Crystalline forms of anti-SARS COV-2 formulations

Abstract

The present invention features crystalline forms of Compound I # imgabs0 #, including polymorphs and pseudopolymorphs, which can be used for the preparation of pharmaceutical compositions.

Description

[0001] Related Applications

[0002] This application claims the benefit of U.S. Provisional Application No. 63 / 413,850, filed Oct. 6, 2022. The entire teachings of the above application are incorporated herein by reference. Technical Field

[0003] The present invention relates to the crystalline polymorphic forms of novel compound I, methods for preparing the same, pharmaceutical compositions comprising the polymorphic forms, methods for preparing pharmaceutical compositions using the polymorphic forms, and methods for treating or preventing coronavirus infections using the polymorphic forms. Background Art

[0004] Coronaviruses are enveloped positive-sense single-stranded RNA viruses. The coronavirus genomic RNA has a 5′-cap structure and a 3′-polyA tail and contains at least six open reading frames (ORFs). The first ORF (ORF 1a / b) directly translates two polyproteins: pp1a and pp1ab. These polyproteins are processed by a 3C-like protease (3CLpro), which is also known as the main protease (Mpro), into 16 non-structural proteins. These non-structural proteins are involved in the production of subgenomic RNAs, which encode four structural proteins, namely, the envelope protein, the membrane protein, the spike protein, and the nucleocapsid protein, as well as other accessory proteins. Thus, it should be understood that the 3C-like protease plays a crucial role in the coronavirus life cycle.

[0005] 3CLpro is a cysteine protease that participates in most cleavage events of the precursor polyprotein. The active 3CLpro is a homodimer containing two protomers and is characterized by a Cys-His dyad located between domains I and II. 3CLpro is conserved among coronaviruses, and the substrates of 3CLpro from different coronaviruses share some common features. Since there is no human homolog of 3CLpro, it is an ideal antiviral target. Although compounds have been reported to inhibit 3CLpro activity, they have not been approved as a treatment for coronaviruses. (See WO 2004101742 A2, US2005 / 0143320A1, US2006 / 0014821 A1, US2009 / 0137818 A1, WO 2013 / 049382 A2, WO 2013 / 166319 A1, WO2018042343, WO2018023054, WO 2022013684, WO 2021252644, WO2022020711, WO 2022020242, US11,174231B1, US11,124497B1, WO2005113580, and WO2006061714).

[0006] There is a need for novel therapeutic agents for treating, ameliorating, or preventing SARS-CoV-2 infection. SUMMARY OF THE INVENTION

[0007] The present invention provides polymorphic and pseudopolymorphic forms of Compound I (chemical name: N-(S)-1-((3R,5'S)-5'-cyano-2-oxospiro[indoline-3,3'-pyrrolidine]-1'-yl)-4-methyl-1-oxopentan-2-yl)-4,6,7-trifluoro-N-methyl-1H-indole-2-carboxamide), the structure of which is as follows.

[0008]

[0009] In certain embodiments, the present invention provides Compound I in polymorphic or pseudopolymorphic forms as disclosed herein.

[0010] In certain embodiments, the present invention provides methods for preparing polymorphs and pseudopolymorphs of Compound I as disclosed herein.

[0011] In certain embodiments, the present invention provides a composition comprising a polymorph or pseudopolymorph of Compound I disclosed herein. In certain embodiments, the composition is a pharmaceutical composition comprising at least one polymorph or pseudopolymorph of Compound I and a pharmaceutically acceptable carrier or excipient. In certain embodiments, the composition is substantially free of other polymorphs or pseudopolymorphs of Compound I.

[0012] In certain embodiments, the present invention provides a method of providing treatment or prophylaxis of coronavirus infection to a subject in need thereof. The method comprises administering to the subject: (a) a therapeutically effective amount of a polymorph or pseudopolymorph of Compound I, (b) a therapeutically effective amount of two or more polymorphic or pseudopolymorphic forms of Compound I, or (c) a therapeutically effective amount of one or more polymorphs and / or pseudopolymorphs of Compound I and an amorphous form of Compound I. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] As shown in the drawings, the above and other objects, features and advantages of the present invention will become apparent from the following more particular description of the preferred embodiments of the present invention, in which like reference numerals refer to the same parts in the various views. The drawings are not necessarily to scale, but emphasis is placed upon illustrating the principles of the invention.

[0014] Figure 1 is an x-ray powder diffraction (XRPD) pattern of the amorphous form of Compound I.

[0015] Figure 2 is a differential scanning calorimetry (DSC) thermogram of the amorphous form of Compound I.

[0016] Figure 3 is an XRPD pattern of Polymorph 1 of Compound I.

[0017] Figure 4 is a DSC thermogram of Polymorph 1 of Compound I.

[0018] Figure 5 is a Fourier-transform infrared (FT-IR) spectrum of Polymorph 1 of Compound I.

[0019] Figure 6 is an ORTEP diagram of the single crystal x-ray structure of Polymorph 1.

[0020] Figure 7 is an XRPD pattern of Polymorph 2 of Compound I.

[0021] Figure 8It is the DSC thermogram of polymorph 2 of Compound I.

[0022] Figure 9 It is the XRPD pattern of polymorph 3 of Compound I.

[0023] Figure 10 It is the DSC thermogram of polymorph 3 of Compound I.

[0024] Figure 11 It is the FT-IR spectrum of polymorph 3 of Compound I.

[0025] Figure 12 It is the XRPD pattern of polymorph 4 of Compound I.

[0026] Figure 13 It is the XRPD pattern of a mixture of polymorph 2 and polymorph 4 of Compound I.

[0027] Figure 14 It is the DSC thermogram of a mixture of polymorph 2 and polymorph 4 of Compound I.

[0028] Figure 15 It is the XRPD pattern of polymorph A of Compound I.

[0029] Figure 16 It is the DSC thermogram of polymorph A of Compound I.

[0030] Figure 17 It is the XRPD pattern of polymorph B of Compound I.

[0031] Figure 18 It is the DSC thermogram of polymorph B of Compound I.

[0032] Figure 19 It is the XRPD pattern of polymorph C of Compound I.

[0033] Figure 20 It is the XRPD pattern of polymorph D of Compound I.

[0034] Figure 21 It is the DSC thermogram of polymorph D of Compound I.

[0035] Figure 22 It is the FT-IR spectrum of polymorph D of Compound I.

[0036] Figure 23 It is the XRPD pattern of polymorph E of Compound I.

[0037] Figure 24 It is the DSC thermogram of polymorph E of Compound I.

[0038] Figure 25 It is the XRPD pattern of polymorph F of Compound I.

[0039] Figure 26 XRPD pattern of Form G of Compound I.

[0040] Figure 27 DSC thermogram of Form G of Compound I.

[0041] Figure 28 XRPD pattern of Form H of Compound I.

[0042] Figure 29 DSC thermogram of Form H of Compound I.

[0043] Figure 30 XRPD pattern of Form H1 of Compound I.

[0044] Figure 31 FT-IR spectrum of Form H1 of Compound I.

[0045] Figure 32 XRPD pattern of Form J of Compound I.

[0046] Figure 33 DSC thermogram of Form J of Compound I.

[0047] Figure 34 FT-IR spectrum of Form J of Compound I. Detailed implementation mode

[0048] Compound I is an effective 3CL protease protein inhibitor, described in patent application WO 2022 / 109363, the content of which is incorporated herein by reference in its entirety. Compound I is a single enantiomer with three chiral centers. The method for preparing Compound I disclosed in WO2022 / 109363 yields the amorphous form of Compound I disclosed herein. Table 1 lists a summary of the processes for generating four polymorphic crystal forms (Form 1, 2, 3, 4), a mixture of Form 2 and Form 4, and ten pseudopolymorphic crystal forms (Form A, B, C, D, E, F, G, H, H1, J).

[0049] Table 1: Processes for preparing polymorphs and pseudopolymorphs of Compound I.

[0050]

[0051]

[0052] By proton nuclear magnetic resonance ( 1Proven by 1H NMR, all polymorphs and pseudopolymorphs are the same in solution. Solid state techniques such as differential scanning calorimetry (DSC), Fourier transform infrared spectroscopy (FT-IR), and powder X-ray diffraction (XRD) reveal the differences between these crystal forms. The drug substance produced by the current synthesis process is an anhydrous, unsolvated amorphous solid, which is stable under the storage conditions described in this application.

