JAK1 pathway inhibitors for treatment of cytokine-related disorders

Abstract

The present disclosure relates to JAK1 pathway inhibitors and their use in the treatment of cytokine-related diseases or disorders, such as cytokine release syndrome (CRS), hemophagic lymphocyte hyperplasia (HLH), macrophage activation syndrome (MAS), and CAR-T cell-associated encephalopathy syndrome (CRES).

Description

[0001] This application is a divisional application of Chinese patent application No. 2019800244505 (filing date: February 14, 2019, invention title: JAK1 pathway inhibitor for treating cytokine-related diseases) Technical Field

[0002] The present disclosure relates to JAK1 pathway inhibitors and their use in treating cytokine-related diseases or disorders. Background Art

[0003] Cytokine-related diseases or disorders are characterized by excessive immune activation and include cytokine release syndrome (CRS), hemophagocytic lymphohistiocytosis (HLH), macrophage activation syndrome (MAS), and CAR-T cell-related encephalopathy syndrome (CRES).

[0004] Cytokine release syndrome (CRS) is a direct result of excessive production of inflammatory cytokines caused by supraphysiological levels of immune activation and manifests as a constellation of clinical symptoms, including fever, nausea, fatigue, myalgia, malaise, hypotension, hypoxia, capillary leakage, leading to potential multi-organ toxicity.

[0005] CRS is a harmful side effect of immune-based therapies for serious illnesses such as cancer. Immune-based therapies that can cause CRS include administering monoclonal antibodies (mAbs) to cancer and adoptive T cell therapy in recent years. Lee et al. Blood. 2014, 124 (2): 188-195. For example, chimeric antigen receptor (CAR) T cell therapy uses altered T cells to target cancer and has been approved by the FDA for certain forms of refractory non-Hodgkin lymphoma and pediatric relapsed lymphoblastic leukemia (ALL).

[0006] The cytokine profile associated with CRS encompasses two major cellular sources: T lymphocyte-derived cytokines, including interferon-γ (IFN)-γ, IL-2, IL-6, soluble IL-6 receptor (IL-6R), and granulocyte-macrophage colony-stimulating factor (GM-CSF); and cytokines secreted primarily by monocytes and / or macrophages, such as IL-1β, IL-6, IL-12, IL-18, and tumor necrosis factor (TNF)-α. Xu XJ, Tang YM. Cancer Lett. 2014;343:172-8. Zhang Y. et al., Sci China Life Sci. 2016;59:379-85. Brentjens R. et al., Mol Ther. 2010;18:666-8.

[0007] Regulation of the amplified cytokine response that causes CRS can provide significant clinical benefits. For example, tocilizumab is an antibody against the IL-6 receptor (IL-6R) that reduces the rate of severe CRS and is FDA-approved for CRS. However, the mechanism of action of tocilizumab is limited to only anti-IL-6R.

[0008] Hemophagocytic lymphohistiocytosis (HLH) is another syndrome of excessive or uncontrolled immune activation that occurs primarily in infants between birth and 18 months of age and can also occur in adults. HLH can be primary (familial) or secondary, meaning it occurs in the setting of other infectious, malignant, rheumatic, or metabolic disorders. Symptoms of HLH include cytopenias, hepatosplenomegaly, and fever. Schram, A. and Berliner, N. Blood. 2005. 125(19), 2908-2914.

[0009] Macrophage activation syndrome (MAS) presents clinically in a manner similar to HLH (and is even considered secondary or acquired HLH) and is an episode of increased inflammation associated with infection, rheumatic disease, or malignancy. Borgia, RE, et al. Arthritis Rheumatol., 2018, doi:10.1002 / art.40417, prior to publication. MAS was initially described in association with juvenile idiopathic arthritis and is also increasingly recognized as a complication of other diseases such as juvenile-onset systemic lupus erythematosus (cSLE). Shimizu M. et al. Clin Immunol. 2013 Feb;146(2):73-6. The development of MAS is characterized by a dramatic increase in many proinflammatory cytokines, i.e., a cytokine storm. Borgia, RE, et al. Arthritis Rheumatol., 2018, doi:10.1002 / art.40417, prior to publication. MAS is a life-threatening condition with a high mortality rate: 8%-22% in pediatric autoimmune diseases in general and 10%-22% in MAS with concurrent cSLE. Borgia, RE, et al., Arthritis Rheumatol., 2018, doi:10.1002 / art.40417, prior to publication.

[0010] CAR-T cell-related encephalopathy syndrome (CRES) is the second most common adverse event associated with CAR-T cell therapy after CRS. CRES is usually characterized by a toxic encephalopathy state, with symptoms of confusion and delirium as well as occasional attacks and cerebral edema. The manifestation of CRES can be biphasic, with symptoms appearing within the first 5 days and / or 3-4 weeks after cellular immunotherapy. It is believed that the pathophysiological mechanism involves the passive diffusion of cytokines into the brain of patients treated with CAR-T cell therapy. Reduction or elimination of this mechanism may benefit such patients. Neelapu et al., Nat Rev Clin Oncol. 2018, 15(1)47-62.

[0011] Therefore, there is a need to develop new therapies for treating cytokine-related diseases or disorders. The present application addresses this need and other needs. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] Figure 1 Depicted is a dose-dependent inhibition of intravascular IL-6 concentrations following administration of Compound 1 during anti-CD3 antibody-induced cytokine release syndrome (see Example B).

[0013] Figures 2A-2C Dose-dependent inhibition of T cell-derived cytokines (ie, IL-6, IFNγ, and GM-CSF) following administration of Compound 1 during Concanavalin A-induced cytokine release syndrome is depicted (see Example C). Figure 2A Inhibition of IL-6 is shown. Figure 2B Inhibition of IFNγ is shown. Figure 2C Inhibition by GM-CSF is shown.

[0014] Figures 3A-3C Dose-dependent inhibition of monocyte- and / or macrophage-derived cytokines (ie, IL-12, IL-1β, and IL-18) following administration of Compound 1 during Concanavalin A-induced cytokine release syndrome is depicted (see Example C). Figure 3A Inhibition of IL-12 is shown. Figure 3B Inhibition of IL-1β is shown. Figure 3C Inhibition of IL-18 is shown.

[0015] Figure 4 It was shown that the cytokine IL-5 was not affected by Compound 1 treatment during Concanavalin A-induced cytokine release syndrome (see Example C). Summary of the Invention

[0016] Provided herein are methods for treating a cytokine-related disease or disorder in a subject in need thereof, comprising administering to the patient a therapeutically effective amount of a JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof.

[0017] Provided herein is a JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof for use in treating a cytokine-related disease or disorder in a subject in need thereof.

[0018] Provided herein is a use of a JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for treating a cytokine-related disease or disorder in a subject in need thereof. DETAILED DESCRIPTION

[0019] The present invention particularly provides a method of treating a cytokine-related disease or disorder in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof.

[0020] The methods described herein utilize JAK1 pathway inhibitors, particularly JAK1 selective inhibitors. A JAK1 selective inhibitor is a compound that preferentially inhibits JAK1 activity over other Janus kinases. JAK1 plays an important role in many cytokine and growth factor signaling pathways that can cause or contribute to disease conditions when dysregulated. For example, IL-6 levels are elevated in rheumatoid arthritis, a disease that has been shown to have adverse effects (Fonesca et al., Autoimmunity Reviews, 8:538-42, 2009). Because IL-6 transmits signals at least in part via JAK1, IL-6 can indirectly produce potential clinical benefits by inhibiting JAK1 (Guschin et al., Embo J 14:1421, 1995; Smolen et al., Lancet 371:987, 2008). In addition, in some cancers, JAK1 mutations lead to constitutively undesirable tumor cell growth and survival (Mullighan, Proc Natl Acad Sci US A. 106: 9414-8, 2009; Flex, J Exp Med. 205: 751-8, 2008). In other autoimmune diseases and cancers, elevated levels of inflammatory cytokines that activate JAK1 throughout the body can also cause the disease and / or associated symptoms. Therefore, patients with such diseases may benefit from JAK1 inhibition. Selective inhibitors of JAK1 may be effective while avoiding the unnecessary and potentially adverse effects of inhibiting other JAK kinases.

