery, and immunotherapy, therapies known to those skilled in the art, to treat myelofibrosis selected from the group consisting of primary myelofibrosis, idiopathic myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of fibrillary / early PMF and fibrotic PMF.

[0103] Methods for treating myeloproliferative neoplasms

[0105] The present invention also relates to a method for treating MPN, comprising administering a BTK inhibitor or a pharmaceutically acceptable salt thereof to a human in need thereof. In one embodiment, the MPN is selected from the group consisting of polycythemia vera, myelofibrosis, primary myelofibrosis, thrombocythemia, essential thrombocythemia, idiopathic systemic mastocytosis (SM), chronic neutrophilic leukemia (CNL), chronic eosinophilic leukemia-not otherwise specified (CEL-NOS), myeloproliferative neoplasm, unclassifiable (MPN-U), myelodysplastic syndrome (MDS), and systemic mast cell disease (SMCD). In one embodiment, the MPN is selected from the group consisting of chronic neutrophilic leukemia (CNL), chronic eosinophilic leukemia, chronic myelomonocytic leukemia (CMML), atypical chronic myelogenous leukemia (aCML), juvenile myelomonocytic leukemia (JMML), hypereosinophilic syndrome (HES), and myelodysplastic / myeloproliferative neoplasm with ringed sideroblasts and thrombocytosis (MDS / MPN-RS-T). In one embodiment, the polycythemia vera is phlebotomy-dependent polycythemia vera. In one embodiment, the human is determined to be hydroxyurea-intolerant (unacceptable side effects). In one embodiment, the human subject is determined to be hydroxyurea-resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy-dependent. In one embodiment, the human subject is phlebotomy-dependent without splenomegaly.

[0104]

[0106] In one embodiment, the human subject is JAK2 inhibitor naive (i.e., has not received therapy with a JAK2 inhibitor). In one embodiment, the human subject is JAK2 inhibitor intolerant. In one embodiment, the human subject is JAK2 inhibitor ineligible due to low platelet count. In one embodiment, the human subject has relapsed after JAK2 inhibitor treatment. In one embodiment, the human subject is refractory to JAK2 inhibitor treatment. In one embodiment, the human subject has failed ruxolitinib or fedratinib therapy. Failure of ruxolitinib or fedratinib therapy includes, but is not limited to, (i) a lack of reduction in the severity or progression of any MPN in a human subject receiving ruxolitinib or fedratinib, or (ii) a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, failure of ruxolitinib or fedratinib therapy is a failure to reduce the severity or progression of any myelofibrosis in a human subject receiving ruxolitinib or fedratinib. In one embodiment, failure of ruxolitinib or fedratinib therapy is a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, the BTK inhibitor is a compound selected from Table 1, and pharmaceutically acceptable salts thereof.

[0105]

[0107] In one embodiment, the present invention relates to a method of treating MPN, comprising administering a BTK inhibitor to a human in need thereof, wherein the BTK inhibitor is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof, and the MPN is selected from the group consisting of polycythemia vera and essential thrombocythemia. In one embodiment, the polycythemia vera is phlebotomy-dependent polycythemia vera. In one embodiment, the human subject is assessed as hydroxyurea intolerant (unacceptable side effects). In one embodiment, the human subject is assessed as hydroxyurea resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy-dependent. In one embodiment, the human subject is phlebotomy-dependent without splenomegaly. In one embodiment, the human subject has failed previous MPN therapy with ruxolitinib or fedratinib.

[0106]

[0108] The present invention also relates to a method for treating blast phase MPN (MPN-BP), comprising administering a BTK inhibitor or a pharmaceutically acceptable salt thereof to a human in need of treatment. In one embodiment, the MPN-BP is selected from the group consisting of blast phase polycythemia vera (BP-PV), blast phase myelofibrosis, blast phase thrombocythemia, blast phase essential thrombocythemia (BP-ET), blast phase systemic mastocytosis (BP-SM), blast phase chronic neutrophilic leukemia (BP-CNL), blast phase myelodysplastic syndrome (BP-MDS), and blast phase systemic mast cell disease (BP-SMCD). In one embodiment, the MPN-BP is selected from the group consisting of blastic phase chronic neutrophilic leukemia (BP-CNL), blastic phase chronic eosinophilic leukemia, blastic phase chronic myelomonocytic leukemia (BP-CMML), blastic phase atypical chronic myelogenous leukemia (BP-aCML), blastic phase juvenile myelomonocytic leukemia (BP-JMML), blastic phase hypereosinophilic syndrome (BP-HES), and blastic phase myelodysplastic / myeloproliferative neoplasm with ringed sideroblasts and thrombocytosis (BP-MDS / MPN-RS-T). In one embodiment, the blastic phase polycythemia vera is phlebotomy-dependent polycythemia vera. In one embodiment, the human is characterized as hydroxyurea intolerant (unacceptable side effects). In one embodiment, the human subject is characterized as hydroxyurea resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy dependent. In one embodiment, the human subject is phlebotomy dependent without splenomegaly.

[0107]

[0109] In one embodiment, the human subject is JAK2 inhibitor naive (i.e., has not received therapy with a JAK2 inhibitor). In one embodiment, the human subject is JAK2 inhibitor intolerant. In one embodiment, the human subject is JAK2 inhibitor ineligible due to low platelet count. In one embodiment, the human subject has relapsed after JAK2 inhibitor treatment. In one embodiment, the human subject is refractory to JAK2 inhibitor treatment. In one embodiment, the human subject has failed ruxolitinib or fedratinib therapy. Failure of ruxolitinib or fedratinib therapy includes, but is not limited to, (i) a lack of reduction in the severity or progression of any MPN in a human subject receiving ruxolitinib or fedratinib, or (ii) a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, failure of ruxolitinib or fedratinib therapy is a failure to reduce the severity or progression of any myelofibrosis in a human subject receiving ruxolitinib or fedratinib. In one embodiment, failure of ruxolitinib or fedratinib therapy is a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, the BTK inhibitor is a compound selected from Table 1 and pharmaceutically acceptable salts thereof. In one embodiment, the BTK inhibitor is administered to a human according to the Dosages and Dosage Regimen section.

[0108]

[0110] In one embodiment, the MPN is characterized by a CALR mutation.

[0109]

[0111] In one embodiment, the MPN is characterized by an MPL mutation.

[0110]

[0112] In one embodiment, the MPN is characterized by the JAK2V617F mutation.

[0111]

[0113] In one embodiment, the MPN is characterized by one or more mutations selected from the group consisting of JAK2V617F, MPL, CALR, and mixtures thereof.

[0112] Combination with BET inhibitors

[0114] Bromodomain and extraterminal domain (BET) proteins are transcriptional regulators required for efficient expression of several growth-promoting, anti-apoptotic genes and cell cycle progression. The BET family includes BRD2, BRD3, BRD4, and BRDT. During transcription, BET proteins are targeted to chromatin via their N-terminal bromodomain (BRD), which recognizes acetylated lysine residues in histones H3 and H4. BET inhibitors disrupt this BET-histone interaction and subsequently downregulate the transcription of oncogenes, including MYC.

[0113]

[0115] MYC and BTK are key regulators of cellular processes and tumor progression. Given the relationship between BET inhibition and MYC downregulation, and the relationship between MYC overexpression and cancer, BET inhibitors are useful for treating MYC-associated diseases. Furthermore, BET inhibitors in combination with BTK inhibitors are useful for treating myeloid cell disorders.

[0114]

[0116] In some embodiments, the present invention relates to a method for treating splenomegaly, extramedullary hematopoiesis, or fibrosis by administering an effective amount of a BTK inhibitor in combination with a BET inhibitor to a human subject in need of such treatment. The disclosure herein demonstrates that desirable therapeutic agents can be selected based on BTK inhibition and BET inhibition. Therefore, without wishing to be bound by a particular mechanism of action, it is expected that BTK inhibition in combination with BET inhibition will alter one or more downstream signaling components (e.g., CXCR-4, CXCL12, VLA-4, VCAM-1) to recruit CD34+ cell migration to the peripheral blood, and will be useful in treating complications associated with myelofibrosis, particularly in treating or preventing one or more myelofibrosis complications, including, but not limited to, splenomegaly, extramedullary hematopoiesis, or fibrosis.

[0115]

[0117] Thus, the present invention relates to a method of treating splenomegaly, extramedullary hematopoiesis, or fibrosis, comprising administering to a human in need thereof a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof, in combination with a BET inhibitor compound selected from Table 2, or a pharmaceutically acceptable salt thereof. In some embodiments, the splenomegaly is secondary to myelofibrosis. In some embodiments, the splenomegaly is associated with primary myelofibrosis, also known as chronic idiopathic myelofibrosis. Primary myelofibrosis is in contrast to myelofibrosis occurring secondary to polycythemia vera or essential thrombocythemia. However, in some embodiments, the present invention encompasses treating splenomegaly due to myelofibrosis occurring secondary to polycythemia vera or essential thrombocythemia. In some embodiments, the BTK inhibitor is a compound selected from Table 1, or a pharmaceutically acceptable salt thereof, in combination with a BET inhibitor compound selected from Table 2, or a pharmaceutically acceptable salt thereof. In one embodiment, the human is determined to be hydroxyurea intolerant (unacceptable side effect). In one embodiment, the human subject is determined to be hydroxyurea resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy dependent. In one embodiment, the human subject is phlebotomy dependent without splenomegaly.

[0116]

[0118] In one embodiment, the human subject is JAK2 inhibitor naive (i.e., has not received therapy with a JAK2 inhibitor). In one embodiment, the human subject is JAK2 inhibitor intolerant. In one embodiment, the human subject is JAK2 inhibitor ineligible due to low platelet count. In one embodiment, the human subject has relapsed after JAK2 inhibitor treatment. In one embodiment, the human subject is refractory to JAK2 inhibitor treatment. In one embodiment, the human subject has failed ruxolitinib or fedratinib therapy. Failure of ruxolitinib or fedratinib therapy includes, but is not limited to, (i) a lack of reduction in the severity or progression of any MPN in a human subject receiving ruxolitinib or fedratinib, or (ii) a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, failure of ruxolitinib or fedratinib therapy is a failure to reduce the severity or progression of any myelofibrosis in a human subject receiving ruxolitinib or fedratinib. In one embodiment, failure of ruxolitinib or fedratinib therapy is a relapse of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, the BTK inhibitor is a compound selected from Table 1 and pharmaceutically acceptable salts thereof. In one embodiment, the human is suffering from splenomegaly, extramedullary hematopoiesis, or fibrosis characterized by one or more mutations selected from the group consisting of JAK2V617F, MPL, CALR, and combinations thereof.

[0117] [Table 2]

[0118]

[0119] The present invention encompasses a method of treating myelofibrosis, comprising administering to a human in need thereof a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof, in combination with a BET inhibitor selected from Table 2, or a pharmaceutically acceptable salt thereof. In some embodiments, the myelofibrosis is primary myelofibrosis, also known as chronic idiopathic myelofibrosis. Primary myelofibrosis is in contrast to myelofibrosis occurring secondary to polycythemia vera or essential thrombocythemia. However, in some embodiments, the present invention encompasses treating myelofibrosis occurring secondary to polycythemia vera or essential thrombocythemia. In some embodiments, the BTK inhibitor is a compound selected from Table 1, or a pharmaceutically acceptable salt thereof, in combination with a BET inhibitor compound selected from Table 2, or a pharmaceutically acceptable salt thereof. In one embodiment, the human is assessed as hydroxyurea intolerant (unacceptable side effects). In one embodiment, the human subject is assessed as hydroxyurea resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy dependent. In one embodiment, the human subject is phlebotomy dependent without splenomegaly.

[0119]

[0120] In one embodiment, the human subject is JAK2 inhibitor naive (i.e., has not received therapy with a JAK2 inhibitor). In one embodiment, the human subject is JAK2 inhibitor intolerant. In one embodiment, the human subject is JAK2 inhibitor ineligible due to low platelet count. In one embodiment, the human subject has relapsed after JAK2 inhibitor treatment. In one embodiment, the human subject is refractory to JAK2 inhibitor treatment. In one embodiment, the human subject has failed ruxolitinib or fedratinib therapy. Failure of ruxolitinib or fedratinib therapy includes, but is not limited to, (i) a lack of reduction in the severity or progression of any MPN in a human subject receiving ruxolitinib or fedratinib, or (ii) a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, failure of ruxolitinib or fedratinib therapy is a failure to reduce the severity or progression of any myelofibrosis in a human subject receiving ruxolitinib or fedratinib. In one embodiment, failure of ruxolitinib or fedratinib therapy is a relapse of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, the BTK inhibitor is a compound selected from Table 1 and pharmaceutically acceptable salts thereof. In one embodiment, the human is suffering from splenomegaly, extramedullary hematopoiesis, or fibrosis characterized by one or more mutations selected from the group consisting of JAK2V617F, MPL, CALR, and combinations thereof.

