Pharmaceutical compound and composition

Description

/ 83 COMPOUND, AND, PHARMACEUTICAL COMPOSITION DIVIDED FROM BR112016028749-5, FILED ON 06 / 11 / 2015 FIELD

[001] This application relates to new stable salt forms of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide, which are suitable for the preparation of pharmaceutical formulations thereof, and for their therapeutic use. FUNDAMENTALS OF THE INVENTION

[002] Janus kinase (JAK) inhibition has been evaluated in the treatment of hyperproliferative diseases. Several JAK inhibitors have been developed: ruxolitinib, tofacitinib, baricitinib, lestaurtinib, pacritinib, fedratinib, XL019, SB1518, and AZD1480 have been developed (Sonbol, Ther. Adv. Hematol. 4: 15-35, 2013). The compound N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide (CYT-0387) is a JAK kinase inhibitor.

[003] In clinical studies, CYT-0387 is effective in treating hyperproliferative diseases such as polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF). In addition, myelofibrosis patients who received CYT-0387 exhibited improved responses to anemia and / or spleen responses (see U.S. Patent No. 8,486,941 and Publication Application No. 2014-0073643, each of which is incorporated herein by reference).

[004] It is desirable to have different forms of the compound that are suitable for the preparation of pharmaceutical formulations containing CYT0387 and their therapeutic use. BRIEF SUMMARY

[005] The present invention is directed to new forms of CYT0387. Petition 870200056639, dated 07 / 05 / 2020, page 14 / 113 / 83

[006] In one aspect, the present invention is directed to form I of the anhydrous monochloride of CYT-0387: which has an x-ray powder diffraction pattern (XRPD) with peaks at approximately 13.5°, 20.9°, 26.1°, 26.6° and 28.3° 2-θ ± 0.2° 2-θ.

[007] In another aspect, the present invention relates to form II of the monohydrate of CYT-0387 dihydrochloride: which has an x-ray powder diffraction pattern (XRPD) with peaks at approximately 7.7°, 19.3°, 24.0°, 25.7° and 29.6° 2-θ ± 0.2° 2-θ.

[008] In another aspect, the present invention is directed to the form CYT-0387 monohydrochloride anhydrous III (form III): which has an x-ray powder diffraction pattern (XRPD) with peaks at approximately 12.7°, 14.6°, 17.8°, 19.7° and 23.3° 2-θ ± 0.2° 2-θ.

[009] The invention also provides compositions, including pharmaceutical compositions, kits that include the compounds, and methods for using and making the pharmaceutical compositions. The pharmaceutical compositions provided herein are useful in the treatment of diseases, disorders, or conditions that are mediated by JAK. In a certain embodiment, the diseases, disorders Petition 870200056639, dated 07 / 05 / 2020, page 15 / 113 / 83 or conditions mediated by JAK are myeloproliferative disorders and cancer.

[0010] In one embodiment, the application is directed to dosage forms comprising form III of CYT-0387, in particular dosage forms as a tablet, and more particularly dosage forms comprising form III of CYT-0387 in an amount equivalent to between 30 and 250 mg of CYT-0387 free base. In another embodiment, the dosage form comprises form II of CYT-0387 in an amount equivalent to 100 to 200 mg of CYT-0387 free base.

[0011] In additional embodiments, the application is directed to dosage forms or pharmaceutical compositions comprising form III of CYT-0387 in an amount equivalent to 200 mg of CYT-0387 free base, which provides a pharmacokinetic profile similar to a dosage form or pharmaceutical composition comprising anhydrous form I of CYT-0387 dihydrochloride in an amount equivalent to 300 mg of CYT-0387 free base.

[0012] In some embodiments, the application is directed to dosage forms comprising form II of CYT-0387 dihydrochloride monohydrate, as a tablet, wherein the dosage forms comprise form II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to between about 100 mg and about 300 mg of CYT-0387 free base. In some embodiments, the application is directed to dosage forms comprising form II of CYT-0387 dihydrochloride monohydrate, as a tablet, wherein the dosage forms comprise form II of CYT0387 dihydrochloride monohydrate in an amount equivalent to approximately 100 mg, approximately 150 mg, approximately 200 mg, approximately 250 mg, or approximately 300 mg of CYT0387 free base. In certain embodiments, the application is directed to dosage forms comprising form II of CYT-0387 dihydrochloride monohydrate, as a tablet, wherein the dosage forms comprise the form Petition 870200056639, dated 07 / 05 / 2020, p. 16 / 113 / 83 II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to approximately 100 mg of CYT-0387 free base. In certain embodiments, the application is directed to dosage forms comprising CYT-0387 dihydrochloride monohydrate II as a tablet, wherein the dosage forms comprise CYT-0387 dihydrochloride monohydrate II in an amount equivalent to approximately 200 mg of CYT-0387 free base. In certain embodiments, the application is directed to dosage forms comprising form II of CYT-0387 dihydrochloride monohydrate, such as a tablet, wherein the dosage forms comprise form II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to approximately 250 mg of CYT-0387 free base.In certain embodiments, the application is directed to dosage forms comprising CYT-0387 dihydrochloride monohydrate form II, as a tablet, wherein the dosage forms comprise CYT-0387 dihydrochloride monohydrate form II in an amount equivalent to approximately 300 mg of CYT-0387 free base. In further embodiments, the application is directed to dosage forms or pharmaceutical compositions comprising CYT-0387 dihydrochloride monohydrate form II in an amount equivalent to 200 mg of CYT-0387 free base, which provides a substantially similar pharmacokinetic profile to a dosage form or pharmaceutical composition comprising CYT-0387 dihydrochloride monohydrate form II in an amount equivalent to 300 mg of CYT-0387 free base.

[0013] A kit is also provided which includes a compound of formula (I) or a pharmaceutically acceptable salt, isomer or a mixture thereof. The kit may further comprise a label and / or instructions for use of the compound in the treatment of a disease, disorder or condition in a human being who requires it. Petition 870200056639, dated 07 / 05 / 2020, page 17 / 113 / 83

[0014] Also provided are articles of manufacture comprising a compound of formula (I) or a pharmaceutically acceptable salt, isomer or a mixture thereof, and a container. In one embodiment, the container may be a bottle, a jar, an ampoule, a pre-filled syringe or an intravenous bag. DESCRIPTION OF THE FIGURES

[0015] Figure 1: XRPD of anhydrous form I of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride (CYT0387).

[0016] Figure 2: DSC from anhydrous form I of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride (CYT0387).

[0017] Figure 3: TGA from anhydrous form I of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride (CYT0387).

[0018] Figure 4: DVS from anhydrous form I of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride (CYT0387).

[0019] Figure 6: Padrão XRPD para a forma II do mono-hidrato do dicloridrato de N-(cianometil)-4-(2-(4-morfolinofenilamino)pirimidin-4il)benzamida.

[0020] Figure 7: Padrão

[0021] Figure 8: XRPD pad for anhydrous form III of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4yl)benzamida monohydrochloride.

[0022] Figure 9: DSC for form II of monohydrate of dichlorohydrate of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pirimidin-4-yl)benzamida. Petition 870200056639, 07 / 05 / 2020, pág. 18 / 113 / 83

[0023] Figure 10: DSC for the anhydrous form I of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pirimidin-4-yl)benzamida monochloridrate.

[0024] Figure 11: DSC for anhydrous form III of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pirimidin-4-yl)benzamida monochloridrate.

[0025] Figure 12: TGA for form II of monohydrate of dichlorohydrate of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pirimidin-4yl)benzamida.

[0026] Figure 13: TGA for the anhydrous form I of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pirimidin-4-yl)benzamide monochloride.

[0027] Figure 14: TGA for anhydrous form III of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pirimidin-4-yl)benzamide monochloride.

[0028] Figure 15: DVS for form II of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4yl)benzamide dihydrochloride monohydrate. DETAILED DESCRIPTION

[0029] The following description sets forth exemplary compositions and methods, parameters and the like. It should be recognized, however, that such description is not intended to be a limitation on the scope of the present description, but rather it is provided as a description of exemplary embodiments.

[0030] JAK inhibitors CYT-0387, N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide, which is described in U.S. Patent No. 8,486,941, with the structure below:

[0031] The anhydrous dihydrochloride form of CYT-0387 is described in PCT application WO 2012 / 071612. Anhydrous CYT-0387 dihydrochloride has the Petition 870200056639, dated 07 / 05 / 2020, page 19 / 113 / 83, structure below:

[0032] XRPD, differential scanning calorimetry (DSC), thermogravimetric analysis (TGA) and dynamic vapor sorption (DVS) of the anhydrous form I of CYT-0387 dihydrochloride (form IV) are shown in figures 1, 2, 3 and 4, respectively.

[0033] The present application provides other new forms of CYT-387: the anhydrous form I of monohydrochloride (form I), the anhydrous form II of dihydrochloride monohydrate (form II) and the anhydrous form III of monohydrochloride (form III).

[0034] As used in the present invention, the terms “form I” or “form I of CYT-0387” are used to refer to the anhydrous form I of CYT-0387 monohydrochloride; the terms “form II” or “form II of CYT0387” are used to refer to the hydrate of CYT-0387 dihydrochloride, the monohydrate of CYT-0387 dihydrochloride, or form II of CYT-0387 dihydrochloride monohydrate; and the terms “form III” or “form III of CYT0387” are used to refer to the anhydrous form III of CYT-0387 monohydrochloride; The terms "form IV" or "form IV of CYT-0387" are used to refer to the anhydrous form I of CYT-0387 dihydrochloride.

[0035] In one embodiment, the form I or form I of CYT-0387 is characterized by the XRPD shown in figure 7, by the DSC in figure 10, or by the TGA in figure 13. In one embodiment, the form I or form I of CYT0387 has an XRPD pattern with peaks at approximately 13.5°, 20.9°, 26.1°, 26.6° and 28.3° 2-θ ± 0.2° 2-θ.

[0036] In some embodiment, form II or form II of CYT-0387 is characterized by the XRPD shown in figure 6, the DSC in figure 9, the TGA in figure 12, or the DVS in figure 15. In one embodiment, form II Petition 870200056639, dated 07 / 05 / 2020, page 20 / 113 / 83 or form II of CYT-0387 has an XRPD pattern with peaks at approximately 7.7°, 19.3°, 24.0°, 25.7° and 29.6° 2-θ ± 0.2° 2-θ.

[0037] In another embodiment, form III or form III of CYT-0387 is characterized by the XRPD shown in figure 8, by the DSC in figure 11, or by the TGA in figure 14. In one embodiment, form IV or form IV of CYT-0387 has an XRPD pattern with peaks at approximately 12.7°, 14.6°, 17.8°, 19.7° and 23.3° 2-θ ± 0.2° 2-θ.

[0038] In a certain embodiment, form IV or form IV of CYT-0387 is characterized by the XRPD shown in figure 8, by the DSC in figure 11 or by the TGA in figure 14. In one embodiment, form IV or form IV of CYT-0387 has an XRPD pattern with peaks at about 5.5°, 10.1°, 14.9°, 25.1° and 26.6° 2-θ ± 0.2° 2-θ.

[0039] The results of the present application have found that CYT-0387 dihydrochloride monohydrate (form II) has greater stability than other salts or forms of CYT-387 under certain conditions. The results described in this document have also found that such properties of CYT-387 form II make it more suitable for development or adoption in various syntheses or processes. In one embodiment, CYT-387 form II is suitable for use in a pharmaceutical composition in tablet form. Similarly, the studies described in this document have shown that the tablet formulation exhibited bioavailability properties similar to those of the capsule formulations.In certain embodiments, a tablet comprising the monohydrate form II of CYT-0387 dihydrochloride in an amount equivalent to 200 mg of CYT-0387 free base provides similar bioavailability as a capsule comprising the anhydrous form I of CYT-0387 dihydrochloride in an amount equivalent to 300 mg of CYT-0387 free base.

[0040] The results of the present application indicate that the anhydrous form I of CYT-0387 dihydrochloride was hygroscopic and physically unstable when Petition 870200056639, dated 07 / 05 / 2020, page 21 / 113 / 83 exposed to moisture. Similarly, the results described below indicate that form II of CYT-0387 dihydrochloride monohydrate is a thermodynamically stable form of the dihydrochloride salt under suitable manufacturing and / or storage conditions. Furthermore, the present application described that the use of propyl gallate (a free radical scavenging oxidant) was effective in inhibiting or preventing the oxidative degradation of form II of CYT-0387 dihydrochloride monohydrate in an aqueous solution and in a tablet formulation. Furthermore, the results suggest that form II of CYT-0387 dihydrochloride monohydrate exhibits greater bioavailability compared to anhydrous form I of CYT-0387 dihydrochloride and the free base of CYT-0387.

[0041] In yet another embodiment, the present invention also provides monohydrate of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride (CYT-0387). In yet another embodiment, the present invention also provides anhydrous amorphous N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride (CYT-0387) and anhydrous amorphous N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride (CYT-0387). In another embodiment, the present invention also provides amorphous N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide (CYT0387) dihydrochloride and amorphous N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide (CYT-0387) monohydrochloride.

[0042] In particular embodiments, form II of the monohydrate of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4yl)benzamide dihydrochloride (CYT-0387) is in a crystalline form.

[0043] In additional embodiments, the anhydrous form I of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4yl)benzamide monochloride (CYT-0387) is in a crystalline form.

[0044] In certain embodiments, the anhydrous form III of the monochloride Petition 870200056639, dated 07 / 05 / 2020, p. 22 / 113 / 83 of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide (CYT-0387) is in a crystalline form.

[0045] In one embodiment, crystal forms are characterized by the gaps in the interconnection plane determined by an X-ray powder diffraction pattern (XRPD). The XRPD diffractogram is typically represented by a diagram that graphically depicts the intensity of the peaks in relation to the peak locations, i.e., the diffraction angle 2θ (two theta) in degrees. The intensities are often given in parentheses, with the following abbreviations: very strong = vst; strong = st; medium = m; weak = w; and very weak = vw. The characteristic peaks of a given XRPD can be selected according to the peak locations and their relative intensity to conveniently distinguish this crystal structure from others.

[0046] Those skilled in the art recognize that measurements of XRPD peak locations and / or the intensity of a given crystalline form of the same compound will vary within a margin of error. The 2θ degree values ​​allow for appropriate margins of error. Typically, the margins of error are represented by “±”. For example, a 2θ degree of about “8.7 ± 0.3” denotes a range of about 8.7 + 0.3, i.e., about 9.0, to about 8.7 - 0.3, i.e., about 8.4. Depending on sample preparation techniques, calibration techniques applied to instruments, human operational variation, etc., those skilled in the art recognize that the appropriate margin error for an XRPD may be ± 0.5; ± 0.4; ± 0.3; ± 0.2; ± 0.1; ± 0.05; or less. In certain embodiments of the invention, the XRPD margin of error is ± 0.2.

[0047] Additional details of the methods and equipment used for XRPD analysis are described in the examples section.

[0048] The XRPD peaks for anhydrous form I of CYT-0387 dihydrochloride, CYT-0387, form II of CYT0387 dihydrochloride monohydrate, anhydrous form I of CYT-0387 monohydrochloride and anhydrous form III of Petition 870200056639, dated 07 / 05 / 2020, page 23 / 113 11 / 83 CYT-0387 monochloride can be found below in Table 1. Table 1: XRPD peaks for the CYT-0387 forms Anhydrous form I of CYT0387 dihydrochloride CYT-0387 dihydrochloride monohydrate form II Anhydrous form I of CYT0387 monohydrochloride Anhydrous form III of CYT0387 monohydrochloride Position [° 2-theta] Relative intensity [%] Position [° 2-theta] Relative intensity [%] Position [° 2-theta] Relative intensity [%] Position [° 2-theta] Relative intensity [%] 5.5 31.0 7.7 33.7 13.5 15.3 12.7 85.0 10.1 100.0 19.3 43.7 20.9 100.0 14.6 50.0 14.9 66.5 24.0 100.0 26.1 20.6 17.8 55.5 25.1 86.7 25.7 79.0 26.6 15.5 19.7 100.0 26.6 69.3 29.6 35.7 28.3 16.6 23.3 60.1

[0049] As used in the present invention, the following words, phrases and symbols are generally intended to have the meanings as defined below, except to the extent that the context in which they are used indicates otherwise.

[0050] Reference to “about” a value or parameter in the present invention includes (and describes) embodiments that are directed to that value or parameter by itself. For example, a description referring to “about X” includes the description of “X”. In certain embodiments, the term “about” includes the stated quantity ± 10%. In other embodiments, the term “about” includes the stated quantity ± 5%. In certain other embodiments, the term “about” includes the stated quantity ± 1%. Similarly, the singular forms “a”, “an”, and “the” include plural references, unless the context clearly specifies otherwise. Thus, for example, reference to “compound” includes a plurality of such antibodies, and reference to “assay” includes reference to one or more assays and equivalents thereof known to those skilled in the art.

[0051] “Pharmaceutically acceptable” or “physiologically acceptable” refers to compounds, salts, compositions, dosage forms and other materials that are useful in the preparation of a pharmaceutical composition that is suitable for veterinary or human pharmaceutical use.

[0052] “Pharmaceutically acceptable salts” or “physiologically acceptable salts” Petition 870200056639, dated 07 / 05 / 2020, page. 24 / 113 / 83 acceptable” refers to salts of pharmaceutical compounds that retain the biological efficacy and properties of the underlying compound and are not undesirable from a biological or other standpoint. There are acid addition salts and base addition salts. Pharmaceutically acceptable acid addition salts can be prepared from organic and inorganic acids. Acids and bases useful for reaction with an underlying compound to form pharmaceutically acceptable salts (acid addition salts or base addition salts, respectively) are known to a person skilled in the art. Similarly, methods of preparing pharmaceutically acceptable salts of an underlying compound (after description) are known to a person skilled in the art and are described, for example, in Berge, et al. Journal of Pharmaceutical Science, January 1977, Vol. 66, No. 1, and other sources.If the compounds described in this document are obtained as an acid addition salt, the free base can be obtained by alkalizing a solution of the acid salt. On the other hand, if the product is a free base, an addition salt, particularly a pharmaceutically acceptable addition salt, it can be produced by dissolving the free base in a suitable organic solvent and treating the solution with an acid, according to conventional procedures for the preparation of acid addition salts from base compounds. “Treatment” or “treating” is an approach to achieving beneficial or desired results, including clinical outcomes.Beneficial or desired clinical outcomes may include one or more of the following: a) inhibition of the disease or condition (e.g., reduction of one or more symptoms resulting from the disease or condition, and / or reduction in the extent of the disease or condition); b) delay or interruption of the development of one or more clinical symptoms associated with the disease or condition (e.g., stabilization of the disease or condition, prevention or delay of worsening or progression of the disease or condition, and / or prevention or delay of spread (e.g., metastasis). Petition 870200056639, dated 07 / 05 / 2020, page 25 / 113 / 83 of the disease or condition); and / or c) relief of the disease, that is, the regression of clinical symptoms (for example, improvement in the disease state, providing total or partial remission of the disease or condition, increased effects of another drug, delay in disease progression, increased quality of life and / or prolonged survival.