[0021] JAK1 pathway inhibitors, particularly compound 1 (i.e., {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinyl]piperidin-4-yl}-3-[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile, see Table 1) achieved highly effective dose-dependent modulation of CRS-associated inflammatory cytokines (see, e.g., Examples B and C and Figure 1 、 2A -2C and 3A-3C). Unexpectedly, the treatment profile covers a variety of pathogenic cytokines and is not limited to the IL-6 / IL-6R axis (unlike, for example, tocilizumab). Efficacy is achieved by suppressing cytokines derived from T cells and monocytes / macrophages that are highly clinically associated with the pathogenesis of CRS. In addition, the data presented herein related to JAK1 inhibitor compound 1 show that therapeutic benefits (as demonstrated by unchanged IL-5 levels) are achieved without extensive cytokine immunosuppression ( Figure 4 ).

[0022] In some embodiments, the cytokine-related disease or disorder is cytokine release syndrome (CRS), hemophagocytic lymphohistiocytosis (HLH), macrophage activation syndrome (MAS), and CAR-T cell-related encephalopathy syndrome (CRES).

[0023] In some embodiments, the cytokine-related disease or disorder is cytokine release syndrome (CRS).

[0024] In some embodiments, the cytokine-related disease or disorder is hemophagocytic lymphohistiocytosis (HLH).

[0025] In some embodiments, the cytokine-related disease or condition is macrophage activation syndrome (MAS). In some embodiments, macrophage activation syndrome is associated with systemic juvenile idiopathic arthritis. In some embodiments, macrophage activation syndrome is associated with pediatric systemic lupus erythematosus.

[0026] In some embodiments, the cytokine-related disease or disorder is CAR-T cell-related encephalopathy syndrome (CRES).

[0027] In some embodiments, the present application provides a method for treating cytokine release syndrome in a subject, the method comprising administering to the subject a CAR-T cell therapy and a JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof. In some embodiments, the treatment is amelioration or inhibition. In some embodiments, the treatment is prevention.

[0028] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered concurrently with CAR-T cell therapy.

[0029] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered after administration of CAR-T cell therapy.

[0030] In some embodiments, the CAR-T cell therapy is axicabtagene ciloleucel.

[0031] In some embodiments, the CAR-T cell therapy is tisagenlecleucel.

[0032] In some embodiments, the subject suffers from a B cell malignancy.

[0033] In some embodiments, the subject has diffuse large B-cell lymphoma (DLBCL), primary mediastinal large B-cell lymphoma, high-grade B-cell lymphoma, transformed follicular lymphoma, or acute lymphoblastic leukemia.

[0034] In some embodiments, a JAK1 pathway inhibitor, or a pharmaceutically acceptable salt thereof, is selective for JAK1 over JAK2, JAK3, and TYK2 (i.e., a JAK1 selective inhibitor). For example, a compound described herein, or a pharmaceutically acceptable salt thereof, preferentially inhibits JAK1 over one or more of JAK2, JAK3, and TYK2. In some embodiments, a compound preferentially inhibits JAK1 over JAK2 (e.g., a JAK2 / JAK1 IC inhibitor). 50 Ratio>1). In some embodiments, the compound or salt is about 10-fold more selective for JAK1 than for JAK2. In some embodiments, the compound or salt is about 10-fold more selective for JAK1 than for JAK2, as measured by IC at 1 mM ATP. 50 The compound or salt may be about 3-fold, about 5-fold, about 10-fold, about 15-fold, or about 20-fold more selective for JAK1 than for JAK2, as calculated (eg, see Example A).

[0035] In some embodiments, the JAK1 pathway inhibitor is a compound of Table 1 or a pharmaceutically acceptable salt thereof. The compounds in Table 1 are selective JAK1 inhibitors (selective over JAK2, JAK3, and TYK2). The IC obtained by the method of Example A at 1 mM ATP is 50 The values ​​are shown in Table 1.

[0036] Table 1

[0037]

[0038]

[0039]

[0040]

[0041]

[0042]

[0043]

[0044]

[0045] + means <10 nM (for assay conditions, see Example A)

[0046] ++ means ≤100 nM (for determination conditions, see Example A)

[0047] +++ means ≤300 nM (for determination conditions, see Example A)

[0048] a Data for enantiomer 1

[0049] b Data for enantiomer 2

[0050] In some embodiments, the JAK1 pathway inhibitor is {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinoyl]piperidin-4-yl}-3[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile or a pharmaceutically acceptable salt thereof.

[0051] In some embodiments, the JAK1 pathway inhibitor is {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinoyl]piperidin-4-yl}-3[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile adipate.

[0052] The synthesis and preparation of {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinoyl]piperidin-4-yl}-3[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile and its adipate salt can be found, for example, in U.S. Patent Publication No. 2011 / 0224190, filed March 9, 2011, U.S. Patent Publication No. 2013 / 0060026, filed September 6, 2012, and U.S. Patent Publication No. 2014 / 0256941, filed March 5, 2014, each of which is herein incorporated by reference in its entirety.

[0053] In some embodiments, the JAK1 pathway inhibitor is 4-[3-(cyanomethyl)-3-(3',5'-dimethyl-1H,1'H-4,4'-bipyrazol-1-yl)azetidin-1-yl]-2,5-difluoro-N-[(1S)-2,2,2-trifluoro-1-methylethyl]benzamide or a pharmaceutically acceptable salt thereof.

[0054] In some embodiments, the JAK1 pathway inhibitor is 4-[3-(cyanomethyl)-3-(3',5'-dimethyl-1H,1'H-4,4'-bipyrazol-1-yl)azetidin-1-yl]-2,5-difluoro-N-[(1S)-2,2,2-trifluoro-1-methylethyl]benzamide phosphate.

[0055] The synthesis and preparation of 4-[3-(cyanomethyl)-3-(3',5'-dimethyl-1H,1'H-4,4'-bipyrazol-1-yl)azetidin-1-yl]-2,5-difluoro-N-[(1S)-2,2,2-trifluoro-1-methylethyl]benzamide and its phosphate salt can be found, for example, in U.S. Patent Publication No. 2014 / 0343030, filed May 16, 2014, which is incorporated herein by reference in its entirety.

[0056] In some embodiments, the JAK1 pathway inhibitor is ((2R,5S)-5-{2-[(1R)-1-hydroxyethyl]-1H-imidazo[4,5-d]thieno[3,2-b]pyridin-1-yl}tetrahydro-2H-pyran-2-yl)acetonitrile or a pharmaceutically acceptable salt thereof.

[0057] In some embodiments, the JAK1 pathway inhibitor is ((2R,5S)-5-{2-[(1R)-1-hydroxyethyl]-1H-imidazo[4,5-d]thieno[3,2-b]pyridin-1-yl}tetrahydro-2H-pyran-2-yl)acetonitrile monohydrate.

[0058] The synthesis of ((2R,5S)-5-{2-[(1R)-1-hydroxyethyl]-1H-imidazo[4,5-d]thieno[3,2-b]pyridin-1-yl}tetrahydro-2H-pyran-2-yl)acetonitrile and the characterization of its anhydrous and monohydrate forms are described in U.S. Patent Publication No. 2014 / 0121198, filed on October 31, 2013, and U.S. Patent Publication No. 2015 / 0344497, filed on April 29, 2015, each of which is herein incorporated by reference in its entirety.

[0059] In some embodiments, the compounds of Table 1 are prepared by the synthetic procedures described in U.S. Patent Publication No. 2011 / 0224190, filed March 9, 2011, U.S. Patent Publication No. 2014 / 0343030, filed May 16, 2014, U.S. Patent Publication No. 2014 / 0121198, filed October 31, 2013, U.S. Patent Publication No. 2010 / 0298334, filed May 21, 2010, U.S. Patent Publication No. 2011 / 0343030, filed May 16, 2014, U.S. Patent Publication No. 2014 / 0121198, filed October 31, 2013, 0059951, U.S. Patent Publication No. 2012 / 0149681 filed on November 18, 2011, U.S. Patent Publication No. 2012 / 0149682 filed on November 18, 2011, U.S. Patent Publication No. 2013 / 0018034 filed on June 19, 2012, U.S. Patent Publication No. 2013 / 0045963 filed on August 17, 2012, and U.S. Patent Publication No. 2014 / 0005166 filed on May 17, 2013, each of which is incorporated herein by reference in its entirety.

