WO2013059548A1

WO2013059548A1 - Compositions and methods for treating cancer using jak2 inhibitor

Info

Publication number: WO2013059548A1
Application number: PCT/US2012/060951
Authority: WO
Inventors: Rita Greco; Francisco Adrian
Original assignee: Sanofi
Priority date: 2011-10-19
Filing date: 2012-10-19
Publication date: 2013-04-25
Also published as: WO2013059548A9

Abstract

Compositions, kits, and methods of treating patients with cancer are provided, wherein the methods comprise administering to the patient an effective amount of a JAK2 inhibitor or a JAK2 inhibitor and an IL-6 antagonist, wherein the patient has a cancer or is at risk of developing a cancer that involves IL-6 signaling.

Description

COMPOSITIONS AND METHODS FOR TREATING CANCER

USING JAK2 INHIBITOR

BACKGROUND

[0001] Inappropriate protein kinase activity, arising from mutation, over-expression, or altered regulation, as well as over or under production of growth factors or cytokines has been implicated in many diseases, including cancer. Protein kinases are families of enzymes that catalyze the phosphorylation of specific residues in proteins, broadly classified into tyrosine and serine/threonine kinases. Inappropriate kinase activity triggers a variety of biological cellular responses relating to cell growth, cell differentiation, survival, apoptosis, mitogenesis, cell cycle control, and/or cell mobility, which play a role in the development and progression of cancer.

[0002] Protein kinases are an important class of enzymes as targets for therapeutic intervention. In particular, the JAK family of cellular protein tyrosine kinases (JAK1, JAK2, JAK3, and Tyk2) is involved in cytokine signaling (Kisseleva et al, Gene, 2002, 285, 1; Yamaoka et al. Genome Biology 2004, 5, 253)). Cytokines are low-molecular-weight, non-antibody proteins that are secreted by various cell types and are involved in cell-to-cell communication between and among cells of the immune and hematopoietic systems. Binding of a cytokine to its receptor activates the associated JAK family member(s) which then triggers a cascade of phosphorylation including phosphorylation of the cytokine receptor and one or more members of the signal transducer and activator of transcription (STAT) family, ultimately leading to gene expression.

[0003] Activation of the JAK2-erythropoietin signaling pathway has been shown to play a significant role in hematological disorders such as myelofibrosis. In particular, a gain-of-function mutation of JAK2 (V617F) was reported to be a causative factor in myelofibrosis, polycythemia vera, and essential thrombocythemia. The mutation has been found in patients with either

"atypical" myeloproliferative disorders and myelodysplasia syndrome (for reviews see Tefferi and Gilliland, Cell Cycle 2005, 4(8), e61; Pesu et. al. Molecular Interventions 2005, 5(4), 211). The V617F JAK2 mutation is associated with constitutive activation of JAK2 and its downstream effectors as well as induction of erythropoietin hypersensitivity in cell based experiments.

Furthermore, mutations in the EPO receptor (EPOR) also results in constitutive activation of the JAK2 pathway leading to myeloproliferative disorders. [0004] N-tert-butyl-3 -[(5 -methyl-2- { [4-(2-pyrrolidin- 1 -ylethoxy)phenyl] amino } pyrimidin-

4-yl)amino]benzenesulfonamide (also referred to herein as Compound 1) is a JAK2 inhibitor that has also been shown to inhibit wild type as well as JAK2 having the V617F mutation in vitro. See WO2007053452, Example 245, the teachings of which are incorporated herein. As described in WO2007053452, N-tert-butyl-3 -[(5 -methyl-2- { [4-(2-pyrrolidin- 1 - ylethoxy)phenyl] amino }pyrimidin-4-yl)amino]benzenesulfonamide inhibits phosphorylation of JAK2V617F, STAT5, and STAT3 in JAK2V617F-expressing Ba/F3 cells in a dose dependent manner. Oral administration of N-tert-butyl-3 - [(5 -methyl-2- { [4-(2-pyrrolidin- 1 - ylethoxy)phenyl] amino }pyrimidin-4-yl)amino]benzenesulfonamide alone has been shown to inhibit tumor growth in a murine model of JAK2V617F-induced polycythemia vera.

[0005] Interleukin 6 (IL-6) is a member of a large group of low-molecular- weight, non- antibody proteins known as cytokines. Cytokines are typically secreted by various cell types and are involved in cell-to-cell communication, such as, for example, coordinating antibody and T cell immune interactions and amplifying immune reactivity. IL-6 (also known as B-cell stimulatory factor-2, interferon- 2, B-cell differentiation factor, hepatocyte stimulatory factor, hybridoma growth factor, and plasmacytoma growth factor) is involved in, for example, modulation of specific immune responses including B- and T-cell differentiation, bone metabolism,

thrombopoiesis, and epidermal proliferation. (Papassotiropoulos, et ah, Neurobiology of Aging, 22:863-871 (2001).

[0006] IL-6 promotes cell signaling by binding to its multi-subunit receptor (IL-6R) which includes a signaling subunit, glycoprotein 130 (gpl30) which does not bind IL-6 per se and an IL- 6 binding subunit (also known as gp80). The IL-6 binding subunit can also be present in a soluble form (sIL-6R or IL-6Ra). Upon binding of IL-6 to the IL-6 binding subunit, the signal- transducing subunit, gpl30, is dimerized. (Jones, J. Immunol. 175:3463-3468 (2005)).

Dimerization of gpl30, Janus Kinases are activated which in turn phosphorylate and activate Signal Transducers and Activators of Transcription (STATs), in particular STAT3.

[0007] Autocrine or paracrine interleukin 6 (IL-6) mediated cell signaling have been shown to provide tumor cells with the ability to proliferate, survive, migrate and metastasize. At the molecular level, IL-6 binding to its receptor results in activation of different signaling cascades that have well established role in cancer, namely PI3K/Akt, MEK/ERK1-2 and JAK1/STAT3 pathways. [0008] Tocilizumab is an anti- IL-6R antibody that is currently approved for use in treating Rheumatoid Arthritis and Systemic Juvenile Idiopathic Arthritis (SJIA), however, because Tocilizumab may decrease the activity of the patient's immune system, treatment with

Tocilizumab may increase the patient's certain cancers. Other IL-6R antibodies are described, for example, in US7582298 (see Table I), US6410691 (Example 11), U5817790 (Examples 1-3), US5795695; and US6670373. The teachings of which are incorporated herein by reference.

[0009] There remains a need for a cancer therapy that is more effective in inhibiting cell proliferation and tumor growth while minimizing patient toxicity and adverse side-effects.

SUMMARY

[0010] Provided herein are compositions, kits, and uses thereof in the treatment of a variety of cancers as described herein.

[0011] In one aspect, provided herein are compositions comprising a JAK2 inhibitor, N-tert- butyl-3 -[(5 -methyl-2- { [4-(2-pyrrolidin- 1 -ylethoxy)phenyl] amino } pyrimidin-4- yl)amino]benzenesulfonamide, or a pharmaceutically acceptable salt thereof, and an IL-6 antagonist.