[0053] The compounds described in this document, in which one or more hydrogen atoms bonded to a carbon atom can be replaced by a deuterium atom or D, where n is the number of hydrogen atoms in the molecule. The deuterium atom is known to be a non-radioactive isotope of the hydrogen atom. Such compounds can increase resistance to metabolism and thus may be useful for increasing the half-life of the compounds described in this document, or pharmaceutically acceptable salts, isomers, prodrugs or solvates thereof, when administered to a mammal. See, for example, Foster, “Deuterium Isotope Effects in Studies of Drug Metabolism”, Trends Pharmacol. Sci., 5(12): 524-527 (1984). Such compounds are synthesized by means known in the art, for example, using starting materials in which one or more hydrogen atoms have been replaced by deuterium.In some embodiments, the compounds described in this document are form II of CYT-0387 dihydrochloride monohydrate, form I anhydrous of CYT-0387 monohydrochloride, form III anhydrous of CYT-0387 monohydrochloride, form IV anhydrous of CYT-0387 dihydrochloride, compound 3, compound 4, compound 8, compound 10, compound 12 and compound 13.

[0054] “Prevention” or “preventing” means any treatment of a disease or condition that causes the clinical symptoms of the disease or condition not to develop. The compounds may, in some modalities, be administered to an individual (including a human being) who is at risk for, or has a family history of, the disease or condition.

[0055] The terms “individual” or “patient” refer to an animal, Petition 870200056639, dated 07 / 05 / 2020, page 26 / 113 / 83 as a mammal (including a human being) that has been or will be the subject of treatment, observation, or experimentation. The methods described herein may be useful in human therapy and / or veterinary applications. In some modalities, the individual is a mammal. In one modality, the individual is a human being. “Human in need” refers to a human being who may have or is suspected of having diseases or conditions that would benefit from certain treatments; for example, being treated with the compounds according to this application. The terms “individual in need” or “patient in need” refer to an individual or a patient who may have, is diagnosed with, or is suspected of having diseases, disorders, or conditions that would benefit from the treatment described in this document.In certain modalities, the individual or patient who (i) has not received any treatment (i.e., treatment-naïve), (ii) has received prior treatment and is unresponsive or does not improve, or (iii) is relapsed or resistant (i.e., refractory) to prior treatment. For example, patients may have received platinum-based chemotherapy or gemcitabine-containing treatment. Additional examples include patients who may be relapsed or refractory to platinum-based chemotherapy or gemcitabine-containing treatment.

[0056] The term “therapeutically effective amount” of a compound of the present application, or a pharmaceutically acceptable salt, isomers, prodrug or solvate thereof, means an amount sufficient to accomplish treatment when administered to an individual to provide a therapeutic benefit, such as improvement of symptoms or retardation of disease progression. The therapeutically effective amount may vary depending on the individual and the disease or condition being treated, the individual’s weight and age, the severity of the disease or condition, and the route of administration, which can be readily determined by a person normally skilled in the art. In one embodiment, the amount Petition 870200056639, dated 07 / 05 / 2020, page 27 / 113 / 83 effective therapeutic use of the compound described in this document is 100 mg, 150 mg, 200 mg, 250 mg or 300 mg.

[0057] The term “inhibition” indicates a decrease in the baseline activity of a biological activity or process. Pharmaceutical Compositions and Administration

[0058] The compounds provided in this document are generally administered in the form of pharmaceutical compositions. Thus, pharmaceutical compositions are provided in this document containing one or more of the compounds of any of the formulas described in this document, or a pharmaceutically acceptable salt, isomers, prodrug or solvate thereof, and one or more pharmaceutically acceptable carriers selected from carriers, adjuvants and excipients. Suitable pharmaceutically acceptable carriers may include, for example, diluents and inert solid fillers, diluents, including a sterile aqueous solution and various organic solvents, permeation enhancers, solubilizers and adjuvants. Such compositions are prepared in a manner well known in the pharmaceutical art. See, for example, Remington's Pharmaceutical Sciences, Mace Publishing Co., Philadelphia, Pa. 17th Ed. (1985); and Modern Pharmaceutics, Marcel Dekker, Inc. 3rd Ed. (GS Banker & CT Rhodes, Eds.).As used in the present invention, a “solvate” is formed by the interaction of a solvent and a compound. Solvates or salts of the compounds of any of the formulas described herein are also provided. Hydrates of the compounds of any of the formulas are also provided. Similarly, a “prodrug” is defined in the pharmaceutical field as a biologically inactive derivative of a drug that, after administration to the human body, is converted into the original biologically active drug according to some chemical or enzymatic pathways.

[0059] Pharmaceutical compositions can be administered in single or multiple doses. The pharmaceutical composition can be Petition 870200056639, dated 07 / 05 / 2020, p. 28 / 113 / 83 administered by various methods, including, for example, rectally, buccally, intranasally and transdermally. In certain embodiments, the pharmaceutical composition may be administered by intra-arterial injection, intravenously, intraperitoneally, parenterally, intramuscularly, subcutaneously, orally, topically or as an inhalant. In some embodiments, the pharmaceutical composition is administered orally.

[0060] One method of administration is parenteral, for example, by injection. Forms in which the pharmaceutical compositions described herein may be incorporated for administration by injection include, for example, aqueous or oily suspensions or emulsions, with sesame oil, corn oil, cottonseed oil or peanut oil, as well as elixirs, mannitol, dextrose or a sterile aqueous solution, and similar pharmaceutical vehicles.

[0061] Oral administration may be another route for administering the compounds described in this document. Administration may be via, for example, capsules or enteric-coated tablets. When making pharmaceutical compositions that include at least one compound of any of the formulas described herein, or a pharmaceutically acceptable salt, prodrug or solvate thereof, the active ingredient is generally diluted by an excipient and / or enclosed within such a vehicle which may be in the form of a capsule, sachet, paper or other container. When the excipient serves as a diluent, it may be in the form of a solid, semi-solid or liquid material that acts as a vehicle, carrier or medium for the active ingredient.Thus, the compositions may be in the form of tablets, pills, powders, sachets, wafers, elixirs, suspensions, emulsions, solutions, syrups, aerosols (as a solid or in liquid medium), ointments containing, for example, up to 10% by weight of the active compound, soft and hard gelatin capsules, sterile injectable solutions and sterile packaged powders. Petition 870200056639, dated 07 / 05 / 2020, page 29 / 113 / 83

[0062] To prepare solid compositions, such as tablets, the main active ingredient may be mixed with a pharmaceutical excipient to form a solid preformulation composition containing a homogeneous mixture of a compound from any of the above formulas, or of a pharmaceutically acceptable salt, prodrug or solvate thereof. When referring to these preformulation compositions as homogeneous, the active ingredient may be uniformly dispersed throughout the composition, so that the composition can be easily subdivided into equally effective unit dosage forms, such as tablets, pills and capsules.

[0063] The tablets or pills of the compounds described in this document may be coated or otherwise composed to provide a dosage form that offers the advantage of prolonged action, or to protect them from the acidic conditions of the stomach. For example, the tablet or pill may include an inner dosage component and an outer dosage component, the latter being in the form of an envelope over the former. The two components may be separated by an enteric coating that serves to resist disintegration in the stomach and allow the inner component to pass intact into the duodenum or to exhibit delayed release. Various materials may be used for such enteric coatings, with such materials including various polymeric acids and mixtures of polymeric acids with such materials, such as shellac, cetyl alcohol, and cellulose acetate.

[0064] The specific dose level of a compound from the formulas described herein for any particular individual will depend on several factors, including the activity of the specific compound used, age, body weight, general health status, sex, diet, time of administration, route of administration and rate of excretion, drug combination, and the severity of the particular disease in the individual undergoing therapy. For example, a dosage Petition 870200056639, dated 07 / 05 / 2020, page 30 / 113 / 83 can be expressed as a number of milligrams of a compound of the formula per kilogram of the individual's body weight (mg / Kg). Dosages between approximately 0.01 and 200 mg / Kg may be adequate. In some embodiments, between approximately 0.01 and 150 mg / Kg may be adequate. In other embodiments, a dosage between 0.05 and 100 mg / Kg may be appropriate. Normalization according to the individual's body weight is particularly useful when adjusting dosages between individuals of widely disparate sizes, as occurs when using the drug in both children and adult humans, or when converting a dosage effective in a non-human individual, such as a dog, to a dosage adequate for a human individual.

[0065] The compounds of this application or compositions thereof may be administered once, twice, three or four times a day, using any appropriate method described above. Similarly, administration or treatment with the compounds according to any of the formulas described herein may be maintained for several days; for example, treatment would commonly continue for at least 7 days, 14 days or 28 days for a treatment cycle. Treatment cycles are generally known and frequently alternated with rest periods of about 1 to 28 days, usually about 7 days or about 14 days between cycles. Treatment cycles in other modalities may also be continuous.

[0066] In some embodiments, the forms or compositions thereof described in this document are formulated for oral administration using pharmaceutically acceptable vehicles. Pharmaceutical compositions formulated for oral administration may be in the form of tablets, pills, capsules, wafers, dragees, trociscos, liquids, gels, syrups, suspensions, elixirs, or powders. Pharmaceutically acceptable vehicles

[0067] The term “vehicle” refers to the diluents or fillers, Petition 870200056639, dated 07 / 05 / 2020, page 31 / 113 / 83 disintegrants, precipitation inhibitors, surfactants, glidants, binders, lubricants, antioxidants and other excipients and vehicles with which the compound is administered. The vehicles are generally described in this document and also in “Remington's Pharmaceutical Sciences” by EW Martin.Examples of carriers include, but are not limited to, aluminum monostearate, aluminum stearate, carboxymethylcellulose, sodium carboxymethylcellulose, crospovidone, glyceryl isostearate, glyceryl monostearate, hydroxyethylcellulose, hydroxyethylcellulose, hydroxymethylcellulose, hydroxyoctacosaniyl hydroxystearate, hydroxypropylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, lactose, lactose monohydrate, magnesium stearate, mannitol, microcrystalline cellulose, poloxamer 124, poloxamer 181, poloxamer 182, poloxamer 188, poloxamer 237, poloxamer 407, povidone, silicon dioxide, colloidal silicon dioxide, silicone, silicone adhesive 4102, and silicone emulsion.It should be understood, however, that the vehicles selected for the pharmaceutical compositions provided in this description, and the quantities of such vehicles in the composition, may vary depending on the formulation method (e.g., dry granulation formulation, solid dispersion formulation).

[0068] The term “diluent” or “filler” generally refers to a substance that is used to dilute the compound of interest before release. Diluents can also serve to stabilize compounds. Examples of diluents may include starch, saccharides, disaccharides, sucrose, lactose, polysaccharides, cellulose, cellulose ethers, hydroxypropylcellulose, sugar alcohols, xylitol, sorbitol, maltitol, microcrystalline cellulose, calcium or sodium carbonate, lactose, lactose monohydrate, dicalcium phosphate, cellulose, compressible sugars, dibasic calcium phosphate dehydrate, mannitol, microcrystalline cellulose, and tribasic calcium phosphate.

[0069] The term “disintegrant” generally refers to a substance Petition 870200056639, dated 07 / 05 / 2020, page 32 / 113 / 83, which, by adding a solid preparation, facilitates its dissolution or disintegration after administration and allows the release of an active ingredient as efficiently as possible to allow its rapid dissolution. Examples of disintegrants may include corn starch, sodium starch glycolate, croscarmellose sodium, crospovidone, microcrystalline cellulose, modified corn starch, sodium carboxymethyl starch, povidone, pregelatinized starch, and alginic acid.

[0070] The term “precipitation inhibitors” generally refers to a substance that prevents or inhibits the precipitation of the active agent. An example of a precipitation inhibitor includes hydroxypropyl methylcellulose.

[0071] The term “surfactants” generally refers to compounds that reduce the surface tension between two liquids or between a liquid and a solid. Examples of surfactants are poloxamer and sodium lauryl sulfate.

[0072] The term “glidant” generally refers to substances used in tablet and capsule formulations to improve flow properties during tablet packing and to produce an anti-caking effect. Examples of glidants may include colloidal silicon dioxide, talc, fumed silica, starch, starch derivatives, and bentonite.

[0073] The term “binder” refers to any pharmaceutically acceptable film that can be used to bind the active and inert components of the vehicle to keep the portions cohesive and separate. Examples of binders may include hydroxypropylcellulose, hydroxypropylmethylcellulose, povidone, copovidone, ethylcellulose, gelatin, and polyethylene glycol.

[0074] The term “lubricant” generally refers to a substance that is added to a powder mixture to prevent the compacted powder mass from sticking to the equipment during the compaction or encapsulation process. A lubricant can help eject the tablet from the Petition 870200056639, dated 07 / 05 / 2020, p. 33 / 113 / 83 matrices, and may improve dust flow. Examples of lubricants may include magnesium stearate, stearic acid, silica, fats, calcium stearate, polyethylene glycol, sodium stearyl fumarate or talc; and solubilizers, such as fatty acids, including lauric acid, oleic acid and C8 / C10 fatty acids.

[0075] The term “antioxidant” generally refers to a substance that inhibits the oxidation of other substances. In certain embodiments of the invention, antioxidants are added to the pharmaceutical composition. Examples of antioxidants may include ethylenediaminetetraacetic acid, disodium salt of ethylenediaminetetraacetic acid, sodium sulfite, sodium metabisulfite, sodium bisulfite, butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), ascorbic acid, ascorbyl palmitate, thioglycerol, thioglycolic acid, tocopherol (vitamin E), Da-tocopherylpolyethylene glycol 1000 succinate (vitamin E TPGS), and propyl gallate. In certain embodiments, the antioxidant is propyl gallate. In one embodiment, the pharmaceutical composition comprises form II of the monohydrate of CYT-0387 dihydrochloride and an antioxidant selected from the group consisting of butylated hydroxyanisole (BHA), ascorbic acid, and propyl gallate.In a certain embodiment, the pharmaceutical composition comprises form II of the monohydrate of CYT-0387 dihydrochloride and an antioxidant, wherein the antioxidant is propyl gallate.

[0076] The antioxidant may be present in an amount sufficient to prevent, inhibit and / or reduce the degradation of the active ingredient (such as form II of CYT-0387). As examples, the antioxidant may be present in an amount of about 0.001%, about 0.002%, about 0.005%, about 0.01%, about 0.02%, about 0.05%, about 0.1%, about 0.2%, about 0.5% or about 1%. In one embodiment, the pharmaceutical composition comprises propyl gallate in an amount of about 0.001%, about 0.01%, about 0.1%, about 0.2%, about Petition 870200056639, dated 07 / 05 / 2020, p. 34 / 113 / 83 0.5% or about 1%. In one embodiment, the pharmaceutical composition comprises form II of CYT-0387 dihydrochloride monohydrate and about 0.2% propyl gallate.

[0077] In certain respects, a pharmaceutical composition is provided comprising at least one active agent (including, for example, the CYT-0387 dihydrochloride monohydrate form II) and one or more of (a) to (e): a) at least one diluent; b) at least one disintegrant; c) at least one glidant; d) at least one lubricant; e) at least one antioxidant.

[0078] In some embodiments, the pharmaceutical composition comprises at least one or at least two diluent(s). In certain embodiments, the pharmaceutical composition comprises one or two diluent(s). In certain embodiments, the diluent is selected from the group consisting of mannitol, microcrystalline cellulose, lactose, glucose, sucrose, ludiflash, F-melt, advantose, GalenIQ or any mixture thereof. In one embodiment, the diluent is mannitol, microcrystalline cellulose or a mixture thereof.

[0079] In some embodiments, the pharmaceutical composition comprises at least one disintegrant. In certain embodiments, the pharmaceutical composition comprises a disintegrant. In one particular embodiment, the disintegrant is sodium starch glycolate. In one embodiment, the disintegrant is croscarmellose sodium. In another embodiment, the disintegrant is crospovidone.

[0080] In some embodiments, the pharmaceutical composition comprises at least one glidant. In certain embodiments, the pharmaceutical composition comprises a glidant. In one embodiment the glidant is colloidal silicon dioxide.

[0081] In some embodiments, the pharmaceutical composition comprises at least one lubricant. In certain embodiments, the pharmaceutical composition comprises a lubricant. In one embodiment, Petition 870200056639, dated 07 / 05 / 2020, page 35 / 113 / 83 the lubricant is magnesium stearate.

[0082] In particular embodiments, the pharmaceutical composition comprises form II of CYT-0387 dihydrochloride monohydrate, at least one diluent, at least one disintegrant, at least one glidant, at least one lubricant, and at least one antioxidant. In further embodiments, at least one diluent is microcrystalline cellulose, at least one disintegrant is sodium starch glycolate, at least one glidant is colloidal silicon dioxide, at least one lubricant is magnesium stearate, and at least one antioxidant is propyl gallate. In still other embodiments, at least one diluent is lactose, at least one disintegrant is sodium starch glycolate, at least one glidant is colloidal silicon dioxide, at least one lubricant is magnesium stearate, and at least one antioxidant is propyl gallate.

[0083] In particular embodiments, the pharmaceutical composition comprises form II of CYT-0387 dihydrochloride monohydrate, at least two diluents, at least one disintegrant, at least one glidant, at least one lubricant and at least one antioxidant. In still other embodiments, the at least two diluents are microcrystalline cellulose and lactose, the at least one disintegrant is sodium starch glycolate, the at least one glidant is colloidal silicon dioxide, the at least one lubricant is magnesium stearate and the at least one antioxidant is propyl gallate.

[0084] In certain embodiments, the pharmaceutical composition comprises CYT-0387, of which at least about 80% is the CYT-0387 dihydrochloride monohydrate form II. In further embodiments, the pharmaceutical composition comprises CYT-0387, of which at least about 85% is the CYT-0387 dihydrochloride monohydrate form II. In still other embodiments, the pharmaceutical composition comprises CYT-0387, of which at least about 90% is the CYT-0387 dihydrochloride monohydrate form II. In still other embodiments, the composition Petition 870200056639, dated 07 / 05 / 2020, page 36 / 113 / 83: The pharmaceutical composition comprises CYT-0387, of which at least about 95% is the CYT-0387 dihydrochloride monohydrate form II. In particular embodiments, the pharmaceutical composition comprises CYT-0387, of which at least about 97% is the CYT-0387 dihydrochloride monohydrate form II. In other embodiments, the pharmaceutical composition comprises CYT-0387, of which at least about 98% is the CYT-0387 dihydrochloride monohydrate form II. In still other embodiments, the pharmaceutical composition comprises CYT-0387, of which at least about 99% is the CYT-0387 dihydrochloride monohydrate form II. In still other embodiments, the pharmaceutical composition comprises CYT-0387, of which at least about 99.5% is the CYT-0387 dihydrochloride monohydrate form II.In particular embodiments, the pharmaceutical composition comprises CYT-0387, of which at least approximately 99.9% is the CYT-0387 dihydrochloride monohydrate form II.

[0085] It should be understood that the pharmaceutical composition comprises the pharmaceutically acceptable vehicles detailed in this document, the same as if each pharmaceutically acceptable vehicle combination were specifically and individually listed. Unit dosage forms

[0086] In some embodiments, the pharmaceutical compositions, as described in the present invention, are formulated in a unit dosage form. The term “unit dosage forms” refers to physically distinct units suitable as unit dosages for individuals (e.g., human individuals and other mammals), with each unit containing a predetermined amount of active material calculated to produce the desired therapeutic effect, in association with a suitable pharmaceutical vehicle.