[0053] In one embodiment, the present invention provides four polymorphic crystal forms (Form 1, 2, 3, 4), a mixture of Form 2 and Form 4, and ten pseudopolymorphs (Form A, B, C, D, E, F, G, H, H1, J), which are characterized by differential scanning calorimetry (DSC). Figure 4 , Figure 8 , Figure 10 , Figure 14 , Figure 16 , Figure 18 , Figure 21 , Figure 24 , Figure 27 , Figure 29 , Figure 33 And Tables 2 and 3 describe the DSC thermal properties of polymorphic crystal forms and pseudopolymorphic crystal forms.

[0054] Table 2: Thermal events (exothermic or endothermic peaks) and related temperatures of polymorphic crystal forms 1, 2, 3, and 4 and the mixture of Form 2 and Form 4

[0055]

[0056]

[0057] Table 3: Thermal events (exothermic or endothermic peaks) and related temperatures of polymorphs of Form A, B, D, E, F, G, H, and J

[0058]

[0059] In one embodiment, the present invention provides four polymorphs (Form 1, 2, 3, 4), a mixture of Form 2 and Form 4, and ten pseudopolymorphs (Form A, B, C, D, E, F, G, H, H1, J), which are further characterized by powder X-ray diffraction (XRPD). The characteristic powder diffraction peaks are expressed in degrees 2θ. The positions (2θ) of the peaks of all crystal forms are different from each other. Tables 4 and 5 give the positions of the strong peaks of polymorphic crystal forms and pseudopolymorphic crystal forms. Although the crystal morphologies are the same, Figure 3 , Figure 7 , Figure 9 , Figure 12 , Figure 13 , Figure 15 , Figure 17 ,Figure 19 , Figure 20 , Figure 23 , Figure 25 , Figure 26 , Figure 28 , Figure 30 and Figure 32 the relative intensities and positions of the strong peaks in may change or shift under certain conditions. Whether a given polymorphic crystal form is the same as one of Figure 3 , Figure 7 , Figure 9 , Figure 12 , Figure 13 , Figure 15 , Figure 17 , Figure 19 , Figure 20 , Figure 23 , Figure 25 , Figure 26 , Figure 28 , Figure 30 , Figure 32 can be easily determined by those of ordinary skill in the art by comparing the peak positions and intensities in their XRPD data.

[0060] It was found that the powder X-ray diffraction patterns of crystalline polymorphic and pseudopolymorphic crystal forms and mixtures are different from each other.

[0061] Table 4: Strong peak positions (2θ) of crystal forms 1, 2, 3, 4 and the mixture of crystal forms 2 and 4.

[0062]

[0063]

[0064]

[0065]

[0066]

[0067]

[0068]

[0069]

[0070]

[0071] Table 5: Strong peak positions (2θ) of the polymorphs of crystal forms A, B, C, D, E, F, G, H, H1, J

[0072]

[0073]

[0074]

[0075]

[0076]

[0077]

[0078]

[0079]

[0080]

[0081]

[0082]

[0083]

[0084]

[0085]

[0086]

[0087]

[0088] In one embodiment, the present invention provides Polymorph 1, Polymorph 3, Polymorph D, Polymorph H1 and Polymorph J, which are further characterized by Fourier Transform Infrared Spectroscopy (FT-IR). The characteristic peaks are expressed in wave numbers (cm -1 ). The position of the characteristic peaks of each polymorph is unique to that polymorph. Table 6 gives the positions of the strong peaks of the polymorphs and pseudopolymorphs. Although the crystal forms are the same, Figure 5 , Figure 11 , Figure 22 , Figure 31 or Figure 34 the relative intensities as well as the positions of the strong peaks in Figure 5 , Figure 11 , Figure 22 , Figure 31 , Figure 34 may change or shift under certain conditions. Whether a given polymorphic form is the same as one of

[0089] The FT-IR spectra of Polymorph 1, Polymorph 3, Polymorph D, Polymorph H1 and Polymorph J are different from each other.

[0090] Table 6: Strong peaks in the FT-IR spectra of Polymorph 1, Polymorph 3, Polymorph D, Polymorph H1 and Polymorph J (unit cm -1 )

[0091]

[0092]

[0093]

[0094]

[0095]

[0096]

[0097]

[0098]

[0099] Polymorph 1 of Compound I provides improved or adjusted physicochemical properties of the compound, including but not limited to solid-state properties (such as crystallinity, hygroscopicity, melting point), pharmaceutical properties (such as stability or compatibility), and crystallization characteristics (such as purity, yield or morphology).

[0100] Polymorph 1 was further characterized by single crystal X-ray diffraction. The crystal parameters obtained therefrom are shown in Table 7.

[0101] Table 7: Crystal parameter data of Polymorph 1

[0102]

[0103]

[0104] Polymorph 1 was further characterized by single crystal X-ray diffraction analysis.

[0105] The results of single crystal X-ray diffraction analysis describe the atomic positions and other structural parameters that determine the structure of Polymorph 1, as shown in Tables 8 and 9, and the bond distances and bond angles of Polymorph 1 are shown in Table 10. The distances are in angstroms and the bond angles are in degrees. The estimated standard deviations of the bond distances and bond angles of the least significant digits are given in parentheses. The molecular structure of the single crystal X-ray diffraction analysis of Polymorph 1 is as Figure 6 shown as an Oak Ridge Thermal Ellipsoid plot (ORTEP).

[0106] Table 8: Fractional Atomic Coordinates and Isotropic or Equivalent Isotropic Displacement Parameters of Polymorph 1

[0107]

[0108]

[0109]

[0110]

[0111] Table 9: Atomic Displacement Parameters of Polymorph 1

[0112]

[0113]

[0114]

[0115] Table 10: Geometric Parameters of Polymorph 1

[0116]

[0117]

[0118]

[0119]

[0120]

[0121] The DSC thermogram of Polymorph 1 is as Figure 4 shown. The DSC thermogram of Polymorph 1 has a characteristic endothermic peak at 251 °C, with an onset temperature of 250 °C.

[0122] The characteristic peaks (2θ) of the X-ray powder diffraction pattern of Polymorph 1 are as Figure 3As shown. The characteristic peaks are located at the following 2θ (two theta) values: 6.88, 8.32, 10.82, 11.90, 13.15, 13.74, 14.54, 15.41, 16.09, 18.19, 19.19, 20.04, 20.56, 21.69, 22.29, 23.87, 24.83, 25.19, 25.31, 26.13, 26.66, 27.63, 27.92, 28.90, 29.29, 30.14, 31.32, 31.32, 31.74, 32.48, 32.86, 34.66, 34.95, 35.75, 36.11, 36.79, 37.12, 37.79, 38.76 and 39.74. The characteristic peaks may alternatively be expressed as (2θ) 6.9, 8.3, 10.8, 11.9, 13.2, 13.7, 14.5, 15.4, 16.1, 18.2, 19.2, 20.0, 20.6, 21.7, 22.3, 23.9, 24.8, 25.3, 26.1, 26.7, 27.6, 27.9, 28.9, 29.3, 30.1, 31.3, 31.3, 31.7, 32.5, 32.9, 34.7, 35.0, 35.8, 36.1, 36.8, 37.1, 37.8, 38.8 and 39.7.

[0123] The FT-IR spectrum of Polymorph 1 is as Figure 5 shown. Polymorph 1 has the following characteristic peaks (in wave numbers (cm -1 -1)): 549, 572, 592, 602, 649, 676, 706, 726, 751, 762, 803, 884, 897, 934, 967, 1016, 1050, 1087, 1111, 1128, 1161, 1185, 1201, 1227, 1243, 1320, 1392, 1447, 1470, 1485, 1528, 1546, 1599, 1673, 1694, 1712, 2868, 2960 and 3199.

[0124] The method for preparing the polymorphs and pseudopolymorphs of the present invention is described below. In each volume / weight ratio described herein, the volume is in milliliters (mL) and the weight is in grams (g).

[0125] The present invention provides a method for preparing Polymorph 1 of Compound I by crystallization. In one embodiment, the method comprises the following steps:

[0126] 1. In the presence of one or more suitable dehydrating agents, a suitable base, and a suitable solvent, a solution of Compound I is prepared by dehydrating (3R,5'S)-1'-(N-methyl-N-(4,6,7-trifluoro-1H-indole-2-carbonyl)-L-leucyl)-2-oxospiro[indoline-3,3'-pyrrolidine]-5'-carboxamide (Compound N).