[0060] In some embodiments, the JAK1 pathway inhibitor is selected from the following compounds or pharmaceutically acceptable salts thereof: U.S. Patent Publication No. 2011 / 0224190 filed on March 9, 2011, U.S. Patent Publication No. 2014 / 0343030 filed on May 16, 2014, U.S. Patent Publication No. 2014 / 0121198 filed on October 31, 2013, U.S. Patent Publication No. 2010 / 0298334 filed on May 21, 2010, U.S. Patent Publication No. 2010 / 0343030 filed on May 16, 2014, 011 / 0059951, U.S. Patent Publication No. 2012 / 0149681 filed on November 18, 2011, U.S. Patent Publication No. 2012 / 0149682 filed on November 18, 2011, U.S. Patent Publication No. 2013 / 0018034 filed on June 19, 2012, U.S. Patent Publication No. 2013 / 0045963 filed on August 17, 2012, and U.S. Patent Publication No. 2014 / 0005166 filed on May 17, 2013, each of which is incorporated herein by reference in its entirety.

[0061] In some embodiments, the JAK1 pathway inhibitor is a compound of formula I

[0062]

[0063] or a pharmaceutically acceptable salt thereof, wherein:

[0064] X is N or CH;

[0065] L is C(=O) or C(=O)NH;

[0066] A is phenyl, pyridyl or pyrimidinyl, each optionally substituted with 1 or 2 independently selected R 1 group substitution; and

[0067] Each R 1 is independently fluoro or trifluoromethyl.

[0068] In some embodiments, the compound of Formula I is {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinoyl]piperidin-4-yl}-3[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile or a pharmaceutically acceptable salt thereof.

[0069] In some embodiments, the compound of Formula I is 4-{3-(cyanomethyl)-3-[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-1-yl}-N-[4-fluoro-2-(trifluoromethyl)phenyl]piperidine-1-carboxamide or a pharmaceutically acceptable salt thereof.

[0070] In some embodiments, the compound of Formula I is [3-[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]-1-(1-{[2-(trifluoromethyl)pyrimidin-4-yl]carbonyl}piperidin-4-yl)azetidin-3-yl]acetonitrile or a pharmaceutically acceptable salt thereof.

[0071] In some embodiments, the JAK1 pathway inhibitor is a compound of formula II

[0072]

[0073] or a pharmaceutically acceptable salt thereof, wherein:

[0074] R 2 C 1-6 Alkyl, C 1-6 Haloalkyl, C 3-6 Cycloalkyl or C 3-6 Cycloalkyl-C 1-3 Alkyl, wherein the C 1-6 Alkyl, C 3-6 Cycloalkyl and C 3-6 Cycloalkyl-C 1-3 The alkyl groups are each optionally substituted with 1, 2 or 3 substituents independently selected from fluoro, -CF3 and methyl;

[0075] R 3 is H or methyl;

[0076] R4 is H, F or Cl;

[0077] R 5 H or F;

[0078] R 6 H or F;

[0079] R 7 H or F;

[0080] R 8 is H or methyl;

[0081] R 9 is H or methyl;

[0082] R 10 is H or methyl; and

[0083] R 11 is H or methyl.

[0084] In some embodiments, the compound of Formula II is 4-[3-(cyanomethyl)-3-(3',5'-dimethyl-1H,1'H-4,4'-bipyrazol-1-yl)azetidin-1-yl]-2,5-difluoro-N-[(1S)-2,2,2-trifluoro-1-methylethyl]benzamide or a pharmaceutically acceptable salt thereof.

[0085] In some embodiments, the JAK1 pathway inhibitor is a compound of formula III

[0086]

[0087] or a pharmaceutically acceptable salt thereof, wherein:

[0088] Cy 4 is a tetrahydro-2H-pyran ring, which is optionally substituted by one or two independently selected from CN, OH, F, Cl, C 1-3 Alkyl, C 1-3 Haloalkyl, cyano-C 1-3 Alkyl, HO-C 1-3 Alkyl, amino, C 1-3 Alkylamino and di(C 1-3 alkyl)amino group, wherein the C 1-3 Alkyl and di(C 1-3 Alkyl)amino is optionally substituted by 1, 2 or 3 independently selected from F, Cl, C 1-3 Alkylaminosulfonyl and C 1-3 The substituent of the alkylsulfonyl group is substituted; and

[0089] R 12It is -CH2-OH, -CH(CH3)-OH or -CH2-NHSO2CH3.

[0090] In some embodiments, the compound of Formula III is ((2R,5S)-5-{2-[(1R)-1-hydroxyethyl]-1H-imidazo[4,5-d]thieno[3,2-b]pyridin-1-yl}tetrahydro-2H-pyran-2-yl)acetonitrile or a pharmaceutically acceptable salt thereof.

[0091] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered in a total daily amount of about 100 mg to about 600 mg calculated as the free base. Thus, in some embodiments, the selective JAK1 pathway inhibitor is administered in a total daily amount of about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, or about 600 mg calculated as the free base.

[0092] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered in a total daily amount of about 200 mg calculated as the free base.

[0093] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered in a total daily amount of about 300 mg calculated as the free base.

[0094] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered in a total daily amount of about 400 mg calculated as the free base.

[0095] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered in a total daily amount of about 500 mg calculated as the free base.

[0096] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered in a total daily amount of about 600 mg calculated as the free base.

[0097] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered once daily in an amount of about 200 mg calculated as the free base.

[0098] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered once daily in an amount of about 300 mg calculated as the free base.

[0099] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered once daily in an amount of about 400 mg calculated as the free base.

[0100] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered once daily in an amount of about 500 mg calculated as the free base.

[0101] In some embodiments, the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered once daily in an amount of about 600 mg calculated as the free base.

[0102] In some embodiments, the JAK1 pathway inhibitor, or a pharmaceutically acceptable salt thereof, is administered as one or more sustained release dosage forms each comprising a JAK1 pathway inhibitor, or a pharmaceutically acceptable salt thereof.

[0103] Provided herein is a method for treating a cytokine-related disease or condition in a subject in need thereof, the method comprising administering to the subject a daily dose of about 100 mg to 600 mg of a JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof as a free base, wherein the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is administered in one or more sustained-release dosage forms comprising the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof.

[0104] The embodiments described herein are intended to be combined in any suitable combination, just as if the examples were multiple dependent claims (e.g., embodiments relating to selective JAK1 pathway inhibitors and dosages thereof, embodiments relating to any salt form of the compounds disclosed herein, embodiments relating to individual types of cytokine-related diseases or disorders, and embodiments relating to compositions and / or administration can be combined in any combination).

[0105] For example, provided herein is a method for treating a cytokine-related disease or condition in a subject, wherein the cytokine-related disease or condition is selected from the group consisting of cytokine release syndrome (CRS), hemophagocytic lymphohistiocytosis (HLH), macrophage activation syndrome (MAS), or CAR-T cell-related encephalopathy syndrome (CRES), the method comprising administering to the subject a once daily dose of about 200 mg of {1-{1-[3-fluoro-2-(trifluoromethyl)isoxazolamide]}(II) as the free base. 1-[3-fluoro-2-(trifluoromethyl)isonicotinoyl]piperidin-4-yl}-3-[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile or a pharmaceutically acceptable salt thereof, wherein the dose comprises one or more sustained release dosage forms each comprising {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinoyl]piperidin-4-yl}-3-[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile.

[0106] Sustained-release dosage forms of {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinyl]piperidin-4-yl}-3[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile or a pharmaceutically acceptable salt thereof (Table 1, Compound 1) can be found in U.S. Publication No. 2015 / 0065484, filed August 6, 2014, which is incorporated herein by reference in its entirety.

[0107] For the sake of brevity only, not all possible combinations are listed individually in this article.

[0108] The compounds described herein may be asymmetric (e.g., having one or more stereocenters). Unless otherwise indicated, all stereoisomers such as enantiomers and diastereomers are intended. Compounds containing asymmetrically substituted carbon atoms can be separated in optically active or racemic forms. Methods for preparing optically active forms from optically inactive starting materials are known in the art, such as by resolving racemic mixtures or by stereoselective synthesis. Many geometric isomers such as alkenes, C=N double bonds, etc. may also be present in the compounds described herein, and all such stable isomers are encompassed in the present invention. The cis and trans geometric isomers of the compounds of the present invention are described and can be separated as isomer mixtures or as separated isomers.

[0109] In some embodiments, the compound has the (R)-configuration. In some embodiments, the compound has the (S)-configuration.