[0012] Provided herein are methods of treating a patient comprising administering a

therapeutically effective amount of a JAK2 inhibitor, N-tert-butyl-3 -[(5 -methyl-2- {[4-(2- pyrrolidin- 1 -ylethoxy)phenyl] amino }pyrimidin-4-yl)amino]benzenesulfonamide, or a

pharmaceutically acceptable salt thereof, wherein the patient has a cancer that involves IL-6 receptor signaling. In some embodiments, the methods comprise administering the JAK2 inhibitor as a monotherapy. In other embodiments, the methods comprise administering a therapeutically effective amount of a JAK2 inhibitor, N-tert-butyl-3 -[(5 -methyl-2- {[4-(2-pyrro lidin-1- ylethoxy)phenyl]amino}pyrimidin-4-yl)amino]benzenesulfonamide or a pharmaceutically acceptable salt thereof, in combination with an IL-6 antagonist, wherein the patient has a cancer that involves IL-6 receptor signaling. In still other embodiments, the methods of treating a patient comprise administering to the patient a first dosage of a JAK2 inhibitor, N-tert-butyl-3 -[(5 -methyl- 2-{[4-(2-pyrrolidin-l-ylethoxy)phenyl]amino}pyrimidin-4-yl)amino]benzenesulfonamide, and a second dosage of an IL-6 antagonist, wherein the patient has a cancer that involves IL-6 receptor signaling.

[0013] In a second aspect, provided herein are kits comprising: (A) a JAK 2 inhibitor, N-tert- butyl-3 -[(5 -methyl-2- { [4-(2-pyrrolidin- 1 -ylethoxy)phenyl] amino } pyrimidin-4- yl)amino]benzenesulfonamide, or a pharmaceutically acceptable salt thereof; and (B) instructions for use of the JAK2 inhibitor to treat a patient having a cancer that involves IL-6 receptor signaling. In some embodiments, the kits provided herein comprise instructions for use of the JAK2 inhibitor as a monotherapy to treat a patient having a cancer that involves IL-6 receptor signaling. In other embodiments, the kits comprise (A) a JAK 2 inhibitor, N-tert-butyl-3-[(5- methyl-2-{[4-(2-pyrrolidin-l-ylethoxy)phenyl]amino}pyrimidin-4-yl)amino]benzenesulfonamide, or a pharmaceutically acceptable salt thereof; and (B) instructions for use of the JAK2 inhibitor in combination with an IL-6 antagonist, wherein the IL-6 antagonist is selected from the group consisting of an anti-IL-6 antibody or IL-6 binding portion thereof and an anti-IL-6 receptor (IL- 6R) antibody or IL-6R binding portion thereof.

[0014] In a third aspect, provided herein are kits comprising (A) an IL-6 antagonist and (B) instructions for use of the IL-6 antagonist in combination with a compound a JAK 2 inhibitor, N- tert-butyl-3 - [(5 -methyl-2- { [4-(2-pyrrolidin- 1 -ylethoxy)phenyl] amino } pyrimidin-4- yl)amino]benzenesulfonamide, or a pharmaceutically acceptable salt thereof to treat a patient having a cancer that involves IL-6 receptor signaling; wherein the IL-6 antagonist is selected from the group consisting of an anti-IL-6 antibody or IL-6 binding portion thereof and an anti-IL-6 receptor (IL-6R) antibody or IL-6R binding portion thereof.

[0015] In a forth aspect provided herein are kits comprising (A) a JAK 2 inhibitor, N-tert-butyl- 3 - [(5 -methyl-2- { [4-(2-pyrrolidin- 1 -ylethoxy)phenyl] amino } pyrimidin-4- yl)amino]benzenesulfonamide, or a pharmaceutically acceptable salt thereof, (B) an IL-6 antagonist; and (C) instructions for use of the JAK2 inhibitor and IL-6 antagonist to treat a patient having a cancer that involves IL-6 receptor signaling, wherein the IL-6 antagonist is selected from the group consisting of an anti-IL-6 antibody or IL-6 binding portion thereof and an anti-IL-6 receptor (IL-6R) antibody or IL-6R binding portion thereof.

[0016] Other objects, features and advantages will become apparent from the following detailed description. The detailed description and specific examples are given for illustration only since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description. Further, the examples demonstrate the principle of the invention and cannot be expected to specifically illustrate the application of this invention to all the examples where it will be obviously useful to those skilled in the prior art.

BRIEF DESCRIPTION OF THE DRAWINGS

[0017] FIG. 1 shows phospho-STAT3 (pSTAT3) levels in cells treated with Compound 1 after no IL-6 stimulation (NS) or after stimulation with recombinant human IL-6 (+ IL-6); A: DU145, B: MDA-MB0468, C: KP4, D: LNCaP, E: BT-474, F: HCT116.

[0018] FIG. 2 shows pSTAT3 levels in the indicated tumor cells treated with A: Compound 1, B: INCB 18424, or C: AC 1480 after no IL-6 stimulation (NS) or after stimulation with recombinant human IL-6 (+ IL-6).

[0019] FIG. 3 shows IC⁵⁰ values for Compound 1 on solid tumor cell lines.

[0020] FIG. 4 shows the results of a clonogenic assay of Compound 1 effect on a panel of hepatocellular carcinoma cell lines.

[0021] FIG. 5 shows tumor volume versus time (A) and Compound 1 plasma concentration versus time (B) for DU145 in a mouse model.

[0022] FIG. 6 shows pEKR levels in DU145 cells treated with A: Compound 1, B: INCB18424, or C: AZD1480 after no IL-6 stimulation (NS) or stimulation with recombinant human IL-6 (+ IL- 6).

[0023] FIG. 7 shows pSTAT3 levels in DU145 cells treated with SAR302503 (Compound 1) after no IL-6 stimulation (NS) or stimulation with recombinant human IL-6 (+ IL-6) and in the presence or absence of anti-IL-6R mAb.

DETAILED DESCRIPTION

[0024] There is an ongoing need in the art for more efficacious methods and compositions in the treatment of cancer. The instant application is directed, generally, to compositions and methods for the treatment of cancer, and more particularly, to compositions, kits, and methods comprising inhibitors of Janus Kinase 2 (JAK2) and inhibitors of IL-6R signaling, separately or in

combination for use in treating cancers that involve IL-6R signaling. [0025] In one aspect, methods for treating patients with cancer are provided. In one embodiment, the methods comprise administering to the patient a therapeutically effective amount of a JAK2 inhibitor, wherein the patient has a cancer that involves IL-6R signaling.

[0026] In another embodiment, the methods comprise administering to the patient a

therapeutically effective amount of a JAK2 inhibitor and a therapeutically effective amount of an IL-6 antagonist, as further described below.