[0087] In a further embodiment, the invention is directed to unit dosage forms comprising form II of the monohydrate of Petition 870200056639, dated 07 / 05 / 2020, page 37 / 113 / 83 CYT-0387 dihydrochloride. In some embodiments, the unit dosage form comprises form II of the CYT-0387 dihydrochloride monohydrate in an amount equivalent to about 10 mg to about 1000 mg, about 10 mg to about 800 mg, about 10 mg to about 700 mg, about 10 mg to about 500 mg, about 10 mg to about 400 mg, about 10 mg to about 300 mg, about 10 mg to about 250 mg, about 10 mg to about 200 mg, about 10 mg to about 150 mg, about 10 mg to about 100 mg, about 10 mg to about 50 mg, about 50 mg to about 1000 mg, about 50 mg to about 800 mg, from about 50 mg to about 700 mg, from about 50 mg to about 500 mg, from about 50 mg to about 400 mg, from about 50 mg to about 300 mg, from about 50 mg to about 250 mg, from about 50 mg to about 200 mg, from about 50 mg to about 150 mg, from about 50 mg to about 100 mg,from about 100 mg to about 1000 mg, from about 100 mg to about 800 mg, from about 100 mg to about 700 mg, from about 100 mg to about 500 mg, from about 100 mg to about 400 mg, from about 100 mg to about 300 mg, from about 100 mg to about 250 mg, from about 100 mg to about 200 mg, from about 150 mg to about 300 mg, from about 150 mg to about 250 mg, from about 150 mg to about 200 mg, from about 200 mg to about 300 mg, from about 200 mg to about 250 mg or from about 200 mg to about 300 mg of free base of CYT-0387.,

[0088] In additional embodiments, the dosage form comprises form II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to approximately 100 mg of CYT-0387 free base, approximately 150 mg of CYT-0387 free base, 200 mg of CYT-0387 free base, approximately 250 mg of CYT-0387 free base, 300 mg of CYT-0387 free base, approximately 400 mg of CYT-0387 free base, or approximately 500 mg of CYT-0387 free base. In certain embodiments, the unit dosage form comprises form II of CYT-0387 dihydrochloride monohydrate in an amount Petition 870200056639, dated 07 / 05 / 2020, page 38 / 113 / 83 equivalent to approximately 50 mg of CYT-0387 free base. In other embodiments, the unit dosage form comprises form II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to approximately 100 mg of CYT-0387 free base. In still other embodiments, the unit dosage form comprises form II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to approximately 150 mg of CYT-0387 free base. In still other embodiments, the dosage form comprises form II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to approximately 200 mg of CYT-0387 free base. In particular embodiments, the dosage form comprises form II of the monohydrate of CYT-0387 dihydrochloride in an amount equivalent to approximately 250 mg of CYT-0387 free base.In further embodiments, the dosage form comprises form II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to approximately 300 mg of CYT-0387 free base. In still other embodiments, the dosage form comprises form II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to approximately 400 mg of CYT-0387 free base. In still other embodiments, the dosage form comprises form II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to approximately 500 mg of CYT-0387 free base. In other embodiments, the pharmaceutical composition is a tablet in a dosage form comprising form II of the monohydrate of CYT-0387 dihydrochloride in an amount equivalent to approximately 300 mg of CYT-0387 free base.

[0089] In further embodiments, the invention is directed to dosage forms comprising form II of CYT-0387 dihydrochloride monohydrate in an amount equivalent to 200 mg of the free base N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide, which provides a pharmacokinetic profile substantially similar to a dosage form comprising anhydrous form I of CYT-0387 dihydrochloride. Petition 870200056639, dated 07 / 05 / 2020, page 39 / 113 / 83 in an amount equivalent to 300 mg of the free base of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide.

[0090] In certain embodiments of the invention, the unit dosage form comprises at least one pharmaceutically acceptable carrier. In other embodiments, the unit dosage form comprises form II of the CYT-0387 dihydrochloride monohydrate, at least two diluents, at least one disintegrant, at least one glidant, at least one lubricant, and at least one antioxidant. In still other embodiments, the unit dosage form comprises about 36% to 44% of form II of the CYT-0387 dihydrochloride monohydrate; about 44% to 58% diluent; about 4% to 8% disintegrant, about 0.25% to 0.75% glidant, about 1.2% to 1.8% lubricant, and about 0.1% to 0.5% antioxidant.In still other embodiments, at least two diluents are microcrystalline cellulose and lactose, at least one disintegrant is sodium starch glycolate, at least one glidant is colloidal silicon dioxide, at least one lubricant is magnesium stearate, and at least one antioxidant is propyl gallate. In further embodiments, the unit dosage form comprises about 36% to 44% of form II of CYT-0387 dihydrochloride monohydrate; about 30% to 38% microcrystalline cellulose; about 14% to 20% lactose, about 4% to 8% sodium starch glycolate, about 0.25% to 0.75% colloidal silicon dioxide, about 1.2% to 1.8% magnesium stearate, and about 0.1% to 0.5% propyl gallate. Manufacturing of pharmaceutical compositions

[0091] The pharmaceutical compositions described in this document may be manufactured using any conventional method, such as, but not limited to, mixing, dissolution, granulation, coating, levigation, emulsification, encapsulation, uptake, rotational solidification with freezing, atomization or lyophilization.

[0092] A knowledgeable professional would recognize methods and techniques. Petition 870200056639, dated 07 / 05 / 2020, page 40 / 113 / 83 suitable for preparing a tablet by conventional formulation. Exemplary methods and techniques for preparing powders for compression into a tablet include dry granulation or wet granulation. Dry granulation generally refers to the process of forming granules without the use of a liquid solution, while wet granulation generally refers to the process of adding a liquid solution to the powders to granulate them. Kits

[0093] Kits are also provided in this document that include a compound of the formulas of the present application, or a pharmaceutically acceptable salt, isomer, prodrug or solvate thereof, and appropriate packaging. In one embodiment, a kit also includes instructions for use. In one aspect, a kit includes a compound of the formulas described herein or a pharmaceutically acceptable salt, isomer, prodrug or solvate thereof, and a label and / or instructions for use of the compounds in the treatment of the indications, including the diseases or conditions, described in this document.

[0094] Also provided in this document are articles of manufacture which include a compound of any of the formulas described herein, or a pharmaceutically acceptable salt, isomer, prodrug or solvate thereof, in a suitable container. The container may be a bottle, a jar, an ampoule, a pre-filled syringe and an intravenous bag. Treatment Methods

[0095] The CYT-0387 forms of the present invention can be used in the treatment of kinase-associated diseases, including JAK kinase-associated diseases, such as immunological and inflammatory diseases, including organ transplants; hyperproliferative diseases, including cancer and myeloproliferative diseases; viral diseases; metabolic diseases; and vascular diseases.

[0096] In addition to primates, such as humans, a variety of Petition 870200056639, dated 07 / 05 / 2020, page 41 / 113 / 83 other mammals can be treated using the compounds, compositions and methods of the present invention. For example, mammals including, but not limited to, cows, sheep, goats, horses, dogs, cats, guinea pigs, rats or other bovine, ovine, equine, canine, feline, rodent or murine species can be treated. However, the invention can also be practiced in other species, such as bird species (e.g., chickens).

[0097] The term “administration” should be understood to mean providing a compound of the invention to an individual who needs treatment.

[0098] The terms “treating” or “treatment” refer to achieving a desired pharmacological and / or physiological effect. The effect may be prophylactic in terms of complete or partial prevention of a disease, and / or therapeutic in terms of a complete or partial cure of a disease, and / or of adverse effects attributable to the disease. Treatment may encompass any treatment of a disease in a mammal, and includes: preventing the disease from occurring in an individual who may be predisposed to the disease but has not yet been diagnosed as having it; inhibiting the disease, i.e., halting its development; or alleviating the disease, i.e., causing regression of the disease. The therapeutic agent may be administered before, during, or after the onset of the disease. Treatment of ongoing disease, where treatment stabilizes or reduces the patient's undesirable clinical symptoms, is of particular interest.Expected progression-free survival times can be measured in months and years, depending on prognostic factors, including the number of relapses, the stage of the disease, and other factors. Prolonged survival includes, without limitation, times of at least 1 month, approximately at least 2 months, approximately at least 3 months, approximately at least 4 months, approximately at least 6 months, approximately at least 1 year, approximately at least 2 years, and so on. Petition 870200056639, dated 07 / 05 / 2020, page 42 / 113 / 83 approximately at least 3 years or more. Overall survival can also be measured in months to years. The patient's symptoms may remain static or may decrease.

[0099] The term “effective amount” refers to an amount that can be effective in eliciting the desired medical or biological response, including the amount of a compound that, when administered to an individual to treat a disease, is sufficient to effect such treatment for the disease. The effective amount will vary depending on the compound, the disease and its severity, and the age, weight, etc. of the individual being treated. The effective amount may include a range of values.

[00100] The term “kinase-associated diseases” refers to a disorder or disorders that result, directly or indirectly, from or are aggravated by aberrant kinase activity, in particular JAK activity, and / or that are relieved by inhibition of one or more of these kinase enzymes.

[00101] In a preferred embodiment, the kinase-associated disease state involves one or more of the JAK kinases, JAK1, JAK2, JAK3, or TYK2. In a particularly preferred embodiment, the disease involves JAK2 kinase. Such diseases include, but are not limited to, those listed in the table below. Activation of the JAK / STAT pathway in various pathologies. Disease Type | Cell Types | Cytokines Involved | JAK kinase Involved | Characteristics | Atopy | Allergic Asthma, Atopic Dermatitis (Eczema), Allergic Rhinitis, CMI | Mast Cells, IL-4, IL-5, IL-6, Eosinophils, IL-7, IL-13 | T Cells, B Cells, JAK1, JAK2, JAK3, Tyk2 | T Cell Activation | B Cells, followed by | IgE-mediated activation of resistant mast cells and eosinophils | Allergic Contact | Hypersensitivity Dermatitis | Pneumonitis | T Cells, B Cells, IL-2, IL-4, IL-5, Macrophages, IL-6, IL-10, Neutrophils | IFNγ, TNF, IL-7, IL-13, JAK1, JAK2, JAK3, Tyk2 | B Cell and / or TDH Cell Activation | Macrophage / Granulocyte Activation Petition 870200056639, dated 07 / 05 / 2020, page 43 / 113 / 83 Disease Type | Cell Types Involved | Cytokines Involved | JAK Kinase Involved | Characteristics | Autoimmune Diseases | Multiple Sclerosis | T Cells, B Cells, IL-2, IL-4, IL-5, JAK1 | Cytokine Production | Glomerulonephritis | Monocytes, IL-6, IL-7, JAK2, (e.g., TNFα / β, IL-1, Systemic Lupus Erythematosus | Macrophages, IL-10, IL-13, JAK3, CSF-1, GM-CSF), T Cell Activation, Neutrophils, IFNγ, TNF, Tyk2 | B Cells, (SLE), (Rheumatoid) | Mast Cells, GM-CSF; G-CSF, Juvenile Arthritis, Sjögren's Syndrome, Scleroderma, Polymyositis, Ankylosing Spondylitis, Psoriatic Arthritis, Transplantation, Eosinophils, JAK / STAT Activation, Allograft Rejection, T Cells, B Cells, IL-2, IL-4, IL-5, JAK1, GvHD-mediated Necrosis, Macrophages, IL-7, IL-13, JAK2, Macrophages / T Cells, TNF, JAK3, Apoptosis, TC Cell-mediated and opsonization / necrosis of foreign graft, B Cell / Ig-mediated, Viral Diseases, Epstein-Barr Virus, Lymphocytes, Viral JAK1, JAK / STAT (EBV), Cytokines, IL-2, JAK2,Mediation Hepatitis B Hepatocytes JAK3 Hepatitis C Hepatocytes HIV Lymphocytes HTLV 1 Lymphocytes Varicella-Zoster Fibroblasts Virus (VZV) Human Papillomavirus (HPV) Epithelial cells Hyperproliferative diseases - cancer Leukemia Leukocytes Various JAK1, Cytokine production, Lymphoma Lymphocytes Autocrine JAK2, JAK / STAT Multiple myeloma Various cytokines, JAK3 Activation Prostate cancer Various Intrinsic breast cancer Various Activation Hodgkin's lymphoma Various Chronic lymphocytic B-cell leukemia Various Lung cancer Various Hepatoma Various Metastatic melanoma Various Glioma Various Petition 870200056639, dated 07 / 05 / 2020, page 44 / 113 / 83 Cytokines Involved JAK kinase involved Characteristics Cell types Disease type involved Myeloproliferative Diseases Polycythemia vera Hematopoietic PV (primary) myelofibrosis, thrombocythemia, Essential thrombocythemia (ET), idiopathic myelofibrosis, chronic myeloid leukemia, systemic mastocytosis (SM), chronic neutrophilic leukemia (CNL), myelodysplastic syndrome (MDS), systemic mastocytosis disease (SMCD) Interleukin-3, erythropoietin thrombopoietin JAK2 Mutation JAK / STAT activation Vascular disease Hypertension, Endothelial cells, Hypertrophy, heart smooth muscle failure, ischemia, including cells including Pulmonary arterial pulmonary artery hypertension smooth muscle cells, cardiac myocytes, fibroblasts, endothelial cells Metabolic disease IL6, angiotensin II, LIF, TNFalpha, serotonin, caveolin1 JAK1, JAK2, TYK2 JAK / STAT activation, obesity, metabolic, adipocytes, pituitary cell syndrome, neurons.Monocytes Leptin JAK2 JAK / STAT activation

[00102] The term “immunological and inflammatory disease” refers to a, Immunological, inflammatory, or autoimmune disease, including, but not limited to, rheumatoid arthritis, polyarthritis, rheumatoid spondylitis, osteoarthritis, gout, asthma, bronchitis, allergic rhinitis, chronic obstructive pulmonary disease, cystic fibrosis, inflammatory bowel disease, irritable bowel syndrome, mucous colitis, ulcerative colitis, diabrotic colitis, Crohn's disease, autoimmune thyroid disorders, gastritis, esophagitis, hepatitis, pancreatitis, nephritis, Petition 870200056639, dated 07 / 05 / 2020, page 45 / 113 / 83 psoriasis, eczema, acne vulgaris, dermatitis, urticaria, multiple sclerosis, Alzheimer's disease, motor neuron disease (Gehrig's disease), Paget's disease, sepsis, conjunctivitis, neuroleptic catarrh, chronic arthrorheumatism, systemic inflammatory response syndrome (SIRS), polymyositis, dermatomyositis (DM), polaritis nodosa (PN), mixed connective tissue disorder (MCTD), Sjögren's syndrome, Crouzon syndrome, achondroplasia, systemic lupus erythematosus, scleroderma, vasculitis, thanatophoric dysplasia, insulin resistance, type I diabetes and complications of diabetes and metabolic syndrome.

[00103] The term “hyperproliferative diseases” includes cancer and myeloproliferative disease states, such as cell proliferation disease states, including, but not limited to: cardiac: sarcoma (angiosarcoma, fibrosarcoma, rhabdomyosarcoma, liposarcoma), myxoma, rhabdomyoma, fibroma, lipoma and teratoma), lung: bronchogenic carcinoma (squamous cell, small undifferentiated cell, large undifferentiated cell, adenocarcinoma), alveolar (bronchiolar) carcinoma, bronchial adenoma, sarcoma, lymphoma, chondromatous hamartoma, ynesothelioma); of the gastrointestinal tract: esophagus (squamous cell carcinoma, adenocarcinoma, leiomyosarcoma, lymphoma), stomach (carcinoma, lymphoma, leiomyosarcoma), pancreas (ductal adenocarcinoma, insulinoma, glucagonoma, gastrinoma, carcinoid tumors, vipoma), small intestine (adenocarcinoma, lymphoma, carcinoid tumors, Kaposi's sarcoma, leiomyoma, hemangioma, lipoma, neurofibroma, fibroma);Large intestine (adenocarcinoma, tubular adenoma, villous adenoma, hamartoma, leiomyoma); Genitourinary tract: kidney (adenocarcinoma, Wilms' tumor, nephroblastoma, lymphoma, leukemia); bladder and urethra (squamous cell carcinoma, transitional cell carcinoma, adenocarcinoma); prostate (adenocarcinoma, sarcoma); testicle (seminoma, teratoma, embryonal carcinoma, teratocarcinoma, choriocarcinoma, sarcoma, cell carcinoma; Petition 870200056639, dated 07 / 05 / 2020, page 46 / 113 / 83 interstitial cells, fibroma, fibroadenoma, adenomatoid tumors, lipoma); liver: hepatoma (hepatocellular carcinoma), cholangiocarcinoma, hepatoblastoma, angiosarcoma, hepatocellular adenoma, hemangioma); Bone (osteogenic sarcoma (osteosarcoma), fibrosarcoma, malignant fibrous histiocytoma, chondrosarcoma, Ewing's sarcoma, malignant lymphoma (reticulum cell sarcoma), multiple myeloma, malignant giant cell tumor, chordoma, osteochondroma (osteocartilaginous exostoses), benign chondroma, chondroblastoma, chondromyxofibroma, osteoid osteoma and giant cell tumors; nervous system: osteoma, hemangioma, granuloma, xanthoma, osteitis deformans), meninges (meningioma, meningiosarcoma, gliomatosis), brain (astrocytoma, medulloblastoma, glioma, ependymoma, germinoma [pinealoma], glioblastoma multiforme, oligodendroglioma, schwannoma, retinoblastoma, congenital tumors), spinal cord: neurofibroma, meningioma, glioma, sarcoma);Gynecological: uterus (endometrial carcinoma), cervix (cervical carcinoma, pre-tumoral cervical dysplasia), ovaries (ovarian carcinoma, serous cystadenocarcinoma, mucinous cystadenocarcinoma, unclassified carcinoma, granulosa-thecal cell tumors, Sertoli-Leydig cell tumors, dysgerminoma, malignant teratoma), vulva (squamous cell carcinoma, intraepithelial carcinoma, adenocarcinoma, fibrosarcoma, melanoma), vagina (clear cell carcinoma, squamous cell carcinoma, botryoid sarcoma [embryonal rhabdomyosarcoma], fallopian tubes (carcinoma)); hematological: blood (myeloid leukemia (acute and chronic), acute lymphoblastic leukemia, chronic lymphocytic leukemia, multiple myeloma, myelodysplastic syndrome), Hodgkin's disease, non-Hodgkin's lymphoma (malignant lymphoma); Skin: malignant melanoma, basal cell carcinoma, squamous cell carcinoma, Karposi's sarcoma, soft dysplastic nevi, lipoma, angioma, dermatofibroma, keloids, psoriasis;Adrenal glands: neuroblastoma; and myeloproliferative diseases, such as polycythemia vera (PV); Petition 870200056639, dated 07 / 05 / 2020, page 47 / 113 / 83 primary myelofibrosis, thrombocythemia, essential thrombocythemia (ET), agonogenic myeloid metaplasia (AMM), also known as idiopathic myelofibrosis (IMF), chronic myeloid leukemia (CML), systemic mastocystosis (SM), chronic neutrophilic leukemia (CNL), myelodysplastic syndrome (MDS) and systemic mastocytosis (SMCD). In a certain embodiment, the myelofibrosis disease is selected from polycythemia vera (PV), primary myelofibrosis, thrombocythemia and essential thrombocythemia (ET). In one embodiment, the pharmaceutical composition of the present application may be suitable for the treatment of myeloproliferative diseases, where the myelofibrosis disease is selected from polycythemia vera (PV), primary myelofibrosis, thrombocythemia and essential thrombocythemia (ET). In one embodiment, the pharmaceutical composition of the present application may be suitable for the treatment of primary myelofibrosis.