[0110] The resolution of the racemic mixture of compound can be carried out by any of many methods known in the art. One exemplary method includes using the chiral resolution acid of the salified organic acid as optically active to carry out fractional crystallization. The resolving agent suitable for fractional crystallization is such as optically active acid, such as tartaric acid, diacetyltartaric acid, dibenzoyltartaric acid, mandelic acid, malic acid, lactic acid or a variety of optically active camphorsulfonic acid, such as D and L forms of β-camphorsulfonic acid. Other resolving agents suitable for fractional crystallization include the stereoisomerically pure form (such as S and R form, or diastereoisomerically pure form) of α-methylbenzylamine, 2-phenylglycinol, norephedrine, ephedrine, N-methylephedrine, cyclohexylethylamine, 1,2-diaminocyclohexane etc.

[0111] The resolution of the racemic mixture can also be carried out by elution on a column filled with an optically active resolving agent (eg dinitrobenzoylphenylglycine). The composition of the appropriate elution solvent can be determined by those skilled in the art.

[0112] The compounds described herein also include tautomeric forms. Tautomeric forms are produced by the exchange of a single bond with an adjacent double bond accompanied by proton migration. Tautomeric forms include proton transfer tautomers, which are isomeric protonation states with the same empirical formula and total charge. Example proton transfer tautomers include keto-enol pairs, amide-imidic acid pairs, lactam-lactim pairs, enamine-imine pairs, and cyclic forms, in which protons can occupy two or more positions in a heterocyclic ring system, such as 1H-imidazole and 3H-imidazole, 1H-1,2,4-triazole, 2H-1,2,4-triazole and 4H-1,2,4-triazole, 1H-isoindole and 2H-isoindole, and 1H-pyrazole and 2H-pyrazole. Tautomeric forms can be in equilibrium or sterically locked into one form by appropriate substitution.

[0113] The compounds described herein may also include isotopically labeled compounds of the present disclosure. An "isotopically" or "radiolabeled" compound is a compound of the present disclosure in which one or more atoms are replaced or substituted with an atom having an atomic mass or mass number different from the atomic mass or mass number normally found in nature (i.e., naturally occurring). Suitable radionuclides that may be incorporated into the compounds of the present disclosure include, but are not limited to 2 H (deuterium is also written as D), 3 H (tritium is also written as T), 11 C. 13 C. 14 C. 13 N. 15 N. 15 O. 17 O. 18 O. 18 F. 35 S. 36 Cl, 82 Br, 75 Br, 76 Br, 77 Br, 123 I. 124 I. 125 I and 131 I. For example, one or more hydrogen atoms in the compounds of the present disclosure may be replaced by a deuterium atom (e.g., C 1-6 One or more hydrogen atoms of an alkyl group may be optionally replaced with a deuterium atom, e.g., -CD3 replacing -CH3). Unless the name indicates a specific stereoisomer, the term "compound" as used herein is intended to include all stereoisomers, geometric isomers, tautomers, and isotopes of the depicted structure. Unless otherwise indicated, a compound identified herein as a particular tautomeric form by name or structure is intended to include other tautomeric forms.

[0114] All compounds and pharmaceutically acceptable salts thereof may be found together with other substances such as water and solvents (eg, hydrates and solvates), or may be isolated.

[0115] In some embodiments, the compounds described herein or their salts are substantially isolated. "Substantially isolated" means that the compound is at least partially or substantially separated from the environment in which it is formed or detected. Partial isolation can include, for example, a composition enriched for the compounds described herein. Substantially isolated can include a composition containing at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% by weight of the compounds described herein or their salts. Methods for isolating compounds and their salts are conventional methods in the art.

[0116] The phrase "pharmaceutically acceptable" as used herein refers to those compounds, substances, compositions and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and animals without excessive toxicity, irritation, allergic response, or other problems or complications, and commensurate with a reasonable benefit / risk ratio.

[0117] As used herein, the expressions "ambient temperature" and "room temperature" or "rt" are understood in the art and generally refer to temperatures, e.g., reaction temperatures, about the temperature of the room in which the reaction is carried out, e.g., about 20°C to about 30°C.

[0118] The present invention also includes pharmaceutically acceptable salts of the compounds described herein. As used herein, "pharmaceutically acceptable salts" refers to derivatives of the disclosed compounds wherein the parent compound is modified by converting an existing acid or base moiety into its salt form. Examples of pharmaceutically acceptable salts include, but are not limited to, inorganic or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; and the like. Pharmaceutically acceptable salts of the present invention include, for example, conventional non-toxic salts of the parent compound formed from non-toxic inorganic or organic acids. Pharmaceutically acceptable salts of the present invention can be synthesized from parent compounds containing a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or free base form of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, non-aqueous media such as diethyl ether, ethyl acetate, alcohols (e.g., methanol, ethanol, isopropanol, or butanol), or acetonitrile (ACN) are preferred. Lists of suitable salts are found in Remington's Pharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, Pa., 1985, p. 1418 and Journal of Pharmaceutical Science, 66, 2 (1977), each of which is herein incorporated by reference in its entirety.

[0119] As used herein, the terms "subject" or "individual" or "patient" are used interchangeably and refer to any animal, including mammals, preferably mice, rats, other rodents, rabbits, dogs, cats, pigs, cattle, sheep, horses, or primates, and most preferably humans. In some embodiments, the "subject" or "patient" is in need of such treatment.

[0120] In some embodiments, the inhibitor is administered in a therapeutically effective amount. As used herein, the phrase "therapeutically effective amount" refers to the amount of an active compound or agent that causes a biological or medical response sought by a researcher, veterinarian, doctor of medicine, or other clinician in a tissue, system, animal, subject, or human. In some embodiments, the dose of the compound or its pharmaceutically acceptable salt administered to a patient or subject is from about 1 mg to about 2 g, from about 1 mg to about 1000 mg, from about 1 mg to about 500 mg, from about 1 mg to about 200 mg, from about 1 mg to about 100 mg, from about 1 mg to 50 mg, or from about 50 mg to about 500 mg. In some embodiments, the dose of the compound or its pharmaceutically acceptable salt is about 200 mg.

[0121] As used herein, the terms "treating" or "treatment" refer to one or more of the following: (1) inhibiting a disease; e.g., inhibiting a disease, disorder, or condition in a subject who is experiencing or exhibiting the lesions or symptoms of the disease, disorder, or condition (i.e., preventing the lesions and / or symptoms from developing further); and (2) ameliorating a disease; e.g., ameliorating a disease, disorder, or condition in a subject who is experiencing or exhibiting the lesions or symptoms of the disease, disorder, or condition (i.e., reversing the lesions and / or symptoms), e.g., reducing the severity of the disease; or (3) preventing a disease, disorder, or condition in a subject who is susceptible to the disease, disorder, or condition but does not yet experience or exhibit the lesions or symptoms of the disease. In some embodiments, treating refers to inhibiting or ameliorating a disease. In some embodiments, treatment is preventing a disease.

[0122] Combination therapy

[0123] The methods described herein may further comprise administering one or more additional therapeutic agents. The one or more additional therapeutic agents may be administered to the patient simultaneously or sequentially.

[0124] In some embodiments, the additional therapeutic agent is an IL-6 antagonist or receptor antagonist. In some embodiments, the IL-6 receptor antagonist is tocilizumab.

[0125] In some embodiments, the additional therapeutic agent is an MCP-1 inhibitor. In some embodiments, the additional therapeutic agent is a MIP1B inhibitor. In some embodiments, the additional therapeutic agent is an IL-2R inhibitor. In some embodiments, the additional therapeutic agent is an IL-1R inhibitor. In some embodiments, the additional therapeutic agent is a TNF-α inhibitor.

[0126] In some embodiments, the additional therapeutic agent is an anti-CD25 antibody. In some embodiments, the anti-CD25 antibody is daclizumab.

[0127] In some embodiments, the additional therapeutic agent is an IL-1β antagonist.

[0128] In some embodiments, the additional therapeutic agent is an IL1 receptor antagonist (IL1Ra). In some embodiments, the IL1 receptor antagonist (IL1Ra) is anakinra.

[0129] In some embodiments, the additional therapeutic agent is a corticosteroid. In some embodiments, the corticosteroid is prednisone.

[0130] In some embodiments, any of the aforementioned additional therapeutic agents is further used in combination with a corticosteroid (eg, prednisone).

[0131] In some embodiments, the additional therapeutic agent comprises tocilizumab and a corticosteroid. In some embodiments, the additional therapeutic agent comprises tocilizumab and prednisone.