[0127] 2. The solution of Compound I in Step 1 is concentrated at a temperature of about 20 °C to about 80 °C, preferably about 45 °C to about 55 °C, more preferably about 50 °C, to a volume (milliliters) / weight (grams) ratio of solvent / Compound N of about 2:1 to about 6:1, preferably about 3.5:1 to about 4.5:1, more preferably about 4:1.

[0128] 3. Optionally, about 0.0001X (X = the weight of Compound N in Step 1) to about 0.1X of polymorph 1 of Compound I, more preferably 0.001X by weight of polymorph 1 of Compound I, is added as a seed crystal to the concentrated solution in Step 2 to induce crystallization of polymorph 1 of Compound I.

[0129] 4. At a temperature of about 20 °C to about 80 °C, preferably about 45 °C to about 55 °C, more preferably about 50 °C, the solvent is exchanged for a suitable solvent to a volume (milliliters) / weight (grams) ratio of solvent / Compound N of about 6:1 to about 10:1, preferably about 7.5:1 to about 8.5:1, more preferably about 8:1. The suitable solvent includes but is not limited to anisole, toluene, xylene, ethyl acetate, isopropyl acetate, and mixtures of two or more thereof. Preferably, the suitable solvent is a mixture of toluene and ethyl acetate.

[0130] 5. Cool to a temperature of about 0 °C to about 50 °C, preferably about 20 °C to about 30 °C, more preferably about 25 °C; and

[0131] 6. Filter the resulting suspension to obtain polymorph 1 of Compound I.

[0132] In one embodiment, Step 1 is carried out at a suitable temperature, such as, for example, from about -10 °C to about 10 °C, preferably from about -5 °C to about 5 °C, more preferably about 0 °C. In one embodiment, the time for preparing the solution of Compound I is about 30 minutes to about 2 hours, preferably about 1 hour.

[0133] In one embodiment of Step 1, suitable solvents include, but are not limited to, acetonitrile, acetone, dichloromethane, dichloroethane, N,N-dimethylformamide, dimethyl sulfoxide, N-methyl-2-pyrrolidone, dioxane, ethyl acetate, heptane, hexane, methyl tert-butyl ether, tetrahydrofuran, toluene, and mixtures of two or more thereof. The preferred solvent is ethyl acetate. Suitable dehydrating agents include, but are not limited to: propylphosphonic anhydride (T3P), trifluoroacetic anhydride (TFAA), N-(triethylaminosulfonyl)carbamate methyl ester (Burgess reagent), phosphorus oxide (P 2 O 5 ). The preferred dehydrating agent is trifluoroacetic anhydride (TFAA). Suitable bases include, but are not limited to: triethylamine, diisopropylethylamine, N-methylmorpholine, 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), imidazole, pyridine, 2,6-lutidine, ethyl nicotinate, N-methylpiperazine, or 1-methylimidazole. The preferred base is triethylamine.

[0134] In certain embodiments, the present invention provides a method for preparing crystalline form 1 of Compound I by precipitation, wherein the method comprises the following steps:

[0135] i. Adding Compound I to a first solvent (such as, but not limited to, ethanol, isopropanol, ethyl acetate, isopropyl acetate, or methyl tert-butyl ether) to form a solution; the Compound I used to form the solution can be a solid crystalline form of the compound or a mixture of two or more solid crystalline forms;

[0136] ii. Removing a portion of the first solvent from the solution, for example, by distillation, and then adding more of the first solvent;

[0137] iii. Repeating the removal of a portion of the first solvent from the solution and adding more of the first solvent until a solution of Compound I is produced, wherein the residual solvent is controlled to be not greater than a predetermined value, such as not greater than 0.1% (wt / wt);

[0138] iv. Adding a second solvent (such as, but not limited to, water or n-heptane) to the solution to precipitate crystalline form 1 of Compound I; and

[0139] v. Separating crystalline form 1 of Compound I by filtration.

[0140] The DSC thermogram of crystalline form 2 is as Figure 8 shown. The DSC thermogram has a characteristic exothermic peak at a temperature of 175 °C, with an onset temperature of 175 °C; and an endothermic peak at a temperature of 251 °C, with an onset temperature of 250 °C.

[0141] In one embodiment, the characteristic peaks (2θ) in the X-ray powder diffraction pattern of crystalline form 2 are as Figure 7As shown. The characteristic peaks are located at the following 2θ (two theta) values: 6.88, 10.82, 11.90, 13.74, 14.54, 16.08, 18.10, 20.04, 20.56, 21.68, 22.29, 23.87, 24.80, 25.18, 26.12, 26.67, 27.93, 28.77, 29.23, 30.13, 31.25, 32.84, 34.28, 34.66, 35.71, 36.07, 37.10 and 39.66. The characteristic peaks may alternatively be expressed as (2θ) 6.9, 10.8, 11.9, 13.7, 14.5, 16.1, 18.1, 20.0, 20.6, 21.7, 22.3, 23.9, 24.8, 25.2, 26.1, 26.7, 27.9, 28.8, 29.2, 30.1, 31.3, 32.8, 34.3, 34.7, 35.77, 36.1, 37.1 and 39.9.

[0142] In one embodiment, the present invention provides a method for preparing polymorph 2 of Compound I. The method comprises the following steps: heating polymorph J of Compound I to a temperature of about 140 °C to about 170 °C, preferably about 150 °C to about 160 °C, more preferably about 155 °C, to provide polymorph 2 of Compound I.

[0143] The DSC thermogram of polymorph 3 is as Figure 10 shown. The DSC thermogram of polymorph 3 has a characteristic endothermic peak at 150 °C with an onset temperature of 142 °C. The DSC thermogram of polymorph 3 also has a characteristic exothermic peak at 176 °C with an onset temperature of 171 °C, and then a characteristic endothermic peak at 252 °C with an onset temperature of 251 °C.

[0144] The characteristic peaks (2θ) in the X-ray powder diffraction pattern of polymorph 3 are as Figure 9As shown. The characteristic peaks are located at the following 2θ (two theta) values: 7.10, 8.71, 8.92, 11.03, 11.70, 13.07, 14.06, 15.23, 15.55, 16.17, 17.14, 17.45, 17.96, 19.16, 19.56, 20.43, 20.68, 21.89, 22.56, 23.45, 23.83, 24.74, 25.79, 26.26, 26.95, 28.12, 28.64, 29.68, 30.44, 31.28, 31.76, 32.34, 32.79, 33.30, 34.67, 35.56, 36.11, 36.47, 37.07, 37.85, and 38.22. The characteristic peaks may alternatively be expressed as (2θ) 7.1, 8.7, 8.9, 11.0, 11.7, 13.1, 14.1, 15.2, 15.6, 16.2, 17.1, 17.5, 18.0, 19.2, 19.6, 20.4, 20.7, 21.9, 22.6, 23.5, 23.8, 24.7, 25.8, 26.3, 27.0, 28.1, 28.6, 29.7, 30.4, 31.3, 31.8, 32.3, 36.5, 37.1, 37.9, and 38.2.

[0145] The characteristic peaks of the Fourier transform infrared (FT-IR) spectrum of polymorph 3 are as Figure 11 shown. The characteristic peaks in terms of wave number (cm -1 ) are: 563, 589, 623, 646, 674, 706, 725, 740, 754, 802, 815, 966, 1023, 1049, 1079, 1096, 1119, 1130, 1158, 1186, 1215, 1254, 1283, 1310, 1352, 1376, 1385, 1394, 1439, 1469, 1484, 1519, 1549, 1598, 1656, 1727, 2878, 2957, 3422, and 3464.

[0146] In one embodiment, the present invention provides a method for preparing polymorph 3 of Compound I. The method comprises the following steps: heating polymorph J of Compound I to a temperature of about 70 °C to about 100 °C, preferably about 75 °C to about 85 °C, more preferably about 80 °C, to provide polymorph 3 of Compound I.