[0120]

[0121] In some embodiments, the BET inhibitor is administered once daily, twice daily, three times daily, four times daily, or five times daily. In some embodiments, the BTK inhibitor is administered at a dosage of about 40 mg / day to about 1000 mg / day. In some embodiments, the BTK inhibitor is administered orally. In some embodiments, the BTK inhibitor and the BET inhibitor are administered simultaneously, sequentially, or intermittently. Doses and administration for the BET inhibitor are as described herein.

[0121]

[0122] In some embodiments, the present invention encompasses a pharmaceutical combination comprising (a) a BTK inhibitor; (b) a BET inhibitor; and (c) a pharmaceutically acceptable excipient. In some embodiments, the combination provides a synergistic therapeutic effect compared to administration of the BTK inhibitor or the BET inhibitor alone. In some embodiments, the combination sensitizes myelofibrosis to the BTK inhibitor. In some embodiments, the BET inhibitor is a compound selected from Table 2 or a pharmaceutically acceptable salt thereof. In some embodiments, the BTK inhibitor is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof. In some embodiments, the combination is in a combined dosage form. In some embodiments, the combination is in separate dosage forms.

[0122]

[0123] In some embodiments, the invention encompasses the use of a therapeutically effective amount of a combination comprising a BTK inhibitor and a BET inhibitor to treat splenomegaly, extramedullary hematopoiesis, or fibrosis in a human subject in need thereof. In some embodiments, the combination provides a synergistic therapeutic effect compared to administration of the BTK inhibitor or the BET inhibitor alone. In some embodiments, the combination sensitizes malignant CD34+ myeloid cells to the BTK inhibitor. In some embodiments, the BET inhibitor comprises a compound from Table 2 or a pharmaceutically acceptable salt thereof. In some embodiments, the splenomegaly, extramedullary hematopoiesis, or fibrosis is associated with primary myelofibrosis, also known as chronic idiopathic myelofibrosis. Primary myelofibrosis is in contrast to myelofibrosis occurring secondary to polycythemia vera or essential thrombocythemia. However, in some embodiments, the invention encompasses treating splenomegaly, extramedullary hematopoiesis, or fibrosis occurring secondary to polycythemia vera or essential thrombocythemia. In one embodiment, the myelofibrosis is selected from the group consisting of primary myelofibrosis, secondary myelofibrosis, myelofibrosis secondary to polycythemia vera, myelofibrosis secondary to essential thrombocythemia, myelofibrosis secondary to chronic myeloid leukemia, and idiopathic myelofibrosis. In one embodiment, the myelofibrosis is selected from the group consisting of primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis. In one embodiment, the primary myelofibrosis is selected from the group consisting of fibrillary / early PMF and fibrotic PMF. In one embodiment, the human is assessed as hydroxyurea intolerant (unacceptable side effects). In one embodiment, the human subject is assessed as hydroxyurea resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy-dependent. In one embodiment, the human subject is phlebotomy-dependent without splenomegaly.

[0123]

[0124] In one embodiment, the human subject is JAK2 inhibitor naive (i.e., has not received therapy with a JAK2 inhibitor). In one embodiment, the human subject is JAK2 inhibitor intolerant. In one embodiment, the human subject is JAK2 inhibitor ineligible due to low platelet count. In one embodiment, the human subject has relapsed after JAK2 inhibitor treatment. In one embodiment, the human subject is refractory to JAK2 inhibitor treatment. In one embodiment, the human subject has failed ruxolitinib or fedratinib therapy. Failure of ruxolitinib or fedratinib therapy includes, but is not limited to, (i) a lack of reduction in the severity or progression of any MPN in a human subject receiving ruxolitinib or fedratinib, or (ii) a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, failure of ruxolitinib or fedratinib therapy is a failure to reduce the severity or progression of any myelofibrosis in a human subject receiving ruxolitinib or fedratinib. In one embodiment, failure of ruxolitinib or fedratinib therapy is a recurrence of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, the BTK inhibitor is a compound selected from Table 1, and pharmaceutically acceptable salts thereof.

[0124]

[0125] In some embodiments, the invention encompasses the use of a therapeutically effective amount of a combination comprising a BTK inhibitor and a BET inhibitor to treat myelofibrosis, comprising administering a therapeutically effective amount of the combination comprising a BTK inhibitor and a BET inhibitor to a human subject in need of treatment. In some embodiments, the combination provides a synergistic therapeutic effect compared to administration of the BTK inhibitor or the BET inhibitor alone. In some embodiments, the combination sensitizes myelofibrosis to the BTK inhibitor. In some embodiments, the BET inhibitor is a compound selected from Table 2 or a pharmaceutically acceptable salt thereof. In some embodiments, the BTK inhibitor is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof. In some embodiments, the myelofibrosis is primary myelofibrosis, also known as chronic idiopathic myelofibrosis. Primary myelofibrosis is in contrast to myelofibrosis occurring secondary to polycythemia vera or essential thrombocythemia. However, in some embodiments, the invention encompasses treating splenomegaly due to myelofibrosis occurring secondary to polycythemia vera or essential thrombocythemia. In some embodiments, the BTK inhibitor is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof in combination with a compound selected from Table 2 or a pharmaceutically acceptable salt thereof. In one embodiment, the human is determined to be hydroxyurea intolerant (unacceptable side effects). In one embodiment, the human subject is determined to be hydroxyurea resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy-dependent. In one embodiment, the human subject is phlebotomy-dependent without splenomegaly. In one embodiment, the human subject is JAK2 inhibitor naive (i.e., has not received therapy with a JAK2 inhibitor). In one embodiment, the human subject is JAK2 inhibitor intolerant. In one embodiment, the human subject is JAK2 inhibitor ineligible due to low platelet count. In one embodiment, the human subject has relapsed after JAK2 inhibitor treatment. In one embodiment, the human subject is refractory to JAK2 inhibitor treatment. In one embodiment, the human subject has failed ruxolitinib or fedratinib therapy.Failure of ruxolitinib or fedratinib therapy includes, but is not limited to, (i) a failure to reduce the severity or progression of any MPN in a human subject receiving ruxolitinib or fedratinib, or (ii) a relapse of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, failure of ruxolitinib or fedratinib therapy is a failure to reduce the severity or progression of any myelofibrosis in a human subject receiving ruxolitinib or fedratinib. In one embodiment, failure of ruxolitinib or fedratinib therapy is a relapse of any myelofibrosis in a human subject after ruxolitinib or fedratinib therapy. In one embodiment, the BTK inhibitor is a compound selected from Table 1 and pharmaceutically acceptable salts thereof. In one embodiment, the human is afflicted with splenomegaly, extramedullary hematopoiesis, or fibrosis characterized by one or more mutations selected from the group consisting of JAK2V617F, MPL, CALR, and combinations thereof.

[0125] Pharmaceutical Composition

[0126] In some embodiments, the present invention provides a pharmaceutical composition comprising a BTK inhibitor compound selected from Table 1 or a pharmaceutically acceptable salt thereof for the treatment of myelofibrosis. In some embodiments, the present invention provides a pharmaceutical composition comprising a BTK inhibitor compound selected from Table 1 or a pharmaceutically acceptable salt thereof for the treatment of myelofibrosis, primary myelofibrosis, or idiopathic myelofibrosis. In one embodiment, the myelofibrosis is selected from primary myelofibrosis, post-polycythemia vera myelofibrosis, and post-essential thrombocythemia myelofibrosis.

[0126]

[0127] In some embodiments, the present invention provides a pharmaceutical composition comprising a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof, for treating polycythemia vera, essential thrombocythemia, or myelofibrosis secondary to chronic myeloid leukemia.

[0127]

[0128] Pharmaceutical compositions are typically formulated to provide a therapeutically effective amount of a BTK inhibitor compound selected from Table 1 or a pharmaceutically acceptable salt thereof. If desired, the pharmaceutical composition contains a pharmaceutically acceptable salt and / or coordination complex thereof, as well as one or more pharmaceutically acceptable excipients, carriers including inert solid diluents and fillers, diluents including sterile aqueous solutions and various organic solvents, permeation enhancers, solubilizers, and adjuvants. If desired, other ingredients in addition to the BTK inhibitor or a pharmaceutically acceptable salt thereof may be mixed into the preparation, or both components may be formulated in separate preparations for use separately or in combination simultaneously.

[0128]

[0129] In selected embodiments, the concentration of the BTK inhibitor or a pharmaceutically acceptable salt thereof provided in the pharmaceutical compositions of the invention may be, for example, 100%, 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.4%, 0.3%, 0.2%, 0.1%, 0.09%, 0.08%, or 100%. %, 0.07%, 0.06%, 0.05%, 0.04%, 0.03%, 0.02%, 0.01%, 0.009%, 0.008%, 0.007%, 0.006%, 0.005%, 0.004%, 0.003%, 0.002%, 0.001%, 0.0009%, 0.0008%, 0.0007%, 0.0006%, 0.0005%, 0.0004%, 0.0003%, 0.0002%, or 0.0001% w / w, w / v, or v / v.

[0129]

[0130] In selected embodiments, the concentration of the BTK inhibitor or a pharmaceutically acceptable salt thereof provided in the pharmaceutical compositions of the invention is independently 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 19.75%, 19.50%, 19.25%, 19%, 18.75%, 18.50%, 18.25%, 18%, 17.75%, 17.50%, 17.25%, 17%, 16.75%, 16.50%, 16.25%, 1 6%, 15.75%, 15.50%, 15.25%, 15%, 14.75%, 14.50%, 14.25%, 14%, 13.75%, 13.50%, 13.25%, 13%, 12.75%, 12.50%, 12.25%, 12%, 11.75%, 11.50%, 11.25%, 11%, 10.75%, 10.50%, 10.25%, 10%, 9.75%, 9.50%, 9.25%, 9%, 8.75%, 8.50%, 8.25% ,8%,7.75%,7.50%,7.25%,7%,6.75%,6.50%,6.25%,6%,5.75%,5.50%,5.25%,5%,4.75%,4.50%,4.25%,4%,3.75%,3.50%,3.25%,3%,2.75%,2.50%,2.25%,2%,1.75%,1.50%,125%,1%,0.5%,0.4%,0.3%,0.2%,0.1%,0.09%,0.08%,0.07 %, 0.06%, 0.05%, 0.04%, 0.03%, 0.02%, 0.01%, 0.009%, 0.008%, 0.007%, 0.006%, 0.005%, 0.004%, 0.003%, 0.002%, 0.001%, 0.0009%, 0.0008%, 0.0007%, 0.0006%, 0.0005%, 0.0004%, 0.0003%, 0.0002%, or 0.0001% w / w, w / v, or v / v.

[0130]

[0131] In selected embodiments, the concentration of the BTK inhibitor or a pharmaceutically acceptable salt thereof is independently between about 0.0001% and about 50%, between about 0.001% and about 40%, between about 0.01% and about 30%, between about 0.02% and about 29%, between about 0.03% and about 28%, between about 0.04% and about 27%, between about 0.05% and about 26%, between about 0.06% and about 25%, between about 0.07% and about 24%, between about 0. The range of values ​​is between about 0.08% and about 23%, between about 0.09% and about 22%, between about 0.1% and about 21%, between about 0.2% and about 20%, between about 0.3% and about 19%, between about 0.4% and about 18%, between about 0.5% and about 17%, between about 0.6% and about 16%, between about 0.7% and about 15%, between about 0.8% and about 14%, between about 0.9% and about 12%, or between about 1% and about 10% w / w, w / v, or v / v.

[0131]

[0132] In selected embodiments, the concentration of the BTK inhibitor or a pharmaceutically acceptable salt thereof is independently in the range of about 0.001% to about 10%, about 0.01% to about 5%, about 0.02% to about 4.5%, about 0.03% to about 4%, about 0.04% to about 3.5%, about 0.05% to about 3%, about 0.06% to about 2.5%, about 0.07% to about 2%, about 0.08% to about 1.5%, about 0.09% to about 1%, or about 0.1% to about 0.9% w / w, w / v, or v / v.