[00104] The term “vascular diseases” refers to diseases including, but not limited to, cardiovascular diseases, hypertension, hypertrophy, hypercholesterolemia, hyperlipidemia, thrombotic disorders, stroke, apoplexy, Raynaud's phenomenon, POEMS syndrome, angina, ischemia, migraine, peripheral arterial disease, heart failure, restenosis, atherosclerosis, left ventricular hypertrophy, myocardial infarction, ischemic diseases of the heart, kidney, liver and brain, and pulmonary arterial hypertension.

[00105] Diseases preferentially treated with selective JAK2 inhibitors include immunological and inflammatory diseases, such as autoimmune diseases, for example, atopic dermatitis, asthma, rheumatoid arthritis, Crohn's disease, psoriasis, Crouzon syndrome, achondroplasia, systemic lupus erythematosus, scleroderma, mixed connective tissue disease, vasculitis, thanatophoric dysplasia and diabetes; hyperproliferative disorders, such as cancer, for example, prostate cancer, colon cancer, breast cancer, liver cancer, such as hepatoma, lung cancer, head and neck cancer, such as glioma, skin cancer, such as metastatic melanoma, leukemia, lymphoma. Petition 870200056639, dated 07 / 05 / 2020, page 48 / 113 / 83 multiple myeloma and myeloproliferative disorders, such as polycythemia vera (PV), myelofibrosis, thrombocythemia, essential thrombocythemia (ET), agonogenic myeloid metaplasia (AMM), also referred to as idiopathic myelofibrosis (IMF) and chronic myelogenous leukemia (CML); and vascular diseases, such as hypertension, hypertrophy, stroke, Raynaud's phenomenon, POEMS syndrome, angina, ischemia, migraine, peripheral arterial disease, heart failure, restenosis, atherosclerosis and pulmonary arterial hypertension.

[00106] In other forms, the disease is a solid tumor. By way of example, solid tumors include, but are not limited to, pancreatic cancer, bladder cancer, colorectal cancer, breast cancer, prostate cancer, renal cancer, hepatocellular carcinoma, lung cancer, ovarian cancer, cervical cancer, rectal cancer, liver cancer, kidney cancer, stomach cancer, skin cancer, gastric cancer, esophageal cancer, head and neck cancer, melanoma, neuroendocrine cancers, CNS cancers (e.g., neuroblastoma), brain tumors (e.g., glioma, anaplastic oligodendroglioma, adult glioblastoma multiforme, and adult anaplastic astrocytoma), bone cancer, or soft tissue sarcoma. In some forms, the solid tumor is non-small cell lung cancer, small cell lung cancer, colon cancer, CNS cancer, melanoma, ovarian cancer, kidney cancer, pancreatic cancer, prostate cancer, or breast cancer.In particular embodiments, the solid tumor is non-small cell lung cancer, colon cancer, pancreatic cancer, or breast cancer. In additional embodiments, the solid tumor is non-small cell lung cancer. In one embodiment, the solid tumor is non-small cell lung cancer (NSCLC). In one embodiment, the solid tumor is modified epidermal growth factor receptor (EGFR), naive metastatic non-small cell lung cancer (NSCLC) tyrosine kinase inhibitor (TKI). By way of example, a. Petition 870200056639, dated 07 / 05 / 2020, p. 49 / 113 / 83 EGFR mutation can be the deletion of exon 19 of EGFR or the exon 21 substitution mutation (L858R). In one embodiment, the solid tumor is non-small cell lung cancer (KRAS) homologous to the metastatic Kirsten kidney sarcoma (NSCLC) viral oncogene. In still other embodiments, the solid tumor is colon cancer. In still other embodiments, the solid tumor is pancreatic cancer. In yet another embodiment, the solid tumor is metastatic pancreatic ductal adenocarcinoma. In still more embodiments, the solid tumor is breast cancer.

[00107] In additional methods of the present invention, CYT-0387 or a form thereof, including form II of CYT-0387 dihydrochloride monohydrate, is used to maintain or raise hemoglobin levels in an individual with anemia or exhibiting decreased hemoglobin levels.Anemic individuals have an endogenous hemoglobin level that is lower than the level that is normal for a healthy individual of equivalent age and gender. Acceptable or “normal” levels are now well established in medical practice. For a human male, anemia is evident when the hemoglobin level is below approximately 13.0 g / dL; for non-pregnant adult women, deficiency is evident when the hemoglobin level is below approximately 12.0 g / dL. Hemoglobin level measurement is performed using well-established techniques. Severe anemia conditions are evident when the hemoglobin level is below 8.0 g / dL.

[00108] In use, CYT-0387 is administered to an anemic individual in an amount effective to maintain or raise the individual's hemoglobin level. Administration of the drug, therefore, has minimal effect in further inhibiting the reduction of hemoglobin levels in the treated individual. More desirably, administration of the drug has the effect of increasing the individual's hemoglobin level.

[00109] Anemic individuals who would benefit from treatment Petition 870200056639, dated 07 / 05 / 2020, page 50 / 113 / 83, concerning CYT-0387, includes individuals who have undergone or are undergoing chemotherapy or radiotherapy, such as cancer patients. A wide variety of chemotherapeutic agents are known to cause a reduction in the level of red blood cell function. Similarly, individuals who are candidates for treatment with CYT-0387 are those afflicted with blood disorders, including blood cancers that result in, or are associated with, a reduction in red blood cell count. In the modalities, the individuals to be treated are individuals with anemia associated with or resulting from such blood conditions as myelodysplastic syndrome. Myelodysplastic syndromes (MDS) is a term used to describe a group of diseases characterized by ineffective hematopoiesis, leading to blood cytopenias and hypercellular bone marrow.MDS has traditionally been considered synonymous with "pre-leukemia" due to the increased risk of transformation into acute myeloid leukemia (AML). Progression to AML and the clinical consequences of cytopenias are the main causes of morbidity and mortality in MDS. Debilitating symptoms of MDS include fatigue, pallor, infection, and bleeding. Anemia, neutropenia, and thrombocytopenia are also common clinical manifestations of MDS. In other modalities, individuals to be treated are those with anemia associated with or resulting from other blood conditions, such as anemias associated with other hematological malignancies, aplastic anemia, anemia of chronic disease affecting red blood cells, and the like.Anemia of chronic disease is associated with conditions such as certain cancers, including lymphomas and Hodgkin's disease; autoimmune diseases, such as rheumatoid arthritis, systemic lupus erythematosus, inflammatory bowel disease, and polymyalgia rheumatica; long-term infections, such as urinary tract infection, HIV, and osteomyelitis; heart failure; and chronic kidney disease. Additionally, patients with anemia resulting from conditions associated with increased destruction and reduced survival of red blood cells may also experience this condition. Petition 870200056639, dated 07 / 05 / 2020, page 51 / 113 / 83 splenic sequestration could also benefit from treatment with CYT-0387. Therefore, patients afflicted with these conditions can be treated to improve their state of decreased or deficient hemoglobin.

[00110] In certain modalities, the individual to be treated is an anemic individual with thalassemia. In other modalities, the individual to be treated is an individual other than an individual with thalassemia.

[00111] In embodiments, a form of CYT-0387 of the present invention, such as form II of the CYT-0387 dihydrochloride monohydrate, is administered to an individual diagnosed with a myeloproliferative disease, such as myeloproliferative neoplasia, to improve the prognosis of the disease and, in other embodiments, particularly to treat hemoglobin deficiency or decline associated with the disease. In other embodiments, a form of CYT-0387 of the present invention, such as form II of the CYT-0387 dihydrochloride monohydrate, is administered to an anemic individual who is different from an anemic individual diagnosed with a myeloproliferative disease. This class of treatable individual presents with anemia unrelated to the myeloproliferative disease. In some embodiments, a form of CYT-0387 of the present application, such as the CYT-0387 dihydrochloride monohydrate form II, is administered to an anemic individual who is diagnosed with cancer.

[00112] “Myeloproliferative diseases” and “myeloproliferative neoplasms (MPNs)”, most notably polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF), are a diverse but interrelated group of clonal disorders of pluripotent hematopoietic stem cells that share a range of biological, pathological, and clinical features, including the relative overproduction of one or more cells of myeloid origin, growth factor-independent colony formation in vitro, bone marrow hypercellularity, hematopoiesis Petition 870200056639, dated 07 / 05 / 2020, page 52 / 113 / 83 extramedullary, splenomegaly and hepatomegaly and thrombotic or hemorrhagic diathesis. An international working group for the research and treatment of myeloproliferative neoplasms (IWG-MRT) was established to delineate and define these conditions (see, for example, Vannucchi et al, CA Cancer J. Clin., 2009, 59: 171-191), and these disease definitions should be applied for the purposes of this descriptive report. Individuals, most notably human patients, with MPN and, particularly, with PMF, are identifiable in the art using the IWG-MRT criteria mentioned above. Individuals “at risk of” a particular form of MPN are likely to have an early form of the disease and may, for example, include individuals with a genetic marker for it, such as the JAK2V617F allele which is associated with PV (> 95%), ET (60%) and PMF (60%).Individuals are also considered to be “at risk of” a form of MFN if they manifest symptoms of an earlier stage form. Thus, individuals presenting with MFN are at risk of post-PV and post-ET, both of which develop after MPN.

[00113] The response of patients with MPN and, particularly, of patients with PMF to CYT-0387 therapy is particularly robust when, according to the present invention, they are patients selected for CYT-0387 therapy based on one or more of the following criteria: i. prior therapy with a selected drug of thalidomide, lenalidomide, pomalidomide and a JAK2 inhibitor other than CYT-0387; ii. a clinical criterion selected from one or both of (1) smaller spleen size and (2) a lower percentage of circulating fragments; iii. a biochemical marker criterion selected from one or more of (1) an elevated level of at least one protein selected from EGF, TNF-α, G-CSF, IFN-α, MP-1p, HGF, MIG and Petition 870200056639, dated 07 / 05 / 2020, p. 53 / 113 / 83 VEGF; (2) a reduced level of eotaxin; and (3) an altered level of at least one selected protein of EPO, hepcidin and BMP2.

[00114] The enhanced outcome of CYT-0387 therapy, resulting from prior patient selection, manifests as a robust improvement in anemia response and / or spleen response. “Anemia response” means an increase in the patient’s hemoglobin level or a patient who was transfusion-dependent becoming transfusion-independent. Ideally, a minimum increase in hemoglobin of 2.0 g / dL, lasting at least 8 weeks, is achieved, which is the level of improvement specified in the International Working Group (IWG) consensus criteria. However, smaller, but clinically significant, increases in hemoglobin levels are also considered to be encompassed by the term “anemia response”.

[00115] By “splenic response” is meant a reduction in the size of the patient’s spleen, as assessed by any palpation of a previously palpable spleen during physical examination or by diagnostic imaging. The IWG consensus criteria specify that there must be a minimum 50% reduction in palpable splenomegaly (enlarged spleen) from a spleen that is at least 10 cm at baseline (before treatment) or from a spleen that is palpable over 5 cm at baseline and becomes non-palpable. However, smaller reductions are also considered to be encompassed by the term “splenic response”.

[00116] In one embodiment, the selected patient is one who has received prior drug therapy. More specifically, patients selected for CYT-0387 therapy include patients who have been treated, or are currently being treated, with thalidomide (CAS number 50-351) or a derivative thereof, particularly lenalidomide (CAS number 191732-72-6). Both of these drugs are used in the treatment of Petition 870200056639, dated 07 / 05 / 2020, page 54 / 113 / 83 multiple myeloma and also appear to be showing some benefit in patients afflicted with the myeloproliferative disorder. To receive the additional benefit resulting from subsequent CYT-0387 therapy, patients will be on treatment with thalidomide, lenalidomide, or pomalidomide or a similar agent, or will be treated with one of these drugs for a period of time relative to the start of CYT-0387 therapy sufficient for the effects of these drugs to manifest. Patients who meet these criteria experience a significant anemia response compared to patients naive to this drug therapy when subsequently treated with CYT-0387. In a preferred embodiment, the patient undergoing CYT-0387 is one who has previously undergone lenalidomide therapy.

[00117] Patients selected for CYT-0387 therapy also include patients who have been treated, or are currently being treated, with a JAK inhibitor other than CYT-0387. It has been particularly observed that patients previously treated with the JAK inhibitor designated INCBO 18424, or with the JAK inhibitor designated TG101348, have a more prominent splenic response to CYT-0387 therapy than patients who are not previously treated with such therapy. In a preferred embodiment, the patient selected for CYT-0387 therapy is one who, in addition to being treated with a JAK inhibitor other than CYT-0387, is also a transfusion-dependent patient. INCBO 18424 is administered in initial doses of 15 or 20 mg orally twice daily (BID), with dose titers of 5 mg BID to 25 mg BID. TG101348 is administered once daily with a maximum tolerated dose (MTD) of 680 mg / day.JAK inhibitors, other than CYT-0387, include any and all other JAK inhibitors and, in particular, other JAK inhibitors with affinity, selectivity, or JAK binding site different from CYT-0387. These properties can be determined using the JAK2 crystal structure and the modeling and activity assay approach described in US 7593820, the total of which... Petition 870200056639, dated 07 / 05 / 2020, page 55 / 113 / 83 description is incorporated in this document by reference. To receive the additional benefit resulting from subsequent CYT-0387 therapy, patients will undergo treatment with the JAK2 inhibitor; others will be treated with such drug for a period of time, relative to the start of CYT-0387 therapy, sufficient for the effects of that JAK2 inhibitor to manifest in the patient.

[00118] Patients selected for CYT-0387 therapy also include patients with altered levels of detectable protein markers. More specifically, patients in whom levels of certain protein markers, including certain cytokines and chemokines, are elevated may experience a significant benefit when treated with CYT-0387, in terms of their response to anemia and / or their spleen response to CYT-0387 therapy. In the modalities, elevated levels of one or more of the following protein markers mean that the patient is a preferred candidate for CYT-0387 therapy: (1) EGF, or epidermal growth factor, the mature form of which comprises residues 971 to 1023 of the sequence, with the designation Swiss-Prot P01133; (2) TNF-α, or tumor necrosis factor alpha, whose mature and soluble form comprises residues 77 to 233 of the sequence, with the designation Swiss-Prot P01375; (3) G-CSF, or granulocyte colony-stimulating factor, whose mature form comprises residues 30 to 207 of the sequence designated Swiss-Prot P09919; (4) IFN-α, or interferon alpha, comprises a family of subtypes whose mature forms are well known in the art; (5) MIP-Ie, or macrophage inflammatory protein 1-beta (now also known as the CC portion of chemokine 4, or CCL4), whose mature form comprises residues 24 to 92 or 26 to 92 of the sequence with the Petition 870200056639, dated 07 / 05 / 2020, p. 56 / 113 / 83 designation Swiss-Prot PI 3236; (6) HGF, or hepatocyte growth factor, whose mature forms are based on the sequence designated Swiss-Prot P14210 and include the alpha chain with residues 32 to 494 and the beta chain with residues 495 to 728; (7) MIG, or interferon-gamma-induced monokine (currently known as CXCL9) is in the family of chemotactic cytokines, whose mature form comprises residues 23 to 125 of the sequence designated Swiss-Prot Q07325; (8) VEGF, or vascular endothelial growth factor A, whose mature form comprises residues 27 to 232 of the sequence designated Swiss-Prot PI 5692.

[00119] Patients undergoing CYT-0387 therapy experience a significant splenic response when they are initially selected based on an elevation in the level of any one or more of the markers indicated above. An elevated level is a level higher than the level in a normal individual.

[00120] Patients presented for CYT-0387 therapy may also experience a significant response when they are initially selected based on a reduction in the level of the protein eotaxin. This protein, also known as eosinophil chemotactic protein, and comprising residues 24 to 97 of the sequence designated Swiss-Prot P51671, functions through interaction with CC3 to promote the accumulation of eosinophils in response to allergens, a prominent feature of allergic inflammatory reactions.

[00121] Other useful markers for selecting patients for CYT-0387 therapy include altered levels of EPO, hepcidin, and BMP-2. Another example of useful markers for selecting patients for CYT-0387 therapy includes BMP-6 levels. Petition 870200056639, dated 07 / 05 / 2020, page 57 / 113 / 83

[00122] Other markers may be used in monitoring therapy or dosage regimens with CYT-0387. By way of example, such markers may include, but are not limited to, compound 3, compound 4, compound 8, compound 10, compound 12, and compound 13. The levels of these markers can be detected by methods that are commonly used, such as those described in the examples in this application.

[00123] The “level” of a given marker is considered altered, i.e., elevated or reduced, when the level measured in a given patient differs statistically significantly from the corresponding level in a normal individual. Patients presenting with marker levels altered sufficiently, desirably, to produce a p-value equal to at least 0.05 or, more significantly, i.e., better, are selected as candidates for therapy with CYT-0387. In modalities, the p-value is equal to at least 0.03, 0.02, or 0.01, and in preferred modalities, the p-value is equal to at least 0.009, 0.007, 0.005, 0.003, 0.001 or better.

[00124] The levels of a given marker can be determined using well-established assays for the detection of the markers indicated above. In embodiments, this is achieved by extracting a biological sample from the candidate patient, such as a whole blood sample or a fraction thereof, such as plasma or serum. The sample is then treated to enrich it for the marker of interest, if desired, and the enriched or pure sample is analyzed, for example, using a detectable ligand for the marker, such as a labeled antibody that selectively binds to the marker. The amount of marker present in the sample can then be determined semi-quantitatively or quantitatively, to obtain a value that is then compared to a reference value that is the normal level for that marker in a healthy individual. As mentioned above, a difference in marker levels Petition 870200056639, dated 07 / 05 / 2020, p. 58 / 113 / 83 sufficient to obtain a p-value that is at least 0.05 indicates a significant level of altered marker, and patients with a high level of that marker (or in the case of eotaxin, a reduced level) are candidates for therapy with CYT-0387.

[00125] Patients who meet certain clinical criteria are also suitable candidates for CYT0387 therapy, including those with a relatively small spleen and those with elevated levels of circulating or peripheral blasts. These patients respond particularly well to CYT-0387 therapy in terms of their splenic response. In one modality, the selected patient is one who has not yet progressed to transfusion dependence. Splenic enlargement is assessed by palpation. Splenic volume and size can also be measured by imaging diagnostics such as ultrasound, CT, or MRI. The normal spleen size is approximately 11.0 cm in crown-rump length.

[00126] Patients with a lower percentage of circulating blasts are also suitable candidates for CYT0387 therapy. Blasts are immature precursor cells that are normally found in the bone marrow and not in the peripheral blood. They normally give rise to mature blood cells. The lowest percentage of circulating blasts is measured by cytomorphological analysis of a peripheral blood smear, as well as by multiparametric flow cytometry and immunohistochemistry. As a prognostic factor, >= 1% blasts are used.