[0027] In one embodiment, the methods and compositions provided herein comprise a JAK2 inhibitor that is N-tert-butyl-3-[(5-methyl-2-{[4-(2-pyrrolidin-l- ylethoxy)phenyl] amino }pyrimidin-4-yl)amino]benzenesulfonamide (also referred to herein as Compound 1.

[0028] The preparation, properties, and JAK2 -inhibiting abilities of Compound 1 are provided in, for example, International Patent Publication No. WO2007053452, particularly in Example 245 therein and PCT application PCT/USlO/56280, filed November 10, 2010. The entire contents of PCT/US 10/56280 are incorporated herein by reference. Neutral and salt forms of Compound 1 are all considered herein.

[0029] In other embodiments, the methods and compositions provided herein comprise an IL-6 antagonist. Suitable IL-6 antagonists are agents that inhibit the IL-6R signaling pathway and include, for example, antibodies that specifically bind to IL-6, IL-6R, or IL-6 complexed with a soluble portion of IL-6R wherein the specific binding results in a reduction in at least one measure of IL-6 pathway signaling. In one embodiment the IL-6 antagonist is an anti- IL-6Ra antibody or IL-6R binding portion thereof. Suitable anti-IL-6R antibodies include, for example anti-IL-6R antibodies described, for example, in US7582298 (see Table I), US6410691 (Example 11), U5817790 (Examples 1-3), US5795695, and US6670373.

[0030] In some embodiments, the IL-6 antagonist is an anti-IL-6 antibody or antigen binding fragment thereof. Suitable anti-IL-6 antibodies or fragments thereof include, for example antibodies described in WO 2011/066371, WO2010056948, and WO 2011/0666378.

[0031] IL-6 is known to sometimes be secreted from cells in a complex with soluble IL-6R IL-6 binding subunit (IL-6Ra). Therefore, in other embodiments, the IL-6 antagonist is an anti-IL-6 antibody or antigen binding fragment thereof that is capable of interacting with the IL-6/IL-6Ra complex in such a way as to prevent the IL-6/IL-6Ra from inducing IL-6 signaling in at least some IL-6R bearing cells. Suitable antibodies or fragments thereof capable of interacting with the IL- 6/IL-6Ra complex include, for example, antibodies disclosed in US20110038877.

[0032] In still other embodiments, the IL-6 antagonist can be an anti-IL-6 receptor (IL-6R) antibody or antigen binding fragment thereof. Suitable anti-IL-6R antibodies or fragments thereof include, for example, antibodies disclosed in US7582298 (see Table I), US6410691, U5817790, US5795,695; and US6670373

[0033] In yet another embodiment, IL-6 antagonists can be one or more siRNA molecules that are capable of down regulating IL-6 or IL-6R expression in cells of a patient. Suitable IL-6 siRNA molecules are described, for example, in WO 2009/070609.

[0034] The preparation and properties of suitable IL-6 antagonists are provided in, for example, in US20110038877, and US7582298 (see Table I), US6410691, U5817790, US5795,695; and US6670373

[0035] In some embodiments, Compound 1 described herein is unsolvated. In other

embodiments, Compound 1 is in solvated form. As known in the art, the solvate can be any of pharmaceutically acceptable solvent, such as water, ethanol, and the like. In general, the presence of a solvate or lack thereof does not have a substantial effect on the efficacy of the JAK2 inhibitor described herein.

[0036] Compound 1 is described in its neutral form, in some embodiments, the compound is used in a pharmaceutically acceptable salt form, for example, as the dihydro chloride monohydrate form. The salt can be obtained by any of the methods well known in the art. A "pharmaceutically acceptable salt" of the compound refers to a salt that is pharmaceutically acceptable and that retains pharmacological activity. It is understood that the pharmaceutically acceptable salts are non-toxic. Additional information on suitable pharmaceutically acceptable salts can be found in Remington 's Pharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, PA, 1985, or S. M. Berge, et al., "Pharmaceutical Salts," J. Pharm. Sci., 1977;66: 1-19, both of which are incorporated herein by reference.

[0037] Examples of pharmaceutically acceptable acid addition salts include those formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, as well as those salts formed with organic acids, such as acetic acid, trifluoroacetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, 3-(4-hydroxybenzoyl)benzoic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1 ,2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, glucoheptonic acid, 4,4'-methylenebis-(3-hydroxy-2-ene-l-carboxylic acid), 3-phenylpropionic acid, trimethylacetic acid, tertiary butylacetic acid, lauryl sulfuric acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, muconic acid, p- toluenesulfonic acid, and salicylic acid.

[0038] In one set of embodiments, the JAK2 inhibitor is administered in a therapeutically effective (i.e., therapeutic) amount or dosage. A "therapeutically effective amount" is an amount of the JAK2 inhibitor that, when administered to a patient by itself, effectively treats the cancer (for example, inhibits tumor growth, stops tumor growth, or causes tumor regression). An amount that proves "therapeutically effective amount" in a given instance, for a particular subject, may not be effective for 100% of subjects similarly treated for the disease or condition under consideration, even though such dosage is deemed a "therapeutically effective amount" by skilled practitioners. The amount of the compound that corresponds to a therapeutically effective amount is strongly dependent on the type of cancer, stage of the cancer, the age of the patient being treated, and other facts. In general, therapeutically effective amounts of these compounds are well-known in the art, such as provided in the supporting references cited above.

[0039] In another set of embodiments, the JAK2 inhibitor is administered in combination with one or more IL-6 antagonist such as inhibitors of IL-6R signaling. In one set of embodiments, the JAK2 inhibitor and/or IL-6 antagonist can be administered at individually therapeutic amounts or dosages. In another set of embodiments, one or both of the JAK2 inhibitor and the IL-6 antagonist are administered in a sub-therapeutically effective amount or dosage. A sub-therapeutically effective amount is an amount of the JAK2 inhibitor or the IL-6 antagonist is one that, when administered to a patient by itself, partially inhibits the biological activity of the intended target.

[0040] Whether administered in therapeutic or sub-therapeutic amounts, the combination of the JAK2 inhibitor and IL-6 antagonist should be effective in treating the cancer. A sub-therapeutic amount of JAK2 inhibitor can be an effective amount if, when combined with the IL-6 antagonist, the combination is effective in the treatment of a cancer. Similarly, a sub-therapeutic amount IL-6 antagonist can be an effective amount if, when combined with the JAK2 inhibitor, the combination is effective in the treatment of a cancer.

[0041] In some embodiments, the combination of JAK2 inhibitor and IL-6 antagonist exhibits a synergistic effect (i.e., greater than additive effect) in treating the cancer, particularly in reducing a tumor volume in the patient. In different embodiments, depending on the combination and the effective amounts used, the combination of compounds can inhibit tumor growth, achieve tumor stasis, or even achieve substantial or complete tumor regression.