[0132]

[0133] In selected embodiments, the amount of BTK inhibitor or a pharmaceutically acceptable salt thereof may be independently 10 g, 9.5 g, 9.0 g, 8.5 g, 8.0 g, 7.5 g, 7.0 g, 6.5 g, 6.0 g, 5.5 g, 5.0 g, 4.5 g, 4.0 g, 3.5 g, 3.0 g, 2.5 g, 2.0 g, 1.5 g, 1.0 g, 0.95 g, 0.9 g, 0.85 g, 0.8 g, 0.75 g, 0.7 g, 0.65 g, 0.6 g, 0.55 g, 0.5 g, 0.45 g, 0.4 g, 0.35 g, 0.3 g, 0.25 g, and is equal to or less than 0.2g, 0.15g, 0.1g, 0.09g, 0.08g, 0.07g, 0.06g, 0.05g, 0.04g, 0.03g, 0.02g, 0.01g, 0.009g, 0.008g, 0.007g, 0.006g, 0.005g, 0.004g, 0.003g, 0.002g, 0.001g, 0.0009g, 0.0008g, 0.0007g, 0.0006g, 0.0005g, 0.0004g, 0.0003g, 0.0002g, or 0.0001g.

[0133]

[0134] In selected embodiments, the amount of BTK inhibitor or a pharmaceutically acceptable salt thereof is independently 0.0001 g, 0.0002 g, 0.0003 g, 0.0004 g, 0.0005 g, 0.0006 g, 0.0007 g, 0.0008 g, 0.0009 g, 0.001 g, 0.0015 g, 0.002 g, , 0.0025g, 0.003g, 0.0035g, 0.004g, 0.0045g, 0.005g, 0.0055g, 0.006g, 0.0065g , 0.007g, 0.0075g, 0.008g, 0.0085g, 0.009g, 0.0095g, 0.01g, 0.015g, 0.02g, 0.02 5g, 0.03g, 0.035g, 0.04g, 0.045g, 0.05g, 0.055g, 0.06g, 0.065g, 0.07g, 0.075g, 0.08g, 0.085g, 0.09g, 0.095g, 0.1g, 0.15g, 0.2g, 0.25g, 0.3g, 0.35g, 0.4g, 0.45g , 0.5g, 0.55g, 0.6g, 0.65g, 0.7g, 0.75g, 0.8g, 0.85g, 0.9g, 0.95g, 1g, 1.5g, 2g, 2.5, 3g, 3.5, 4g, 4.5g, 5g, 5.5g, 6g, 6.5g, 7g, 7.5g, 8g, 8.5g, 9g, 9.5g, or more than 10g.

[0134]

[0135] The BTK inhibitor compounds and their pharmaceutically acceptable salts in Table 1 are effective over a wide dosage range. For example, in the treatment of adult humans, dosages ranging from 0.01 to 1000 mg per day, 0.5 to 100 mg per day, 1 to 50 mg per day, and 5 to 40 mg per day, independently, are examples of dosages that may be used. The exact dosage will depend on the route of administration, the form in which the compound is administered, the sex and age of the subject to be treated, the weight of the subject to be treated, and the preference and experience of the attending physician.

[0135]

[0136] Non-limiting exemplary pharmaceutical compositions and methods for preparing pharmaceutical compositions are described below.

[0136] Pharmaceutical Composition for Oral Administration

[0137] In selected embodiments, the present invention provides a pharmaceutical composition for oral administration comprising a BTK inhibitor compound selected from Table 1, or a pharmaceutically acceptable salt thereof, and a pharmaceutical excipient suitable for oral administration.

[0137]

[0138] In selected embodiments, the present invention provides a solid pharmaceutical composition for oral administration containing (i) an effective amount of a BTK inhibitor or a pharmaceutically acceptable salt thereof in combination and (ii) a pharmaceutical excipient suitable for oral administration. In selected embodiments, the composition further contains (iii) an effective amount of at least one additional active ingredient.

[0138]

[0139] In selected embodiments, the pharmaceutical composition may be a liquid pharmaceutical composition suitable for oral consumption. Pharmaceutical compositions of the present invention suitable for oral administration may be presented as discrete dosage forms such as capsules, cachets, or tablets, or as liquids or aerosol sprays, each containing a predetermined amount of the active ingredient in an aqueous or non-aqueous liquid, an oil-in-water emulsion, or a water-in-oil liquid emulsion, as a powder, or in the form of granules, solution, or suspension. Such dosage forms may be prepared by any method, but all methods include the step of bringing the active ingredient(s) into association with a carrier that constitutes one or more necessary ingredients. Generally, the compositions are prepared by uniformly and intimately admixing the active ingredient(s) with a liquid carrier or finely divided solid carrier, or both, and then, if necessary, shaping the product into the desired presentation. For example, a tablet may be prepared by compression or molding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredients in a free-flowing form such as powder or granules, optionally mixed with excipients such as, but not limited to, binders, lubricants, inert diluents, and / or surface active agents or dispersing agents. Molded tablets may be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent.

[0139]

[0140] Because water can promote the degradation of some compounds, the present invention further encompasses anhydrous pharmaceutical compositions and dosage forms. For example, in the pharmaceutical arts, water can be added (e.g., 5%) as a means of simulating long-term storage to determine characteristics such as shelf life or the stability of a formulation over time. Anhydrous pharmaceutical compositions and dosage forms of the present invention can be prepared using anhydrous or low-moisture containing ingredients and low-moisture or low-humidity conditions. Pharmaceutical compositions and dosage forms of the present invention containing lactose can be made anhydrous if substantial contact with moisture and / or humidity is expected during manufacturing, packaging, and / or storage. Anhydrous pharmaceutical compositions can be prepared and stored such that the anhydrous nature of the anhydrous pharmaceutical composition is maintained. Thus, anhydrous compositions can be packaged using materials known to prevent exposure to water, allowing the anhydrous compositions to be included in suitable formulary kits. Examples of suitable packaging include, but are not limited to, hermetically sealed foils, plastics, unit-dose containers, blister packs, and strip packs.

[0140]

[0141] The BTK inhibitor or a pharmaceutically acceptable salt thereof can be combined in intimate admixture with a pharmaceutical carrier according to conventional pharmaceutical compounding techniques. Carriers can take a wide variety of forms, depending on the form of preparation desired for administration. When preparing compositions for oral dosage forms, any of the usual pharmaceutical media, such as water, glycols, oils, alcohols, flavoring agents, preservatives, coloring agents, and the like, can be employed as the carrier for oral liquid preparations (such as suspensions, solutions, and elixirs) or aerosols; or, for oral solid preparations, in some embodiments, lactose can be omitted, and carriers such as starch, sugars, microcrystalline cellulose, diluents, granulating agents, lubricants, binders, and disintegrants can be used. For example, for solid oral preparations, suitable carriers include powders, capsules, and tablets. If desired, tablets can be coated using standard aqueous or nonaqueous techniques.

[0141]

[0142] Binders suitable for use in pharmaceutical compositions and dosage forms include, but are not limited to, corn starch, potato starch, or other starches, gelatin, natural and synthetic gums such as acacia, sodium alginate, alginic acid, other alginates, powdered tragacanth, guar gum, cellulose and its derivatives (e.g., ethyl cellulose, cellulose acetate, calcium carboxymethylcellulose, sodium carboxymethylcellulose), polyvinylpyrrolidone, methylcellulose, pregelatinized starch, hydroxypropyl methylcellulose, microcrystalline cellulose, and mixtures thereof.

[0142]

[0143] Examples of suitable fillers for use in the pharmaceutical compositions and dosage forms disclosed herein include, but are not limited to, talc, calcium carbonate (e.g., granules or powder), microcrystalline cellulose, powdered cellulose, dextrates, kaolin, mannitol, silicic acid, sorbitol, starch, pregelatinized starch, and mixtures thereof.

[0143]

[0144] Disintegrants can be used in the compositions of the present invention to provide tablets that disintegrate when exposed to an aqueous environment. Excessive disintegrant can result in tablets that disintegrate in the bottle. Too little can be insufficient for disintegration to occur, thereby altering the rate and extent of release of the active ingredient from the dosage form. Thus, dosage forms of the compounds disclosed herein can be formed using a sufficient amount of disintegrant that is neither too little nor too much to adversely alter the release of the active ingredient(s). The amount of disintegrant used can vary based on the type of formulation and mode of administration and can be readily discerned by one of ordinary skill in the art. About 0.5 to about 15 weight percent of disintegrant, or about 1 to about 5 weight percent of disintegrant, can be used in the pharmaceutical composition. Disintegrants that can be used to form pharmaceutical compositions and dosage forms of the invention include, but are not limited to, agar, alginic acid, calcium carbonate, microcrystalline cellulose, croscarmellose sodium, crospovidone, polacrilin potassium, sodium starch glycolate, potato or tapioca starch, other starches, pregelatinized starch, other starches, clays, other algins, other celluloses, gums, or mixtures thereof.

[0144]

[0145] Lubricants that can be used to form pharmaceutical compositions and dosage forms of the present invention include, but are not limited to, calcium stearate, magnesium stearate, mineral oil, light mineral oil, glycerin, sorbitol, mannitol, polyethylene glycol, other glycols, stearic acid, sodium lauryl sulfate, talc, hydrogenated vegetable oils (e.g., peanut oil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, and soybean oil), zinc stearate, ethyl oleate, ethyl laurate, agar, or mixtures thereof. Additional lubricants include, for example, syloid silica gel, coagulated aerosol of synthetic silica, or mixtures thereof. Lubricants can optionally be added in an amount of less than about 1 weight percent of the pharmaceutical composition.

[0145]

[0146] When aqueous suspensions and / or elixirs are desired for oral administration, the essential active ingredients in the aqueous suspensions and / or elixirs may be combined with diluents such as water, ethanol, propylene glycol, glycerin, and various combinations thereof, as well as various sweetening or flavoring agents, coloring substances, or dyes, and, if desired, emulsifying and / or suspending agents.

[0146]

[0147] Tablets can be uncoated or coated by known techniques to delay disintegration and absorption in the gastrointestinal tract, thereby providing a sustained action over a longer period. For example, a time-delay material such as glyceryl monostearate or glyceryl distearate can be employed. Formulations for oral use can be presented as hard gelatin capsules in which the active ingredient is mixed with an inert solid diluent, such as calcium carbonate, calcium phosphate, or kaolin, or as soft gelatin capsules in which the active ingredient is mixed with water or an oil medium, such as peanut oil, liquid paraffin, or olive oil.

[0147]

[0148] Surfactants that can be used to form the pharmaceutical compositions and dosage forms of the present invention include, but are not limited to, hydrophilic surfactants, lipophilic surfactants, and mixtures thereof, i.e., a mixture of hydrophilic surfactants can be employed, a mixture of lipophilic surfactants can be employed, or a mixture of at least one hydrophilic surfactant and at least one lipophilic surfactant can be employed.

[0148]

[0149] Suitable hydrophilic surfactants may generally have an HLB value of at least 10, while suitable lipophilic surfactants may generally have an HLB value of about 10 or less. The hydrophilic-lipophilic balance ("HLB" value) is an empirical parameter used to characterize the relative hydrophilicity and hydrophobicity of nonionic amphiphilic compounds. Surfactants with lower HLB values ​​are more lipophilic or hydrophobic and have greater solubility in oils, while surfactants with higher HLB values ​​are more hydrophilic and have greater solubility in aqueous solutions. Hydrophilic surfactants are generally considered to be those compounds with an HLB value greater than about 10, as well as anionic, cationic, or zwitterionic compounds for which the HLB scale is not generally applicable. Similarly, lipophilic (i.e., hydrophobic) surfactants are compounds with an HLB value of about 10 or less. However, the HLB value of a surfactant is merely a rough guideline commonly used to enable the formulation of industrial, pharmaceutical, and cosmetic emulsions.

[0149]

[0150] The hydrophilic surfactant may be either ionic or nonionic. Suitable ionic surfactants include, but are not limited to, alkylammonium salts, fusidate salts, fatty acid derivatives of amino acids, oligopeptides, and polypeptides, glyceride derivatives of amino acids, oligopeptides, and polypeptides, lecithin and hydrogenated lecithin, lysolecithin and hydrogenated lysolecithin, phospholipids and derivatives thereof, lysophospholipids and derivatives thereof, carnitine fatty acid ester salts, salts of alkyl sulfates, fatty acid salts, docusate sodium, acylactylates, mono- and di-acetylated tartaric acid esters of mono- and di-glycerides, succinylated mono- and di-glycerides, citrate esters of mono- and di-glycerides, and mixtures thereof.

[0150]

[0151] Of the aforementioned group, ionic surfactants include, by way of example, lecithin, lysolecithin, phospholipids, lysophospholipids, and derivatives thereof; carnitine fatty acid ester salts; salts of alkyl sulfates; fatty acid salts; sodium docusate; acyl lactylates; mono- and di-acetylated tartaric acid esters of mono- and di-glycerides; succinylated mono- and di-glycerides; citrate esters of mono- and di-glycerides; and mixtures thereof.