[00127] The compounds according to the present application may be used in combination with one or more additional therapeutic agents. The therapeutic agents may be in the form of compounds, antibodies, polypeptides or polynucleotides. The therapeutic agent includes, but is not limited to, a chemotherapeutic agent, an immunotherapeutic agent, a Petition 870200056639, dated 07 / 05 / 2020, page 59 / 113 / 83 radiotherapeutic agent, an antineoplastic agent, an anticancer agent, an antiproliferative agent, an antifibrotic agent, an antiangiogenic agent, a therapeutic antibody or any combination thereof.

[00128] In one embodiment, the application provides a formulation described herein and an additional therapeutic agent, as a combination preparation for simultaneous, sequential or separate use in therapy, for example, a method of treating a disease, disorder or condition that is JAK-mediated. The therapeutic agents may be those that inhibit or modulate the activities of Bruton's tyrosine kinase, spleen tyrosine kinase, apoptosis signal regulator kinase, Janus kinase, lysyl oxidase, lysyl oxidase-like proteins, matrix metallopeptidase, bromodomain-containing proteins, adenosine receptor A2B, isocitrate dehydrogenase, serine / threonine kinase TPL2, discoidin domain receptor, serine / threonine protein kinases, IKK, MEK, EGFR, histone deacetylase, protein kinase C or any combination thereof. In certain modalities, the therapeutic agent may be selected from a PI3K inhibitor (including PI3Ky, PI3KA, PI3Ke, PI3Ka and / or pan-PI3K).A JAK (Janus kinase, including JAK1, JAK2 and / or JAK3) inhibitor, a SYK (spleen tyrosine kinase) inhibitor, a BTK (Bruton's tyrosine kinase) inhibitor, an A2B (adenosine receptor A2B) inhibitor, an ACK (CDC-activated kinase, including ACK1) inhibitor, an ASK (apoptosis signaling kinase regulator, including ASK1) inhibitor, an Aurora kinase inhibitor, a BRD (bromodomain-containing protein, including BRD4) inhibitor, a Bcl (B cell CLL / lymphoma, including Bcl-1 and / or Bcl-2) inhibitor, a CAK (CDK-activating kinase) inhibitor, a CaMK (calmodulin-dependent protein kinase) inhibitor, a CDK (cyclin-dependent kinase, including CDK1, 2, 3, 4 or 6) inhibitor, an inhibitor of (casein kinase, including CK1 and / or CK2) CK, an inhibitor of (disc receptor domain, including DDR1 or DDR2) DDR, an EGFR inhibitor, an FXR (farnesoid, Petition 870200056639, dated 07 / 05 / 2020, page. 60 / 113 / 83 x receptor) inhibitor, a FAK (focal adhesion kinase) inhibitor, a GSK (glycogen synthase kinase) inhibitor, an HDAC (histone deacetylase) inhibitor, an IDO (indoleamine 2,3-dioxygenase) inhibitor, an IDH (isocitrate dehydrogenase, including IDH1) inhibitor, an IKK (I-Kappa-B kinase) inhibitor, a KDM5 (lysine demethylase) inhibitor, an LCK (lymphocyte-specific protein tyrosine kinase) inhibitor, a LOX (lysyl oxidase) inhibitor, a LOXL (lysyl oxidase-like protein, including LOXL1, LOXL2, LOXL3, LOXL4, or LOXL5) inhibitor, an MTH (mut T homolog) inhibitor, a MEK (mitogen-activated protein kinase) inhibitor, a matrix metalloproteinase (MMP, including MMP2 and / or MMP9) inhibitor, a MAPK (mapped protein kinase) inhibitor activated by mitogen, a PD-1 inhibitor (programmed cell death protein 1), a PD-L1 inhibitor (programmed cell death ligand 1),a PDGF (platelet-derived growth factor) inhibitor, a phosphorylase kinase (PK) inhibitor, a PLK (pole-type kinase) inhibitor, including PLK1, 2, 3, a protein kinase (PK) inhibitor, including protein kinase A, B, C, an STK (serine / threonine kinase) inhibitor, a STAT (signal transduction and transcription) inhibitor, a serine / threonine kinase inhibitor, a TBK (tank kinase) inhibitor, a TLR (toll-like receptor modulator) inhibitor, including TLR-1, TLR-2, TLR-3, TLR-4, TLR-5, TLR-6, ​​TLR-7, TLR-8, TLR-9, TLR-10, TLR-11, TLR-12, and / or TLR-13), a tyrosine (tyrosine kinase) inhibitor, a TPL2 (serine / threonine) inhibitor kinase), a NEK9 inhibitor, an Abl inhibitor, a p38 kinase inhibitor, a PYK inhibitor, a PYK inhibitor, a c-Kit inhibitor, an NPM-ALK inhibitor, a Flt-3 inhibitor, a c-Met inhibitor, a KDR inhibitor, a TIE-2 inhibitor, a VEGFR inhibitor, an SRC inhibitor,an HCK inhibitor, a LYN inhibitor, an FYN inhibitor, a SIM inhibitor, a chemotherapeutic agent, an immunotherapeutic agent, a radiotherapeutic agent, an antineoplastic agent, an anticancer agent, an agent, Petition 870200056639, dated 07 / 05 / 2020, p. 61 / 113 / 83 antiproliferative agent, an antifibrotic agent, an antiangiogenic agent, a therapeutic antibody, or any combination thereof. In some embodiments, the PBK-δ inhibitor is (S)-2-(1-((9H-purin-6-yl)amino)propyl)5-fluoro-3-phenylquinazolin-4(3H)-one, as named by ChemDraw (may also be referred to as 5-fluoro-3-phenyl-2-[(1S)-1-(9H-purin-6ylamino)propyl]quinazolin-4(3H)-one) and may be synthesized by the methods described in U.S. Patent No. 7,932,260. In a certain embodiment, the SyK inhibitor is 6-(1H-indazol-6-yl)-N-(4-morpholinophenyl)imidazo[1,2-a]pyrazine-8-amine, as named by ChemDraw (it may also be referred to as 6-(1H-indazol-6-yl)-N-[4(morpholin-4-yl)phenyl]imidazo[1,2-a]pyrazine-8-amine) and can be synthesized by the methods described in U.S. Patent No. 8,450,321.In other embodiments, the BTK inhibitor is (S)-6-amino-9-(1-(but-2-inoyl)pyrrolidin-3-yl)-7-(4-phenoxyphenyl)-7H-purin-8(9H)-one, as named by ChemDraw (it may also be 6-amino-9-[(3R)-1-(2-butinoyl)3-pyrrolidinyl]-7-(4-phenoxyphenyl)-7,9-dihydro-8H-purin-8-one) and can be synthesized by the methods in U.S. Patent No. 8,557,803. In one embodiment, the MEK inhibitor is trametinib. In another embodiment, the EGFR inhibitor is erlotinib. In one embodiment, the formulation of the present application can be used together with a PI3K inhibitor, a MEK inhibitor, a TBK inhibitor, an EGFR inhibitor, or a combination thereof. In an additional embodiment, the formulation of this application can be used in conjunction with trametinib. In another embodiment, the formulation of this application can be used in conjunction with erlotinib.

[00129] Chemotherapeutic agents can be classified according to their mechanisms of action, for example, into the following groups: antimetabolite / anticancer agents, such as pyrimidine analogs (floxuridine, capecitabine and cytarabine); purine analogs, folate antagonists and Petition 870200056639, dated 07 / 05 / 2020, page 62 / 113 / 83 related inhibitors, antimitotic / antiproliferative agents, including natural products such as vinca alkaloids (vinblastine, vincristine) and microtubules such as taxanes (paclitaxel, docetaxel), vinblastine, nocodazole, epothilones and navelbine, epidipodophyllotoxins (etoposide, teniposide); Agents that cause DNA damage (actinomycin, amsacrine, busulfan, carboplatin, chlorambucil, cisplatin, cyclophosphamide, cytoxane, dactinomycin, daunorubicin, doxorubicin, epirubicin, ifosfamide, melphalan, merchlorethamine, mitomycin, mitoxantrone, nitrosourea, procarbazine, taxol, taxotere, teniposide, etoposide, triethylenethiophosphoramide); antibiotics such as dactinomycin (actinomycin D), daunorubicin, doxorubicin (adriamycin), idarubicin, anthracyclines, mitoxantrone, bleomycins, plicamycin (mitramycin), and mitomycin;enzymes (L-asparaginase, which systemically metabolizes L-asparagine and deprives cells that lack the ability to synthesize their own asparagine); antiplatelet agents; antiproliferative / antimitotic alkylating agents, such as nitrogen mustards (cyclophosphamide and analogues, melphalan, chlorambucil) and (hexamethylmelamine and thiotepa), alkyl nitrosoureas (BCNU) and analogues, streptozocin), trazenosdacarbazinine (DTIC); antiproliferative / antimitotic antimetabolites, such as folic acid analogues (methotrexate); platinum coordination complexes (cisplatin, oxyloplatin, carboplatin), procarbazine, hydroxyurea, mitotane, aminoglutethimide; hormones, hormone analogues (estrogen, tamoxifen, goserelin, bicalutamide, nilutamide) and aromatase inhibitors (letrozole, anastrozole); anticoagulants (heparin, synthetic heparin salts and other thrombin inhibitors);fibrinolytic agents (such as tissue plasminogen activator, streptokinase and urokinase), aspirin, dipyridamole, ticlopidine, clopidogrel; antimigration agents; antisecretion agents (breveldine); immunosuppressants tacrolimus, sirolimus, azathioprine, mycophenolate; compounds (TNP-470, genistein) and growth factor inhibitors (vascular endothelial growth factor inhibitors, inhibitors; Petition 870200056639, dated 07 / 05 / 2020, page 63 / 113 / 83 of fibroblast growth factor); angiotensin receptor blocker, nitric oxide donors; antisense oligonucleotides; antibodies (trastuzumab, rituximab); cell cycle inhibitors and differentiation inducers (tretinoin); topoisomerase inhibitors (doxorubicin (adriamycin), daunorubicin, dactinomycin, eniposide, epirubicin, etoposide, idarubicin, irinotecan and mitoxantrone, topotecan, irinotecan, camptotesin), corticosteroids (cortisone, dexamethasone, hydrocortisone, methylprednisolone, prednisone and prenisolone); growth factor kinase signal transduction inhibitors; Inducers of dysfunction, toxins such as cholera toxin, ricin, Pseudomonas exotoxin, Bordetella pertussis adenylate cyclase toxin or diphtheria toxin and caspase activators; and chromatin.

[00130] As used in the present invention, the term “chemotherapeutic agent” or “chemotherapeutic” (or chemotherapy, in the case of treatment with a chemotherapeutic agent) is intended to encompass any non-protein (i.e., non-peptide) chemical compound useful in the treatment of cancer.Examples of chemotherapeutic agents include alkylating agents, such as thiotepa and cyclophosphamide (CYTOXAN); alkyl sulfonates, such as busulfan, improsulfan, and piposulfan; aziridines, such as benzodopa, carboquone, meturedopa, and uredopa; emilerumins and memylamelamines, including alfetamine, triemylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide, and trimemylenemelamine; acetogenins (especially bulatacin and bulatacinone); camptothecin (including the synthetic analogue topotecan); bryostatin; calistatin; CC-1065 (including its synthetic analogues adozelesin, carzelesin, and bizelesin); cryptophycins (particularly cryptophycin 1 and cryptophycin 8); dolastatin; duocarmycin (including the synthetic analogues KW-2189 and CBI-TMI); eleutherobin; pancratistatin; a sarcodictin; spongistatin; nitrogen mustards, such as chlorambucil, chlornaphazine, colofosfamide, estramustine. Petition 870200056639, dated 07 / 05 / 2020, page 64 / 113 / 83 ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembin, fenesterin, prednimustine, trofosfamide, uracil mustard; nitrosoureas such as carmustine, chlorozotocin, foramustine, lomustine, nimustine, ranimustine; antibiotics such as enedine antibiotics (e.g., calicheamicin, especially calicheamicin gamma II and calicheamicin phiI1, see, for example, Agnew, Chem. Intl. Ed. Engl, 33:183-186 (1994); dynecin, including dynecin A; bisphosphonates, such as clodronate; esperamycin;as well as the neocarzinostatin chromophore and related enedin chromoprotein antibiotic chromophores), aclacinomycins, actinomycin, autramycin, azaserine, bleomycins, cactinomycin, carabicin, carninomycin, carzinophilin, cromomycin, dactinomycin, daunorubicin, detorrubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin (including morpholino-doxorubicin, cyanomorpholinodoxorubicin, 2-pyrrolino-doxorubicin and deoxyxorubicin), epirubicin, esorubicin, idarubicin, marcelomycin, mitomycins such as mitomycin C, mycophenolic acid, nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin, chelamicin, rodorrubicin, Streptonigrin, streptozocin, tubercidine, ubenimex, zinostatin, zorubicin; antimetabolites, such as methotrexate and 5-fluorouracil (5-FU); folic acid analogs, such as demopterin, methotrexate, pteropterin, trimetrexate; purine analogs, such as fludarabine, 6-mercaptopurine, tiamiprine, thioguanine;Pyrimidine analogs, such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxyfluridine, enocitabine, floxuridine; androgens, such as calusterone, dromostanolone propionate, epitiostanol, mepitiostane, testolactone; antiadrenals, such as aminoglutethimide, mitotane, trilostane; folic acid supplements, such as frolinic acid; aceglathone; aldofosfamide glycoside; aminolevulinic acid; enyluracil; amsacrine; hestrabucil; bisanthrene; edatraxate; defofamine; demecolcine; diaziquone; elformin; elliptinium acetate; epothilone; etoglucoside; gallium nitrate; Petition 870200056639, dated 07 / 05 / 2020, page 65 / 113 / 83 hydroxyurea; lentinan; leucovorin; lonidamine; maytansinoids, such as maytansine and ansamitocins; mitoguazone; mitoxantrone; mopidamol; nitracrine; pentostatin; fenamet; pirarubicin; losoxantrone; fluoropyrimidine; folinic acid; podophyllinic acid; 2-ethylhydrazide; procarbazine; PSK(r); razoxane; rhizoxine; sizofirane; spirogermanium; tenuazonic acid; triaziquone; 2,2',2''-trichlorotriemylamine; trichothecenes (especially T-2 toxin, verracurin A, roridine A and anguidin); urethane; vindesine; dacarbazine; manomustine; mitobronitol; mitolactol; pipobromane; gacytosine; arabinoside (“Ara-C”); cyclophosphamide; thiopeta; taxoids, for example, paclitaxel (TAXOL®) and docetaxel (TAXOTERE®); chlorambucil; gemcitabine (Gemzar®); 6-thioguanine; mercaptopurine; methotrexate; platinum analogs, such as cisplatin and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitroxantrone; vancristine;vinorelbine (Navelbine®); novantron; teniposide; edatrexate; daunomycin; aminopterin; xeoloda; ibandronate; CPT-11; topoisomerase inhibitor RFS 2000; difluoromethylornithine (DMFO); retinoids, such as retinoic acid; capecitabine; FOLFIRI (fluorouracil, leucovorin and irinotecan) and pharmaceutically acceptable salts, acids or derivatives of any of the above. One or more chemotherapeutic agents are used or included in this application. For example, gemcitabine, nab-paclitaxel, gemcitabine / nab-paclitaxel, capecitabine, oxaliplatin and capecitabine / oxaliplatin may be used with the formulation of this application.

[00131] Also included in the definition of “chemotherapeutic agent” are anti-hormonal agents that act to regulate or inhibit the action of the hormone on tumors, such as antiestrogens and selective estrogen receptor modulators (SERMs), including, for example, tamoxifen (including Nolvadex™), raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, queoxifene, LY117018, onapristone and Petition 870200056639, dated 07 / 05 / 2020, page 66 / 113 / 83 toremifene (Fareston®); aromatase enzyme inhibitors, which regulate estrogen production in the adrenal glands, such as, for example, 4(5)imidazoles, aminoglutethimide, megestrol acetate (Megace®), exemestane, formestane, fadrozole, vorozole (Rivisor®), letrozole (Femara®) and anastrozole (Arimidex®); and antiandrogens, such as flutamide, bicalutamide, nilutamide, leuprolide and goserelin; and pharmaceutically acceptable salts, acids or derivatives of any of the above described.

[00132] Antiangiogenic agents include, but are not limited to, retinoic acid and its derivatives, 2-methoxyestradiol, ANGIOSTATIN(r), ENDOSTATIN(r), suramin, squalamine, tissue metalloproteinase inhibitor-1, tissue metalloproteinase inhibitor-2, plasminogen activator inhibitor-1, plasminogen activator inhibitor-2, cartilage-derived inhibitor, paclitaxel (nab-paclitaxel), platelet factor 4, protamine sulfate (clupein), sulfated chitin derivatives (prepared from queen crab shells), sulfated peptidoglycan polysaccharide complex (sp-pg), staurosporine, matrix metabolism modulators, including, for example, proline analogs (1-azetidino-2-carboxylic acid (LACA), cis-hydroxyproline, d,I-3,4dehydroproline, tiaproline, alpha-dipyridyl, betaaminopropionitrile fumarate, 4-propyl-5-(4-pyridinyl)-2(3h)-oxazolone;methotrexate, mitoxantrone, heparin, interferons, serum 2-macroglobulin, chimp-3, chymostatin, beta-cyclodextrin tetradecasulfate, eponemycin; fumagillin, sodium gold thiomalate, d-penicillamine (CDPT), serum beta-1 anticollagenase, alpha-2-antiplasmin, bisanthrene, lobenzarit disodium, disodium n2-carboxyphenyl-4-chloroanthonylic acid or “CCA”, thalidomide; angiostatic steroid, carboxinaminolmidazole; metalloproteinase inhibitors, such as BB94. Other antiangiogenic agents include antibodies, preferably monoclonal antibodies against these angiogenic growth factors: betaFGF, alpha-FGF, FGF-5, VEGF isoforms, VEGF-C, HGF / SF and Ang; Petition 870200056639, dated 07 / 05 / 2020, page 67 / 113 / 83 1 / Ang-2. See Ferrara N. and Alitalo, K. “Clinical application of angiogenic growth factors and their inhibitors” (1999) Nature Medicine 5:1359-1364.