[0042] When used in combination with an IL-6 antagonist, the JAK2 inhibitor can be

administered simultaneously (also referred to herein as concomitantly) with the IL-6 antagonist. Simultaneous administration typically means that both compounds enter the patient at precisely the same time. However, simultaneous administration also includes the possibility that the JAK2 inhibitor and the IL-6 antagonist enter the patient at different times, but the difference in time is sufficiently short such that the first administered compound is not provided the time to take effect on the patient before entry of the second administered compound. Such differences in

administration times typically correspond to less than 1 minute, and more typically, less than 30 seconds.

[0043] In one example, wherein the JAK2 inhibitor and the IL-6 antagonist are in solution, simultaneous administration can be achieved by administering a solution containing the combination of the JAK2 inhibitor and the IL-6 antagonist. In another example, separate solutions, one of which contains the JAK2 inhibitor and the other of which contains the IL-6 antagonist, can be administered simultaneously.

[0044] In another example, wherein the JAK2 inhibitor and the IL-6 antagonist are in solid form, simultaneous administration can be achieved by administering a composition containing the combination of the JAK2 inhibitor and the IL-6 antagonist. In another example, separate compositions, one of which contains the JAK2 inhibitor and the other of which contains the IL-6 antagonist, can be administered simultaneously.

[0045] In another example, wherein either the JAK2 inhibitor or IL-6 antagonist are in solid form and the other of either the JAK2 inhibitor or the IL-6 antagonist are in solution, simultaneous administration can be achieved by administering the solution and the solid form at the same time as described above. [0046] In other embodiments the JAK2 inhibitor and the IL-6 antagonist are not simultaneously administered. In one embodiment, the first compound (either the JAK2 inhibitor or the IL-6 antagonist) is administered, and the other compound can be administered after allowing sufficient time for the fist compound to have some therapeutic effect in the patient. Generally, the difference in time does not extend beyond the time for the first administered compound to complete its effect in the patient, or beyond the time the first administered compound is completely or substantially eliminated or deactivated in the patient.

[0047] In one embodiment, the JAK2 compound is administered first. In another embodiment, the IL-6 antagonist is administered first. The time difference in non-simultaneous administrations is typically greater than 1 minute, and can be, for example, precisely, at least, up to, or less than 5 minutes, 10 minutes, 15 minutes, 30 minutes, 45 minutes, 60 minutes, two hours, three hours, six hours, nine hours, 12 hours, 24 hours, 36 hours, 48 hours, 1 week, 2 weeks, 3 weeks, 4 weeks.

[0048] In some embodiments, the JAK2 inhibitor is Compound 1 and is administered at a dosage of about 240 mg per day to about 680 mg per day, and wherein the specified weight is the free base moiety weight of the compound. In some embodiments, the compound is administered at a dose of about 300 mg per day to about 500 mg per day (e.g., about 300 mg per day to about 400 mg per day, or about 400 mg per day to about 500 mg per day), and wherein the specified weight is the free base moiety weight of the compound. In some embodiments, the compound is administered at a dose of about any of 240 mg per day, 250 mg per day, 300 mg per day, 350 mg per day, 400 mg per day, 450 mg per day, 500 mg per day, 550 mg per day, 600 mg per day, 650 mg per day, or 680 mg per day, and wherein the specified weight is the free base moiety weight of the compound.

[0049] In some embodiments, the IL-6 antagonist is administered at a dosage of about 0.5 mg/kg to about 10 mg/kg per dose. In some embodiments, the IL-6 antagonist is administered at a dose of about 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, or 10 mg/kg.

[0050] As used herein, the term "about" generally indicates a possible variation of no more than 10%, 5%, or 1% of a value. For example, "about 25 mg/kg" will generally indicate, in its broadest sense, a value of 22.5-27.5 mg/kg, i.e., 25 ± 10 mg/kg. [0051] Where the JAK2 inhibitor is used in combination with a second therapeutic agent, the amounts of JAK2 inhibitor and the second therapeutic agent should result in the effective treatment of a cancer, the amounts, when combined, are preferably not excessively toxic to the patient (i.e., the amounts are preferably within toxicity limits as established by medical guidelines). In some embodiments, either to prevent excessive toxicity and/or provide a more efficacious treatment of the cancer, a limitation on the total administered dosage is provided. Typically, the amounts considered herein are per day; however, half-day and two-day or three-day cycles also are considered herein.

[0052] Where the JAK2 inhibitor is used in combination with an IL-6 antagonist, the amounts of JAK2 inhibitor and IL-6 antagonist should result in the effective treatment of a cancer, the amounts, when combined, are preferably not excessively toxic to the patient (i.e., the amounts are preferably within toxicity limits as established by medical guidelines). In some embodiments, either to prevent excessive toxicity and/or provide a more efficacious treatment of the cancer, a limitation on the total administered dosage is provided. Typically, the amounts considered herein are per day; however, half-day and two-day or three-day cycles also are considered herein.

[0053] Different dosage regimens may be used to treat the cancer. In some embodiments, a daily dosage, such as any of the exemplary dosages described above, is administered once, twice, three times, or four times a day for three, four, five, six, seven, eight, nine, or ten days. Depending on the stage and severity of the cancer, a shorter treatment time (e.g., up to five days) may be employed along with a high dosage, or a longer treatment time (e.g., ten or more days, or weeks, or a month, or longer) may be employed along with a low dosage. In some embodiments, a once- or twice-daily dosage is administered every other day.

[0054] In some embodiments, each dosage contains the JAK2 inhibitor as the sole active ingredient. In some embodiments, each dosage contains the JAK2 inhibitor and the second therapeutic agent, while in other embodiments, each dosage contains either the JAK2 inhibitor or second therapeutic agent. In yet other embodiments, some of the dosages contain both the JAK2 inhibitor and the second therapeutic agent, while other dosages contain only the JAK2 or the second therapeutic agent. In some embodiments, each dosage contains both the JAK2 inhibitor and the IL-6 antagonist, while in other embodiments, each dosage contains either the JAK2 inhibitor or IL-6 antagonist. In yet other embodiments, some of the dosages contain both the JAK2 inhibitor and the IL-6 antagonist, while other dosages contain only the JAK2 or the IL-6 antagonist.

[0055] Examples of types of cancers to be treated with the present invention include, but are not limited to cancers that have involvement of IL-6 autocrine or paracrine signaling. Such cancers include for example, prostate cancer, breast cancer, lung cancer, colorectal cancer, and pancreatic cancer. In some embodiments, the patient has a cancer that has a mutation in a ras gene. In some embodiments, the cancer having a mutant ras gene is a colorectal or pancreatic cancer.

[0056] The patient considered herein is typically a human. However, the patient can be any mammal for which cancer treatment is desired. Thus, the methods described herein can be applied to both human and veterinary applications.