[0132] Drug preparations and dosage forms

[0133] When used as medicine, JAK1 pathway inhibitors or pharmaceutically acceptable salts thereof can be administered in the form of pharmaceutical compositions. These compositions can be prepared by methods well known in the pharmaceutical field and can be administered by a variety of routes, depending on whether local or systemic treatment is required and the area to be treated. Administration can be topical (including transdermal, epidermal, ocular and mucosal, including intranasal, vaginal and rectal delivery), pulmonary (e.g., by inhalation or insufflation of powder or aerosol, including through a nebulizer; intra-airway or intranasal), oral or parenteral. Parenteral administration includes intravenous, intra-arterial, subcutaneous, intraperitoneal, intramuscular administration or injection or infusion; or intracranial, such as intrathecal or intraventricular administration. Parenteral administration can be in the form of a single bolus dose, or can be, for example, by a continuous infusion pump. Pharmaceutical compositions and preparations for topical administration may include transdermal patches, ointments, lotions, creams, gels, drops, suppositories, sprays, liquids and powders. Conventional pharmaceutical carriers, aqueous, powder or oily bases, thickeners, etc. may be needed or desirable.

[0134] The present invention also includes pharmaceutical compositions containing a JAK1 pathway inhibitor as described herein or a pharmaceutically acceptable salt thereof as an active ingredient, in combination with one or more pharmaceutically acceptable carriers (excipients). In some embodiments, the composition is suitable for topical administration. When manufacturing the composition, the active ingredient is typically mixed with an excipient, diluted by an excipient, or loaded into such a carrier, in the form of, for example, a capsule, a sachet, a paper, or other container. When an excipient acts as a diluent, it may be a solid, semisolid, or liquid substance, acting as a vehicle, carrier, or medium for the active ingredient. Therefore, the composition may be in the form of tablets, pills, powders, lozenges, sachets, cachets, elixirs, suspensions, emulsions, solutions, syrups, aerosols (in solid form or in a liquid medium), ointments containing, for example, up to 10% by weight of the active compound, soft and hard gelatin capsules, suppositories, sterile injectable solutions, and sterile packaged powders.

[0135] In preparing the formulation, the active compound may be ground before combining with the other ingredients to provide an appropriate particle size. If the active compound is substantially insoluble, it may be ground to a particle size of less than 200 mesh. If the active compound is substantially soluble in water, the particle size may be adjusted by grinding to provide a substantially uniform distribution in the formulation, for example, about 40 mesh.

[0136] JAK1 pathway inhibitors can be ground using known milling procedures, such as wet milling, to obtain a particle size suitable for forming tablets and other formulation types. Fine powder (nanoparticle) formulations of JAK1 selective inhibitors can be prepared by methods known in the art, for example, see International Application No. WO 2002 / 000196.

[0137] Some examples of suitable excipients include lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginates, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, polyvinyl pyrrolidone, cellulose, water, syrup, and methylcellulose. The formulation may additionally include: lubricants such as talc, magnesium stearate, and mineral oil; wetting agents; emulsifiers and suspending agents; preservatives such as methylhydroxy-benzoate and propylhydroxy-benzoate; sweeteners; and flavorings. The compositions of the present invention can be formulated to provide rapid, sustained, or delayed release of the active ingredient after administration to a patient by employing procedures known in the art.

[0138] The compositions can be formulated in unit dosage form, each dosage containing from about 5 to about 1000 mg (1 g), more usually from about 100 to about 500 mg, of the active ingredient. The term "unit dosage form" refers to physically discrete units suitable as unitary dosages for human subjects and other mammals, each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect, in association with a suitable pharmaceutical excipient.

[0139] In some embodiments, the compositions of the present invention contain about 5 to about 50 mg of active ingredient. One of ordinary skill in the art will appreciate that this embodies compositions containing about 5 to about 10 mg, about 10 to about 15 mg, about 15 to about 20 mg, about 20 to about 25 mg, about 25 to about 30 mg, about 30 to about 35 mg, about 35 to about 40 mg, about 40 to about 45 mg, or about 45 to about 50 mg of active ingredient.

[0140] In some embodiments, the compositions of the present invention contain from about 50 to about 500 mg of active ingredient. One of ordinary skill in the art will appreciate that this embodies compositions containing from about 50 to about 100 mg, from about 100 to about 150 mg, from about 150 to about 200 mg, from about 200 to about 250 mg, from about 250 to about 300 mg, from about 350 to about 400 mg, or from about 450 to about 500 mg of active ingredient.

[0141] In some embodiments, the compositions of the present invention contain from about 500 to about 1000 mg of active ingredient. One of ordinary skill in the art will appreciate that this embodies compositions containing from about 500 to about 550 mg, from about 550 to about 600 mg, from about 600 to about 650 mg, from about 650 to about 700 mg, from about 700 to about 750 mg, from about 750 to about 800 mg, from about 800 to about 850 mg, from about 850 to about 900 mg, from about 900 to about 950 mg, or from about 950 to about 1000 mg of active ingredient.

[0142] Similar dosages of the compounds described herein can be used in the methods and uses of the present invention.

[0143] The active compound can be effective over a wide dosage range and is generally administered in a pharmaceutically effective amount. However, it should be understood that the actual amount of compound administered will generally be determined by the physician based on the relevant circumstances, including the condition to be treated, the selected route of administration, the actual compound administered, the age, weight and response of the individual patient, the severity of the patient's symptoms, etc.

[0144] To prepare solid compositions such as tablets, the principal active ingredient is mixed with a pharmaceutical excipient to form a solid preformulation composition containing a homogeneous mixture of a compound of the invention. When these preformulation compositions are referred to as homogeneous, the active ingredient is typically dispersed evenly throughout the composition so that the composition can be readily subdivided into equally effective unit dosage forms, such as tablets, pills, and capsules. This solid preformulation is then subdivided into unit dosage forms of the type described above, containing, for example, from about 0.1 to about 1000 mg of the active ingredient of the invention.

[0145] The tablets or pills of the present invention can be coated or otherwise compounded to provide a dosage form that provides the advantage of long-term action. For example, the tablet or pill can include an inner formulation component and an outer formulation component, the latter being in the form of a film coating on the former. The two components can be separated by an enteric layer that is used to resist disintegration in the stomach and allows the inner component to enter the duodenum intact or to be delayed in release. A variety of materials can be used for such enteric layers or coatings, including many polymeric acids and mixtures of polymeric acids with materials such as shellac, cetyl alcohol, and cellulose acetate.

[0146] Liquid forms in which the compounds and compositions of the present invention can be incorporated for oral administration or injection include aqueous solutions, suitably flavored syrups, aqueous or oily suspensions, and emulsions flavored with edible oils such as cottonseed oil, sesame oil, coconut oil, or peanut oil, as well as elixirs and similar pharmaceutical vehicles.

[0147] Compositions for inhalation or insufflation include solutions and suspensions and powders in pharmaceutically acceptable aqueous or organic solvents or mixtures thereof. Liquid or solid compositions may contain suitable pharmaceutically acceptable excipients as described above. In some embodiments, the composition is administered by oral or nasal respiratory route to achieve local or systemic effects. The composition can be atomized using an inert gas. The atomized solution can be breathed directly from the atomizing device, or the atomizing device can be connected to a mask, tent, or intermittent positive pressure breathing machine. The solution, suspension, or powder composition can be administered orally or nasally from a device that delivers the formulation in an appropriate manner.

[0148] Topical preparations can contain one or more conventional carriers. In some embodiments, ointments can contain water and one or more hydrophobic carriers selected from, for example, liquid paraffin, polyoxyethylene alkyl ether, propylene glycol, white petrolatum. The carrier composition of the cream can be based on water with glycerol and one or more other components (for example, glyceryl monostearate, PEG-glyceryl monostearate and cetearyl alcohol) combination. Gels can use isopropyl alcohol and water, suitably formulated in combination with other components such as glycerol, hydroxyethyl cellulose and its analogs. In some embodiments, topical preparations contain at least about 0.1wt%, at least about 0.25wt%, at least about 0.5wt%, at least about 1wt%, at least about 2wt% or at least about 5wt% compound as described herein. Topical preparations can be, for example, suitably packaged in tubes with 100g, and the tubes are optionally connected to instructions for treating, for example, psoriasis or other skin disorders.

[0149] The amount of compound or composition administered to a patient will vary depending on the substance being administered, the purpose of administration (e.g., prevention or treatment), the patient's condition, the mode of administration, and the like. In therapeutic applications, the composition can be administered to a patient already suffering from a disease in an amount sufficient to cure or at least partially prevent the symptoms of the disease and its complications. The effective dose will depend on the condition being treated and the judgment of the attending clinician, depending on factors such as the severity of the disease, the patient's age, weight, and overall condition.