[0147] The characteristic peaks (2θ) of the X-ray powder diffraction pattern of polymorph 4 are as Figure 12As shown. Polymorph 4 has characteristic peaks in the X-ray powder diffraction (XRPD) pattern at the following 2θ (two theta) values: 4.39, 5.14, 7.06, 7.50, 8.47, 8.92, 9.75, 10.08, 10.33, 11.03, 11.71, 12.45, 13.09, 13.83, 14.06, 14.62, 14.84, 15.38, 15.78, 15.95, 16.91, 17.45, 17.93, 18.54, 19.23, 19.66, 20.07, 20.96, 21.84, 22.50, 23.42, 24.19, 24.91, 25.13, 26.18, 26.54, 27.81, 28.54, 29.41 and 30.99. The characteristic peaks may alternatively be expressed as (2θ) 4.4, 5.1, 7.1, 7.5, 8.5, 8.9, 9.8, 10.1, 10.3, 11.0, 11.7, 12.5, 13.1, 13.8, 14.1, 14.6, 14.8, 15.4, 15.8, 16.0, 16.9, 17.5, 17.9, 18.5, 19.2, 19.7, 20.1, 21.0, 21.8, 22.5, 23.4, 24.2, 24.9, 25.1, 26.2, 26.5, 27.8, 28.5, 29.4 and 31.0.

[0148] In one embodiment, the present invention provides a method for preparing polymorph 4 of Compound I. The method comprises the following steps: heating polymorph J of Compound I to a temperature of about 40 °C to about 70 °C, preferably about 55 °C to about 65 °C, more preferably about 60 °C, to provide polymorph 4 of Compound I.

[0149] The DSC thermogram of a mixture of polymorph 2 and polymorph 4 is as Figure 14 shown. The DSC thermogram has a characteristic melting endothermic peak at 175 °C, with an onset temperature of 166 °C, and a characteristic endothermic peak at 139 °C, with an onset temperature of 132 °C. The DSC thermogram also has a characteristic exothermic peak at 151 °C, with an onset temperature of 147 °C, and then an endothermic peak at 251 °C, with an onset temperature of 250 °C.

[0150] The characteristic peaks (2θ) in the X-ray powder diffraction pattern of a mixture of polymorph 2 and polymorph 4 are as Figure 13As shown. The characteristic peaks (2θ) are 4.36, 6.87, 7.06, 8.46, 8.71, 8.92, 9.77, 10.80, 11.04, 11.72, 11.89, 12.43, 12.58, 13.73, 14.06, 14.53, 15.81, 16.07, 18.08, 18.55, 20.03, 20.56, 21.06, 21.68, 21.87, 22.30, 22.58, 23.44, 23.86, 24.21, 24.79, 25.18, 26.11, 26.66, 27.90, 30.11, 31.25, 31.72, 32.88 and 35.72 respectively. The characteristic peaks may alternatively be expressed as (2θ) 4.4, 6.9, 7.1, 8.5, 8.7, 8.9, 9.8, 10.8, 11.0, 11.7, 11.9, 12.4, 12.6, 13.7, 14.1, 14.5, 15.8, 16.1, 18.1, 18.6, 20.0, 20.6, 21.1, 21.7, 21.9, 22.3, 22.6, 23.4, 23.9, 24.2, 24.2, 26.1, 26.7, 27.9, 30.1, 31.3, 31.7, 32.9 and 35.7.

[0151] In one embodiment, the present invention provides a method for preparing a mixture of crystalline form 2 and crystalline form 4 of compound I, the method comprising the steps of: heating crystalline form J of compound I to a temperature of about 100 °C to about 140 °C, preferably about 120 °C to about 130 °C, more preferably about 125 °C, to provide a mixture of crystalline form 2 and crystalline form 4 of compound I.

[0152] The DSC thermogram of crystalline form A of compound I is as Figure 16 shown. The DSC of crystalline form A has a characteristic endothermic peak at 109 °C, with an onset temperature of 107 °C.

[0153] The characteristic peaks (2θ) of the X-ray powder diffraction pattern of crystalline form A are as Figure 15 shown. The characteristic peaks are located at the following 2θ (two theta) values: 6.96, 8.05, 13.00, 13.97 and 21.03. The characteristic peaks may alternatively be expressed as (2θ) 7.0, 8.1, 13.0, 14.0 and 21.0.

[0154] In one embodiment, the present invention provides a method for preparing crystalline form A of compound I, the method comprising the steps of:

[0155] i. preparing a suspension of the amorphous form of compound I in a mixture of benzyl alcohol and toluene (the volume ratio thereof is about 1:1 to about 4:1);

[0156] ii. Heat the suspension between about 5 °C and 50 °C and then cool it for multiple cycles for about 1 day to 20 days, preferably about 12 days, to obtain a suspension of crystalline form A of Compound I; and

[0157] iii. Filter the suspension of crystalline form A of Compound I from step ii at a temperature of about 10 °C to 30 °C, preferably about 15 °C to 25 °C, more preferably about 22 °C, to provide crystalline form A of Compound I.

[0158] The DSC thermogram of crystalline form B of Compound I is as Figure 18 shown. The DSC thermogram of crystalline form B has a characteristic endothermic peak at 108 °C, with an onset temperature of 93 °C.

[0159] The characteristic peaks (2θ) of the X-ray powder diffraction pattern of crystalline form B are as Figure 17 shown. The characteristic peaks are at the following 2θ (two theta) values: 7.17, 14.33, and 21.55. The characteristic peaks may alternatively be designated as (2θ) 7.2, 14.3, and 21.6.

[0160] In one embodiment, the present invention provides a method for preparing crystalline form B of Compound I, the method comprising the following steps:

[0161] i. Prepare a suspension of the amorphous form of Compound I in anisole;

[0162] ii. Heat and cool the suspension between about 5 °C and 50 °C repeatedly for about 1 day to 10 days, preferably about 5 days, to obtain a suspension of crystalline form B of Compound I; and

[0163] iii. Filter the suspension at a temperature of about 10 °C to 30 °C, preferably about 15 °C to about 25 °C, more preferably about 22 °C, to provide crystalline form B of Compound I.

[0164] The characteristic peaks (2θ) in the X-ray powder diffraction pattern of crystalline form C are as Figure 19As shown. The characteristic peaks are located at the following 2θ (two theta) values: 4.99, 6.01, 6.72, 7.83, 8.70, 9.33, 10.68, 10.98, 11.66, 11.90, 12.96, 13.43, 13.71, 14.01, 14.92, 15.19, 15.53, 16.28, 17.73, 18.22, 18.96, 20.44, 21.11, 21.99, 23.09, 24.06, 25.33, 25.86, 27.91 and 28.52. The characteristic peaks may alternatively be expressed as (2θ) 5.0, 6.0, 6.7, 7.8, 8.7, 9.3, 10.7, 11.0, 11.7, 11.9, 13.0, 13.4, 13.7, 14.0, 14.9, 15.2, 15.5, 16.3, 17.7, 18.2, 19.0, 20.4, 21.1, 22.0, 23.1, 24.1, 25.3, 25.9, 27.9 and 28.5.

[0165] In one embodiment, the present invention provides a method for preparing crystalline form C of compound I, the method comprising the following steps:

[0166] i. Prepare a clear solution of the amorphous form of compound I in anisole (about 6X by volume);

[0167] ii. Remove anisole by slow evaporation over about 1 day to 10 days, preferably about 6 days, at about 10 °C to 30 °C, preferably about 15 °C to about 25 °C, more preferably about 22 °C, to obtain a suspension, and

[0168] iii. Filter the suspension to provide crystalline form C of compound I.

[0169] The DSC thermogram of crystalline form D of compound I is as Figure 21 shown. The DSC thermogram of crystalline form D has a characteristic endothermic peak at 118 °C, with an onset temperature of 117 °C.

[0170] In one embodiment, the characteristic peaks (2θ) in the X-ray powder diffraction pattern of crystalline form D are as Figure 20As shown. The characteristic peaks are located at the following 2θ (two theta) values: 7.10, 7.99, 8.27, 8.98, 9.89, 11.88, 13, 14, 13.27, 14.16, 16.21, 17.55, 18.45, 18.77, 20.97, 21.32, 21.79, 23.11, 24.72, 26.71 and 27.72. The characteristic peaks may alternatively be expressed as (2θ) 7.1, 8.0, 8.3, 9.0, 9.9, 11.9, 13, 1, 13.3, 14.2, 16.2, 17.6, 18.5, 18.8, 21.0, 21.3, 21.8, 23.1, 24.7, 26.7 and 27.7.

[0171] In one embodiment, the Fourier transform infrared (FT-IR) spectrum of polymorph D is as Figure 22 shown. The FT-IR spectrum of polymorph D has characteristic peaks (in cm -1 -1) at: 591, 626, 671, 696, 707, 731, 752, 810, 968, 1050, 1079, 1094, 1114, 1130, 1161, 1202, 1329, 1376, 1387, 1431, 1469, 1485, 1524, 1546, 1595, 1649, 1727, 2958 and 3231.