[00133] Antifibrotic agents include, but are not limited to, compounds such as beta-aminopropionitrile (BAPN), as well as the compounds described in U.S. Patent No. 4,965,288 to Palfreyman et al., issued October 23, 1990, entitled “Inhibitors of lysyl oxidase,” relating to lysyl oxidase inhibitors and their use in the treatment of diseases and conditions associated with abnormal collagen deposition; U.S. Patent No. 4,997,854 to Kagan et al., issued March 5, 1991, entitled “Anti-fibrotic agents and methods for inhibiting the activity of lysyl oxidase in situ using adjacently positioned diamine analogue substrate,” relating to compounds that inhibit LOX for the treatment of various pathological fibrotic states, which are incorporated herein by reference. Additional exemplary inhibitors are described in U.S. Patent No. 4,943,593, by Palfreyman, et al., issued on July 24, 1990, entitled “Inhibitors of lysyl oxidase,” relating to compounds such as 2-isobutyl-3-fluorine, chlorine, or bromoallylamine; as well as, for example, U.S. Patent No. 5,021,456; U.S. Patent No. 5,505,9714; U.S. Patent No. 5,120,764; U.S. Patent No. 5,182,297; U.S. Patent No. 5,252,608 (relating to 2-(1-naphthyloxymemyl)-3-fluoroallylamine); and U.S. Patent Application No. 2004 / 0248871, which are incorporated herein by reference. Exemplary antifibrotic agents also include primary amines that react with the carbonyl group of the active site of lysyl oxidases and, more particularly, those that produce, after binding to the carbonyl, a resonance-stabilized product, such as the following primary amines: phenylemamine, hydrazine, phenylhydrazine and its derivatives, semicarbazide and urea derivatives, aminonitriles, such as beta-aminopropionitrile (BAPN), or. Petition 870200056639, dated 07 / 05 / 2020, page 68 / 113 / 83 2-nitroethylamine, saturated or unsaturated haloamines, such as 2-bromoethylamine, 2-chloroethylamine, 2-trifluoroethylamine, 3-bromopropylamine, p-halobenzylamines, and selenohomocysteine ​​lactone. In addition, antifibrotic agents are copper chelating agents, which may or may not penetrate cells. Exemplary compounds include indirect inhibitors, such as compounds that block aldehyde derivatives resulting from the oxidative deamination of lysyl and hydroxylysyl residues by lysyl oxidases, such as thiolamines, in particular D-penicillamine, or their analogues, such as 2-amino-5-mercapto-5-methylhexanoic acid, D-2-amino-3-methyl-3-((2-acetamidoethyl)dithio)butanoic acid, p-2-amino-3-methyl-3-((2-aminoethyl)dithio)butanoic acid, sodium sulfate of 4-((p-1-dimethyl-2-amino-2-carboxyethyl)dithio)butane, 2-acetamidoethyl-2-acetamidoethanethiol sulfonate and sodium trihydrate of 4-mercaptobutanesulfinate.

[00134] Immunotherapeutic agents include, but are not limited to, therapeutic antibodies suitable for treating patients, such as those for treating patients;como abagovomabe, adecatumumabe, affutuzumabe, alemtuzumabe, altumomabe, amatuximabe, anatumomabe, arcitumomabe, bavituximabe, bectumomabe, bevacizumabe, bivatuximabe, blinatumomabe, brentuximabe, cezu cantuximabe, cituzumabe cixutumabe, clivatuzumabe, conatumabe, daritumabe, drozitumabe, duligoumabe, dusigitumabe, detumomabe, dacetuzumabe, dalotuzumabe, ecromeximabe, elotuzumabe, ensituximabe, ertumaxomabe, etaracituzumabe,berie,fizumabe, faromeximabe flanvomabe, future, speakers, leaders, leaders, writers, leaders, governors, leaders, leaders, leaders, leaders, filmmakers, leaders, leaders, leaders, leaders, leaders, leaders matumumabe, matuzumabe, milatuzumabe, minretumabe, mitumoma, moxetumabe, narnamama, naptumomabe, necimatumabe, nimotuzumabe, nofetumomabn, ocaratuzumabe,; Petition 870200056639, dated 07 / 05 / 2020, page. 69 / 113 / 83 ofatumumab, olaratumab, onartuzumab, oportuzumab, oregovomab, panitumumab, parsatuzumab, patritumab, pemtumomab, pertuzumab, pintumomab, pritumumab, racotumomab, radretumab, rilotumumab, rituximab, robatumumab, satumomab, sibrotuzumab, siltuximab, simtuzumab, solitomab, tacatuzumab, taplitumomab, tenatumomab, teprotumumab, tigatuzumab, tositumomab, trastuzumab, tucotuzumab, ublituximab, veltuzumab, vorsetuzumab, votumumab, zalutumumab, obinutuzumab, CC49 and 3F8. The therapeutic antibodies exemplified may also be labeled or combined with a radioisotope particle, such as indium In 111, yttrium Y 90, and iodine I-131.

[00135] The application also provides a method for treating an individual who is undergoing one or more standard therapies, such as chemotherapy, radiotherapy, immunotherapy, surgery, or a combination thereof. Therefore, one or more therapeutic agents or inhibitors may be administered before, during, or after the administration of chemotherapy, radiotherapy, immunotherapy, surgery, or a combination thereof.

[00136] Other examples of chemotherapy treatments (including standard or experimental chemotherapies) are described below. In addition, the treatment of certain lymphomas is reviewed in Cheson, BD, Leonard, JP, “Monoclonal Antibody Therapy for B-Cell Non-Hodgkin's Lymphoma” The New England Journal of Medicine 2008, 359(6), p. 613-626; and Wierda, WG, “Current and Investigational Therapies for Patients with CLL” Hematology 2006, p. 285-294. Lymphoma incidence patterns in the United States are profiled in Morton, LM, et al. “Lymphoma Incidence Patterns by WHO Subtype in the United States, 1992-2001” Blood 2006, 107(1), p. 265-276.

[00137] Examples of immunotherapeutic agents include, but are not limited to, rituximab (as Rituxan), alemtuzumab (as Campath, Petition 870200056639, dated 07 / 05 / 2020, pages 70 / 113 / 83 MabCampath), anti-CD19 antibodies, anti-CD20 antibodies, anti-MN-14 antibodies, anti-TRAIL antibodies, anti-TRAIL DR4 and DR5 antibodies, anti-CD74 antibodies, apolizumab, bevacizumab, CHIR-12.12, epratuzumab (humanized hLL2-anti-CD22 antibody), galiximab, ha20, ibritumomab tiuxetane, lumiliximab, milatuzumab, ofatumumab, PRO131921, SGN-40, WT-1 analog peptide vaccine, WT1 peptide vaccine 126-134, autologous human tumor-derived HSPPC-96, veltuzumab. Additional immunotherapy agents include the use of cancer vaccines based on the genetic makeup of an individual patient's tumor, for example, the lymphoma vaccine GTOP-99 (MyVax®).

[00138] Examples of chemotherapeutic agents include aldesleukin, alvocidib, antineoplaston AS2-1, antineoplaston A10, antithymocyte globulin, amifostine trihydrate, aminocamptothecin, arsenic trioxide, beta-alethin, Bcl-2 protein family inhibitor ABT-263, ABT-199, BMS345541, bortezomib (Velcade®), briostatin 1, busulfan, carboplatin, campat-1H, CC-5103, carmustine, caspofungin acetate, clofarabine, cisplatin, cladribine (Leustarin), chlorambucil (Leukeran), curcumin, cyclosporine, cyclophosphamide (Cyloxan, Endoxan, Endoxana, Cyclostin), dexamethasone, DT PACE, docetaxel, dolastatin 10, doxorubicin (Adriamycin® Adriblastine), doxorubicin hydrochloride, enzastaurin, epoetin alfa, etoposide, everolimus (RAD001), fenretinide, filgrastim, melphalan, mesna, flavopiridol, fludarabine (Fludara), geldanamycin (17 AAG), ifosfamide, irinotecan hydrochloride, ixabepilone, lenalidomide (Revlimid®, CC-5013), lymphokine-activated killer cells, melphalan,methotrexate, mitoxantrone hydrochloride, motexafin, gadolinium, mycophenolate mofetil, nelarabine, oblimersen (Genasense), Obatoclax (GX15-070), oblimersen, octreotide acetate, omega-3 fatty acids, oxaliplatin, paclitaxel, PD0332991, pegylated liposomal doxorubicin hydrochloride, pegfilgrastim, pentastatin (Nipent), perifosine, prednisolone, prednisone, R, Petition 870200056639, dated 07 / 05 / 2020, page. 71 / 113 / 83 roscovitine (Selicilib, CYC202), recombinant interferon-alpha, recombinant interleukin-12, recombinant interleukin-11, recombinant flt3 ligand, recombinant human thrombopoietin, rituximab, sargramostim, sildenafil citrate, simvastatin, sirolimus, styryl sulfones, tacrolimus, tanespimycin, torisel (CCl-779), thalidomide, allogeneic lymphocyte therapies, thiotepa, tipifarnib, Velcade® (bortezomib or PS-341), vincristine (BACOD), vincristine sulfate, vinorelbine ditartrate, vorinostat (SANTOS), vorinostat and FR (fludarabine, rituximab), CHOP (cyclophosphamide, doxorubicin, vincristine, prednisone), CVP (cyclophosphamide, vincristine and prednisone), FCM (fludarabine, cyclophosphamide, mitoxantrone), FCR (fludarabine, cyclophosphamide, rituximab), hyperCVAD (hyperfractionated cyclophosphamide, vincristine, doxorubicin, dexamethasone, methotrexate, cytarabine), ICE (ifosfamide, carboplatin and etoposide), MCP (mitoxantrone,Chlorambucil and prednisolone), R-CHOP (rituximab plus CHOP), R-CVP (rituximab plus FCM), R-FCM (rituximab plus FCM), R-ICE (rituximab ICE), and R-MCP (R-MCP). For example, oxaliplatin may be used in conjunction with this application.

[00139] Therapeutic treatments may be supplemented or combined with any of the above therapies with transplantation or stem cell treatment. An example of a modified approach is radioimmunotherapy, in which a monoclonal antibody is combined with a radioisotope particle, such as indium In 111, yttrium Y 90, and iodine I-131. Examples of combination therapies include, but are not limited to, iodine-131 tositumomab (Bexxar®), yttrium-90 ibritumomab tiuxetan (Zevalin®), Bexxar® with CHOP.

[00140] Other therapeutic procedures include peripheral blood stem cell transplantation, autologous hematopoietic stem cell transplantation, autologous bone marrow transplantation, antibody therapy, biological therapy, enzyme inhibitor therapy, irradiation Petition 870200056639, dated 07 / 05 / 2020, page 72 / 113 / 83 total body, stem cell infusion, bone marrow ablation with stem cell support, in vitro treated peripheral blood stem cell transplantation, umbilical cord blood transplantation, immunoenzymatic technique, pharmacological study, low-LET cobalt-60 gamma ray therapy, bleomycin, conventional surgery, radiotherapy and non-myeloablative allogeneic hematopoietic stem cell transplantation.

[00141] This application provides a formulation, alone or in combination with one or more therapeutic agents, for the treatment of myeloproliferative and cancerous diseases. This application also provides a method for treating a disease comprising administering the formulation described herein (including form II of CYT-0387 dihydrochloride monohydrate in a tablet format) to a patient in need thereof, wherein the disease is selected from the group consisting of polycythemia vera (PV), myelofibrosis, thrombocythemia, essential thrombocythemia (ET), pancreatic cancer, metastatic pancreatic ductal adenocarcinoma, breast cancer, colon cancer, non-small cell lung cancer (NSCLC) and metastatic NSCLC, including EGFR-modified NSCLC, metastatic EGFR TKI-naive NSCLC and metastatic KRAS-modified NSCLC.Furthermore, the present invention provides a method for treating a disease comprising administering the formulation described herein (including form II of CYT-0387 dihydrochloride monohydrate in a tablet format) and optionally administering one or more therapeutic agents to a patient in need thereof, wherein the one or more therapeutic agents are selected from the group consisting of trametinib, erlotinib, gemcitabine, nab-paclitaxel, oxaliplatin, capecitabine or a combination thereof.

[00142] It should be understood that the examples below illustrate certain aspects of the present application. It is also understood that the values ​​and parameters shown in the examples may be modified within reasonable variation, and that various modifications may be made within the scope of the present application. Petition 870200056639, dated 07 / 05 / 2020, page 73 / 113 / 83 Example 1. Methods for Producing

[00143] N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4yl)benzamide (CYT-0387) can be synthesized as described in U.S. Patent No. 8,486,941 and PCT application WO 2012 / 071612. CYT-0387 dihydrochloride monohydrate form II from CYT-0387 dihydrochloride anhydrous form I

[00144] To a suspension of the anhydrous form I of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride (CYT0387) in methanol, an excess molar of hydrochloric acid in water was added. The resulting solids were isolated and washed with methanol and aqueous hydrochloric acid to yield the form II of the CYT-0387 dihydrochloride monohydrate. CYT-0387 dihydrochloride monohydrate form II from CYT-0387 free base

[00145] To a free base suspension of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide (CYT-0387) in methanol, an excess molar of concentrated hydrochloric acid was added. The resulting suspension was optionally seeded with form II of the N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4yl)benzamide (CYT-0387) hydrochloride monohydrate, and water was added. Form II of the N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4yl)benzamide (CYT-0387) dihydrochloride monohydrate can be prepared as described above. The resulting solids were isolated and washed with methanol and aqueous hydrochloric acid to produce form II of the CYT0387 dihydrochloride monohydrate. CYT-0387 monohydrochloride anhydrous form I from CYT-0387 free base

[00146] A freebase suspension of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide (CYT-0387) in methanol was Petition 870200056639, dated 07 / 05 / 2020, p. 74 / 113 / 83 added 1.0 molar equivalent of concentrated hydrochloric acid. The resulting solids were isolated and washed with methanol to produce the anhydrous form I of CYT-0387 monochloride. Anhydrous form III of CYT-0387 monohydrochloride from anhydrous form I of CYT-0387 monohydrochloride

[00147] A suspension of anhydrous form I of CYT0387 monochloride is mixed in water / THF (30% water; v / v). The resulting solids were isolated and washed with a water / THF mixture to produce anhydrous form III of CYT-0387 monochloride. CYT-0387 monohydrochloride anhydrous form III from CYT-0387 dihydrochloride monohydrate form II

[00148] A suspension of form II of CYT-0387 dihydrochloride monohydrate is stirred in methanol / water (30% water; v / v). The resulting solids were isolated and washed with a methanol / water mixture to produce the anhydrous form III of CYT-0387 hydrochloride.

[00149] The above shapes were characterized by various analytical techniques, including X-ray powder diffraction pattern (XPPD), differential scanning calorimetry (DSC), thermographic analysis (TGA), and dynamic vapor sorption (DVS) using the procedures described below.

[00150] X-ray powder diffraction: XRPD analysis was performed on a diffractometer (PANanalytical XPERT-PRO, PANanalytical BV, Almelo, Netherlands) using copper radiation (Cu Kα, λ = 1.5418 Å). Samples were prepared for analysis by depositing the powder sample in the center of an aluminum holder equipped with a zero-backed signal plate. The generator was operated at a voltage of 45 kV and an amperage of 40 mA. The slits used were Soller 0.02 rad, antiscatter 1.0°, and divergence. The sample rotation speed was 2 s. Scans were performed from 2 to 40° 2θ for 15 min, with a step size of 0.0167° 2θ. Data analysis was performed using X'Pert Highscore version 2.2c (PANalytical). Petition 870200056639, dated 07 / 05 / 2020, pages 75 / 113 / 83 (PANalytical BV, Almelo, Netherlands) and by X'Pert data viewer version 1.2d (PANalytical BV, Almelo, Netherlands).

[00151] The XRPD standard for form II of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4yl)benzamide dihydrochloride monohydrate is shown in Figure 6.

[00152] The XRPD standard for the anhydrous form I of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride is shown in Figure 7.

[00153] The XRPD standard for the anhydrous form III of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride is shown in Figure 8.

[00154] The XRPD peaks of the various forms of CYT-0387 are shown in Table 1 above.

[00155] Differential scanning calorimetry: The thermal properties of form II of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride monohydrate were evaluated using a differential scanning calorimetry (DSC) instrument (TA Q1000, TA Instruments, Newcastle, DE, USA). Approximately 5 to 10 mg of solid sample were placed in a standard aluminum pan with a hole for each experiment and heated at a rate of 10°C / min under 50 mL / min of nitrogen purge. Data analysis was performed using Universal Analysis 2000, version 4.7A (TA Instruments, Newcastle, DE, USA). The heat of fusion analysis was performed by sigmoidal integration of the endothermic fusion peak.

[00156] The DSC for form II of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride monohydrate is represented in Figure 9.

[00157] The DSC for anhydrous form I of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride is Petition 870200056639, dated 07 / 05 / 2020, page 76 / 113 / 83 represented in figure 10.

[00158] The DSC for anhydrous form III of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride is represented in figure 11.

[00159] Thermogravimetric analysis: Thermogravimetric analysis (TGA) of form II of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride monohydrate was performed on a TGA instrument (TA Q500, TA Instruments, Newcastle, DE, USA). Approximately 5 to 10 mg of solid sample were placed in an open aluminum pan for each experiment and heated at a rate of 10°C / min under 60 mL / min of nitrogen purge. Data analysis was performed using Universal Analysis 2000, version 4.7A (TA Instruments, Newcastle, DE, USA).

[00160] The TGA for form II of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride monohydrate is represented in Figure 12.

[00161] The TGA for anhydrous form I of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride is represented in figure 13.

[00162] The TGA for anhydrous form III of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride is represented in figure 14.

[00163] Dynamic vapor sorption: The hygroscopicity of form II of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride monohydrate was evaluated at room temperature using a dynamic vapor sorption (DVS) instrument (TGA Q5000 TA Instruments, Newcastle, DE). Water adsorption and desorption were studied as a function of relative humidity (RH) in the range of 0 to 90% at room temperature. The humidity in the chamber increased from the initial level of Petition 870200056639, dated 07 / 05 / 2020, p. 77 / 113 65 / 83 The relative humidity (RH) was increased from 50% to 60% RH and maintained until the solid and atmosphere reached equilibrium. The equilibrium test continued until it passed, or ended after 10 hours. At this point, the RH was increased by 10% more and the process was repeated until 90% RH was reached and equilibrium was achieved. During this period, water absorption was monitored. For desorption, the relative humidity decreased similarly to measure a complete adsorption / desorption cycle. All experiments were performed in dm / dt (change in mass over time) mode to determine the equilibrium parameter. Approximately 4 mg of solid CYT-0387 was used. Data analysis was performed using Universal Analysis 2000, version 4.7A (TA Instruments, Newcastle, DE, USA).

[00164] The DVS for form II of N(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride monohydrate is represented in Figure 15.