[0150] The composition administered to the patient may be in the form of a pharmaceutical composition as described above. These compositions may be sterilized by conventional sterilization techniques or may be sterile filtered. The aqueous solution may be packaged for use as is or lyophilized, the lyophilized formulation being combined with a sterile aqueous carrier prior to administration. The pH of the compound formulation will generally be between 3 and 11, more preferably between 5 and 9 and most preferably between 7 and 8. It will be understood that the use of some of the above-mentioned excipients, carriers, or stabilizers will result in the formation of pharmaceutical salts.

[0151] The therapeutic dose of the compounds of the present invention may vary according to, for example, the specific use of the treatment, the mode of administration of the compound, the health and condition of the patient, and the judgment of the prescribing physician. The ratio or concentration of the compounds described herein in the pharmaceutical composition may vary depending on many factors including dosage, chemical properties (e.g., hydrophobicity), and route of administration. For example, the compounds described herein may be provided in an aqueous physiological buffer solution containing about 0.1 to about 10% w / v of the compound for parenteral administration. Some typical dosage ranges are from about 1 μg to about 1 g per kilogram of body weight per day. In some embodiments, the dosage range is from about 0.01 mg to about 100 mg per kilogram of body weight per day. The dosage may depend on, for example, the type and degree of progression of the disease or condition, the overall health status of the particular patient, the relative biological efficacy of the selected compound, the formulation of the excipient, and the variable of its route of administration. The effective dose can be extrapolated from a dose-response curve derived from an in vitro or animal model test system.

[0152] The compositions of the present invention may also include one or more additional agents, such as chemotherapeutic agents, steroids, anti-inflammatory compounds, or immunosuppressants, examples of which are listed herein.

[0153] medicine box

[0154] The present invention also includes a kit that can be used, for example, to treat and / or prevent cytokine-related diseases or conditions, such as CRS, comprising one or more containers containing a pharmaceutical composition comprising a therapeutically effective amount of a compound described herein. If necessary, such kits may also include one or more of a variety of conventional kit components, such as containers with one or more pharmaceutically acceptable carriers, other containers, etc., as will be apparent to those skilled in the art. Instructions, administration instructions, and / or component mixing instructions indicating the amount of the component to be administered, as an insert or as a label, may also be included in the kit.

[0155] Example

[0156] The present invention will be described in more detail with reference to specific examples. The following examples are provided for illustrative purposes and are not intended to limit the present invention in any way. One skilled in the art will readily recognize a number of non-critical parameters that can be varied or modified to yield substantially the same results. The compounds of the examples have been found to be JAK inhibitors according to at least one of the assays described herein.

[0157] Example A: In vitro JAK kinase assay

[0158] According to the following in vitro assay described in Park et al., Analytical Biochemistry 1999, 269, 94-104, the inhibitory activity of JAK1 pathway inhibitors that can be used to treat cytokine-related diseases or disorders on JAK targets was tested. The catalytic domains of human JAK1 (aa837-1142), JAK2 (aa828-1132) and JAK3 (aa781-1124) with an N-terminal His tag were expressed in insect cells using baculovirus and purified. The catalytic activity of JAK1, JAK2 or JAK3 was determined by measuring the phosphorylation of biotinylated peptides. The phosphorylated peptides were detected by homogeneous time-resolved fluorescence (HTRF). The IC of the compound against each kinase was measured in 40 microliters of reaction containing enzyme, ATP and 500nM peptide in 50mM Tris (pH 7.8) buffer with 100mM NaCl, 5mM DTT and 0.1mg / mL (0.01%) BSA. 50 For 1mM IC 50 The ATP concentration in the reaction was 1 mM. The reaction was carried out at room temperature for 1 hour and then terminated with 20 μL of 45 mM EDTA, 300 nM SA-APC, 6 nM Eu-Py20 in an assay buffer (Perkin Elmer, Boston, MA). The binding to the europium-labeled antibody was carried out for 40 minutes and the HTRF signal was measured on a Fusion plate reader (Perkin Elmer, Boston, MA). The compounds in Table 1 were tested in this assay and shown to have IC values ​​also found in Table 1. 50 value.

[0159] Example B: Anti-CD3 Antibody-Induced Cytokine Release Syndrome in BALB / c Mice

[0160] According to the in vivo assay described in Ferran, C. et al. Clin. Exp. Immunol. 1991, 86, 537-543, the efficacy of JAK1 pathway inhibitors against CRS can be tested. Specifically, this study can test the ability of compounds to reduce or improve cytokine release syndrome (CRS) induced by anti-CD3 antibodies in BALB / c mice. Antibody clone 145-2C11 is an immunoglobulin G (IgG) hamster MoAb specific for the epsilon chain of the CD3 murine molecule (Léo, O. et al., Proc. Natl. Acad. Sci. USA, 1987, 34, 1374). Treatment with 145-2C11 induces high-affinity IL-2 receptors on the surface of splenic T cells and causes the release of several cytokines, such as tumor necrosis factor (TNF-α), IL-2, IL-3, IL-6, and interferon-γ (IFN-γ) (Ferran et al., Eur. J. Immunol. 1990, 20, 509-515 and Algre, M. et al., Eur. J. Immunol., 1990, 707). The release of these cytokines causes behavioral changes in animals (e.g., inactivity, piloerection, etc.).

[0161] A. Materials and Methods

[0162] Species / Strain: Mouse: Male BALB / c

[0163] Physiological status: Normal

[0164] Age / weight range at study entry: 6-8 weeks

[0165] Animal supplier: Charles River Laboratories

[0166] Number of animals / sex: 32 male mice in total

[0167] Randomization: Mice were randomized into four (4) groups of eight (8) mice each before the start of the study. Mice.

[0168] Rationale: In the literature, it has been shown that injection of anti-CD3 antibody (clone 145-2C11) induces

[0169] Cytokine release syndrome and its use as a tool for testing potential therapeutics Effective model.

[0170] Replacement No animals were replaced during the study.

[0171] Anti-CD3ε

[0172] Identity and Lot Number: Anti-CD3ε Clone 145-2C11

[0173] Source: BioXCell

[0174] Storage conditions: 4℃

[0175] Vehicle: Sterile saline

[0176] Dosage: 10 μg

[0177] Administration route / volume IV, 100 μL per animal

[0178] Compound: Compound 1 (Jak1 inhibitor) A

[0179] Storage conditions: Room temperature (prepared at room temperature on a tube rotator) Vehicle: 0.5% methylcellulose

[0180] Dosage: 60 mg / kg and 120 mg / kg

[0181] Route / volume: PO, 0.1 mL / 20 g (5 mL / kg)

[0182] Frequency and duration of dosing: QD on Day 0

[0183] A The synthesis and preparation of Compound 1 of Table 1 or {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinoyl]piperidin-4-yl}-3[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile and its adipate salt can be found, for example, in U.S. Patent Publication No. 2011 / 0224190, filed March 9, 2011, U.S. Patent Publication No. 2013 / 0060026, filed September 6, 2012, and U.S. Patent Publication No. 2014 / 0256941, filed March 5, 2014, each of which is incorporated herein by reference in its entirety.

[0184] B. Experimental Design

[0185] The primary objective of this study was to test the ability of JAK1 pathway inhibitors (e.g., Compound 1) to reduce or ameliorate anti-CD3 antibody-induced cytokine release syndrome (CRS) in BALB / c mice. A total of thirty-two (32) BALB / c mice were used for this one-day study. Animals were weighed before test article administration and monitored throughout the experimental period. On day 0, one hour prior to administration of anti-CD3 antibody (1), animals in Groups 2-4 were given a single dose of vehicle (0.5% methylcellulose) or Compound 1 via oral gavage (PO), as detailed in Table 1A. Group 1 served as an untreated control and was not treated. After pretreatment with vehicle or Compound 1 for 1 hour, animals in Groups 2-4 were administered 10 μg of anti-CD3ε antibody (clone 145-2C11) via intravenous injection (IV) to induce CRS. All animals were euthanized by CO2 inhalation 1.5 hours after administration of anti-CD3. Whole blood was collected via cardiac puncture into K2EDTA tubes and stored on ice until plasma processing. Plasma was collected and stored at -80°C until cytokine multiplex analysis.