[0172] In one embodiment, the present invention includes providing a method for preparing polymorph D of compound I. The method comprises the following steps:

[0173] i. Preparing a suspension of polymorph A of compound I in a mixed solvent comprising acetone and water;

[0174] ii. Stirring the suspension at a temperature of about 10 °C to 40 °C, preferably about 20 °C to about 30 °C, more preferably about 25 °C;

[0175] iii. Filtering the suspension to provide polymorph D of compound I.

[0176] The DSC thermogram of polymorph E is as Figure 24 shown. The DSC thermogram of polymorph E has a characteristic endothermic peak at 142 °C, with an onset temperature of 138 °C.

[0177] The characteristic peaks (2θ) of the X-ray powder diffraction pattern of polymorph E are as Figure 23As shown. The characteristic peaks are at the following 2θ (two theta) values: 5.85, 8.52, 9.26, 11.08, 12.69, 12.95, 13.22, 14.71, 15.01, 16.01, 16.83, 17.04, 17.56, 18.54, 18.83, 19.18, 19.73, 20.19, 20.83, 21.09, 21.38, 21.99, 22.67, 23.19, 23.77, 24.67, 25.10, 25.84, 26.68, 27.40, 27.69, 28.41, 28.97, 29.60, 30.51, 30.83, 31.99, 32.56, and 33.52. The characteristic peaks may alternatively be expressed as being at the following 2θ (two theta) values: 5.9, 8.5, 9.3, 11.1, 12.7, 13.0, 13.2, 14.7, 15.0, 16.0, 16.8, 17.0, 17.6, 18.5, 18.8, 19.2, 19.7, 20.2, 20.8, 21.1, 21.4, 22.0, 22.7, 23.2, 23.8, 24.7, 25.1, 25.8, 26.7, 27.4, 27.7, 28.4, 29.0, 29.6, 30.5, 30.8, 32.0, 32.6, and 33.5.

[0178] In one embodiment, the present invention provides a method for preparing crystalline form E of Compound I. The method comprises the following steps:

[0179] i. Prepare a suspension of crystalline form D of Compound I in a solvent mixture containing dimethyl sulfoxide (the ratio of the volume of dimethyl sulfoxide to the weight of Compound I is about 1 / 1) and water (the ratio of the volume of water to the weight of Compound I is about 1 / 1);

[0180] ii. Preferably add from about 0.0001X (X = the weight of Compound I in Step 1) to about 0.1X of crystalline form E of Compound I, more preferably 0.001X by weight of crystalline form E of Compound I as a seed crystal;

[0181] iii. Heat and cool the suspension repeatedly preferably between about 5°C and 50°C for preferably about 15 to 25 times, more preferably about 20 times; and

[0182] iv. Filter the resulting suspension to provide crystalline form E of Compound I.

[0183] The characteristic peaks (2θ) of the X-ray powder diffraction pattern of crystalline form F are as Figure 25 shown. The characteristic peaks are at the following 2θ (two theta) values: 6.87, 13.07, and 20.63. The characteristic peaks may alternatively be expressed as (2θ) 6.9, 13.1, and 20.6.

[0184] In one embodiment, the present invention provides a method for preparing polymorph F of Compound I. The method comprises the following steps:

[0185] i. Preparing a suspension of polymorph D of Compound I in a solvent mixture containing polyethylene glycol (the ratio of the volume of polyethylene glycol to the weight of Compound I is about 1 / 1) and water (the ratio of the volume of water to the weight of Compound I is about 1 / 1);

[0186] ii. Repeatedly heating and cooling the suspension preferably between about 5°C and 50°C for preferably about 5 to 15 times, more preferably about 10 times; and

[0187] iii. Filtering the suspension to provide polymorph F of Compound I.

[0188] The DSC thermogram of polymorph G of Compound I is as Figure 27 shown. The DSC thermogram has a characteristic endothermic peak at 130°C with an onset temperature of 109°C.

[0189] The characteristic peaks (2θ) in the X-ray powder diffraction pattern of polymorph G are as Figure 26 shown. The characteristic peaks are located at the following 2θ (two theta) values: 4, 18, 6.86, 8.35, 9.03, 9.44, 10.34, 11.08, 11.39, 12.53, 12.86, 13.28, 13.52, 14.31, 14.75, 15.14, 15.73, 16.22, 16.72, 16.96, 17.48, 17.62, 17.97, 18.75, 19.08, 19.68, 20.16, 20.60, 20.90, 21.13, 22.02, 22.39, 23.01, 23.38, 23.57, 23.98, 24.31, 24.64, 25.07, 25.38, 25.84, 26.63, 27.21, 28.29, 28.72, 29.01, 29.91 and 31.30. The characteristic peaks may alternatively be expressed as (2θ) 4, 2, 6.9, 8.4, 9.0, 9.4, 10.3, 11.1, 11.4, 12.5, 12.9, 13.3, 13.6, 14.3, 14.8, 15.1, 15.7, 16.2, 16.7, 17.0, 17.5, 17.6, 18.0, 18.8, 19.1, 19.7, 20.2, 20.6, 20.9, 21.1, 22.0, 22.4, 23.0, 23.4, 23.6, 24.0, 24.3, 24.6, 25.1, 25.4, 25.8, 26.6, 27.2, 28.3, 28.7, 29.0, 29.9 and 31.3.

[0190] In one embodiment, the present invention provides a method for preparing polymorph G of Compound 1. The method comprises the following steps:

[0191] i. Prepare a suspension of polymorph D of Compound I in a solvent mixture containing methyl ethyl ketone (the ratio of the volume of methyl ethyl ketone to the weight of Compound I is about 1 / 1) and heptane (the ratio of the volume of heptane to the weight of Compound I is about 1 / 1);

[0192] ii. Preferably add about 0.0001X (X = the weight of Compound I in Step 1) to about 0.1X of polymorph G of Compound I, and more preferably 0.001X by weight of polymorph G of Compound I as a seed crystal;

[0193] iii. Repeatedly heat and cool the suspension at a temperature preferably between 5°C and 50°C; the heating and cooling cycles are preferably carried out about 5 to 15 times, more preferably about 10 times; and

[0194] iv. Filter the suspension to provide polymorph G of Compound I.

[0195] The DSC thermogram of polymorph H of Compound I is as Figure 29 shown. The DSC thermogram of polymorph H has a characteristic endothermic peak at 74°C, with an onset temperature of 60°C, and then a characteristic endothermic peak at 157°C, with an onset temperature of 151°C.

[0196] The characteristic peaks (2θ) of the X-ray powder diffraction pattern of polymorph H are as Figure 28As shown, the characteristic peaks are located at the following 2θ (two theta) values: 3.83, 7.04, 7.61, 7.79, 8.94, 9.37, 11.46, 11.68, 12.32, 12.81, 13.21, 13.74, 14.06, 14.56, 14.80, 15.14, 15.47, 15.93, 16.15, 16.63, 17.17, 17.44, 18.01, 18.64, 19.03, 19.13, 19.59, 20.46, 20.82, 21.21, 22.20, 23.02, 23.22, 3.56, 23.80, 24.14, 25.85, 26.76, 26.96 and 27.56. The characteristic peaks may alternatively be expressed as (2θ) 3.8, 7.0, 7.6, 7.8, 8.9, 9.4, 11.5, 11.7, 12.3, 12.8, 13.2, 13.7, 14.1, 14.6, 14.8, 15.1, 15.5, 15.2, 16.6, 17.2, 17.4, 18.0, 18.6, 19.0, 19.1, 19.6, 20.5, 20.8, 21.2, 22.2, 23.0, 23.2, 23.6, 23.8, 24.1, 25.9, 26.8, 27.0 and 27.6.

[0197] In one embodiment, the present invention provides a method for preparing crystalline form H of Compound I. The method comprises the following steps:

[0198] i. Preparing a suspension of crystalline form D of Compound I in a solvent mixture containing isopropanol (the ratio of the volume of isopropanol to the weight of Compound I is about 1 / 1) and water (the ratio of the volume of water to the weight of Compound I is about 1 / 1);

[0199] ii. Preferably adding about 0.0001X (X = the weight of Compound I in step 1) to about 0.1X of crystalline form H of Compound I, more preferably 0.001X by weight of crystalline form H of Compound I as a seed crystal;

[0200] iii. Stirring the suspension at a temperature between about 10°C and 40°C, preferably between about 20°C and 30°C, more preferably about 25°C for a period of preferably about 5 days to about 15 days, more preferably about 10 days; and

[0201] iv. Filtering the suspension to provide crystalline form H of Compound I.