[0151]

[0152] Ionic surfactants include lecithin, lysolecithin, phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, phosphatidic acid, phosphatidylserine, lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylglycerol, lysophosphatidic acid, lysophosphatidylserine, PEG-phosphatidylethanolamine, PVP-phosphatidylethanolamine, and lactylic acid esters of fatty acids (lactylic ester), stearoyl-2-lactylate, stearoyl lactylate, succinylated monoglycerides, mono / diacetylated tartaric acid esters of mono / diglycerides, citrate esters of mono / diglycerides, cholylsarcosine, caproate, caprylate, caprate, laurate, myristate, palmitate, oleate, ricinoleate, linolenate, linolenate, stearate, lauryl sulfate, teracecyl sulfate, docusate, lauroylcarnitine, palmitoylcarnitine, myristoylcarnitine, and salts and mixtures thereof.

[0152]

[0153] The hydrophilic nonionic surfactant may include, but is not limited to, alkyl glucosides; alkyl maltosides; alkyl thioglucosides; lauryl macrogol glycerides; polyoxyalkylene alkyl ethers such as polyethylene glycol alkyl ethers; polyoxyalkylene alkylphenols such as polyethylene glycol alkylphenols; polyoxyalkylene alkylphenol fatty acid esters such as polyethylene glycol fatty acid monoesters and polyethylene glycol fatty acid diesters; polyethylene glycol glycerol fatty acid esters; polyglycerol fatty acid esters; polyoxyalkylene sorbitan fatty acid esters such as polyethylene glycol sorbitan fatty acid esters; hydrophilic transesterification products of polyols with at least one member of the group consisting of glycerides, vegetable oils, hydrogenated vegetable oils, fatty acids, and sterols; polyoxyethylene sterols, derivatives, and analogs thereof; polyoxyethylated vitamins and derivatives thereof; polyoxyethylene-polyoxypropylene block copolymers; and mixtures thereof; polyethylene glycol sorbitan fatty acid esters and hydrophilic transesterification products of polyols with at least one member of the group consisting of triglycerides, vegetable oils, and hydrogenated vegetable oils. The polyol may be glycerol, ethylene glycol, polyethylene glycol, sorbitol, propylene glycol, pentaerythritol, or a saccharide.

[0153]

[0154] Other hydrophilic nonionic surfactants include, but are not limited to, PEG-10 laurate, PEG-12 laurate, PEG-20 laurate, PEG-32 laurate, PEG-32 dilaurate, PEG-12 oleate, PEG-15 oleate, PEG-20 oleate, PEG-20 dioleate, PEG-32 oleate, PEG-200 oleate, PEG-400 oleate, PEG-15 stearate, PEG-32 distearate, PEG-40 stearate, PEG-100 stearate, PEG-20 dilaurate, PEG-25 glyceryl trioleate, PEG-32 dioleate, PEG-20 glyceryl laurate, PEG-30 glyceryl laurate, PEG-20 glyceryl stearate, PEG-20 glyceryl oleate, PEG-30 glyceryl oleate, PEG-30 glyceryl laurate, PEG-40 glyceryl laurate, PE G-40 Palm Kernel Oil, PEG-50 Hydrogenated Castor Oil, PEG-40 Castor Oil, PEG-35 Castor Oil, PEG-60 Castor Oil, PEG-40 Hydrogenated Castor Oil, PEG-60 Hydrogenated Castor Oil, PEG-60 Corn Oil, PEG-6 Capric / Caprylic Glycerides, PEG-8 Capric / Caprylic Glycerides, Polyglyceryl-10 Laurate, PEG-30 Cholesterol, PEG-25 Phytosterols, PEG-30 Soybean Sterols PEG-20 trioleate, PEG-40 sorbitan oleate, PEG-80 sorbitan laurate, polysorbate 20, polysorbate 80, POE-9 lauryl ether, POE-23 lauryl ether, POE-10 oleyl ether, POE-20 oleyl ether, POE-20 stearyl ether, tocopheryl PEG-100 succinate, PEG-24 cholesterol, polyglyceryl-10 oleate, Tween 40, Tween 60, sucrose monostearate, sucrose monolaurate, sucrose monopalmitate, PEG 10-100 nonylphenol series, PEG 15-100 octylphenol series, and poloxamer.

[0154]

[0155] Suitable lipophilic surfactants include, by way of example only, fatty alcohols; glycerol fatty acid esters; acetylated glycerol fatty acid esters; lower alcohol fatty acid esters; propylene glycol fatty acid esters; sorbitan fatty acid esters; polyethylene glycol sorbitan fatty acid esters; sterols and sterol derivatives; polyoxyethylated sterols and sterol derivatives; polyethylene glycol alkyl ethers; sugar esters; sugar ethers; lactic acid derivatives of mono- and diglycerides; hydrophobic transesterification products of polyols with at least one member of the group consisting of glycerides, vegetable oils, hydrogenated vegetable oils, fatty acids, and sterols; oil-soluble vitamins / vitamin derivatives; and mixtures thereof. Of this group, preferred lipophilic surfactants include glycerol fatty acid esters, propylene glycol fatty acid esters, and mixtures thereof, or hydrophobic transesterification products of polyols with at least one member of the group consisting of vegetable oils, hydrogenated vegetable oils, and triglycerides.

[0155]

[0156] In one embodiment, the composition may contain a solubilizer to ensure good solubilization and / or dissolution of the compound of the present invention and to minimize precipitation of the compound of the present invention. The inclusion of a solubilizer may be particularly important for compositions for parenteral use, such as compositions for injection. A solubilizer can be added to increase the solubility of hydrophilic drugs and / or other components, such as surfactants, or to maintain the composition as a stable or homogeneous solution or dispersion.

[0156]

[0157] Examples of suitable solubilizers are: alcohols and polyols such as ethanol, isopropanol, butanol, benzyl alcohol, ethylene glycol, propylene glycol, butanediol and its isomers, glycerol, pentaerythritol, sorbitol, mannitol, transcutol, dimethyl isosorbide, polyethylene glycol, polypropylene glycol, polyvinyl alcohol, hydroxypropyl methylcellulose and other cellulose derivatives, cyclodextrin and cyclodextrin derivatives; ethers of polyethylene glycol having an average molecular weight of about 200 to about 6000, such as tetrahydrofurfuryl alcohol PEG ether (glycofurol) or methoxy PEG; 2-pyrrolidone, 2-piperidone, ε-caprolactam, N-alkylpyrrolidone, N-hydroxybenzoates, ... amides and other nitrogen-containing compounds such as hydroxyalkylpyrrolidone, N-alkylpiperidone, N-alkylcaprolactam, dimethylacetamide, and polyvinylpyrrolidone; esters such as ethyl propionate, tributyl citrate, acetyl triethyl citrate, acetyl tributyl citrate, triethyl citrate, ethyl oleate, ethyl caprylate, ethyl butyrate, triacetin, propylene glycol monoacetate, propylene glycol diacetate, epsilon-caprolactone and its isomers, δ-valerolactone and its isomers, β-butyrolactone and its isomers; and other solubilizing agents known in the art, such as, but not limited to, dimethylacetamide, dimethyl isosorbide, N-methylpyrrolidone, monooctanoin, diethylene glycol monoethyl ether, and water.

[0157]

[0158] Mixtures of solubilizers may also be used. Examples include, but are not limited to, triacetin, triethyl citrate, ethyl oleate, ethyl caprylate, dimethylacetamide, N-methylpyrrolidone, N-hydroxyethylpyrrolidone, polyvinylpyrrolidone, hydroxypropyl methylcellulose, hydroxypropyl cyclodextrin, ethanol, polyethylene glycol 200-100, glycofurol, transcutol, propylene glycol, and dimethyl isosorbide. Particularly preferred solubilizers include sorbitol, glycerol, triacetin, ethyl alcohol, PEG-400, glycofurol, and propylene glycol.

[0158]

[0159] The amount of solubilizer that can be included is not particularly limited. The amount of a given solubilizer may be limited to a bioacceptable amount, which can be easily determined by one of ordinary skill in the art. In some situations, for example, to maximize drug concentration, it may be advantageous to include an amount of solubilizer that far exceeds the bioacceptable amount and remove the excess solubilizer using conventional techniques such as distillation or evaporation before providing the composition to a patient. Thus, when present, the solubilizer may be present in a weight ratio of 10%, 25%, 50%, 100%, or up to about 200% by weight, based on the combined weight of the drug and other excipients. If desired, very small amounts of solubilizer, such as 5%, 2%, 1%, or even less, may also be used. Typically, the solubilizer may be present in an amount of about 1% to about 100% by weight, more typically about 5% to about 25% by weight.

[0159]

[0160] The composition may further comprise one or more pharmaceutically acceptable additives and excipients, including, but not limited to, detackifiers, antifoaming agents, buffers, polymers, antioxidants, preservatives, chelating agents, viscomodulators, tonicifiers, flavorants, colorants, odorants, opacifiers, suspending agents, binders, fillers, plasticizers, lubricants, and mixtures thereof.

[0160]

[0161] Additionally, acids or bases may be incorporated into the compositions to facilitate processing, enhance stability, or for other reasons. Examples of pharmaceutically acceptable bases include amino acids, amino acid esters, ammonium hydroxide, potassium hydroxide, sodium hydroxide, sodium bicarbonate, aluminum hydroxide, calcium carbonate, magnesium hydroxide, magnesium aluminum silicate, synthetic aluminum silicate, synthetic hydrocalcite, magnesium aluminum hydroxide, diisopropylethylamine, ethanolamine, ethylenediamine, triethanolamine, triethylamine, triisopropanolamine, trimethylamine, tris(hydroxymethyl)aminomethane (TRIS), and the like. Also suitable are bases that are salts of pharmaceutically acceptable acids, such as acetic acid, acrylic acid, adipic acid, alginic acid, alkanesulfonic acid, amino acids, ascorbic acid, benzoic acid, boric acid, butyric acid, carbonic acid, citric acid, fatty acids, formic acid, fumaric acid, gluconic acid, hydroquinosulfonic acid, isoascorbic acid, lactic acid, maleic acid, oxalic acid, parabromophenylsulfonic acid, propionic acid, p-toluenesulfonic acid, salicylic acid, stearic acid, succinic acid, tannic acid, tartaric acid, thioglycolic acid, toluenesulfonic acid, and uric acid. Salts of polybasic acids, such as sodium phosphate, disodium hydrogen phosphate, and sodium dihydrogen phosphate, may also be used. When the base is a salt, the cation can be any convenient and pharmaceutically acceptable cation, such as ammonium, alkali metals, and alkaline earth metals. Examples may include, but are not limited to, sodium, potassium, lithium, magnesium, calcium, and ammonium.

[0161]

[0162] Suitable acids are pharmaceutically acceptable organic or inorganic acids. Examples of suitable inorganic acids include hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, boric acid, phosphoric acid, etc. Examples of suitable organic acids include acetic acid, acrylic acid, adipic acid, alginic acid, alkanesulfonic acid, amino acids, ascorbic acid, benzoic acid, boric acid, butyric acid, carbonic acid, citric acid, fatty acids, formic acid, fumaric acid, gluconic acid, hydroquinonesulfonic acid, isoascorbic acid, lactic acid, maleic acid, methanesulfonic acid, oxalic acid, parabromophenylsulfonic acid, propionic acid, p-toluenesulfonic acid, salicylic acid, stearic acid, succinic acid, tannic acid, tartaric acid, thioglycolic acid, toluenesulfonic acid, and uric acid.

[0162] Injectable pharmaceutical composition

[0163] In selected embodiments, the present invention provides a pharmaceutical composition for injection comprising a BTK inhibitor compound selected from Table 1 or a pharmaceutically acceptable salt thereof, and a pharmaceutical excipient suitable for injection. The components and amounts of agents in the composition are as described herein.

[0163]

[0164] Forms into which the compositions of the present invention can be incorporated for administration by injection include aqueous or oily suspensions, or emulsions with sesame oil, corn oil, cottonseed oil, or peanut oil, as well as elixirs, mannitol, dextrose, or sterile aqueous solutions, and similar pharmaceutical vehicles.

[0164]

[0165] Aqueous solutions in physiological saline are also conventionally used for injection. Ethanol, glycerol, propylene glycol and liquid polyethylene glycol (and suitable mixtures thereof), cyclodextrin derivatives, and vegetable oils may also be employed. Proper fluidity can be maintained, for example, by the use of a coating such as lecithin to maintain the required particle size in the case of dispersions, and by the use of surfactants. Prevention of the action of microorganisms can be brought about by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, and thimerosal.