[0186] Table 1A. Study design

[0187]

[0188] C. Experimental Procedure

[0189] I. Test Article Pretreatment

[0190] On day 0, animals were given vehicle or test article or Compound 1 as shown in Table 1A. Group 2 received a single dose of vehicle (0.5% methylcellulose) at 0.1 mL / 20 g via PO. Group 3 received a single dose of 60 mg / kg Compound 1 at 0.1 mL / 20 g via PO. Group 4 received a single dose of 120 mg / kg Compound 1 at 0.1 mL / 20 g via PO. Group 1 served as an untreated control and was not treated.

[0191] II. Anti-CD3ε Antibody Administration

[0192] One (1) hour after test article administration, anti-CD3ε antibody (clone 145-2C11) was administered to Groups 2-4 via IV injection. Each animal in Groups 2-4 received 10 μg of anti-CD3ε antibody in 0.1 mL.

[0193] III. Lifelong Monitoring

[0194] After administration of anti-CD3 antibodies, animals were closely monitored for signs of distress caused by the resulting systemic inflammatory response. Animals that failed to recover, were cold to the touch, or were moribund were euthanized. Moribund animals were euthanized by CO inhalation, and blood was collected via cardiac puncture and plasma was retained.

[0195] IV. Execution

[0196] One point and one-half (1.5) hours after administration of anti-CD3 antibodies, all animals were euthanized by CO2 inhalation.

[0197] V. Sample Collection

[0198] After sacrifice, whole blood was collected from each animal via cardiac puncture into K2EDTA tubes. The blood was centrifuged and plasma was collected in cryovials. The plasma was frozen and stored at -80°C for downstream cytokine multiplex analysis.

[0199] VI. Cytokine Multiplex Analysis

[0200] Plasma samples were thawed on ice and used for cytokine multiplex analysis according to the manufacturer's protocol (ThermoFisher).

[0201] D.Results

[0202] Compound 1 dose-dependently inhibited the IL-6 concentration in the blood compartment ( Figure 1 This served as confirmation of the bioactivity observed in the Con A preclinical model described in Example C below. Unpaired one-way analysis of variance (ANOVA) with Sidak's multiple test comparison was performed using GraphPad Prism (Version 4.00; GraphPad Software, San Diego, California, USA). P values ​​< 0.05 were considered significant.

[0203] Example C: Concanavalin A-induced cytokine release syndrome

[0204] Concanavalin A (Con A) is a selective T lymphocyte mitogen that causes extensive inflammatory cytokine release and CD4 and CD8 T cell proliferation. Injection of Con-A has been shown in the literature to induce cytokine release syndrome and to be used as a model for testing the efficacy of cytokine release syndrome therapy (Gantner, F. et al. Hepatology, 1995, 21, 190-198). Mitogen response depends on the expression of T cell receptors. Animals exhibit behavioral changes, such as fever, malaise, hypotension, hypoxia, capillary leakage and potential multi-organ toxicity.

[0205] A. Materials and Methods Species / Strain: Mouse: Female BALB / c

[0206] Physiological status: Normal Age / weight range at study entry: 6-8 weeks Animal Supplier: Taconic

[0207] Number of animals / sex: 40 mice in total Randomization: Mice were randomized into five (5) groups before the start of the study.

[0208] There were eight (8) mice per group.

[0209] Rationale: Con-A injection has been shown in the literature to induce cytokine release syndrome and is used as a model to test the efficacy of potential therapies.

[0210] B. Experimental Design

[0211] In detail, this study tested the ability of selective JAK1 inhibitors (e.g., Compound 1 of Table 1) to reduce or improve Con A-induced cytokine release syndrome (CRS) in BALB / c mice. A total of forty (40) BALB / c mice were used for this one-day study. The animals were weighed before the test article was administered and monitored throughout the experimental period. On Day 0, sixty (60) minutes before Con A administration, animals in Groups 2-4 were given a single dose of vehicle (0.5% methylcellulose) or Compound 1 (60 and 120 mg / kg) via oral gavage (PO), as detailed in Table 2A. Group 1 served as an untreated control and was not treated. After 45 minutes of pretreatment with vehicle or Compound 1, animals in Groups 2-4 were administered 20 mg / kg Con A via intravenous injection (IV) to induce CRS. All animals were euthanized by inhalation of CO2 two hours after Con A administration. Whole blood was collected via cardiac puncture into K2EDTA tubes and stored on ice until plasma processing. Plasma was collected and stored at -80°C until cytokine multiplex analysis.

[0212] Table 2A. Study design

[0213]

[0214]

[0215] C. Experimental Procedure

[0216] Day -1

[0217] The animals were weighed and the Con-A dose (20 mg / kg) was calculated.

[0218] Vehicle and compound 1 at corresponding doses were prepared.

[0219] Day 0

[0220] I. Test Article Pretreatment

[0221] On day 0, animals were given vehicle or compound 1, as shown in Table 2A. Group 1 served as an untreated control and received no treatment. Group 2 received a single dose of vehicle (0.5% methylcellulose) at 0.1 mL / 20 g PO. Group 3 received a single dose of 60 mg / kg compound 1 at 0.1 mL / 20 g PO. Group 4 received a single dose of 120 mg / kg compound 1 at 0.1 mL / 20 g PO.

[0222] II. Con-A Administration

[0223] Sixty (60) minutes after administration of the test article, Con-A was administered via IV injection to Groups 2-4. Each animal in Groups 2-4 received 20 mg / kg Con-A in 0.2 mL.

[0224] III. Lifelong Monitoring

[0225] Following Con-A administration, animals were closely monitored for signs of distress resulting from the resulting systemic inflammatory response.

[0226] IV. Execution

[0227] Two hours after Con-A administration, all animals were euthanized by CO2 inhalation.

[0228] V. Sample Collection

[0229] After sacrifice, whole blood was collected from each animal via cardiac puncture into K2EDTA tubes. The blood was centrifuged and plasma was collected in cryovials. The plasma was frozen and stored at -80°C for downstream cytokine multiplex analysis.

[0230] VI. Cytokine Multiplex Analysis

[0231] Plasma samples were thawed on ice and used for cytokine multiplex analysis according to the manufacturer's protocol (ThermoFisher).

[0232] D.Results

[0233] Compound 1 dose-dependently inhibited the IL-6 concentration in the blood compartment ( Figure 2AThis cytokine is key in mediating the pathophysiology of CRS. T cell-derived IFNγ and GM-CSF cytokines were also significantly inhibited, suggesting that compound 1 has therapeutic potential beyond the limited mechanism of action of tocilizumab (only against IL-6R) ( Figure 2B and 2C ).

[0234] Monocyte and / or macrophage derived cytokines were also reduced. A statistically significant dose-dependent reduction in IL-12 was observed ( Figure 3A ), and IL-1β ( Figure 3B ) and IL-18( Figure 3C ), suggesting that JAK1-specific inhibition has therapeutic potential in immune cell types associated with CRS lesions.

[0235] Importantly, the cytokine IL-5 ( Figure 4 ) was not affected by compound 1 treatment, was independent of JAK1, and was not associated with CRS lesions. This data suggests that the efficacy of compound 1 is not mediated through broad, nonspecific immunosuppression.

[0236] Unpaired one-way analysis of variance (ANOVA) with Sidak multivariate comparisons was performed using GraphPad Prism (version 4.00; GraphPad Software, San Diego California, USA). p values ​​< 0.05 were considered significant.

[0237] Example D: Preparation of sustained-release formulation of compound 1

[0238] Sustained-release tablets containing Compound 1 were prepared using the excipients in the amounts shown in the table below. Protocol A was used for SR1 tablets, Protocol B was used for SR2 tablets, Protocol C was used for SR3 tablets and 25 mg SR tablets, and Protocol D was used for SR4 tablets. These procedures are disclosed in U.S. Patent Publication No. 2015 / 0065484, which is directed to sustained-release dosage forms of Compound 1.

[0239] Option A:

[0240] Step 1. Adipate salt of Compound 1, microcrystalline cellulose, hydroxypropyl methylcellulose (Methocel K100LV and Methocel K4M) and lactose monohydrate were sieved individually.

[0241] Step 2. Transfer the screened material from step 1 to a suitable blender and mix.

[0242] Step 3. Transfer the blend from step 2 to a suitable granulator and mix.

[0243] Step 4. Add purified water while mixing.

[0244] Step 5. Transfer the granules from step 4 to a suitable dryer and dry until the LOD is less than 3%.

[0245] Step 6. Screen the granules from step 5.

[0246] Step 7. Mix the screened magnesium stearate with the granules from step 6 in a suitable blender.