[0202] In one embodiment, the characteristic peaks (2θ) in the X-ray powder diffraction pattern of crystalline form H1 are as Figure 30As shown. The characteristic peaks are located at the following 2θ (two theta) values: 3.91, 6.45, 7.08, 7.58, 7.90, 8.14, 9.28, 9.76, 10.32, 10.68, 11.63, 12.51, 12.95, 13.27, 14.15, 14.78, 15.12, 15.86, 16.57, 17.56, 17.98, 18.43, 18.67, 19.41, 20.25, 20.79, 21.25, 21.42, 21.74, 22.93, 23.36, 23.86, 24.09, 24.69, 25.40, 26.24, 26.94, 27.24, 28.16, 28.67, 29.67, 30.48, 31.12, 31.75, and 33.71. The characteristic peaks may alternatively be expressed as (2θ) 3.9, 6.5, 7.1, 7.6, 7.9, 8.1, 9.3, 9.8, 10.3, 10.7, 11.6, 12.5, 13.0, 13.3, 14.2, 14.8, 15.1, 15.9, 16.6, 17.6, 18.0, 18.4, 18.7, 19.4, 20.3, 20.8, 21.3, 21.4, 21.7, 22.9, 23.4, 23.9, 24.1, 25.4, 26.2, 26.9, 27.2, 28.2, 28.7, 29.7, 30.5, 31.1, 31.8, and 33.7.

[0203] In one embodiment, the Fourier transform infrared (FT-IR) spectrum of crystalline form H1 is as Figure 31 shown. The FT-IR spectrum has characteristic peaks (in cm -1 -1) at: 537, 590, 627, 674, 708, 732, 744, 755, 791, 811, 969, 1053, 1084, 1110, 1132, 1161, 1203, 1234, 1284, 1325, 1388, 1445, 1470, 1525, 1598, 1638, 1658, 1711, and 3245.

[0204] The present invention provides a method for preparing crystalline form H1 of Compound I. The method comprises storing crystalline form H of Compound I at between about -15 °C and 20 °C, preferably between about 0 °C and 10 °C, more preferably at about 25 °C, preferably for about 1 month to 12 months, more preferably for about 6 months, to provide crystalline form H1 of Compound I.

[0205] The DSC thermogram of crystalline form J of Compound I is as Figure 33As shown, the DSC thermogram has characteristic endothermic peaks at 110 °C with an onset temperature of 83 °C; and an endothermic peak at 140 °C with an onset temperature of 133 °C. The DSC thermogram also has a characteristic exothermic peak at 179 °C with an onset temperature of 133 °C, and then an endothermic peak at 251 °C with an onset temperature of 250 °C.

[0206] The characteristic peaks (2θ) of the X-ray powder diffraction pattern of polymorph J are as Figure 32 shown. The characteristic peaks are located at the following 2θ (two theta) values: 4.36, 6.92, 7.05, 7.44, 8.21, 8.45, 9.47, 9.76, 10.22, 10.80, 11.26, 11.49, 12.02, 12.41, 12.90, 13.48, 13.74, 14.09, 14.53, 14.82, 15.24, 15.63, 15.78, 16.07, 16.41, 16.61, 17.60, 18.64, 19.13, 19.43, 19.72, 20.03, 20.26, 20.36, 20.51, 21.11, 21.87, 22.32, 22.97, 23.07, 23.35, 23.80, 24.14, 24.75, 24.99, 25.16, 25.42, 26.11, 26.69, 27.12, 27.42, 27.89, 28.51, 28.88, 32.33, 32.88, 33.67, 34.13, 34.38, 35.03, 35.73, 36.53, 37.47 and 38.84. The characteristic peaks may alternatively be expressed as (2θ) 4.4, 6.9, 7.1, 7.4, 8.2, 8.5, 9.5, 9.8, 10.2, 10.8, 11.3, 11.5, 12.0, 12.4, 12.9, 13.5, 13.7, 14.1, 14.5, 14.8, 15.2, 15.6, 15.8, 16.1, 16.4, 16.6, 17.6, 18.0, 18.2, 18.6, 19.1, 19.4, 19.7, 20.0, 20.3, 20.4, 20.5, 21.1, 21.9, 22.3, 23.0, 23.1, 23.4, 23.8, 24.1, 24.8, 25.0, 25.2, 25.4, 26.1, 26.7, 27.1, 27.4, 27.9, 28.5, 28.7, 29.2, 29.9, 30.1, 30.5, 31.1, 31.4, 32.0, 32.3, 32.9, 33.7, 34.1, 34.4, 35.0, 35.7, 36.5, 37.5 and 38.8.

[0207] The Fourier transform infrared (FT-IR) spectrum of polymorph J is as Figure 34As shown. FT-IR has characteristic peaks at the following (in cm -1 -1): 564, 598, 620, 648, 676, 706, 737, 749, 813, 910, 968, 1005, 1019, 1065, 1089, 1120, 1133, 1156, 1209, 1249, 1267, 1306, 1326, 1353, 1377, 1394, 1426, 1439, 1471, 1484, 1522, 1546, 1598, 1657, 1723, 2870, 2957, 3196, 3335.

[0208] In one embodiment, the present invention provides a method for preparing polymorph J of compound I. The method comprises the following steps:

[0209] i. Preferably between about 10 °C and 40 °C, more preferably between about 22 °C and 25 °C, prepare a solution of compound I in about 2.3X (X = weight of compound I) of tetrahydrofuran by volume;

[0210] ii. Preferably add about 0.001X to about 0.1X of polymorph J of compound I as a seed crystal, and add about 3X of heptane by volume to provide a suspension of polymorph J of compound I; and

[0211] iii. Filter the suspension to provide polymorph J of compound I.

[0212] In another aspect, the present invention features a crystal form (polymorph or pseudopolymorph) of compound I, which has, in an X-ray powder diffraction (XRPD) pattern, as Figure 3 , Figure 7 , Figure 9 , Figure 12 , Figure 13 , Figure 15 , Figure 17 , Figure 19 , Figure 20 , Figure 23 , Figure 25 , Figure 26 , Figure 28 , Figure 30 , Figure 32The characteristic peaks shown are substantially pure. As used herein, the term "substantially pure", when referring to a given crystalline form, means that the purity of the crystalline form is at least about 90%. This means that no more than about 10% of any other form of Compound I is present in the crystalline form. Preferably, the term "substantially pure" means that the purity of the crystalline form of Compound I is at least about 95%. This means that no more than about 5% of any other form of Compound I is present in this crystalline form of Compound I. More preferably, the term "substantially pure" means that the purity of the crystalline form of Compound I is at least about 97%. This means that no more than about 3% of any other form of Compound I is present in this crystalline form of Compound I.

[0213] For each polymorph and pseudopolymorph disclosed herein 1 the 1H NMR spectrum is mainly consistent with Compound I.

[0214] Table 11 lists the instruments and related methods used to characterize the polymorphs and pseudopolymorphs of Compound I. The results of these studies are shown in Figures 1 - 34 . As is known in the art, although the polymorphs and pseudopolymorphs are the same, Figures 1 - 34 the relative intensities of the peaks in [it] may change or shift under certain conditions. By comparing the XRPD pattern of a given polymorph or pseudopolymorph with those of Figure 3 , Figure 7 , Figure 9 , Figure 12 , Figure 13 , Figure 15 , Figure 17 , Figure 19 , Figure 20 , Figure 23 , Figure 25 , Figure 26 , Figure 28 , Figure 30 and Figure 32 ; comparing its DSC thermogram with those of DSC of Figure 4 , Figure 8 , Figure 10 , Figure 14 , Figure 16 , Figure 18 , Figure 21 , Figure 24 , Figure 27 , Figure 29 and Figure 33 ; comparing its FTIR spectrum with those of Figure 5 , Figure 11 , Figure 22 , Figure 31 and Figure 34 , one of ordinary skill in the art can readily determine whether the polymorph or pseudopolymorph is the same as the polymorph or pseudopolymorph described herein.