[00165] X-ray crystallography data for the single crystal form II of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide dihydrochloride monohydrate CYT-0387 and the anhydrous form I of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride (CYT-0387) are summarized in Table 2 below. Data from the additional characterization of the crystals are summarized in Table 3 below. Table 2: Single-crystal X-ray crystallography data Shape and composition Unit cell dimensions Distance (Â) Angle (°) Form APF Water: Solvent abcd β γ Form II of CYT-0387 dihydrochloride monohydrate 1:1:0 10.2837(6) 10.4981(6) 11.5143(7) 83.297(2) 87.649(2) 67.445(2) Form I of anhydrous CYT0387 monohydrochloride 1:0:0 9.4255(3) 11.6729(4) 19.7561(6) 85.3940(10) 88.103(2) 83.821(2) Table 3: Crystal data and structure refinement Property Form II of CYT-0387 dihydrochloride monohydrate Anhydrous form I of CYT-0387 monohydrochloride Empirical formula C23H26Cl2N6O3 C23H23CIN6O2 Petition 870200056639, dated 07 / 05 / 2020, pp. 78 / 113 66 / 83 Formula weight 505.40 450.92 Temperature 100(2) K 100(2) K Wavelength 1.54178 Å 1.54178 Å Crystal system Triclinic Triclinic Spacer group Pl Pl Volume 1140.14(12) Å3 2153.36(12) Å3 Z 2 4 Density (calculated) 1.472 g / cm3 1.472 g / cm3 100(2) K represents 100 ± 2 °Κ Microscopic images of the various forms of the invention were acquired using an Olympus polarizing microscope (BX-51, Olympus, Center Valley, PA, USA). Samples were dispersed in mineral oil and examined under cross-polarized light using a 530 nm waveplate (results not shown). Example 2

[00166] Tablets comprising form II of CYT-0387 dihydrochloride monohydrate in amounts equivalent to 100 mg, 150 mg and 200 mg of CYT-0387 free base may be prepared according to the process described in this document. Tablets comprising form II of CYT-0387 dihydrochloride monohydrate in amounts equivalent to 100 mg, 150 mg, 200 mg, 250 mg and 300 mg of CYT-0387 free base have been prepared. Table 4 below summarizes the formulations of such tablets. Table 4: Tablet Formulation Quantity per dosage unit Component Quantity (% w / w) Concentration of 100 mg (mg) Concentration of 150 mg (mg) Concentration of 200 mg (mg) Concentration of 250 mg (mg) Concentration of 300 mg (mg) Form II of CYT-0387 dihydrochloride monohydrate 40.65 121.94 182.91 243.88 304.88 365.85 Propyl gallate 0.20 0.60 0.90 1.20 1.50 1.80 Microcrystalline cellulose (PH 105) 34.23 102.70 154.05 205.40 256.73 308.07 Lactose (Fast Flo 316) 16.92 50.76 76.14 101.52 126.9 152.28 Sodium starch glycolate 6.00 18.00 27.00 36.00 45.00 54.00 Colloidal silicon dioxide 0.50 1.50 2.25 3.00 3.75 4.50 Magnesium stearate 1.50* 4.50 6.750 9.00 11.25 13.50 Total of the tablet with core 100.00 300.00 450.00 600.00 750.00 900.00 Opadry II Brown 85F165010 4.00 12.00 18.00 24.00 30.00 36.00 Film-coated tablet total 104.00 312.00 468.00 624.00 780.00 936.00 Petition 870200056639, dated 07 / 05 / 2020, page 79 / 113 / 83 * 0.75% intragranular, 0.75% extragranular of Mg stearate

[00167] The tablet formulation in Table 4 comprised propyl gallate, which decreased the extent or levels of oxidative degradation of CYT-0387 form II and resulted in increased stability of CYT-0387 form II. This was determined from the study that investigated the potential effects of different antioxidants on inhibiting or preventing the degradation of CYT-0387 dihydrochloride monohydrate form II. In the initial study, five antioxidants with three different mechanisms of action were examined: free radical scavenging antioxidants (propyl gallate, butylated hydroxyanisole (BHA), and butylated hydroxytoluene (BHT)), a sacrificial reducing agent (ascorbic acid), and an oxygen scavenger (sodium metabisulfite).

[00168] Aqueous solutions containing 20 pg / mL of CYT-0387 dihydrochloride monohydrate form II in 70% 50 mM (v / v) acetate buffer (pH 4.0) and 30% (v / v) methanol were incubated in the absence (which were used as a control) and in the presence of 0.1% (w / v) antioxidant at 60°C for up to 7 days. On days 0, 5, and 7, the solution was analyzed using reversed-phase HPLC with a Zorbax SB-C8 column (Phenomenex, Torrance, CA). As shown in Table 7, the presence of propyl gallate, BHA, or ascorbic acid at the 0.1% (w / v) level inhibited or prevented the degradation of CYT-0387 form II relative to BHT and sodium metabisulfite at the same level. Less than one percent of the CYT-0387 form II was degraded in the presence of propyl gallate or BHA. Although total degradation products could not be determined for 0.1% ascorbic acid due to interference, the major oxidative degradation products were not observed (data not shown).

[00169] In further studies, the effects of lower concentrations of 0.01% and 0.001% (w / v) of propyl gallate, BHA, and ascorbic acid on the stability of CYT-0387 were examined under the same conditions. The results of these studies are summarized in Table 7. The results showed that, at the antioxidant level of 0.01% (w / v), a Petition 870200056639, dated 07 / 05 / 2020, p. 80 / 113 / 83 greater stability of form II of CYT-0387 was observed in the presence of propyl gallate or ascorbic acid at 60°C for up to 7 days. Furthermore, at the 0.001% (w / v) antioxidant level, the presence of propyl gallate decreased or inhibited the degradation of CYT-0387 compared to BHA and ascorbic acid (Table 7). These results indicate that, among the antioxidants tested, propyl gallate was the most effective in inhibiting or preventing the degradation of form II of CYT0387 dihydrochloride monohydrate. Table 7: Effects of antioxidants on the degradation of CYT form II 0387 in aqueous buffer at pH 4 at 60°C. Antioxidant Quantity (% w / v) Time Point (days) Total Degradation at (%) Control 0 0 0.29 5 17.24 7 21.84 Butylated hydroxytoluene (BHT) 0.1 0 0.06 5 1.71 7 4.35 Sodium metabisulfite 0.1 0 0.33 5 43.83 7 46.31 Propyl gallate 0.1 0 0.29 5 0.31 7 0.32 0.01 0 0.22 5 0.38 7 0.72 0.001 0 0.22 5 1.98 7 4.32 Butylated hydroxyanisole (BHA) 0.1 0 0.45 5 0.50 7 1.24 0.01 0 0.55 5 1.75 7 3.04 0.001 0 0.30 5 7.64 7 13.76 Ascorbic acid 0.1 0 N / Ab 5 N / Ab 7 N / Ab 0.01 0 0.20 5 1.58 7 1.78 0.001 0 0.51 5 17.95 7 21.28 aIncludes impurity that was present in the formulation. bN / A: Not available Petition 870200056639, dated 07 / 05 / 2020, page 81 / 113 / 83

[00170] Next, the stability of the 100 mg tablet formulation comprising form II of CYT-0387 was examined in the presence of 0%, 0.2%, 0.5% or 1.0% propyl gallate at 25°C / 60% RH (relative humidity) or 40°C / 75% RH for up to 6 months. Degradation profiles were determined at months 0, 1, 3, and 6. Results from studies at 40°C / 75% RH for up to 6 months are summarized in Table 8. Results showed that, at 40°C / 75% RH, the 100 mg CYT-0387 form II tablet formulation with 0.2% propyl gallate exhibited greater stability compared to the 100 mg CYT-0387 form II tablet formulation with 0%, 0.5%, or 1.0% propyl gallate. Results from the study at 25°C / 60% RH showed that degradation of CYT-0387 form II was also reduced by 0.2%, 0.5%, and 1.0% propyl gallate (data not shown).The observed trend was similar for the degradation profiles at 25°C / 60% RH compared to that observed at 40°C / 75% RH, i.e., greater stability of form II of CYT-0387 in the tablet formulation was observed at 0.2% propyl gallate compared to 0%, 0.5%, and 1% propyl gallate (data not shown). Taken together, these results indicate that, among the antioxidants and percentages examined in these studies, 0.2% propyl gallate provided the preferred level of stability for form II of CYT-0387 dihydrochloride monohydrate. Table 8: Effects of propyl gallate levels on the degradation of CYT-0387 form II in the 100 mg tablet formulation. Propyl gallate (%) Time point (months) Total degradation (%) 0 0 0.10 1 0.68 3 1.01 6 1.28 0.2 0 0.10 1 0.34 3 0.67 6 0.80 0.5 0 0.11 1 0.39 3 0.67 6 0.95 Petition 870200056639, dated 07 / 05 / 2020, pp. 82 / 113 70 / 83 1.0 0 0.14 1 0.44 3 0.87 6 1.34 Example 3

[00171] Tablets comprising the CYT-0387 dihydrochloride monohydrate form II (doses equivalent to 100, 150, 200 and 300 mg of free base) and capsules comprising the anhydrous form I of CYT-0387 dihydrochloride (dose equivalent to 300 mg of free base) were evaluated in a single-dose phase I study in healthy subjects.

[00172] Intensive PK and PD sampling occurred from 0.5 hours to 36 hours post-dose. Safety was monitored throughout the study. A parametric analysis of variance (ANOVA) using a mixed-effects model was used to adjust for the natural logarithmic transformation of the PK parameters (AUC and Cmax). 90% confidence intervals were constructed for the ratio of geometric means of CYT-0387 dihydrochloride monohydrate monohydrate tablets II at 100, 150, 200, and 300 mg vs. CYT-0387 dihydrochloride anhydrous form I capsules at 300 mg, using equivalence linkages of 70% to 143% for AUC and Cmax. Pharmacokinetic data are presented in Table 5. Table 5: Pharmacokinetic data for the tablet formulations of form II of CYT-0387 dihydrochloride monohydrate and the capsule formulation of form I anhydrous CYT-0387 dihydrochloride. CYT-0387 Dihydrochloride Monohydrate II Tablet Dose Plasma PK Parameters Tablet: Mean (SD) CYT-0387 Dihydrochloride Anhydrous Form I 300 mg Capsule Mean (SD) GMR (%) (90% CI) 100 mg AUCinf (h-ng / mL) 1360 (497.9) 2813 (1984) 55.8 (42.1, 73.9) Cmax (ng / mL) 166.5 (73.3) 388.8 (225.0) 47.3 (35.4, 63.2) 150 mg AUCmf (h-ng / mL) 3018 (1532) 4285 (1923) 69.9 (59.6, 81.9) Cmax (ng / mL) 354.8 (150.6) 549.3 (259.9) 65.6 (55.2, 77.8) 200 mg AUCmf (h-ng / mL) 2572(1671) 2672 (1993) 101.9 (87.7, 118.4) Cmax (ng / mL) 323.7 (188.3) 356.1 (195.6) 92.0 (79.0, 107.1) Petition 870200056639, dated 07 / 05 / 2020, page 83 / 113 / 83 300 mg AUCinf (h-ng / mL) 3194 (1445) 2586 (1481) 136.2 (107.5, 172.4) Cmax (ng / mL) 415.3 (183.2) 381.8 (200.8) 115.7 (87.7, 152.7) Example 4

[00173] CYT-0387 is a selective small molecule inhibitor of Janus kinase 1 and 2 (JAK1 / JAK2) currently under investigation for the treatment of myelofibrosis. This study evaluated the mass balance / recovery, metabolite profile, pharmacokinetics, and safety of radiolabeled CYT-0387 in humans.

[00174] Six healthy individuals (subjects) received a single oral dose of 200 mg of CYT-0387 containing ~100 pCi of [14C]-CYT-0387. Blood samples were collected for up to 21 days or until plasma radioactivity in 2 consecutive samples fell below the detection limit or urine and stool sampling was discontinued. Urine / stool samples were collected for up to 21 days or until >90% of the administered dose was recovered in stool and urine, and radioactivity in 2 consecutive sampling intervals was <1% of the administered dose. Plasma concentrations of CYT-0387 and metabolites were measured using LC-MS / MS, and total radioactivity was assessed by liquid scintillation counting. Metabolite profiling was performed on selected plasma, stool, and urine samples. Safety assessments were conducted throughout the study.

[00175] Results: CYT-0387 was well tolerated. No grade 3 or 4 AEs, SAEs, or AEs leading to study discontinuation were reported. The most frequently reported AEs were dizziness, headache, and nausea. The peak concentration of drug-derived radioactivity in plasma was observed 2.5 hours post-dose. Mean blood-to-plasma concentration ratios ranged from 0.7 to 0.9 over 24 hours post-dose, indicating low association of radioactivity with blood cells. Overall radioactivity recovery was 96.7% (feces: 69.3%; urine: 27.5%). Circulating radioactivity consisted primarily of Petition 870200056639, dated 07 / 05 / 2020, p. 84 / 113 / 83 metabolite M21 (64.2%), CYT-0387 (17.3%) and metabolites (M8: 5.8%; metabolite M19: 5.2%; M5: 2.7%; M28: 2.5%; and M20: 2.3%). The main component excreted in feces was M14 (21.4% of the dose), along with CYT-0387 (12.6% of the dose) and other metabolites (M21: 12.7% of the dose; M19 / M33 co-eluted: 7.1% of the dose). The remaining 10 metabolites identified in feces represented less than 5% of the dose. In urine, the metabolite M21 was the main species (11.5% of the dose), with low levels of minor metabolites observed.

[00176] After oral administration in healthy individuals, [14C]CYT-0387 was primarily eliminated in the feces as a combination of unchanged metabolites and parent drug. Example 5

[00177] CYT-0387 is a selective small molecule inhibitor of Janus kinase 1 and 2 (JAK1 / JAK2) and is currently under investigation for the treatment of myelofibrosis. In a phase 1 / 2 study in patients with myelofibrosis, CYT-0387 at 300 mg capsule once daily was selected as the phase 3 dose based on a favorable benefit:risk profile. An immediate-release tablet formulation (CYT-0387 tablet) was developed for further clinical evaluations. The relative bioavailability of the tablet vs. the capsule of CYT-0387 was evaluated in this study to identify the phase 3 dose of the CYT-0387 tablet.

[00178] The pharmacokinetics (PK) of the CYT-0387 tablet (100 to 300 mg) versus the capsule (300 mg) after a single dose was evaluated in healthy subjects. The PK of the CYT-0387 tablet at supratherapeutic doses (400 and 800 mg), under fasted and fed conditions, and with an acid-reducing agent (i.e., omeprazole) was also evaluated. Intensive PK sampling occurred up to 36 hours post-dose. Safety was monitored throughout the study. A parametric analysis of variance was performed. Petition 870200056639, dated 07 / 05 / 2020, p. 85 / 113 / 83. An ANOVA using a mixed-effects model was used to adjust for the natural logarithmic transformation of the PK parameters (AUC and Cmax). 90% confidence intervals were constructed for the ratio of the geometric means of the PK of the CYT-0387 tablet at 100, 150, 200, and 300 mg vs. the CYT-0387 capsule at 300 mg, using equivalence linkages of 70% to 143% for AUC and Cmax. A similar approach was used to assess the effect of food and omeprazole.

[00179] The 200 mg CYT-0387 tablet provided plasma exposures equivalent to the 300 mg CYT-0387 capsule (Table 6). Plasma exposures to CYT-0387 increased less than the dose-proportional from 100 to 800 mg. Ingestion of light and high-fat meals modestly increased Cmax (38% and 28% increase for light and high-fat meals, respectively) and AUCinf (16% and 28% increase for light and high-fat meals, respectively) for the CYT-0387 tablet. Omeprazole decreased CYT-0387 tablet exposure by 36% for Cmax and 33% for AUCinf. These differences were not considered clinically relevant.

[00180] The CYT-0387 200 mg tablet provides comparable exposure to the CYT-0387 300 mg capsule. Plasma exposures to the CYT-0387 tablet increased less proportionally to the dose. No clinically relevant effects of food or acid-reducing agents were observed on the PK of the CYT-0387 tablet. Table 6: Relative bioavailability of the tablet compared to the capsule of CYT-0387 in phase 3 after administration of a single dose of CYT-0387 PK Parameters 200 mg Tablet: Mean (% CV) 300 mg Capsule: Mean (% CV) GMR [%] (90% CI) Cmax (ng / mL) 323.7 (58.2) 356.1 (54.9) 91.99 (78.98, 107.15) AUCinf (ng / mL) 2549.7 (66.1) 2665.5 (74.9) 101.69 (87.45, 118.26) AUClast (h· ng / mL) 2324.5 (65.3) 2443.9 (71.3) 100.35 (86.73, 116.10) % CV = % coefficient of variation; CI = confidence interval; data rounded as applicable and shown as three significant figures. Petition 870200056639, dated 07 / 05 / 2020, p. 86 / 113 / 83 Example 6 This example described the preparation of M14 (compound 3), M8 (compound 4), M20 (compound 12), M21 (compound 13), compound 8, and compound 10.

[00181] In a flask, 4-(2-chloropyrimidin-4-yl)benzoic acid (3.0 g, 12.8 mmol), 4-morpholoneaniline (2.7 g, 14.0 mmol, 1.1 equiv) and NMP (30 mL) were placed. The resulting solution was stirred at 120°C. After completion, the reaction was cooled and 30 mL of aqueous NaHCO3 solution was added. The resulting fluid paste was filtered, washed with water and dried under vacuum at 45°C to produce 4-(2-((4-(3oxomorpholino)phenyl)amino)pyrimidin-4-yl)benzoic acid (compound 3) with the structure below: 1H NMR (400 MHz, DMSO-d6): δ = 13.21 (s, 1H), 9.85 (s, 1H), 8.62 (d, J = 5.2 Hz, 1H), 8.28 (d, J = 8.2 Hz, 2H), 8.10 (d, J = 8.2 Hz, 2H), 7.85 (d, J = 8.4 Hz, 2H), 7.49 (d, J = 5.2 Hz, 1H), 7.33 (d, J = 9.0 Hz, 2H), 4.19 (s, 2H), 3.98 (m, 2H), 3.71 (m, 2H)

[00182] Compound 3 (1.0 g, 2.43 mmol), TBTU (1.0 g, 3.15 mmol, 1.3 equiv.), glycinamide hydrochloride (0.32 g, 1.2 equiv.), DMSO (9 mL), and i-Pr2NEt (0.65 g, 2.92 mmol, 1.2 equiv.) were loaded into a flask. After the reaction was complete, water (7.7 mL) was added, and the resulting fluid paste was filtered and washed with DMSO / water (2:1) and water. The isolated solids were refluxed in 10 mL of MeOH, filtered, washed with MeOH, and dried in a vacuum oven at 45°C to produce N-(2-amino-2-oxoethyl)-4-(2-((4-(3-oxomorpholino)phenyl)amino)pyrimidin-4-yl)benzamide (compound 4) with the structure below: Petition 870200056639, dated 07 / 05 / 2020, p. 87 / 113 / 83 RMN 1H (400 MHz, DMSO-dô): δ = 9,83 (s, 1H), 8,81 (t, J = 6,0 Hz, 1H), 8,60 (d, J = 2,9 Hz, 1H), 8,27 (d, J = 8,5 Hz, 2H), 8,05 (d, J = 8,4 Hz, 2H), 7,85 (d, J = 6,9 Hz, 2H), 7,51 (d, J = 4,8 Hz, 1H), 7,41 (bs, 1H), 7,32 (d, J = 8,9 Hz, 2H), 7,06 (bs, 1H), 4,19 (s, 2H), 3,98 (m, 2H), 3,85 (d, J = 5,8 Hz, 2H), 3,72 (m, 2H); HRMS (ESI+): calc. para C23H23N6O4 [M+H]+: 447,18 encontrado: 447,19.