[0057] The term "treating" or "treatment", as used herein, indicates that the method has, at the least, mitigated abnormal cellular proliferation. For example, the method can reduce the rate of tumor growth in a patient, or prevent the continued growth of a tumor, or even reduce the size of a tumor. As used herein, an effective amount of the JAK2 inhibitor or JAK2 inhibitor in

combination with the IL-6 antagonist are amounts that can, for example, inhibit tumor growth, achieve tumor stasis, and/or achieve substantial or complete tumor regression

[0058] In another aspect, methods for preventing cancer in an animal, wherein the cancer involves IL-6 signaling are provided. In this regard, prevention denotes causing the clinical symptoms of the disease not to develop in an animal that may be predisposed to the disease but does not yet experience or display symptoms of the disease. In one embodiment, the methods comprise administering to the patient a JAK2 inhibitor as described herein. In one example, a method of preventing cancer in an animal comprises administering to the animal Compound 1 , or a pharmaceutically acceptable salt thereof, wherein the animal is at risk for developing a cancer that involves IL-6 signaling. In another embodiment, the methods comprise administering to the patient a JAK2 inhibitor and an IL-6 antagonist, as described herein. In one embodiment, a method of preventing cancer in an animal comprises administering to the animal Compound 1 , or a pharmaceutically acceptable salt thereof, in combination with a compound selected from the group consisting of an anti-IL-6 antibody or IL-6 binding portion thereof, and anti-IL-6R antibody or IL- 6R binding portion thereof, or an anti-IL-6/IL-6Ra antibody or IL-6/IL-6Ra binding portion thereof. [0059] The JAK2 inhibiting compound, or their pharmaceutically acceptable salts or solvate forms, in pure form or in an appropriate pharmaceutical composition, can be administered via any of the accepted modes of administration or agents known in the art. The compound can be administered, for example, orally, nasally, parenterally (intravenous, intramuscular, or

subcutaneous), topically, transdermally, intravaginally, intravesically, intracistemally, or rectally. The dosage form can be, for example, a solid, semi-solid, lyophilized powder, or liquid dosage forms, such as for example, tablets, pills, soft elastic or hard gelatin capsules, powders, solutions, suspensions, suppositories, aerosols, or the like, preferably in unit dosage forms suitable for simple administration of precise dosages. A particular route of administration is oral, particularly one in which a convenient daily dosage regimen can be adjusted according to the degree of severity of the disease to be treated.

[0060] The IL-6 antagonist, in an appropriate pharmaceutical composition, can be administered via any of the accepted modes of administration or agents known in the art. The compound can be administered, for example, orally, nasally, parenterally (intravenous, intramuscular, or

subcutaneous), topically, transdermally, intravaginally, intravesically, intracistemally, or rectally. The dosage form can be, for example, a solid, semi-solid, lyophilized powder, or liquid dosage forms, such as for example, tablets, pills, soft elastic or hard gelatin capsules, powders, solutions, suspensions, suppositories, aerosols, or the like, preferably in unit dosage forms suitable for simple administration of precise dosages. A particular route of administration is intravenous,

subcutaneous, or intramuscular.

[0061] In another aspect, the instant application is directed to a composition that includes the Compound 1 and an IL-6 antagonist. In some embodiments, the composition is in the form of a solid (e.g., a powder or tablet) including Compound 1 and an IL-6 antagonist in solid form, and optionally, one or more auxiliary (e.g., adjuvant) or pharmaceutically active compounds in solid form. In other embodiments, the composition further includes any one or combination of pharmaceutically acceptable carriers (i.e., vehicles or excipients) known in the art, thereby providing a liquid dosage form.

[0062] Auxiliary and adjuvant agents may include, for example, preserving, wetting, suspending, sweetening, flavoring, perfuming, emulsifying, and dispensing agents. Prevention of the action of microorganisms is generally provided by various antibacterial and antifungal agents, such as, parabens, chlorobutanol, phenol, sorbic acid, and the like. Isotonic agents, such as sugars, sodium chloride, and the like, may also be included. Prolonged absorption of an injectable pharmaceutical form can be brought about by the use of agents delaying absorption, for example, aluminum monostearate and gelatin. The auxiliary agents also can include wetting agents, emulsifying agents, pH buffering agents, and antioxidants, such as, for example, citric acid, sorbitan monolaurate, triethanolamine oleate, butylated hydroxytoluene, and the like.

[0063] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is admixed with at least one inert customary excipient (or carrier) such as sodium citrate or dicalcium phosphate or (a) fillers or extenders, as for example, starches, lactose, sucrose, glucose, mannitol, and silicic acid, (b) binders, as for example, cellulose derivatives, starch, alignates, gelatin, polyvinylpyrrolidone, sucrose, and gum acacia, (c) humectants, as for example, glycerol, (d) disintegrating agents, as for example, agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, croscarmellose sodium, complex silicates, and sodium carbonate, (e) solution retarders, as for example paraffin, (f) absorption accelerators, as for example, quaternary ammonium compounds, (g) wetting agents, as for example, cetyl alcohol, and glycerol monostearate, magnesium stearate and the like (h) adsorbents, as for example, kaolin and bentonite, and (i) lubricants, as for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. In the case of capsules, tablets, and pills, the dosage forms also may comprise buffering agents.

[0064] Solid dosage forms as described above can be prepared with coatings and shells, such as enteric coatings and others well-known in the art. They can contain pacifying agents and can be of such composition that they release the active compound or compounds in a certain part of the intestinal tract in a delayed manner. Examples of embedded compositions that can be used are polymeric substances and waxes. The active compounds also can be in microencapsulated form, if appropriate, with one or more of the above-mentioned excipients.

[0065] In another aspect, the instant application is directed to a composition that includes one or both of the Compound 1 and an IL-6 antagonist in a form suitable for parenteral injection.

Dosage forms suitable for parenteral injection may comprise physiologically acceptable sterile aqueous or nonaqueous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Examples of suitable aqueous and nonaqueous carriers, diluents, solvents or vehicles include water, ethanol, polyols (propyleneglycol, polyethyleneglycol, glycerol, and the like), suitable mixtures thereof, vegetable oils (such as olive oil) and injectable organic esters such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersions and by the use of surfactants.

[0066] In another aspect, the instant application is directed to a composition that includes one or both of the Compound 1 and an IL-6 antagonist in a liquid dosage form. Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs. Such dosage forms are prepared, for example, by dissolving, dispersing, etc.,

Compound 1 a pharmaceutically acceptable salt thereof and/or an IL-6 antagonist and optional pharmaceutical adjuvants in a carrier, such as, for example, water, saline, aqueous dextrose, glycerol, ethanol and the like; solubilizing agents and emulsifiers, as for example, ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propyleneglycol, 1,3- butyleneglycol, dimethyl formamide; oils, in particular, cottonseed oil, groundnut oil, corn germ oil, olive oil, castor oil and sesame oil, glycerol, tetrahydrofurfuryl alcohol,

polyethyleneglycols and fatty acid esters of sorbitan; or mixtures of these substances, and the like, to thereby form a solution or suspension.