[0215] Table 11 Instrumental Methods of XRPD, DSC, NMR, FT-IR and Single Crystal X-ray Diffraction

[0216]

[0217]

[0218] Differential Scanning Calorimeter (DSC)

[0219]

[0220] Nuclear Magnetic Resonance (NMR)

[0221]

[0222] Single Crystal X-ray Diffractometer (SCXRD) Method

[0223]

[0224]

[0225] In one embodiment, the methods or compositions of the invention described herein (including any methods or compositions described in any aspect, embodiment, example or preference) use or comprise substantially pure Form 1 of Compound I. For example, the purity of Form 1 can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0226] In one embodiment, the methods or compositions of the invention described herein (including any methods or compositions described in any aspect, embodiment, example or preference) use or comprise substantially pure Form 2 of Compound I. For example, the purity of Form 2 can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0227] In one embodiment, the methods or compositions of the invention described herein (including any methods or compositions described in any aspect, embodiment, example or preference) use or comprise substantially pure Form 3 of Compound I. For example, the purity of Form 3 can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0228] In one embodiment, the above-described methods or compositions of the invention (including any methods or compositions described in any aspect, embodiment, example or preference) use substantially pure Form 4 of Compound I. For example, the purity of Form 4 can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0229] In one embodiment, the method of the present invention as described above (including any method described in any aspect, embodiment, example or preference) uses substantially pure Form A of Compound I. For example, the purity of Form A can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0230] In one embodiment, the method of the present invention as described above (including any method described in any aspect, embodiment, example or preference) uses substantially pure Form B of Compound I. For example, the purity of Form B can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0231] In one embodiment, the method of the present invention as described above (including any method described in any aspect, embodiment, example or preference) uses substantially pure Form C of Compound I. For example, the purity of Form C can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0232] In one embodiment, the method of the present invention as described above (including any method described in any aspect, embodiment, example or preference) uses substantially pure Form D of Compound I. For example, the purity of Form D can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0233] In one embodiment, the method of the present invention as described above (including any method described in any aspect, embodiment, example or preference) uses substantially pure Form E of Compound I. For example, the purity of Form E can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0234] In one embodiment, the method of the present invention as described above (including any method described in any aspect, embodiment, example or preference) uses substantially pure Form F of Compound I. For example, the purity of Form F can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0235] In one embodiment, the method of the present invention as described above (including any method described in any aspect, embodiment, example or preference) uses substantially pure Form G of Compound I. For example, the purity of Form G can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0236] In one embodiment, the method of the present invention as described above (including any method described in any aspect, embodiment, example or preference) uses substantially pure Form H of Compound I. For example, the purity of Form H can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0237] In one embodiment, the method of the present invention (including any method described in any aspect, embodiment, example or preference) uses polymorphic form H1 of substantially pure Compound I. For example, the purity of polymorphic form H1 can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0238] In one embodiment, the method of the present invention (including any method described in any aspect, embodiment, example or preference) uses polymorphic form J of substantially pure Compound I. For example, the purity of polymorphic form J can be at least 90%, preferably at least 95%, or more preferably at least 97%.

[0239] Unless otherwise defined, all technical and scientific terms used herein have the meanings commonly known to those of ordinary skill in the art. All references cited herein, whether in print, electronic, computer-readable storage medium or other forms, are hereby expressly incorporated by reference in their entirety, including but not limited to abstracts, articles, journals, publications, texts, dissertations, Internet websites, databases, patents and patent publications.

[0240] Pharmaceutical composition

[0241] The pharmaceutical composition of the present invention comprises a therapeutically effective amount of a polymorph or pseudopolymorph of Compound I of the present invention or a combination of two or more such polymorphs or pseudopolymorphs of Compound I, formulated together with one or more pharmaceutically acceptable carriers or excipients.

[0242] As used herein, the term "pharmaceutically acceptable carrier or excipient" refers to a non-toxic, inert solid, semi-solid or liquid filler, diluent, encapsulating material or any type of formulation aid. Some examples of materials that can serve as pharmaceutically acceptable carriers are sugars (such as lactose, glucose and sucrose), starches (such as corn starch and potato starch), cellulose and its derivatives (such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate), powdered tragacanth, malt, gelatin, talc, excipients (such as cocoa butter and suppository wax), oils (such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, soybean oil), alcohols (such as propylene glycol), esters (such as ethyl oleate, ethyl laurate), agar, buffers (such as magnesium hydroxide, aluminum hydroxide), alginic acid, pyrogen-free water, isotonic saline, Ringer's solution, ethanol and phosphate buffer solutions, and other non-toxic compatible lubricants (such as sodium lauryl sulfate and magnesium stearate), and coloring agents, release agents, coating agents, sweetening agents, flavoring agents and fragrances, preservatives and antioxidants may also be present in the composition, as determined by the formulator's judgment.

[0243] The pharmaceutical composition of the present invention can be administered orally, parenterally, by inhalation spray, topically, rectally, nasally, orally, vaginally or by implantation of a reservoir, preferably by oral administration or by injection. The pharmaceutical composition of the present invention may contain any conventional non-toxic pharmaceutically acceptable carrier, adjuvant or vehicle. In some cases, the pH value of the formulation can be adjusted with pharmaceutically acceptable acids, bases or buffers to enhance the stability of the formulated compound or its delivery form. As used herein, the term parenteral includes subcutaneous injection, intradermal injection, intravenous injection, intramuscular injection, intra-articular injection, intra-arterial injection, intra-synovial injection, intra-sternal injection, intrathecal injection, intra-lesional injection and intracranial injection or infusion techniques.

[0244] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active compound, the liquid dosage forms may contain inert diluents commonly used in the art, such as water or other solvents, solubilizers and emulsifiers, such as ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butanediol, dimethylformamide, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil and sesame oil), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycol and sorbitan fatty acid esters, and mixtures thereof. In addition to the inert diluent, the oral compositions may also include adjuvants such as wetting agents, emulsifying agents and suspending agents, sweetening agents, flavoring agents and fragrances.

[0245] Injectable preparations, such as sterile injectable aqueous or oily suspensions, can be formulated according to known techniques using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation can also be a sterile injectable solution, suspension or emulsion in a non-toxic parenterally acceptable diluent or solvent, such as a 1,3-butanediol solution. Acceptable vehicles and solvents can be water, Ringer's solution, U.S.P. and isotonic sodium chloride solution. In addition, a sterile fixed oil is commonly used as a solvent or suspending medium. For this purpose, any mild fixed oil can be used, including synthetic monoglycerides or diglycerides. In addition, fatty acids such as oleic acid can also be used in injectable preparations.

[0246] The injectable preparation can be sterilized, for example, by filtration through a bacteria-retaining filter, or by the addition of a sterilizing agent in the form of a sterile solid composition that is soluble or dispersible in sterile water or other sterile injectable medium before use.

[0247] To prolong the action of a drug, it is often necessary to slow its absorption by subcutaneous or intramuscular injection. This can be achieved by using a liquid suspension of a crystalline or amorphous substance with poor water solubility. Then the absorption rate of the drug depends on its dissolution rate, which in turn can depend on crystal size and crystal form. Alternatively, delayed absorption in a parenteral dosage form is achieved by dissolving or suspending the drug in an oil vehicle. Injectable depot forms are made by forming a microcapsule matrix of the drug in a biodegradable polymer such as poly(lactide-co-glycolide). Depending on the ratio of the drug to the polymer and the nature of the specific polymer used, the drug release rate can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Sustained-release injectable formulations can also be prepared by encapsulating the drug in liposomes or microemulsions that are compatible with body tissues.

[0248] Compositions for rectal or vaginal administration are preferably suppositories, which can be prepared by mixing a compound of the present invention with a suitable non-irritating excipient or carrier, such as cocoa butter, polyethylene glycol, or suppository wax, which is solid at ambient temperature but liquid at body temperature, and thus melts and releases the active compound in the rectum or vagina.

[0249] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is admixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or calcium phosphate dibasic and / or with: a) fillers or bulking agents such as starch, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and acacia, c) humectants such as glycerol, d) disintegrating agents such as agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) solution retarders such as paraffin, f) absorption promoters such as quaternary ammonium compound; g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate; h) adsorbents such as kaolin and bentonite clays; and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets, and pills, the dosage form may also include buffering agents.

[0250] Solid compositions of a similar type can also use excipients such as lactose or milk sugar and high molecular weight polyethylene glycol as fillers in soft and hard gelatin capsules.

[0251] Solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells, such as enteric coatings and other coatings well known in the pharmaceutical formulation art. They can optionally contain emulsifying agents and can also be compositions that optionally release the active ingredient only in a certain part of the intestine or preferentially in a certain part of the intestine in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes.