[00183] A suspension of {4-[(cyanomethyl)carbamoyl]phenyl}boronic acid (4.2 g, 20.6 mmol), 2,4-dichloropyrimidine (4.3 g, 28.8 mmol), potassium carbonate (2.8 kg, 20.6 mmol) and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) complex with dichloromethane (1:1) (84 mg, 0.10 mmol) in acetonitrile (21 mL) and water (11 mL) was sprayed with N2 for 30 minutes. The mixture was heated to 75°C until the reaction was complete. The mixture was cooled to 60°C and the layers were separated. An aqueous solution of N-acetylcysteine ​​(6 mL) was added, followed by the addition of water (15 mL). The mixture was cooled to 20°C. The solids were filtered, washed with H2O / CH3CN (3:1) and dried at 50°C to produce 4-(2-chloropyrimidin-4-yl)-N-(cyanomethyl)benzamide, with the structure below: 1H RMN (300 MHz, DMSO-d6): δ 4.36 (d, J = 5.5 Hz, 2H), 8.05 (m, J = 8.5 Hz, 2H), 8.24 (d, J = 5.3 Hz, 1H), 8.2, 8.9 (m, J = 8), J 5.2 Hz, 1H), 9.39 (t, J = 5.5 Hz, 1H). HRMS (ESI+): calc. for CbHkjCHINESE [M+1]: 273.15 found 273.25. Petition 870200056639, dated 07 / 05 / 2020, p. 88 / 113 / 83

[00184] In a flask, 4-(2-chloropyrimidin-4-yl)-N(cyanomethyl)benzamide (6.7 g, 24.6 mmol), 2-((4-aminophenyl)amino)ethanol (7.5 g, 49.3 mmol, 2.0 equiv), i-Pr2NEt (4.8 g, 36.9 mmol, 1.5 equiv.) and DMSO (20 mL) were placed. The resulting solution was stirred at 100°C. After the reaction was complete, the solution was cooled to 20°C and then 135 mL of water was added. The resulting fluid paste was filtered and washed with 70 mL of water. The solids were refluidified in i-PrOH (70 mL). The resulting fluid paste was filtered and washed with i-PrOH. The solids were dried under vacuum and dissolved in 30 mL of THF and heated to 50°C. Water (85 mL) was slowly added and the fluid paste was cooled to 20°C. The resulting solids were isolated by filtration, washed with THF / water (1:3) and water, and dried at 40°C to give N-(cyanomethyl)-4-(2-((4-((2-hydroxyethyl)amino)phenyl)amino)pyrimidin-4-yl)benzamide (compound 8) with the structure below: 1H NMR (DMSO-d6): 9.33 (t, J = 5.5 Hz, 1H), 9.24 (s, 1H), 8.48 (d, J = 5.1 Hz, 1H), 8.24 (d, J = 8.4 Hz, 2H), 8.01 (d, J = 8.5 Hz, 2H), 7.46 (d, J = 8.8 Hz, 2H), 7.33 (d, J = 5.2 Hz, 1H), 6.58 (d, J = 8.9 Hz, 2H), 5.20 (t, J = 5.8 Hz, 1H), 4.66 (t, J = 5.4 Hz, 1H), 4.35 (d, J = 5.4 Hz, 2H), 3.57 (q, J = 5.8 Hz, 2H), 3.08 (q, J = 5.8 Hz, 2H); HRMS (ESI+): calc. for C21H21N6O2 [M+1]: 389.17 found: 389.27.

[00185] In a flask, 4-(2-chloropyrimidin-4-yl)-N(cyanomethyl)benzamide (4.0 g, 14.7 mmol), phenylenediamine (3.2 g, 29.3 mmol, 2.0 equiv), i-Pr2NEt (2.9 g, 22.1 mmol, 1.5 equiv) and DMSO (12 mL) were placed. The resulting solution was stirred at 60°C. After the reaction was complete, the solution was cooled to 20°C and then 50 mL of water was added. The resulting fluid paste was filtered and washed with water, followed by i-PrOH. The Petition 870200056639, dated 07 / 05 / 2020, p. 89 / 113 / 83 solids were refluidified in i-PrOH (50 mL), filtered and washed with iPrOH and dried at 40°C to produce 4-(2-((4-aminophenyl)amino)pyrimidin-4yl)-N-(cyanomethyl)benzamide (compound 10) with the structure below: 1H NMR (400 MHz, DMSO-dô): δ = 9.33 (t, J = 5.6 Hz, 1H), 9.21 (s, 1H), 8.47 (d, J = 5.2 Hz, 1H), 8.24 (d, J = 8.5 Hz, 2H), 8.01 (d, J = 8.2 Hz, 2H); HRMS (ESI+): calc. for C19H17N6O [M+H]: 345.15 found: 345.28.

[00186] In a flask, 4-(2-chloropyrimidin-4-yl)-N(cyanomethyl)benzamide (4.0 g, 14.7 mmol), 4'-aminoacetanilide (2.6 g, 17.6 mmol, 1.2 equiv), i-Pr2NEt (2.9 g, 22.1 mmol, 1.5 equiv.) and DMSO (12 mL) were placed. The resulting solution was stirred at 120°C. After the reaction was complete, the mixture was cooled to 20°C and MeOH (30 mL) was slowly added. The resulting fluid paste was filtered and washed with MeOH. The solids were refluxed in 40 mL of MeOH, filtered, washed with MeOH, and dried at 40°C to produce 4-(2-((4-acetamidophenyl)amino)pyrimidin-4-yl)-N-(cyanomethyl)benzamide (compound 12) with the structure below: 1H NMR (DMSO-do): 9.82 (s, 1H), 9.64 (s, 1H), 9.33 (t, J = 5.5 Hz, 1H), 8.57 (d, J = 5.1 Hz, 1H), 8.28 (d, J = 8.1 Hz, 2H), (8.04, J = 8.5 Hz, 2H); HRMS (ESI+): calc. for C21H19N6O2 [M+1]: 387.16 found: 387.28. Petition 870200056639, dated 07 / 05 / 2020, pp. 90 / 113 / 83

[00187] A mixture of 4-(2-chloropyrimidin-4-yl)-N-(cyanomethyl)benzamide (2.0 g, 7.2 mmol), 4-(4-aminophenyl)morpholin-3-one (1.4 g, 7.2 mmol) and zinc dichloride (98 mg, 0.72 mmol) in N-methylpyrrolidinone (10 mL) was sprayed with N2 for 10 minutes and then heated to 90°C until the reaction was considered complete. The mixture was cooled to 50°C and water (15 mL) was then slowly added to the reaction mixture. The resulting fluid paste was cooled to 20°C and the solids were filtered, washed with water and dried. The solids were dissolved in 15 mL of DMSO and heated to 50°C. Methanol (25 mL) was added to the mixture and then cooled to 20°C. The resulting solids were filtered, washed with MeOH, and dried at 60°C under vacuum to produce N-(cyanomethyl)-4-(2-{[4-(3-oxomorpholin-4-yl)phenyl]amino}pyrimidin-4-yl)benzamido-(cyanomethyl)-4-(2-{[4-(3-oxomorpholin-4-yl)phenyl]amino}pyrimidin-4-yl)benzamide (compound 13) with the structure below: 1H NMR (300 MHz, DMSO-dô): δ 9.83 (s, 1H), 9.34 (t, J = 5.5 Hz, 1H), 8.62 (d, J = 5.2 Hz, 1H), 8.30 (m, J = 8.6 Hz, 2H), 8.04 (m, J = 8.6 Hz, 2H), 7.85 (m, J = 8.9 Hz, 2H), 7.51 (d, J = 5.2 Hz, 1H), 7.33 (m, J = 8.9 Hz, 2H), 4.36 (d, J = 5.4 Hz, 2H), 4.20 (s, 2H), 3.98 (dd, J = 5.9, 4.19 Hz, 2H), 3.62 - 3.79 (m, 2H). HRMS (ESI+): calc. for C23H21N6O3[M+1]: 429.17, found: 429.0. Example 7

[00188] This study characterized the effect of CYT-0387 on hepcidin production in HepG2 cells, a hepatocellular carcinoma cell line. Bone morphogenetic proteins (BMPs) were shown to be involved in the transcriptional induction of hepcidin in hepatocytes, facilitating Petition 870200056639, dated 07 / 05 / 2020, p. 91 / 113 / 83 the association of constitutively active BMP receptor type II kinases (BMPR-kinase) with type I BMPR-kinase (Andriopoulos, et al. Nat Genet, 2009. 41(4): p482-7; Zhao, et al., J Clin Invest, 2013. 123(6): p2337-43). This results in the phosphorylation and activation of type I BMPR-kinases and the subsequent downstream activation of SMAD effector proteins (SMAD1 / 5 / 8), followed by nuclear translocation in association with SMAD4 (Wrana, Cold Spring Harb Perspect Biol, 2013. 5:a011197).

[00189] HepG2 cells were pre-incubated for 2 hours with CYT-0387 (ranging from 0 μM to 10 μM) in the presence of 1% FBS, then stimulated for 6 hours with 10 ng / mL of BMP6. Total RNA was isolated from the cells and analyzed for hepcidin levels by qRT-PCR. GUSB (glucuronidase, beta) was used as a maintenance control to normalize the levels measured by qRT-PCR. The percentage of hepcidin induction times change was calculated (100% equals hepcidin induction in vehicle-treated cells) and the results are summarized in Table 9. The results showed that CYT-0387 resulted in a dose-dependent inhibition of BMP6-mediated hepcidin induction.

[00190] HepG2 cells were pre-incubated for 2 hours with increasing concentrations of CYT-0387 (from 0.02 to 10 μM of CYT387) in the presence of 1% FBS, and then stimulated for 30 minutes with 10 ng / mL of BMP6. Protein was extracted from the lysed cells and analyzed using immunoblot analysis with antibodies specific for phospho-SMAD1 (Ser463 / 465), phospho-SMAD5 (Ser463 / 465), and phosphoSMAD8 (Ser465 / 467) and β-actin. Crude phospho-SMAD1 / 5 / 8 levels were quantified using densitometry software (Image Studio) and normalized to β-actin levels. The percentage of phosphoSMAD1 / 5 / 8 levels was calculated (100% equals the phosphoSMAD1 / 5 / 8 levels in vehicle-treated cells stimulated with 10 ng / mL of BMP6) and is summarized in Table 9. The results showed that CYT-0387 resulted in Petition 870200056639, dated 07 / 05 / 2020, page 92 / 113 / 83 a dose-dependent inhibition of phospho-SMAD1 / 5 / 8 levels mediated by BMP6. Table 9: Normalized percentage change in induction times and phospho-SMAD1 / 5 / 8 levels in HepG2 cells stimulated with BMP6 in the presence of CYT-0387. CYT-0387 (μM) induction in change in times of hepcidin at DPb levels of phosphoSMAD1 / 5 / 8 c DP b 0 100 0 100 0 0.020 115 40 83 28 0.039 91 28 80 11 0.078 74 28 91 14 0.156 86 21 96 10 0.313 89 43 88 18 0.625 45 24 91 16 1.25 28 21 65 14 2.5 6 13 52 15 5.0 -4 6 40 9 10.0 -3 4 31 11 a) Average induction for n = 2. b) Standard Deviation (SD) = Average induction for n = 6.

[00191] In addition, biochemical binding assays (DiscoveRx) and in vitro enzyme inhibition assays (LanthaScreen, Life Technologies) were performed to determine the affinity and binding inhibition activities of CYT-0387 to type I BMPR kinases (ALK2, ALK3, and ALK6). Transforming growth factor receptor beta-1 (TGFBR1, ALK5) was used as a control to determine selectivity to type I BMPR kinases. The results are summarized in Table 10 and showed that CYT-0387 had higher affinity and inhibitory activities for ALK2 and ALK6 compared to ALK3. Table 10: Biochemical Kd and IC50 values ​​of CYT-0387 for the BMPR-kinases. CYT-0387 Kd (nM) IC50 (nM) AVGa DPb ALK2 (Acvr1) 25 8 2 ALK3 (BMPR1a) 1000 405 131 ALK6 (BMPR1B) 19 107 8 TGFBR1 670 205 37 aAVG: average value for n = 3. bSD = Standard Deviation Example 8 Petition 870200056639, dated 07 / 05 / 2020, pages 93 / 113 / 83

[00192] Studies are conducted to investigate the effects of the CYT-0387 formulation in the treatment of non-small cell lung cancer (NSCLC) or metastatic pancreatic ductal adenocarcinoma (PDA). In one study, patients with KRAS-modified NSCLC (metastatic Kirsten syndrome viral oncogene homolog) who failed platinum-based chemotherapy received either the CYT-0387 formulation alone, trametinib alone, or the combination of CYT-0387 and trametinib, for at least one treatment cycle over 28 days.The CYT-0387 formulation (which may comprise the CYT-0387 dihydrochloride monohydrate form II in a tablet format) may be administered orally once or twice daily at doses of 100 mg, 150 mg, 200 mg, 250 mg, or 300 mg; and trametinib (which may be referred to by the chemical name N-(3-{3-cyclopropyl-5-[(2-fluoro-4-iodophenyl)amino]-6,8-dimethyl-2,4,7-trioxo-3,4,6,7-tetrahydropyrido[4,3-d]pyrimidin-1(2H)yl}phenyl)acetamide) may be administered orally once daily at doses of 0.5 mg, 1 mg, or 2 mg. The study enrolls patients with metastatic or recurrent non-small cell lung cancer with a modified KRAS score, who have received prior treatment with platinum-based chemotherapy or up to two prior lines of chemotherapy, with measurable disease as RECIST v 1.1 and an Eastern Oncology Cooperative Group (ECOG) performance status of 0 or 1.

[00193] In another study, patients with metastatic NSCLC with modified epidermal growth factor receptor (EGFR) and naive EGFR tyrosine kinase inhibitor (TKI) received erlotinib alone or a combination of CYT-0387 and erlotinib for at least one 28-day treatment cycle. The CYT-0387 formulation (which may comprise the CYT-0387 dihydrochloride monohydrate form II in a tablet format) may be administered orally once or twice daily at doses of 100 mg, 150 mg, 200 mg, 250 mg, or 300 mg; and erlotinib Petition 870200056639, dated 07 / 05 / 2020, p. 94 / 113 / 83 (which may be referred to by the chemical name N-(3-ethinylphenyl)-6,7-bis(2-methoxyethoxy)4-quinazolinamine) may be administered orally once daily at a dose of 25 mg, 100 mg, or 150 mg. This study involves patients with metastatic NSCLC with EGFR exon 19 deletion or exon 21 (L858R) substitution mutation who have not received prior treatment or platinum-based chemotherapy, with an Eastern Oncology Cooperative Group (ECOG) performance status of 0, 1, or 2.

[00194] In an additional study, patients with relapsed or refractory metastatic pancreatic adenocarcinoma receive the combination of capecitabine and CYT-0387 (with the CYT-0387 dihydrochloride monohydrate form II in a tablet format) or the combination of oxaliplatin, capecitabine, and CYT-0387, for at least one 21-day treatment cycle. The CYT-0387 formulation (which may comprise the CYT-0387 dihydrochloride monohydrate form II in a tablet format) may be administered orally once or twice daily at doses of 100 mg, 150 mg, 200 mg, 250 mg, or 300 mg.Capecitabine (which may be referred to by the chemical name [1-(3,4-dihydroxy-5-methyltetrahydrofuran-2-yl)-5-fluoro-2-oxo-1H-pyrimidin-4-yl]carbamate) pentyl can be administered orally twice daily for 14 days, followed by a 7-day break, until the end of treatment; oxaliplatin (which may be referred to by the chemical name [(1R,2R)-cyclohexane-1,2-diamine](ethanedioate-O,O')platinum(II)) can be administered intravenously over 120 minutes on day 1 of each 21-day treatment cycle. Patients have relapsed or refractory metastatic pancreatic adenocarcinoma and have received prior treatment with a gemcitabine-containing regimen, with measurable disease by RECIST v 1.1 and an Eastern Oncology Cooperative Group (ECOG) performance status of 0 or 1.

[00195] Studies monitor several factors, including, but not limited to, safety, toxicity, tolerability, complete response (CR) Petition 870200056639, dated 07 / 05 / 2020, page 95 / 113 / 83 or partial response (PR) or stable disease (SD), as assessed by the Response Evaluation Criteria in Solid Tumor (RECIST) v 1.1, overall survival (i.e., interval from treatment to death, from any cause), progression-free survival (i.e., the interval from the date of the first dose to the first documented death or definitive disease progression based on RECIST v 1.1 criteria), and / or total response range (i.e., the proportion of patients who received a complete response or a partial response), at various treatment points.

[00196] Each of the references, including all patents, patent applications and publications cited in this application, is incorporated herein by reference in its entirety, as if each one were individually incorporated. Furthermore, it would be considered that, in teaching the above invention, a person skilled in the art could make certain alterations or modifications to the invention, and these equivalents would still be within the scope of the invention as defined by the appended claims of the application. Petition 870200056639, dated 07 / 05 / 2020, pp. 96 / 113

Claims

1 / 2 CLAIMS 1. Compound, characterized in that it is selected from the group consisting of: Anhydrous form I of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride; exhibiting an X-ray powder diffraction (XRPD) pattern with peaks at 13.5°, 20.9°, 26.1° and 28.3° 2-θ ± 0.2° 2-θ as determined by copper irradiation (Cu Kα, λ = 1.5418 Å); and anhydrous Form III of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride, which exhibits an X-ray powder diffraction (XRPD) pattern with peaks at 12.7°, 14.6°, 17.8°, 19.7° and 23.3° 2-θ ± 0.2° 2-θ as determined by copper irradiation (Cu Kα, λ = 1.5418 Å).

2. Compound according to claim 1, characterized in that the compound is anhydrous Form I of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride; which exhibits an X-ray powder diffraction (XRPD) pattern with peaks at 13.5°, 20.9°, 26.1° and 28.3° 2± 0.2° 2-θ as determined by copper irradiation (Cu Κα, λ = 1.5418 Å).

3. Compound according to claim 1, characterized in that the compound is anhydrous Form III of N-(cyanomethyl)4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride, which exhibits an X-ray powder diffraction (XRPD) pattern with peaks at 12.7°, 14.6°, 17.8°, 19.7° and 23.3° 2-θ ± 0.2° 2-θ as determined by copper irradiation (Cu Kα, λ = 1.5418 Å).

4. Pharmaceutical composition, characterized by comprising the compound as defined in any one of claims 1 to 3.

5. Pharmaceutical composition according to claim 4, Petition 870200056639, dated 07 / 05 / 2020, page 97 / 113 2 / 2 characterized in that the compound Form I anhydrous of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride is present in an amount equivalent to 50 mg, 100 mg, 150 mg or 200 mg of free base of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4yl)benzamide.

6. Pharmaceutical composition according to claim 4, characterized in that the anhydrous Form III compound of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide monohydrochloride is present in an amount equivalent to 50 mg, 100 mg, 150 mg or 200 mg of free base of N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4yl)benzamide.

7. Pharmaceutical composition according to any one of claims 4 to 6, characterized in that it is in the form of a tablet.

8. Pharmaceutical composition according to any one of claims 4 to 6, characterized in that it is in the form of a capsule.

9. Pharmaceutical composition according to claim 4, characterized in that it is for use in a method for treating a disease associated with Janus kinase (JAK), wherein the disease is a myeloproliferative disease selected from the group consisting of polycythemia vera (PV), myelofibrosis, thrombocythemia, essential thrombocythemia (ET), idiopathic myelofibrosis, chronic myelogenous leukemia, systemic mastocytosis (SM), chronic neutrophilic leukemia (CNL), myelodysplastic syndrome (MDS) and systemic mastocytosis (SMCD).

10. Pharmaceutical composition according to any one of claims 4 to 8, characterized in that it is for use in a method for treating primary myelofibrosis. Petition 870200056639, dated 07 / 05 / 2020, pp. 98 / 113

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