[0067] Suspensions, in addition to the active compounds, may contain suspending agents, as for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters,

microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth, or mixtures of these substances, and the like.

[0068] Compositions for rectal administrations are, for example, suppositories that can be prepared by mixing the compounds described herein with, for example, suitable non- irritating excipients or carriers such as cocoa butter, polyethyleneglycol or a suppository wax, which are solid at ordinary temperatures but liquid at body temperature and therefore, melt while in a suitable body cavity and release the active component therein.

[0069] Dosage forms for topical administration may include, for example, ointments, powders, sprays, and inhalants. The active component is admixed under sterile conditions with a

physiologically acceptable carrier and any preservatives, buffers, or propellants as can be required. Ophthalmic formulations, eye ointments, powders, and solutions also can be employed. [0070] Generally, depending on the intended mode of administration, the pharmaceutically acceptable compositions will contain about 1% to about 99% by weight of the compounds described herein, or a pharmaceutically acceptable salt thereof, and 99% to 1% by weight of a pharmaceutically acceptable excipient. In one example, the composition will be between about 5% and about 75% by weight of a compounds described herein, or a pharmaceutically acceptable salt thereof, with the rest being suitable pharmaceutical excipients.

[0071] Actual methods of preparing such dosage forms are known, or will be apparent, to those skilled in this art. Reference is made, for example, to Remington's Pharmaceutical Sciences, 18th Ed., (Mack Publishing Company, Easton, Pa., 1990).

[0072] In some embodiments, the composition does not include one or more other anti-cancer compounds. In other embodiments, the composition includes one or more other anti-cancer compounds. For example, administered compositions can comprise standard of care agents for the type of tumors selected for treatment.

[0073] In another aspect, kits are provided. In some embodiments, kits according to the invention include package(s) comprising compounds or compositions of the invention. In one embodiment, kits comprise Compound 1 , or a pharmaceutically acceptable salt thereof, and instructions for using the compound to treat a patient having or at risk of developing a cancer that involves IL-6 signaling. In another embodiment, kits comprise Compound 1, or a

pharmaceutically acceptable salt thereof, and an IL-6 antagonist or a pharmaceutically acceptable composition thereof.

[0074] The phrase "package" means any vessel containing compounds or compositions presented herein. In some embodiments, the package can be a box or wrapping. Packaging materials for use in packaging pharmaceutical products are well-known to those of skill in the art. Examples of pharmaceutical packaging materials include, but are not limited to, bottles, tubes, inhalers, pumps, bags, vials, containers, syringes, bottles, and any packaging material suitable for a selected formulation and intended mode of administration and treatment.

[0075] The kit also can contain items that are not contained within the package but are attached to the outside of the package, for example, pipettes.

[0076] Kits can contain instructions for administering compounds or compositions provided herein to a patient. Kits also can comprise instructions for approved uses of compounds herein by regulatory agencies, such as the United States Food and Drug Administration. Kits also can contain labeling or product inserts for the compositions provided herein. The package(s) and/or any product insert(s) may themselves be approved by regulatory agencies. The kits can include compounds in the solid phase or in a liquid phase (such as buffers provided) in a package. The kits also can include buffers for preparing solutions for conducting the methods, and pipettes for transferring liquids from one container to another.

[0077] Examples have been set forth below for the purpose of illustration and to describe certain specific embodiments of the invention. However, the scope of the claims is not to be in any way limited by the examples set forth herein.

[0078] EXAMPLE 1. Inhibition of Stat3 phosphorylation

[0079] DU145 MDA-MB0468, KP4, LNCaP, BT-474, or HCT116 cells were obtained from ATCC and grown according to vendor recommendations. 500,000 cells were plated in 6 well plates in 2 ml of medium supplemented with 10% fetal bovine serum (Gibco).

[0080] Cells were stimulated with 10 ng/mL recombinant human IL-6 (rhIL6) for 15 min. or were not exposed to the exogenous rhIL6 (unstimulated). The stimulated and stimulated cells were treated with Compound 1 for 60 min. Cells were washed in cold phosphate buffered saline (PBS) and lysed using RIPA lysis buffer (Santa Cruz Biotechnology). The lysate was cleared by centrifugation, the debris discarded and the protein concentration measured using a colorimetric assay. 10 μg of cleared protein lysate was resolved on an SDS polyacrylamide gel, transferred to and immobilized onto a nitrocellulose membrane. pSTAT3 level from lysate was measured using anti-pSTAT3 (Y705) antibody (Cell Signaling Technologies) according to the manufacturer's instructions. As shown in FIG. 1, Compound 1 inhibits both unstimulated (NS) and hrIL6 stimulated (+ IL-6) STAT3 phosphorylation in multiple solid tumor cell lines including, DU145 MDA-MB0468, KP4, LNCaP, BT-474, HCT116.

[0081] EXAMPLE 2. Inhibition of Stat3 phosphorylation

[0082] Cells were plated as described in Example 1.

[0083] DU145 MDA-MB0468, LNCaP, and BT-474, F cells were stimulated with rhIL6 or were not exposed to the exogenous rhIL6 (unstimulated). The unstimulated and stimulated cells were treated with the indicated levels of Compound 1 (A), INCB 18424 (B), or AZD1480 (C) for 60 min. pSTAT3 level was measured from cell lysates as described in Example 1.

[0084] As shown in FIG. 2, Compound 1 inhibits both unstimulated (NS) and hrIL6 stimulated (+ IL-6) STAT3 phosphorylation to a similar extent as INCB 18424 and more potently than AZD1480.

[0085] EXAMPLE 3. Antiproliferative activity on solid tumor cell lines

[0086] A panel of 240 cancer cell lines representing about 30 cancer types were exposed to Compound 1 at 0.0003-10 μΜ and the anti-pro liferative, pro-apoptotic, and anti-mitotic activity was measured. Cells were grown in RPMI1640 or DMEM, 10% FBS, 2 mM L-alanyl-L- Glutamine, 1 mM Na Pyruvate in a humidified atmosphere of 5% C0₂ at 37° C. Cells were seeded into 384-well plates and incubated in a humidified atmosphere of 5% C0₂ at 37° C. Compounds were added 24 h post cell seeding. At the same time, a time zero untreated cell plate was generated. After a 72 h incubation period, cells were fixed and stained with fluorescently labeled antibodies and nuclear dye to allow visualization of nuclei, apoptotic cells and mitotic cells.

Apoptotic cells were detected using an anti-active caspase-3 antibody. Mitotic cells were detected using an anti phospho-histone-3 antibody. Compounds were serially diluted 3 -fold and assayed over 10 concentrations in a final assay concentration of 0.1% DMSO from the highest test concentration specified in the sample information chapter. Automated fluorescence microscopy was carried out using a GE Healthcare IN Cell Analyzer 1000, and images were collected with a 4X objective. Twelve bit tiff images were acquired using the InCell Analyzer 1000 3.2 and analyzed with Developer Toolbox 1.6 software. EC₅₀ and IC₅₀ values were calculated using nonlinear regression to fit data to a sigmoidal 4 point, 4 parameter One-Site dose response model, where: y (fit) = A + [(B - A)/(l + ((C/x) ^Λ D))]. Curve-fitting, EC₅₀ / IC₅₀ calculations and report generation are performed using a custom data reduction engine MathlQ based software (AIM).