[0165]

[0166] Sterile injectable solutions are prepared by incorporating the required amount of the BTK inhibitor or its pharmaceutically acceptable salt in an appropriate solvent, along with various other ingredients listed above, as required, followed by filtration sterilization. Generally, dispersions are prepared by incorporating various sterilized active ingredients into a sterile vehicle containing a basic dispersion medium and the required other ingredients listed above. In the case of sterile powders for the preparation of sterile injectable solutions, certain preferred methods of preparation are vacuum drying and freeze-drying techniques, which produce a powder of the active ingredient plus any additional desired ingredients from a previously sterile-filtered solution of the active ingredient plus any additional desired ingredients.

[0166]

[0167] Administration of the BTK inhibitor or pharmaceutically acceptable salt thereof, or pharmaceutical compositions of these compounds, can be achieved by any method that allows delivery of the compound to the site of action. These methods include oral routes, intraduodenal routes, parenteral injection (including intravenous, intraarterial, subcutaneous, intramuscular, intravascular, intraperitoneal, or infusion), topical (e.g., transdermal application), rectal administration, local delivery via catheter or stent, or by inhalation. Combinations of compounds can also be administered intraadiposally or intrathecally.

[0167]

[0168] Exemplary parenteral dosage forms include solutions or suspensions of the active compounds in sterile aqueous solutions, for example, aqueous propylene glycol or dextrose solutions. Such dosage forms may be suitably buffered, if desired.

[0168]

[0169] The present invention also provides kits. The kits include a BTK inhibitor compound selected from Table 1 or a pharmaceutically acceptable salt thereof, alone or in combination in suitable packaging, and written materials, which may include instructions for use, a discussion of clinical studies, and a list of side effects. Such kits may also include information such as scientific literature references, package insert materials, clinical trial results, and / or summaries of this information, which demonstrate or define the activity and / or benefits of the composition and / or describe dosing, administration, side effects, drug interactions, or other information useful to healthcare providers. Such information may be based on the results of various studies, for example, studies using experimental animals, including in vivo models, and studies based on human clinical trials. The kits may further contain another active pharmaceutical ingredient. Appropriate packaging and additional items for use (e.g., measuring cups for liquid preparations, foil wrapping to minimize exposure to air, etc.) are known in the art and may be included in the kits. The kits described herein may be provided, sold, and / or promoted to healthcare providers, including physicians, nurses, pharmacists, formulary personnel, etc. Kits may, in selected embodiments, be sold directly to consumers. In one embodiment, the invention provides kits comprising a BTK inhibitor, or a pharmaceutically acceptable salt thereof, for use in treating myelofibrosis as described herein.

[0169] Dosage and Dosage Regimen

[0170] The amount of BTK inhibitor or its pharmaceutically acceptable salt administered will depend on the individual being treated, the severity of the disorder or condition, the rate of administration, the pharmacokinetics of the compound, and the discretion of the prescribing physician. However, effective dosages range from about 0.001 to about 100 mg per kg of body weight per day, e.g., about 1 to about 35 mg / kg / day, in single or divided doses. For a 70 kg individual, an effective dosage would be about 0.05 to 7 g / day, e.g., about 0.05 to about 2.5 g / day. In some situations, dosage levels below the lower end of the aforementioned range may be more than sufficient, while in other cases, even larger doses can be employed without causing any toxic side effects, e.g., by dividing such larger doses into several smaller doses for administration throughout the day.

[0170]

[0171] In some embodiments, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered in a single dose. Multiple daily administration, e.g., twice daily, is also embodied. Typically, such administration is oral; however, other routes may be used as appropriate.

[0171]

[0172] In some embodiments, the BTK inhibitor or pharmaceutically acceptable salt thereof is administered in multiple doses to treat myelofibrosis. In one embodiment, the BTK inhibitor or pharmaceutically acceptable salt thereof is administered in multiple doses. In one embodiment, dosing may be once, twice, three times, or four times per day. In one embodiment, dosing may be selected from the group consisting of once daily, twice daily, three times daily, or four times daily, every other day, once weekly, twice weekly, three times weekly, four times weekly, every other week, and once monthly. In other embodiments, the BTK inhibitor or pharmaceutically acceptable salt thereof is administered from about once per day to about four times per day. In some embodiments, the BTK inhibitor or pharmaceutically acceptable salt thereof is administered once daily, while in other embodiments, the BTK inhibitor or pharmaceutically acceptable salt thereof is administered twice daily, and in other embodiments, the BTK inhibitor or pharmaceutically acceptable salt thereof is administered three times daily. In some embodiments, the BTK inhibitor, or a pharmaceutically acceptable salt thereof, is administered three times a week, including every Monday, Wednesday, and Friday.

[0172]

[0173] Administration of the BTK inhibitor or pharmaceutically acceptable salt thereof may continue for as long as necessary. In some embodiments, the BTK inhibitor or pharmaceutically acceptable salt thereof is administered for more than 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, or more days. In some embodiments, the BTK inhibitor or pharmaceutically acceptable salt thereof is administered for less than 28, 14, 7, 6, 5, 4, 3, 2, or 1 day. In some embodiments, the BTK inhibitor or pharmaceutically acceptable salt thereof is administered for about 14 days, about 21 days, about 28 days, about 35 days, about 42 days, about 49 days, or about 56 days. In some embodiments, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered continuously and chronically, e.g., for the treatment of chronic effects. In another embodiment, administration of the MDM2 inhibitor and the BTK inhibitor continues for less than about 7 days. In yet another embodiment, administration continues for more than about 6, 10, 14, 28 days, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, or 1 year. In some embodiments, administration continues for more than about 1 year, 2 years, 3 years, 4 years, or 5 years. In some embodiments, continuous dosing is achieved and maintained as long as necessary.

[0173]

[0174] In some embodiments, the effective dosage of the BTK inhibitor or a pharmaceutically acceptable salt thereof is about 1 mg to about 600 mg, about 10 mg to about 500 mg, about 20 mg to about 450 mg, about 25 mg to about 200 mg, about 10 mg to about 200 mg, about 20 mg to about 150 mg, about 30 mg to about 120 mg, about 10 mg to about 90 mg, about 20 mg to about 80 mg, about 30 mg to about 70 mg, about 40 mg to about 60 mg, about 45 mg to about 55 mg, or about 4 The value range is 8 mg to about 52 mg, about 50 mg to about 150 mg, about 60 mg to about 140 mg, about 70 mg to about 130 mg, about 80 mg to about 120 mg, about 90 mg to about 110 mg, about 95 mg to about 105 mg, about 150 mg to about 250 mg, about 160 mg to about 240 mg, about 170 mg to about 230 mg, about 180 mg to about 220 mg, about 190 mg to about 210 mg, about 195 mg to about 205 mg, or about 198 mg to about 202 mg. In some embodiments, the effective dosage of the BTK inhibitor or a pharmaceutically acceptable salt thereof is about 15 mg, about 25 mg, about 30 mg, about 50 mg, about 50 mg, about 75 mg, about 90 mg, about 100 mg, about 120 mg, about 125 mg, about 150 mg, about 175 mg, about 180 mg, about 200 mg, about 225 mg, about 240 mg, about 250 mg, about 275 mg, about 300 mg, about 325 mg, about 350 mg, about 360 mg, about 375 mg, about 400 mg, about 425 mg, about 450 mg, about 475 mg, about 480 mg, or about 500 mg.

[0174]

[0175] In some embodiments, the effective dosage of the BTK inhibitor or a pharmaceutically acceptable salt thereof is 15 mg, 25 mg, 30 mg, 50 mg, 60 mg, 75 mg, 90 mg, 100 mg, 120 mg, 150 mg, 175 mg, 180 mg, 200 mg, 225 mg, 240 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 360 mg, 375 mg, and 480 mg.

[0175]

[0176] In some embodiments, the effective dosage of the BTK inhibitor or a pharmaceutically acceptable salt thereof is from about 0.01 mg / kg to about 4.3 mg / kg, from about 0.15 mg / kg to about 3.6 mg / kg, from about 0.3 mg / kg to about 3.2 mg / kg, from about 0.35 mg / kg to about 2.85 mg / kg, from about 0.15 mg / kg to about 2.85 mg / kg, from about 0.3 mg to about 2.15 mg / kg, from about 0.45 mg / kg to about 1.7 mg / kg, or from about 0.15 mg / kg to about 1. 3mg / kg, about 0.3mg / kg~about 1.15mg / kg, about 0.45mg / kg~about 1mg / kg, about 0.55mg / kg~about 0.85mg / kg, about 0.65mg / kg~about 0.8mg / kg, about 0.7mg / kg~ Approximately 0.75mg / kg, approximately 0.7mg / kg to approximately 2.15mg / kg, approximately 0.85mg / kg to approximately 2mg / kg, approximately 1mg / kg to approximately 1.85mg / kg, approximately 1.15mg / kg to approximately 1.7mg / kg, approximately 1.3mg / kg In some embodiments, the effective dosage of the BTK inhibitor or a pharmaceutically acceptable salt thereof is about 0.35 mg / kg, about 0.7 mg / kg, about 1 mg / kg, about 1.4 mg / kg, about 1.8 mg / kg, about 2.1 mg / kg, about 2.5 mg / kg, about 2.85 mg / kg, about 3.2 mg / kg, or about 3.6 mg / kg.

[0176]

[0177] In some embodiments, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered at a dosage of 10 to 500 mg BID, including dosages of 15 mg, 25 mg, 30 mg, 50 mg, 60 mg, 75 mg, 90 mg, 100 mg, 120 mg, 150 mg, 175 mg, 180 mg, 200 mg, 225 mg, 240 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 360 mg, 375 mg, and 480 mg BID.

[0177]

[0178] In some embodiments, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered in a dosage of 10-600 mg QD, including dosages of 15 mg, 25 mg, 30 mg, 50 mg, 60 mg, 75 mg, 90 mg, 100 mg, 120 mg, 150 mg, 175 mg, 180 mg, 200 mg, 225 mg, 240 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 360 mg, 375 mg, and 480 mg QD.

[0178]

[0179] An effective amount of the BTK inhibitor or a pharmaceutically acceptable salt thereof may be administered in either single or multiple doses by any of the accepted modes of administration of agents, including buccal, sublingual, and transdermal routes, by intra-arterial injection, intravenously, parenterally, intramuscularly, subcutaneously, or orally, with similar utility.

[0179]

[0180] In some embodiments, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to a subject intermittently, known as intermittent administration. By "intermittent administration," the term refers to administration of a therapeutically effective dose of the BTK inhibitor or a pharmaceutically acceptable salt thereof for a period of time, followed by a period of discontinuation, then followed by another administration period, etc. Within each administration period, the dosing frequency can be independently selected from three times per day, twice per day, daily, once per week, twice per week, three times per week, four times per week, five times per week, six times per week, or once per month. In one embodiment, the BTK inhibitor is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof.

[0180]

[0181] By "period of discontinuance" or "interruption period" or "rest period," this term refers to the length of time during which administration of a BTK inhibitor or a pharmaceutically acceptable salt thereof is discontinued. The time of interruption may be longer or shorter than the administration period, or may be the same as the administration period. During the interruption period, other therapeutic agents besides the BTK inhibitor or a pharmaceutically acceptable salt thereof may be administered. A interruption period may be necessary to alleviate any toxic effects associated with a particular BTK inhibitor compound.

[0181]

[0182] In one embodiment, a BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to a human subject in need thereof to treat myelofibrosis in a first dosing period, followed by a withdrawal period, then a second dosing period, etc. The first dosing period, second dosing period, and discontinuation period are independently selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 1 month, 5 weeks, 6 weeks, 7 weeks, 2 months, 9 weeks, 10 weeks, 11 (eleven) weeks, 3 months, 13 weeks, 14 weeks, 15 weeks, a period of more than 4 months, and more days, and the BTK inhibitor or pharmaceutically acceptable salt thereof is administered to the subject three times per day, twice per day, once daily, once per week, twice per week, three times per week, four times per week, five times per week, six times per week, or once per month. In one embodiment, the first administration period is the same length as the second administration period. In one embodiment, the first administration period is shorter than the second administration period. In one embodiment, the first administration period is longer than the second administration period. In one embodiment, the first administration period and the second administration period are about 1 week, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 3 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 3 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject once a week; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 4 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first dosing period and the second dosing period are about 4 weeks, and the BTK inhibitor or pharmaceutically acceptable salt thereof is administered to the subject once a week; and the discontinuation period is about 2 weeks. In one embodiment, the BTK inhibitor is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof.