[0247] Step 8. Compress the final blend from step 7 on a suitable rotary tablet press.

[0248] Option B:

[0249] Step 1. Compound 1 adipate salt, microcrystalline cellulose, hydroxypropyl methylcellulose and pregelatinized starch were sieved individually.

[0250] Step 2. Transfer the screened material from step 1 to a suitable blender and mix.

[0251] Step 3. Transfer the blend from step 2 to a suitable granulator and mix.

[0252] Step 4. Add purified water while mixing.

[0253] Step 5. Transfer the granules from step 4 to a suitable dryer and dry until the LOD is less than 3%.

[0254] Step 6. Screen the granules from step 5.

[0255] Step 7. Polyox, butylated hydroxytoluene and colloidal silica were sieved individually.

[0256] Step 8. Transfer the granules from step 6 and the material from step 7 to a suitable blender and mix.

[0257] Step 9. Add the screened magnesium stearate to the material in step 8 and continue blending.

[0258] Step 10. Compress the final blend from step 9 on a suitable rotary tablet press.

[0259] Option C:

[0260] Step 1. Sieve lactose monohydrate, adipate salt of Compound 1, microcrystalline cellulose and hydroxypropyl methylcellulose individually through a suitable sieve.

[0261] Step 2. Transfer the screened material from step 1 to a suitable blender and mix.

[0262] Step 3. Transfer the blend from step 2 to a suitable granulator and mix.

[0263] Step 4. Add purified water while mixing.

[0264] Step 5. Sieve the wet granules through a suitable sieve.

[0265] Step 6. Transfer the granules from step 5 to a suitable dryer and dry until the LOD is less than 3%.

[0266] Step 7. Grind the granules from step 6.

[0267] Step 8. Mix the screened magnesium stearate with the granules from step 7 in a suitable blender.

[0268] Step 9. Compress the final blend from step 8 on a suitable rotary tablet press.

[0269] Plan D:

[0270] Step 1. Pregelatinized starch, adipate salt of Compound 1, hydroxypropyl methylcellulose and a portion of the desired microcrystalline cellulose were sieved individually through a suitable sieve.

[0271] Step 2. Transfer the screened material from step 1 to a suitable blender and mix.

[0272] Step 3. Transfer the blend from step 2 to a suitable granulator and mix.

[0273] Step 4. Add purified water while mixing.

[0274] Step 5. Sieve the wet granules through a suitable sieve.

[0275] Step 6. Transfer the granules from step 5 to a suitable dryer and dry until the LOD is less than 3%.

[0276] Step 7. Grind the granules from step 6.

[0277] Step 8. Sieve the remaining portion of microcrystalline cellulose and half of the sodium bicarbonate.

[0278] Step 9. Transfer the milled granules from step 7 and the screened material from step 8 into a suitable blender and mix.

[0279] Step 10. The remaining portion of sodium bicarbonate is sieved and mixed with the blend from step 9.

[0280] Step 11. Magnesium stearate is sieved and mixed with the blend in step 10.

[0281] Step 12. Compress the final blend from step 11 on a suitable rotary tablet press.

[0282] Composition of SR1: 100 mg sustained-release tablets

[0283]

[0284]

[0285] a The conversion factor from adipate to free base is 0.7911 b Add after granulation

[0286] c Removed during processing

[0287] Composition of SR2: 100 mg sustained-release tablets

[0288]

[0289] a The conversion factor from adipate to free base is 0.7911 b Add after granulation

[0290] c Removed during processing

[0291] SR3 (100 mg): Composition of 100 mg sustained-release tablets

[0292]

[0293] a The conversion factor from adipate to free base is 0.7911 b Add after granulation

[0294] c Removed during processing

[0295] Composition of SR4: 100 mg sustained-release tablets

[0296]

[0297]

[0298] a The conversion factor from adipate to free base is 0.7911 b Add after granulation

[0299] c Removed during processing

[0300] d 25 mg SR: Composition of 25 mg sustained-release tablets added partially before granulation and partially after granulation

[0301]

[0302] a The conversion factor from adipate to free base is 0.7911 b Add after granulation

[0303] c Removed during processing

[0304] In addition to those described herein, various modifications of the present invention will be apparent to those skilled in the art from the foregoing description. Such modifications are also intended to be within the scope of the appended claims. Each reference cited in this application, including all patents, patent applications, and publications, is incorporated herein by reference in its entirety.

[0305] In summary, the present invention includes but is not limited to the following:

[0306] 1. A method for treating a cytokine-related disease or disorder in a subject, the method comprising administering to the subject a JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof.

[0307] 2. The method of claim 1 , wherein the JAK1 pathway inhibitor or a pharmaceutically acceptable salt thereof is selective for JAK1 over JAK2, JAK3, and Tyk2.

[0308] 3. The method of claim 1 or 2, wherein the cytokine-related disease or condition is cytokine release syndrome (CRS), hemophagocytic lymphohistiocytosis (HLH), macrophage activation syndrome (MAS) and CAR-T cell-associated encephalopathy syndrome (CRES).

[0309] 4. The method of claim 3, wherein the cytokine-related disease or condition is cytokine release syndrome (CRS).

[0310] 5. The method of claim 3, wherein the cytokine-related disease or disorder is hemophagocytic lymphohistiocytosis (HLH).

[0311] 6. The method of claim 3, wherein the cytokine-related disease or disorder is macrophage activation syndrome (MAS).

[0312] 7. The method of claim 6, wherein the macrophage activation syndrome (MAS) is associated with systemic juvenile idiopathic arthritis.

[0313] 8. The method of claim 3, wherein the cytokine-related disease or condition is CAR-T cell-related encephalopathy syndrome (CRES).

[0314] 9. A method as described in any one of items 1-8, wherein the JAK1 pathway inhibitor is {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinoyl]piperidin-4-yl}-3[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile or a pharmaceutically acceptable salt thereof.

[0315] 10. The method of any one of items 1-8, wherein the JAK1 pathway inhibitor is {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinoyl]piperidin-4-yl}-3[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile adipate.

[0316] 11. A method as described in any one of items 1-8, wherein the JAK1 pathway inhibitor is 4-[3-(cyanomethyl)-3-(3',5'-dimethyl-1H,1'H-4,4'-bipyrazol-1-yl)azetidin-1-yl]-2,5-difluoro-N-[(1S)-2,2,2-trifluoro-1-methylethyl]benzamide or a pharmaceutically acceptable salt thereof.

[0317] 12. A method as described in any one of items 1-8, wherein the JAK1 pathway inhibitor is 4-[3-(cyanomethyl)-3-(3',5'-dimethyl-1H,1'H-4,4'-bipyrazol-1-yl)azetidin-1-yl]-2,5-difluoro-N-[(1S)-2,2,2-trifluoro-1-methylethyl]benzamide phosphate.

[0318] 13. A method as described in any one of items 1-8, wherein the JAK1 pathway inhibitor is ((2R,5S)-5-{2-[(1R)-1-hydroxyethyl]-1H-imidazo[4,5-d]thieno[3,2-b]pyridin-1-yl}tetrahydro-2H-pyran-2-yl)acetonitrile or a pharmaceutically acceptable salt thereof.

[0319] 14. The method of any one of items 1-8, wherein the JAK1 pathway inhibitor is ((2R,5S)-5-{2-[(1R)-1-hydroxyethyl]-1H-imidazo[4,5-d]thieno[3,2-b]pyridin-1-yl}tetrahydro-2H-pyran-2-yl)acetonitrile monohydrate.

[0320] 15. The method of any one of items 1-14, further comprising administering tocilizumab to the subject.

[0321] 16. The method of any one of items 1-14, further comprising administering a corticosteroid to the subject.

[0322] 17. The method of any one of items 1-14, further comprising administering prednisone to the subject.

[0323] 18. The method of any one of items 1-14, further comprising administering tocilizumab and a corticosteroid to the subject.

Claims

1. Use of a JAK1 selective pathway inhibitor or a pharmaceutically acceptable salt thereof in the preparation of a medicament for preventing cytokine release syndrome in a subject in need thereof, wherein the JAK1 selective pathway inhibitor is {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinyl]piperidin-4-yl}-3[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile or a pharmaceutically acceptable salt thereof.

2. The use according to claim 1, wherein the JAK1 selective pathway inhibitor is {1-{1-[3-fluoro-2-(trifluoromethyl)isonicotinyl]piperidin-4-yl}-3[4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl]azetidin-3-yl}acetonitrile adipate.

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