[0087] FIG. 3 is a bar graph showing cancer cell lines grouped by cancer type, where the number indicates the number of cancer cell lines in the given cancer type, versus the IC₅₀. As shown in FIG. 3, Compound 1 demonstrated an IC₅₀ of less than 2 μΜ for several different cell lines of solid tumors including cancers from bladder, brain/CNS, breast, colon, endocrine, head and neck, kidney, liver, lung, melanoma, pancreas, prostate, sarcoma, skin, and stomach.

[0088] EXAMPLE 4. Antiproliferative activity on solid tumor cell lines, 2d cultures [0089] Cells were seeded at different densities (ranging from 1000 to 5000) in 384 well plates in a final 50 μΐ volume of the recommended medium and treated for 72 h with Compound 1 or INCBO 18424. Cell viability was measured as a function of the ATP concentration using Cell Titer Glo (Promega) at day 0 and day 3. IC₅₀ was determined by normalizing data to baseline

(dimethylsulfoxide (DMSO) treated) and plotted as a function of percent inhibition versus concentration using dose response model 205 of XLfit version 4.2.

[0090] As shown in Table I, Compound 1 exhibits a varying degree of

antiproliferative/cytostatic activity (IC₅₀: 0.82-3.7 μΜ) in different solid tumor cell lines, while INCBO 18424 is inactive up to 10 μΜ in the same tumor cell lines.

TABLE 1

IC50 (μΜ, n=3)

SAR302503 INCBO 18424

Set-2 0.199 ± 0.027 0.026 ± 0.006

Ovarian cancer MDA-H2774 0.934 ± 0.014 >10

Colorectal cancer HCT116 1.259 ± 0.021 >10

Pancreatic cancer KP4 3.714 ± 0.657 >10

Pancreatic cancer Mia-PACA2 0.828 + 0.075 >10

Prostate Cancer DU145 0.882 ± 0.076 >10

Prostate Cancer LNCaP 1.212 ± 0.131 >10

Breast cancer MDA-MB-468 2.336 ± 0.740 >10

Breast cancer SK-BR3 3.206 ± 0.127 >10

NSCLC NCI-H1975 1.288 + 0.347 >10

NSCLC NCI-H1650 \ .156 ± 0.190 >10

NSCLC NCI-H1299 1.616 + 0.09% >10

NSCLC A549 1.815 + 0.152 >10

[0091] EXAMPLE 5. Antiproliferative activity on solid tumor cell lines, Ras mutant cell lines

[0092] As shown in Table 2, Compound 1 inhibits colony formation of Ras mutant cell lines more potently than other JAK inhibitors. A soft agar assay was performed in 384 well plates using 2000 cells/well/50 uL of 0.4% Noble agar (Difco) in vendor recommended media supplemented with 10% fetal bovine serum (GIBCO) and then incubated at 37° C, 5% C0₂. At 72 h incubation, the indicated compound was added to designated wells at a dilution range of 15 uM-7 nM at a three-fold dose response and incubated for an additional 72 h under the above mentioned incubation conditions. In addition to the JAK2 inhibitors SAR302503, INCB018424, and

AZD1480, a pan-JAK inhibitor, and the EGFR inhibitor, AG490, (Selleckbio) were included as a control. At day 7, 10 uL of Alamar blue was dispensed to each well and relative fluorescence was read and recorded for each well using an Envision 2104 plate reader (Perkin Elmer). IC₅₀ curve data was generated as a % of maximum inhibition normalized to DMSO (Sigma) and Brij L23 (Sigma) treated samples.

[0093] TABLE 2

IC50 (μΜ)

Cancer Type SAR302503 INCB018424 AZD1480 pan-Jak inh AG490

Colorectal DLD1 0.372 >15 10.0 0.829 15.0

Pancreatic PANC-1 2.7 >15 12.3 >15 >15

Colonrectal HCT116 2.4 >15 13.0 >15 >15

Pancreatic CFPAC 3.4 >15 >15 1.6 >15

Kidney ACHN (wt) 0.368 >15 1.2 6.0 >15

[0094] EXAMPLE 5. Antiproliferative activity on solid tumor cell lines, hepatocellular carcinoma cell lines

[0095] Compound 1 was tested in a clonogenic assay on a panel of hepatocellular carcinoma (HCC) cell lines. All HCC cells were seeded in six well plates at 1000 cells per well in 1 mL 10% fetal bovine supplemented media (GIBCO) according to the vendor specifications. At twenty four hours, the compound was added at a tenfold serial dilution range (10 - 0.5081 uM ). Following a three-week incubation period at 37° C, 5% C0₂, media was removed and cells were stained with crystal violet (Sigma) in 3.7% formaldehyde (Fisher) for two hours at room temperature. Cells were washed with distilled water and colonies were imaged on a Canon 8800 scanner. As shown in FIG. 4, Compound 1 inhibits colony formation of HCC cell lines HLF (A), SK-Hep-1 (B), FILE (C), and HuCCTl (D) at concentrations below 1 μΜ.

[0096] Similar results were obtained with 5 additional HCC lines (SNU182, SNU449, SNU423, SNU475, PLC/PPvF/5 and HepG2).

[0097] EXAMPLE 6. Antiproliferative activity in xenograft models

[0098] FIG. 5A and 5B show the results of male SCID mice injected with DU145 cells and treated with Compound 1 at 100 mg/kg (A), 75 mg/kg (T), or 50 mg/kg (♦), twice daily. FIG. 5 A is a graph of tumor volume over time in days. As shown in FIG. 5 A, 17 days after inoculation with tumor cells, the mice were treated orally with vehicle alone (■) or with the indicated amounts of Compound I. As shown in FIG. 5A, oral Compound 1 resulted in dose dependent inhibition of tumor growth. FIG. 5B is a graph of Compound 1 concentration in tumor tissue (— ) and plasma (— -) over time in hours. As shown in FIG. 5B, plasma and tumor concentrations increased with increasing dose; with higher concentrations reached in tumor tissue compared to plasma.

[0099] EXAMPLE 7. Combination with IL-6 antagonists

[00100] As demonstrated herein, Compound 1 is able to block IL-6 stimulated pathways in a panel of tumor cell lines representing different cancer types (prostate, breast, lung, colorectal, pancreas, etc).

[00101] DU145 cells were plated as described in Example 1. pSTAT3, pER l/2, and ER levels measured from cell lysates as described in Example 1 using the appropriate antibody (Cell Signaling Technologies) according to the vendor's instructions.