[0182]

[0183] In one embodiment, to treat myelofibrosis secondary to polycythemia vera, a BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to a human subject in need thereof for a first dosing period, followed by a withdrawal period, then a second dosing period, etc. The first dosing period, second dosing period, and discontinuation period are independently selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 1 month, 5 weeks, 6 weeks, 7 weeks, 2 months, 9 weeks, 10 weeks, 11 weeks, 3 months, 13 weeks, 14 weeks, 15 weeks, a period of more than 4 months, and more days, and the BTK inhibitor or pharmaceutically acceptable salt thereof is administered to the subject three times per day, twice per day, once daily, once per week, twice per week, three times per week, four times per week, five times per week, six times per week, or once per month. In one embodiment, the first administration period is the same length as the second administration period. In one embodiment, the first administration period is shorter than the second administration period. In one embodiment, the first administration period is longer than the second administration period. In one embodiment, the first administration period and the second administration period are about 1 week, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 3 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 3 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject once a week; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 4 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first dosing period and the second dosing period are about 4 weeks, and the BTK inhibitor or pharmaceutically acceptable salt thereof is administered to the subject once a week; and the discontinuation period is about 2 weeks. In one embodiment, the BTK inhibitor is a compound selected from Table 1, and pharmaceutically acceptable salts thereof.

[0183]

[0184] In one embodiment, to treat myelofibrosis secondary to essential thrombocythemia, a BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to a human subject in need thereof for a first dosing period, followed by a withdrawal period, then a second dosing period, etc. The first dosing period, second dosing period, and discontinuation period are independently selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 1 month, 5 weeks, 6 weeks, 7 weeks, 2 months, 9 weeks, 10 weeks, 11 weeks, 3 months, 13 weeks, 14 weeks, 15 weeks, a period of more than 4 months, and more days, and the BTK inhibitor or pharmaceutically acceptable salt thereof is administered to the subject three times per day, twice per day, once daily, once per week, twice per week, three times per week, four times per week, five times per week, six times per week, or once per month. In one embodiment, the first administration period is the same length as the second administration period. In one embodiment, the first administration period is shorter than the second administration period. In one embodiment, the first administration period is longer than the second administration period. In one embodiment, the first administration period and the second administration period are about 1 week, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 3 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 3 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject once a week; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 4 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first dosing period and the second dosing period are about 4 weeks, and the BTK inhibitor or pharmaceutically acceptable salt thereof is administered to the subject once a week; and the discontinuation period is about 2 weeks. In one embodiment, the BTK inhibitor is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof.

[0184]

[0185] In one embodiment, to treat myelofibrosis secondary to chronic myeloid leukemia, a BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to a human subject in need thereof for a first dosing period, followed by a withdrawal period, then a second dosing period, etc. The first dosing period, second dosing period, and discontinuation period are independently selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 1 month, 5 weeks, 6 weeks, 7 weeks, 2 months, 9 weeks, 10 weeks, 11 weeks, 3 months, 13 weeks, 14 weeks, 15 weeks, a period of more than 4 months, and more days, and the BTK inhibitor or pharmaceutically acceptable salt thereof is administered to the subject three times per day, twice per day, once daily, once per week, twice per week, three times per week, four times per week, five times per week, six times per week, or once per month. In one embodiment, the first administration period is the same length as the second administration period. In one embodiment, the first administration period is shorter than the second administration period. In one embodiment, the first administration period is longer than the second administration period. In one embodiment, the first administration period and the second administration period are about 1 week, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 3 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 3 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject once a week; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 4 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first dosing period and the second dosing period are about 4 weeks, and the BTK inhibitor or pharmaceutically acceptable salt thereof is administered to the subject once a week; and the discontinuation period is about 2 weeks. In one embodiment, the BTK inhibitor is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof.

[0185]

[0186] In one embodiment, a BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to a human subject in need of treatment to treat primary myelofibrosis for a first administration period, then followed by a discontinuation period, then followed by a second administration period, etc. The first dosing period, second dosing period, and discontinuation period are independently selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 1 month, 5 weeks, 6 weeks, 7 weeks, 2 months, 9 weeks, 10 weeks, 11 weeks, 3 months, 13 weeks, 14 weeks, 15 weeks, a period of more than 4 months, and more days, and the BTK inhibitor or pharmaceutically acceptable salt thereof is administered to the subject three times per day, twice per day, once daily, once per week, twice per week, three times per week, four times per week, five times per week, six times per week, or once per month. In one embodiment, the first administration period is the same length as the second administration period. In one embodiment, the first administration period is shorter than the second administration period. In one embodiment, the first administration period is longer than the second administration period. In one embodiment, the first administration period and the second administration period are about 1 week, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 3 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 3 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject once a week; and the discontinuation period is about 2 weeks. In one embodiment, the first administration period and the second administration period are about 4 weeks, the BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to the subject daily; and the discontinuation period is about 2 weeks. In one embodiment, the first dosing period and the second dosing period are about 4 weeks, and the BTK inhibitor or pharmaceutically acceptable salt thereof is administered to the subject once a week; and the discontinuation period is about 2 weeks. In one embodiment, the BTK inhibitor is a compound selected from Table 1 or a pharmaceutically acceptable salt thereof.

[0186]

[0187] In one embodiment, a BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to treat myelofibrosis for 3 weeks, 6 weeks, 9 weeks, 12 weeks, 15 weeks, 18 weeks, 21 weeks, 24 weeks, 27 weeks, 30 weeks, 33 weeks, 36 weeks, 39 weeks, 42 weeks, 45 weeks, 48 ​​weeks, 51 weeks, 54 weeks, 57 weeks, 60 weeks, 63 weeks, 66 weeks, 69 weeks, 72 weeks, 75 weeks, 78 weeks, 81 weeks, 84 weeks, 87 weeks, 90 weeks, 93 weeks, or 100 weeks. , 96 weeks, 99 weeks, 102 weeks, 105 weeks, 108 weeks, 111 weeks, 114 weeks, 117 weeks, 120 weeks, 123 weeks, 126 weeks, 129 weeks, 132 weeks, 135 weeks, 138 weeks, 141 weeks, 144 weeks, 147 weeks, 150 weeks, 153 weeks, and 156 weeks to a subject in need thereof, wherein the BTK inhibitor is selected from any of the compounds in Table 1 or a pharmaceutically acceptable salt thereof. In one embodiment, the BTK inhibitor is administered orally at a dose of 100 mg twice daily.

[0187]

[0188] In one embodiment, a BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to treat primary myelofibrosis for 3 weeks, 6 weeks, 9 weeks, 12 weeks, 15 weeks, 18 weeks, 21 weeks, 24 weeks, 27 weeks, 30 weeks, 33 weeks, 36 weeks, 39 weeks, 42 weeks, 45 weeks, 48 ​​weeks, 51 weeks, 54 weeks, 57 weeks, 60 weeks, 63 weeks, 66 weeks, 69 weeks, 72 weeks, 75 weeks, 78 weeks, 81 weeks, 84 weeks, 87 weeks, 90 weeks, 93 weeks, 94 weeks, 95 weeks, 96 weeks, 97 weeks, 98 weeks, 99 weeks, 100 weeks, 101 weeks, 102 weeks, 103 weeks, 104 weeks, 105 weeks, 106 weeks, 107 weeks, 108 weeks, 109 weeks, 110 weeks, 111 weeks, 112 weeks, 113 weeks, 114 weeks, 115 weeks, 116 weeks, 117 weeks, 118 weeks, 119 weeks, 120 weeks, 121 weeks, 122 weeks, 123 weeks, 124 weeks, 125 weeks, 126 weeks, 127 weeks, 128 weeks, 129 weeks, 130 weeks, 131 weeks, 132 weeks, 133 weeks, 134 weeks, 135 weeks, 136 weeks, 137 weeks, 138 weeks, 139 weeks, 140 weeks, 141 weeks, 142 weeks, 143 weeks, 144 weeks, 145 weeks, 146 weeks, 147 weeks, 148 weeks, 149 weeks, 1 The BTK inhibitor is administered to a subject in need thereof for a period selected from 96 weeks, 99 weeks, 102 weeks, 105 weeks, 108 weeks, 111 weeks, 114 weeks, 117 weeks, 120 weeks, 123 weeks, 126 weeks, 129 weeks, 132 weeks, 135 weeks, 138 weeks, 141 weeks, 144 weeks, 147 weeks, 150 weeks, 153 weeks, and 156 weeks, wherein the BTK inhibitor is selected from any of the compounds in Table 1 or a pharmaceutically acceptable salt thereof. In one embodiment, the human subject is hydroxyurea-resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy-dependent. In one embodiment, the human subject is phlebotomy-dependent without splenomegaly. In one embodiment, the human subject has failed ruxolitinib or fedratinib therapy.

[0188]

[0189] In one embodiment, for the treatment of myelofibrosis secondary to polycythemia vera, a BTK inhibitor or a pharmaceutically acceptable salt thereof is administered for 3 weeks, 6 weeks, 9 weeks, 12 weeks, 15 weeks, 18 weeks, 21 weeks, 24 weeks, 27 weeks, 30 weeks, 33 weeks, 36 weeks, 39 weeks, 42 weeks, 45 weeks, 48 ​​weeks, 51 weeks, 54 weeks, 57 weeks, 60 weeks, 63 weeks, 66 weeks, 69 weeks, 72 weeks, 75 weeks, 78 weeks, 81 weeks, 84 weeks, 87 weeks, 89 weeks, 90 weeks, 91 weeks, 92 weeks, 93 weeks, 94 weeks, 95 weeks, 96 weeks, 97 weeks, 98 weeks, 99 weeks, 100 weeks, 101 weeks, 102 weeks, 103 weeks, 104 weeks, 105 weeks, 106 weeks, 107 weeks, 108 weeks, 109 weeks, 110 weeks, 111 weeks, 112 weeks, 113 weeks, 114 weeks, 115 weeks, 116 weeks, 117 weeks, 118 weeks, 119 weeks, 120 weeks, 121 weeks, 122 weeks, 123 weeks, 124 weeks, 125 weeks, 126 weeks, 127 weeks, 128 weeks, 129 weeks, 130 weeks, 131 weeks, 132 weeks, 133 weeks, 134 weeks, 135 weeks, 136 weeks, 137 weeks, 138 weeks, 139 weeks, 140 weeks, 141 weeks, 142 weeks, 143 weeks, 144 weeks, 145 weeks, weeks, 90 weeks, 93 weeks, 96 weeks, 99 weeks, 102 weeks, 105 weeks, 108 weeks, 111 weeks, 114 weeks, 117 weeks, 120 weeks, 123 weeks, 126 weeks, 129 weeks, 132 weeks, 135 weeks, 138 weeks, 141 weeks, 144 weeks, 147 weeks, 150 weeks, 153 weeks, and 156 weeks, and the BTK inhibitor compound is selected from Table 1 and pharmaceutically acceptable salts thereof.

[0189]

[0190] In one embodiment, for treating myelofibrosis secondary to essential thrombocythemia, a BTK inhibitor or a pharmaceutically acceptable salt thereof is administered for 3 weeks, 6 weeks, 9 weeks, 12 weeks, 15 weeks, 18 weeks, 21 weeks, 24 weeks, 27 weeks, 30 weeks, 33 weeks, 36 weeks, 39 weeks, 42 weeks, 45 weeks, 48 ​​weeks, 51 weeks, 54 weeks, 57 weeks, 60 weeks, 63 weeks, 66 weeks, 69 weeks, 72 weeks, 75 weeks, 78 weeks, 81 weeks, 84 weeks, 87 weeks, 89 weeks, 90 weeks, 91 weeks, 92 weeks, 93 weeks, 94 weeks, 95 weeks, 96 weeks, 97 weeks, 98 weeks, 99 weeks, 100 weeks, 101 weeks, 102 weeks, 103 weeks, 104 weeks, 105 weeks, 106 weeks, 107 weeks, 108 weeks, 109 weeks, 110 weeks, 111 weeks, 112 weeks, 113 weeks, 114 weeks, 115 weeks, 116 weeks, 117 weeks, 118 weeks, 119 weeks, 120 weeks, 121 weeks, 122 weeks, 123 weeks, 124 weeks, 125 weeks, 126 weeks, 127 weeks, 128 weeks, 129 weeks, 130 weeks, 131 weeks, 132 weeks, 133 weeks, 134 weeks, 135 weeks, 136 weeks, 137 weeks, 138 weeks, 139 weeks, 140 weeks, 141 weeks, 142 weeks, 143 weeks, 144 weeks, 145 weeks, weeks, 90 weeks, 93 weeks, 96 weeks, 99 weeks, 102 weeks, 105 weeks, 108 weeks, 111 weeks, 114 weeks, 117 weeks, 120 weeks, 123 weeks, 126 weeks, 129 weeks, 132 weeks, 135 weeks, 138 weeks, 141 weeks, 144 weeks, 147 weeks, 150 weeks, 153 weeks, and 156 weeks, and the BTK inhibitor compound is selected from Table 1 and pharmaceutically acceptable salts thereof.