[00102] In all the cell lines shown herein, Compound 1 efficiently blocks basal and IL-6 induced STAT3 tyrosine phosphorylation. However as shown in FIG. 6, while ER 1/2 phosphorylation in DU145 cells that had not been stimulated with hrIL-6 (NS) was inhibited by Compound 1 , ER 1/2 phosphorylation upon stimulation with hrIL-6 was not blocked in the presence of 1 or 0.1 uM of Compound 1 (FIG. 6A). FIG. 6B and 6C show the effect of

INCB18424 (B) and AZD1480 (C), respectively, on the level of pERK in DU145 cells stimulated with rhIL-6 or not stimulated with rhIL-6 (NS). Therefore, a more complete anti-proliferative and anti-tumor activity will likely be achieved by combining Compound 1 with an IL-6 antagonist, such as an anti-IL-6R antibody that is able to bind IL-6R and block IL-6 intracellular signaling.

[00103] DU145 cells were plated and treated with the indicated amounts of Compound 1 and rhIL6 as described in Example 1. Anti-IL-6R mAb (Sarilumab) was added at 1 ug/mL or 10 ug/mL to samples as shown in FIG.7. As shown in FIG. 7, combined treatment with Compound 1 and anti-IL-6R mAb at 1 ug/mL or 10 ug/mL resulted in increased inhibition of STAT3 phosphorylation compared to samples that had not been treated with anti-IL-6R mAb (compare +IL6 samples treated with 1, 0.5, and 0.1 uM SAR302503 and 0, 1, and 10 ug/mL anti-IL-6R mAb.

[00104] While there have been shown and described what are at present considered the preferred embodiments of the invention, those skilled in the art may make various changes and modifications which remain within the scope of the appended claims.

Claims

WE CLAIM:

1. A composition comprising a JAK2 inhibitor, N-tert-butyl-3-[(5-methyl-2-{[4-(2-pyrrolidin- l-ylethoxy)phenyl] amino }pyrimidin-4-yl)amino]benzenesulfonamide or a pharmaceutically acceptable salt thereof, and an IL-6 antagonist.

2. The composition of claim 1, further comprising a pharmaceutically acceptable carrier.

3. The composition of claim 1, wherein the JAK2 inhibitor and IL-6 antagonist are in amounts that produce a synergistic effect in alleviating a cancer symptom in a patient when said composition is administered to the patient.

4. The composition of any one of claims 1-3, wherein the IL-6 antagonist is an anti-IL-6R antibody or IL-6R binding portion thereof.

5. A method of treating a patient comprising administering a therapeutically effective amount of a JAK2 inhibitor, N-tert-butyl-3-[(5-methyl-2-{[4-(2-pyrrolidin-l- ylethoxy)phenyl]amino}pyrimidin-4-yl)amino]benzenesulfonamide or a pharmaceutically acceptable salt thereof, wherein the patient has a cancer that involves IL-6 receptor signaling.

6. A method of treating a patient comprising administering a first dosage of a JAK2 inhibitor, N-tert-butyl-3 -[(5 -methyl-2- { [4-(2-pyrrolidin- 1 -ylethoxy)phenyl] amino } pyrimidin-4- yl)amino]benzenesulfonamide and a second dosage of an IL-6 antagonist, wherein the patient has a cancer that involves IL-6 receptor signaling.

7. The method of claim 5 or 6, wherein administration results in tumor stasis in said patient.

8. The method of claim 5 or 6, wherein said first and second dosages exhibit a combined synergistic effect in alleviating a cancer symptom in said patient.

9. The method of claim 5 or 6, wherein said first and second dosages cause a combined effect of tumor stasis in said patient.

10. The method of claim 6, wherein the IL-6 antagonist is selected from the group consisting of an anti-IL-6 antibody or IL-6 binding portion thereof and an anti-IL-6 receptor (IL-6R) antibody or IL-6R binding portion thereof.

11. The method of claim 10, wherein the IL-6 antagonist is an anti-IL-6R antibody or IL-6R binding portion thereof.

12. The method of claim 6, wherein the dosage of the JAK2 inhibitor is administered first.

13. The method of claim 6, wherein the dosage of the IL-6 antagonist is administered first.

14. The method of any one of claims 5-13, wherein the patient has a cancer selected from the group consisting of prostate, ovarian, pancreatic, breast, colorectal, kidney, and hepatocellular cancer.

15. The method of claim 14, wherein the patient has a cancer that has a mutation in a ras gene.

16. The method of claim 15, wherein the cancer is selected from the group consisting of colorectal and pancreatic cancer.

17. A kit comprising: (A) a JAK 2 inhibitor, N-tert-butyl-3-[(5-methyl-2-{[4-(2-pyrrolidin-l- ylethoxy)phenyl]amino}pyrimidin-4-yl)amino]benzenesulfonamide or a pharmaceutically acceptable salt thereof; and (B) instructions for using said compound to treat a patient having a cancer that involves IL-6 signaling.

18. A kit comprising : (A) a JAK 2 inhibitor, N-tert-butyl-3 - [(5 -methyl-2- { [4-(2-pyrrolidin- 1 - ylethoxy)phenyl]amino}pyrimidin-4-yl)amino]benzenesulfonamide or a pharmaceutically acceptable salt thereof; (B) an IL-6 antagonist; and (C) instructions for use, wherein the IL-6 antagonist is selected from the group consisting of an anti-IL-6 antibody or IL-6 binding portion thereof and an anti-IL-6 receptor (IL-6R) antibody or IL-6R binding portion thereof.

19. A kit comprising: (A) a JAK 2 inhibitor, N-tert-butyl-3 -[(5 -methyl-2- {[4-(2-pyrro lidin-1- ylethoxy)phenyl]amino}pyrimidin-4-yl)amino]benzenesulfonamide or a pharmaceutically acceptable salt thereof; and (B) instructions for use of the JAK2 inhibitor in combination with an IL-6 antagonist, wherein the IL-6 antagonist is selected from the group consisting of an anti-IL-6 antibody or IL-6 binding portion thereof and an anti-IL-6 receptor (IL-6R) antibody or IL-6R binding portion thereof.

20. A kit comprising: (A) an IL-6 antagonist and (B) instructions for use of the IL-6 antagonist in combination with a compound a JAK 2 inhibitor, N-tert-butyl-3-[(5-methyl-2-{[4-(2-pyrrolidin- l-ylethoxy)phenyl] amino }pyrimidin-4-yl)amino]benzenesulfonamide or a pharmaceutically acceptable salt thereof; wherein the IL-6 antagonist is selected from the group consisting of an anti-IL-6 antibody or IL-6 binding portion thereof and an anti-IL-6 receptor (IL-6R) antibody or IL-6R binding portion thereof.

21. The kit of any one of claims 17-20, wherein the IL-6 antagonist is an anti-IL-6R antibody or IL-6R binding portion thereof.