[0190]

[0191] In one embodiment, a BTK inhibitor or a pharmaceutically acceptable salt thereof is administered to treat myelofibrosis secondary to chronic myeloid leukemia for 3 weeks, 6 weeks, 9 weeks, 12 weeks, 15 weeks, 18 weeks, 21 weeks, 24 weeks, 27 weeks, 30 weeks, 33 weeks, 36 weeks, 39 weeks, 42 weeks, 45 weeks, 48 ​​weeks, 51 weeks, 54 weeks, 57 weeks, 60 weeks, 63 weeks, 66 weeks, 69 weeks, 72 weeks, 75 weeks, 78 weeks, 81 weeks, 84 weeks, 87 weeks, or 90 weeks. and 156 weeks, wherein the BTK inhibitor compound is selected from the group consisting of a BTK inhibitor compound selected from the group consisting of a BTK inhibitor compound of Table 1 and a pharmaceutically acceptable salt thereof.

[0191]

[0192] In one embodiment, the human subject is hydroxyurea-resistant (inadequate response). In one embodiment, the human subject has splenomegaly. In one embodiment, the human subject has splenomegaly and is phlebotomy-dependent. In one embodiment, the human subject is phlebotomy-dependent without splenomegaly. In one embodiment, the human subject has failed ruxolitinib or fedratinib therapy. [Example]

[0192]

[0193] Embodiments encompassed herein will now be described with reference to the following examples, which are provided for illustrative purposes only, and the disclosure encompassed herein should in no way be construed as being limited to these examples, but rather as encompassing any and all variations that become evident as a result of the teachings provided herein.

[0193] Example 1: BTK inhibitor monotherapy for patients with myelofibrosis

[0194] The purpose of this study is to investigate the safety and efficacy of BTK inhibitor compounds in patients with myelofibrosis. Thirty MF patients will participate in the study and will receive 200 or 300 mg of the BTK inhibitor compound once daily. Participation criteria are: (1) no prior treatment with at least one other agent (hydroxyurea, interferon, anagrelide); (2) age ≥ 18 years; (3) acceptable pre-study organ function during screening, defined as total bilirubin ≤ 1.5 times the upper limit of normal (ULN), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) ≤ 2.5 times the ULN, and serum creatinine ≤ 1.5 x ULN, except when due to Gilbert's disease or hemolysis; and (4) women and men of childbearing age must agree to use adequate contraception (i.e., hormonal or barrier methods of birth control; abstinence) prior to study entry and throughout their study participation. If a female subject becomes pregnant while participating in this study or if she is suspected to be pregnant, she should be immediately removed from the study.

[0194]

[0195] During or at the end of the study, each MF patient will be evaluated for the following: (1) hematologic response; (2) reduction in JAK2V617F allele burden; (3) change in bone marrow histopathological abnormalities; (4) reduction in baseline reticulin / collagen fibrosis; (5) incidence of venous and arterial thrombosis; and (6) change in MF-related symptoms to determine the safety and efficacy of the BTK inhibitor compound.

[0195] Example 2: Open-Label Phase 2a / 2b Study of a BTK Inhibitor Compound

[0196] There is a significant unmet need for improved therapy in patients with myelofibrosis who have primary resistance to treatment with ruxolitinib, a suboptimal response, or who relapse after treatment with ruxolitinib. BTK inhibitor compounds are orally bioavailable small molecule cytotoxic chemotherapeutic agents that bind to BTK.

[0196] Study design

[0197] The study is an open-label, two-part (Part A and Part B) Phase 2a / 2b study of the compound of Formula (I) in subjects with PMF, post PV-MF, or post ET-MF who have failed ruxolitinib therapy. Approximately 190 subjects will be enrolled in the study (90 in Part A and 100 in Part B).

[0197]

[0198] Part A (N=90): In Part A of the study, subjects will be randomly assigned to one of three treatment groups: Cohort 1, N=30 subjects: 280 mg of the BTK inhibitor compound once daily. Cohort 2, N=30 subjects: 420 mg of the BTK inhibitor compound once daily. Cohort 3, N=30 subjects: 560 mg of the BTK inhibitor compound once daily.

[0198]

[0199] Part B (N=100): Approximately 100 subjects will be enrolled in Part B and treated with the recommended dose and schedule from Part A. A Data Monitoring Committee (DMC) will meet every 3 months for Parts A and B during the conduct of the study to review safety data related to the clinical study. The DMC will also meet after all subjects in Part A have had an opportunity to complete the 24-week assessment. The DMC will determine the recommended dose and schedule of the BTK inhibitor compound based on the efficacy and safety data from Part A. In Parts A and B, subjects will receive the BTK inhibitor compound orally (PO) once daily in 28-day cycles. Dose reductions for hematologic and non-hematologic toxicity will be permitted. All subjects should be treated until disease progression or lack of tolerability. Disease progression will be defined as imaging and modified ELN criteria: a spleen volume increase of ≥ 25% from the nadir during the study by MRI (or CT) by central imaging review, lasting at least 2 weeks, and a ≥ 1 x 10 9 Based on leukemic transformation confirmed by ≥ 20% bone marrow blasts or ≥ 20% peripheral blood blast content associated with an absolute blast count of ≥ 10 cells / L.

[0199] Research purpose [Table 3]

[0200] Participation criteria Subjects in both Part A and Part B must meet all of the following criteria to be eligible for the study: 1. Adults >18 years of age; 2. Palpable splenomegaly at least 5 cm below the left costal margin; 3. Confirmed diagnosis of PMF, post-PV-MF, or post-ET-MF as assessed by a treating physician according to World Health Organization (WHO) criteria; 4. High-risk, intermediate-2 risk, or intermediate-1 risk as defined by the Dynamic International Prognostic System (DIPSS); ECOG performance status of 5.0 to 2; 6. Adequate hematologic, hepatic, and renal organ function (as defined by protocol and within 14 days prior to the first dose of the BTK inhibitor compound). Hematologic: ANC ≥ 1.0 x 10 in the absence of growth factors for the preceding 7 days. 9 Platelets / L; Platelet count ≧100×10 9 cells / L; peripheral blood blast count <10%. Liver: total bilirubin ≤ 2.0 times the upper limit of normal (ULN) except in cases of Gilbert syndrome; aspartate transaminase / serum glutamic oxaloacetic transaminase (AST / SGOT) and alanine transaminase / serum glutamic pyruvic transaminase (ALT / SGPT) ≤ 2.5ULN. Renal: Cockcroft-Gault estimated creatinine clearance > 45 mL / min.

[0201]

number

[0202] 7. Women of childbearing potential and men with partners of childbearing potential must agree to use effective contraception during the study. In addition, men must continue using contraception for 3 months after the last dose of study drug, and men must continue using contraception for 1 week after the last dose of study drug. Effective birth control includes: (a) combined estrogen- and progestogen-containing hormonal contraception (oral, intravaginal, transdermal); (b) progestogen-only hormonal contraception (oral, injectable, implantable); (c) intrauterine device; (d) intrauterine hormone-releasing system; (e) bilateral tubal occlusion; (f) vasectomized partner; and (g) sexual abstinence.

[0203]

[0202] Subjects in Part A must meet the following criteria for ruxolitinib treatment failure to be eligible for the study: Ruxolitinib treatment failure in Part A must meet either of the following criteria (a) or (b): (a) Patients who have received at least 12 weeks of ruxolitinib treatment and have both: persistent splenomegaly on physical examination that is palpable ≥5 cm below the lower costal margin (LCM) and a TSS of >10 on the MPN-SAF TSS 2.0, or a single symptom score of >5 or two symptoms of >3, including only symptoms of left upper quadrant pain, bone pain, pruritus, or night sweats. (b) Or progressive disease at any time during ruxolitinib treatment, defined by any one of the following: a ≥ 25% increase in splenic volume from nadir as assessed by MRI or CT; the appearance of new splenomegaly that is palpable at least 5 cm below the LCM; a ≥ 100% increase in palpable distance below the LCM relative to baseline splenomegaly of 5-10 cm; or a ≥ 50% increase in palpable distance below the LCM relative to baseline splenomegaly of > 10 cm.

[0204] Subjects in Part B must meet the following ruxolitinib treatment failure criteria to be eligible for the study: Ruxolitinib treatment failure in Part B must meet either criterion (a) or (b) below: (a) For subjects who have received at least 12 weeks of ruxolitinib treatment and have the following: failure to have at least a ≥ 35% reduction in spleen volume; baseline splenomegaly before ruxolitinib treatment that is palpable 5-10 cm below the LCM but remains palpable; baseline splenomegaly before ruxolitinib treatment that is palpable > 10 cm below the LCM but remains palpable; (b) lack of splenic response, defined as having at least one of the following: baseline splenomegaly not reduced by 50%, baseline splenomegaly pre-ruxolitinib treatment that is palpable <5 cm below the LCM, ineligible for consideration as a ruxolitinib treatment failure; (b) or progressive disease at any time during ruxolitinib treatment, defined by any one of the following: a spleen volume increase of ≥25% from nadir, as assessed by MRI or CT, the appearance of new splenomegaly that is palpable at least 5 cm below the LCM, a ≥100% increase in palpable distance below the LCM from baseline splenomegaly of 5-10 cm, or a ≥50% increase in palpable distance below the LCM from baseline splenomegaly of >10 cm.

[0205] Exclusion criteria Subjects in both Part A and Part B who meet any of the following criteria will not be eligible for the study: 1. Participation in another interventional clinical trial within 4 weeks prior to the first dose of a compound of Formula (I) (participation in observational studies is allowed). 2. Recent / contemporaneous treatment such as major surgery, chemotherapy, immunomodulating therapy, biologic therapy, radiation therapy, or investigational therapy within 4 weeks or approximately 5 half-lives of the first dose of a compound of Formula (I). 3. Prior splenectomy. 4. Splenic irradiation within 3 months prior to the first dose of a compound of Formula (I). 5. Prior allogeneic stem cell transplant or eligible for allogeneic stem cell transplant. 6. Prior treatment with a histone deacetylase (HDAC) inhibitor or BCL-2 inhibitor. 7. Prior BTK inhibitor therapy. 8. Pregnant or lactating women. 9. History of major organ transplant. 10. Acute hepatitis A; known history of human immunodeficiency virus (HIV) positivity; clinically significant cardiac disease (New York Heart Association class III or IV); symptomatic congestive heart failure; unstable angina; ventricular arrhythmia; or uncontrolled intercurrent illness, including but not limited to psychiatric illness / social circumstances that would limit compliance with study requirements. 11. Subjects with clinically significant bacterial, fungal, parasitic, or viral infections requiring therapy. Subjects with acute bacterial infections requiring antibiotic use should delay screening / enrollment until the course of antibiotic therapy is completed. Other malignancies within the past 3 years other than curatively treated basal or squamous cell skin cancer, carcinoma in situ of the cervix, organconfined or treated non-metastatic prostate cancer with normal prostate-specific antigen, in situ breast carcinoma after complete surgical resection, or superficial transitional cell bladder carcinoma. 13. Grade 2 or higher QTc prolongation (>480 milliseconds according to NCI-CTCAE criteria version 5.0). 14. Hematopoietic growth factors (i.e., erythropoietin (Epo), granulocyte colony-stimulating factor (GCSF), romiplostim) within 28 days prior to receiving the first dose of a compound of formula (I). 15. Active or chronic bleeding within 4 weeks prior to receiving the first dose of a compound of formula (I).

[0206] Randomization procedure Part A: Subjects will be randomized to one of three treatment cohorts using a 1:1:1 allocation scheme. A contracted clinical service provider will develop the Part A randomization schedule, and the actual randomization assignment will be performed by a robust Interactive Response Technology (IRT) system. Part B: Subjects will be randomized to the BTK inhibitor compound dose and schedule recommended by the DMC.

[0207] statistical analysis The DMC will meet every 3 months for Parts A and B during the conduct of the study to review safety data related to the clinical study. The DMC will also meet after all subjects in Part A have had an opportunity to complete the 24-week assessment. The DMC will determine the recommended dose and schedule of the BTK inhibitor compound for Part B based on the efficacy and safety data from Part A. Results of statistical analyses, descriptive summary statistics, and supportive listings will be presented by study part (A or B) and within Part A (by cohort).

[0208] Research duration

[0207] The study will be considered for a full two years after the last subject is enrolled, at which time subjects on study treatment will be evaluated for eligibility to enroll in the rollover study.

[0209] Example 3: BTK Inhibition in Healthy B Cells and Myeloid Cells Inhibition of BTK phosphorylation (activation) by Compound No. 128 (1-(4-(((6-amino-5-(4-phenoxyphenyl)pyrimidin-4-yl)amino)methyl)-4-fluoropiperidin-1-yl)prop-2-en-1-one) was assessed in healthy B cells and in the myeloid cell lines MOLM-13 basal and He1-92 basal. Compound No. 128 was incubated with the myeloid cell lines without stimulation for 2 hours and with healthy PBMCs stimulated with 10 minutes of αIgM and HO for 2 hours. As shown in Table 3, the efficacy of Compound No. 128 in myeloid cells is similar to that of Compound No. 128 in healthy B cells.

[0210] [Table 4]

[0211] Example 4: Inhibition of cell migration towards SDF-1 (CXCL12) In MF, the spleen has elevated levels of SDF-1 compared to the blood. As a chemoattractant chemokine, SDF-1 attracts cells along a concentration gradient. In vitro inhibition of cell migration through a permeable membrane by Compound No. 128 and ruxolitinib (Rux) was assessed using He1-92 (V617F mut) cells. As shown in Figures 1A and 1B, both Compound No. 128 and ruxolitinib inhibit cell migration toward SDF-1.

[0212] Example 5: Fibronectin release Fibronectin is an important component of the microenvironment extracellular matrix and attaches to cells through VLA-4 (integrin α4β1). 6Cell detachment from fibronectin after exposure to Compound No. 128 was assessed by seeding MV-411 cells (in triplicate). Cells were allowed to adhere overnight at 37°C, and then unadhered cells were washed away. The adhered cells were then treated with DMSO or Compound No. 128 (1, 2, or 5 μM) for 2 hours at 37°C. Detached cells were then harvested, and cell number was determined using CellTiter Glo. As shown in Figure 2, cells detached from fibronectin after exposure to Compound No. 128.

[0213] Example 6: Expression of surface molecules involved in cell adhesion Decreased expression of surface molecules involved in cell adhesion to the cellular microenvironment can inhibit the protection of malignant cells in various compartments. Monocytes from the blood of COVID-19 patients treated with Compound No. 128 were assessed for levels of the surface molecules CD11a (LFA-1), CD62L (L-selectin), and CD49d (VLA-4). As shown in Figure 3, treatment with Compound No. 128 reduced the levels of surface molecules in some patients.

[0214] Example 7: Cytokine production The effect on in vitro cytokine production after exposure to Compound No. 128 was assessed in stimulated whole blood after 24 hours of incubation. As shown in Figure 4, Compound No. 128 reduced cytokine and chemokine production upon cell stimulation. Further embodiments are as follows. [Embodiment 1] 1. A method of treating splenomegaly in a human subject in need thereof, comprising administering to the human subject a Bruton's tyrosine kinase (BTK) inhibitor. [Embodiment 2] 2. The method of embodiment 1, wherein said human subject has an accumulation of malignant CD34+ myeloid cells in their spleen. [Embodiment 3] 3. The method of embodiment 2, wherein said malignant CD34+ bone marrow cells have decreased expression of CXCR4 compared to normal bone marrow cells. [Embodiment 4] 3. The method of embodiment 2, wherein the BTK inhibitor is administered in an amount sufficient to stimulate migration of the malignant CD34+ myeloid cells into the peripheral blood of the human subject. [Embodiment 5] 3. The method of embodiment 2, wherein the BTK inhibitor is administered in an amount sufficient to reduce activity of VLA-4 in the malignant CD34+ myeloid cells. [Embodiment 6] 3. The method of embodiment 2, wherein the BTK inhibitor is administered in an amount sufficient to reduce expression of VLA-4 on the malignant CD34+ myeloid cells. [Embodiment 7] 7. The method of any one of embodiments 1 to 6, wherein the human subject is suffering from myelofibrosis. [Embodiment 8] 1. A method of stimulating migration of malignant CD34+ myeloid cells from the spleen to peripheral blood in a human subject suffering from splenomegaly, comprising administering to the human subject a BTK inhibitor. [Embodiment 9] 9. The method of embodiment 8, wherein the human subject has an accumulation of malignant CD34+ myeloid cells in the spleen. [Embodiment 10] 9. The method of embodiment 8, wherein said malignant CD34+ bone marrow cells have decreased expression of CXCR4 compared to normal bone marrow cells. [Embodiment 11] 9. The method of embodiment 8, wherein the human subject is suffering from myelofibrosis. [Embodiment 12] 12. The method of embodiment 7 or 11, wherein the myelofibrosis is selected from the group consisting of primary myelofibrosis (PMF), post-polycythemia vera myelofibrosis (post PV-MF), and post-essential thrombocythemia myelofibrosis (post ET-MF). [Embodiment 13] The method of any one of embodiments 1-12, wherein the human subject has not responded to ruxolitinib therapy. [Embodiment 14] The method of any one of embodiments 1 to 13, wherein the human subject has a JAK2V617F mutation. [Embodiment 15] 15. The method of embodiment 14, wherein the human subject has acute myeloid leukemia (AML) secondary to a myeloproliferative neoplasm (MPN). [Embodiment 16] The method of any one of embodiments 1 to 13, wherein the human subject does not have a JAK2V617F mutation. [Embodiment 17] 17. The method of embodiment 16, wherein the human subject has acute myeloid leukemia secondary to a myeloproliferative neoplasm. [Embodiment 18] The method of any one of embodiments 1-17, wherein the human subject has not been treated with a JAK2 inhibitor. [Embodiment 19] The method of any one of embodiments 1-18, wherein the human subject is intolerant to a JAK2 inhibitor. [Embodiment 20] The method of any one of embodiments 1-19, wherein the human subject is ineligible for treatment with a JAK2 inhibitor. [Embodiment 21] The method of any one of embodiments 1-20, wherein the human subject relapses after JAK2 inhibitor treatment or is refractory to JAK2 inhibitor treatment. [Embodiment 22] 22. The method of any one of embodiments 1-21, wherein the BTK inhibitor is administered once daily at a dose selected from the group consisting of 15 mg, 25 mg, 30 mg, 50 mg, 60 mg, 75 mg, 90 mg, 100 mg, 120 mg, 150 mg, 175 mg, 180 mg, 200 mg, 225 mg, 240 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 360 mg, 375 mg, 480 mg, and 560 mg. [Embodiment 23] 23. The method of any one of embodiments 1-22, wherein the BTK inhibitor is administered twice daily at a dose selected from the group consisting of 15 mg, 25 mg, 30 mg, 50 mg, 60 mg, 75 mg, 90 mg, 100 mg, 120 mg, 150 mg, 175 mg, 180 mg, 200 mg, 225 mg, 240 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 360 mg, 375 mg, 480 mg, and 560 mg. [Embodiment 24] 24. The method of any one of embodiments 1-23, wherein the BTK inhibitor is administered orally. [Embodiment 25] 25. The method of any one of embodiments 1-24, wherein the BTK inhibitor is a covalent BTK inhibitor. [Embodiment 26] 26. The method of any one of embodiments 1-25, wherein the BTK inhibitor is a non-covalent BTK inhibitor. [Embodiment 27] 27. The method of any one of embodiments 1-26, wherein said BTK inhibitor is a compound selected from Table 1, or a pharmaceutically acceptable salt thereof.

Claims

1. 1. A pharmaceutical composition for treating myelofibrosis in a human subject in need thereof, comprising a Bruton's tyrosine kinase (BTK) inhibitor, the BTK inhibitor is 1-(4-(((6-amino-5-(4-phenoxyphenyl)pyrimidin-4-yl)amino)methyl)-4-fluoropiperidin-1-yl)prop-2-en-1-one or a pharmaceutically acceptable salt thereof; The pharmaceutical composition, wherein the human subject has an accumulation of malignant CD34+ myeloid cells in the hematopoietic tissues.

2. 2. The pharmaceutical composition of claim 1, wherein the malignant CD34+ bone marrow cells have decreased expression of CXCR4 compared to normal bone marrow cells.

3. 10. The pharmaceutical composition of claim 1, comprising the BTK inhibitor in an amount sufficient to stimulate migration of the malignant CD34+ bone marrow cells into the peripheral blood of the human subject.

4. 10. The pharmaceutical composition of claim 1, comprising the BTK inhibitor in an amount sufficient to reduce activity of VLA-4 in the malignant CD34+ myeloid cells.

5. 10. The pharmaceutical composition of claim 1, comprising the BTK inhibitor in an amount sufficient to reduce VLA-4 expression in the malignant CD34+ myeloid cells.

6. 1. A pharmaceutical composition for stimulating the migration of malignant CD34+ myeloid cells from hematopoietic tissues to peripheral blood in a human subject suffering from myelofibrosis, comprising a BTK inhibitor, 1. A pharmaceutical composition, wherein the BTK inhibitor is 1-(4-(((6-amino-5-(4-phenoxyphenyl)pyrimidin-4-yl)amino)methyl)-4-fluoropiperidin-1-yl)prop-2-en-1-one or a pharmaceutically acceptable salt thereof.

7. The pharmaceutical composition of claim 6, wherein the human subject has an accumulation of malignant CD34+ myeloid cells in the spleen.

8. 7. The pharmaceutical composition of claim 6, wherein the malignant CD34+ bone marrow cells have reduced expression of CXCR4 compared to normal bone marrow cells.

9. 7. The pharmaceutical composition according to claim 1, wherein the myelofibrosis is selected from the group consisting of primary myelofibrosis (PMF), post-polycythemia vera myelofibrosis (post-PV-MF), and post-essential thrombocythemia myelofibrosis (post-ET-MF).

10. The pharmaceutical composition of any one of claims 1 to 9, wherein the human subject has failed to respond to ruxolitinib therapy.

11. The pharmaceutical composition of any one of claims 1 to 10, wherein the human subject has a JAK2V617F mutation.

12. 12. The pharmaceutical composition of claim 11, wherein the human subject has acute myeloid leukemia (AML) secondary to myelofibrosis.

13. The pharmaceutical composition of any one of claims 1 to 10, wherein the human subject does not have the JAK2V617F mutation.

14. 14. The pharmaceutical composition of claim 13, wherein the human subject has acute myeloid leukemia secondary to myelofibrosis.

15. The pharmaceutical composition of any one of claims 1 to 14, wherein the human subject has not been treated with a JAK2 inhibitor.

16. The pharmaceutical composition of any one of claims 1 to 15, wherein the human subject is intolerant to a JAK2 inhibitor.

17. The pharmaceutical composition of any one of claims 1 to 16, wherein the human subject is ineligible for treatment with a JAK2 inhibitor.

18. 18. The pharmaceutical composition of any one of claims 1 to 17, wherein the human subject relapses after or is refractory to JAK2 inhibitor treatment.

19. 19. The pharmaceutical composition of any one of claims 1-18, wherein the BTK inhibitor is administered once daily at a dose selected from the group consisting of 15 mg, 25 mg, 30 mg, 50 mg, 60 mg, 75 mg, 90 mg, 100 mg, 120 mg, 150 mg, 175 mg, 180 mg, 200 mg, 225 mg, 240 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 360 mg, 375 mg, 480 mg, and 560 mg.

20. 19. The pharmaceutical composition of any one of claims 1-18, wherein the BTK inhibitor is administered twice daily at a dose selected from the group consisting of 15 mg, 25 mg, 30 mg, 50 mg, 60 mg, 75 mg, 90 mg, 100 mg, 120 mg, 150 mg, 175 mg, 180 mg, 200 mg, 225 mg, 240 mg, 250 mg, 275 mg, 300 mg, 325 mg, 350 mg, 360 mg, 375 mg, 480 mg, and 560 mg.

21. 21. The pharmaceutical composition of any one of claims 1 to 20, wherein the BTK inhibitor is administered orally.

22. The pharmaceutical composition according to claim 1 or 6, wherein the hematopoietic tissue is extramedullary hematopoietic tissue.

23. The pharmaceutical composition of claim 1 or 6, wherein the hematopoietic tissue is bone marrow.

24. The pharmaceutical composition of claim 1 or 6, wherein the hematopoietic tissue is the spleen.

25. The pharmaceutical composition of claim 1 or 6, wherein the hematopoietic tissue is the liver.

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