[0252] Dosage forms for topical or transdermal administration of the compounds of the present invention include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants, or patches. The active ingredient is admixed with a pharmaceutically acceptable carrier and any desired preservatives or buffering agents that may be required under sterile conditions. Ophthalmic preparations, ear drops, eye ointments, powders, and solutions are also considered to be within the scope of the present invention.

[0253] In addition to the active compounds of the present invention, ointments, pastes, creams, and gels can also contain excipients such as animal and vegetable fats, oils, waxes, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonite, silicic acid, talc, and zinc oxide, or mixtures thereof.

[0254] In addition to the compounds of the present invention, powders and sprays can also contain excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicate, and polyamide powder, or mixtures of these substances. Sprays can also contain conventional propellants such as chlorofluorocarbons.

[0255] An additional advantage of transdermal patches is that they can provide controlled delivery of the compound to the human body. Such dosage forms can be prepared by dissolving or dispensing the compound in a suitable medium. Absorption enhancers can also be used to increase the flux of the compound through the skin. The rate can be controlled by providing a rate-controlling membrane or by dispersing the compound in a polymer matrix or gel.

[0256] For pulmonary delivery, the therapeutic compositions of the present invention are formulated and administered directly, for example, by inhalation into the respiratory system, in the form of solid or liquid particles to the patient. Solid or liquid particle dosage forms of the active compounds prepared for the practice of the present invention include respirable-sized particles: i.e., particles that are small enough to pass through the mouth and larynx upon inhalation and enter the bronchi and alveoli of the lungs. The delivery of nebulized therapeutic agents, particularly nebulized antibiotics, is known in the art (see, for example, U.S. Patent No. 5,767,068 to Van Devanter et al., U.S. Patent No. 5,508,269 to Smith et al., and WO 98 / 43650 to Montgomery, all incorporated herein by reference).

[0257] Antiviral activity

[0258] The present invention provides a prevention or treatment of viral infection in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a polymorph or pseudopolymorph of Compound I described herein or a combination of two or more thereof. The viral infection is preferably a coronavirus infection. In certain embodiments, the coronavirus is SARS-CoV-1, SARS-CoV-2 or MERS-CoV. Preferably, the coronavirus is SARS-CoV-2.

[0259] The viral inhibitory amount or dosage range of Compound I of the present invention can be from about 0.01 mg / Kg to about 500 mg / Kg, or from about 1 mg / Kg to about 50 mg / Kg. The inhibitory amount or dosage will also vary depending on the route of administration and the possibility of co-use with other agents.

[0260] According to the treatment method of the present invention, a therapeutically effective amount of the compound of the present invention is administered to a patient (such as a human or other animal) to treat or prevent viral infection in the patient in an amount and time required to achieve the desired effect.

[0261] The "therapeutically effective amount" of the compound of the present invention refers to the amount of the compound that confers a therapeutic effect on a treated subject at a reasonable benefit / risk ratio applicable to any medical treatment. The therapeutic effect can be objective (i.e., measurable by certain tests or markers), or subjective (i.e., the subject shows signs of the effect or feels the effect). The range of the therapeutically effective amount of the above compound can be, for example, from about 0.1 mg / Kg to about 500 mg / Kg, preferably from about 1 mg / Kg to about 50 mg / Kg. The effective dosage will also vary depending on the route of administration and the possibility of co-use with other agents. However, it is understood that the total daily dosage of the compounds and compositions of the present invention will be determined by the attending physician within the scope of reasonable medical judgment. The specific therapeutically effective dosage level for any particular patient will depend on a variety of factors, including the disease being treated and the severity of the disease; the activity of the particular compound used; the particular composition used; the age, weight, general health, sex and diet of the patient; the time of administration, the route of administration and the excretion rate of the particular compound used; the duration of treatment; the drugs used in combination with or simultaneously with the particular compound used; and similar factors well known in the medical art.

[0262] The total daily dosage of the compound of the present invention administered to a human or other animal in a single or divided dose can be, for example, from 0.01 mg / kg body weight to 50 mg / kg body weight, or generally an amount from 0.1 mg / kg body weight to 25 mg / kg body weight. A single-dose composition can contain such a dosage or a sub-dose thereof to constitute the daily dosage. Generally, the treatment regimen according to the present invention comprises administering about 10 mg to about 1000 mg of the compound of the present invention to a patient in need of treatment in a single dose or multiple doses per day.

[0263] The compounds of the invention described herein can be administered, for example, by injection (intravenous, intraarterial, subdermal, intraperitoneal, intramuscular or subcutaneous); or orally, buccally, nasally, transmucosally, topically, ophthalmically or by inhalation, in a dosage range of from about 0.1 mg / kg body weight to 500 mg / kg body weight, or a dose between 1 mg / dose to 1000 mg / dose, administered once every 4 hours to 120 hours, or as required for a particular drug. The methods described herein contemplate administering an effective amount of the compound or compound composition to achieve the desired or stated effect. Generally, the pharmaceutical compositions of the invention will be administered about 1 to about 6 times per day, or as a continuous infusion. Such administration can be used as a chronic or acute therapy. The amount of active ingredient that can be combined with a pharmaceutical excipient or carrier to form a single dosage form will vary depending on the host being treated and the particular mode of administration. Typical formulations will contain from about 5% to about 95% active compound (w / w). Alternatively, such formulations can contain from about 20% to about 80% active compound. Lower or higher doses than those described above may be required. The specific dosage and treatment regimen for any particular patient will depend on a variety of factors, including the activity of the particular compound used, age, weight, general health, sex, diet, time of administration, rate of excretion, drug combination, disease, condition or severity and course of symptoms, the patient's disposition to the disease, condition or symptoms, and the judgment of the attending physician.

[0264] After the patient's condition improves, a maintenance dose of the compound, composition or combination of the invention can be administered if necessary. Subsequently, depending on the change in symptoms, the dosage or dosing frequency or both the dosage and dosing frequency can be reduced until the condition is maintained at an improved level when the symptoms are alleviated to the desired extent. However, when the symptoms of the disease recur, the patient may require long-term intermittent treatment.

[0265] Combination and alternative therapies

[0266] The compounds of the present invention can be used in combination with one or more antiviral therapeutic agents or anti-inflammatory agents that can be used for the prevention or treatment of viral diseases or related pathophysiological changes. Therefore, the compounds of the present invention and their salts, solvents or other pharmaceutically acceptable derivatives can be used alone or in combination with other antiviral or anti-inflammatory therapeutic agents. The compounds and pharmaceutically acceptable salts thereof described herein can be used in combination with one or more other agents that can be used for the prevention or treatment of respiratory diseases, inflammatory diseases, autoimmune diseases, such as: antihistamines, corticosteroids (such as fluticasone propionate, fluticasone furoate, beclomethasone dipropionate, budesonide, ciclesonide, mometasone furoate, triamcinolone acetonide, flunisolide), non-steroidal anti-inflammatory drugs, leukotriene regulators (such as montelukast, zafirlukast, pranlukast), tryptase inhibitors, IKK2 inhibitors, p38 inhibitors, Syk inhibitors, protease inhibitors (such as elastase inhibitors), integrin antagonists (such as β-2 integrin antagonists), adenosine A2a agonists, mediator release inhibitors (such as sodium cromoglycate), 5-lipoxygenase inhibitors (zyflo), DP1 antagonists, DP2 antagonists, PI3Kδ inhibitors, ITK inhibitors, LP (lysophosphatidic acid) inhibitors or FLAP (5-lipoxygenase activating protein) inhibitors (such as sodium 3-(3-(tert-butylthio)-1-(4-(6-ethoxypyridin-3-yl)benzyl)-5-((5-ethylpyridin-2-yl)methoxy)-1H-indol-2-yl)-2,2-dimethylpropionate), bronchodilators (such as muscarinic antagonists, β-2 agonists), methotrexate and similar preparations; monoclonal antibody therapies, such as anti-lgE, anti-TNF, anti-IL-5, anti-IL-6, anti-IL-12, anti-IL-1 and similar preparations; cytokine receptor therapies, such as etanercept and similar preparations; antigen-nonspecific immunotherapies (such as interferons or other cytokines / chemokines, chemokine receptor modulators such as CCR3, CCR4 or CXCR2 antagonists, other cytokine / chemokine agonists or antagonists, TLR agonists and similar agents), suitable anti-infective agents, including antibacterial agents, antifungal agents, anthelmintics, antimalarials, antiprotozoals, antituberculosis agents and antiviral agents, including those listed at https: / / www.drugs.com / drug-class / anti-infectives.html. Generally, combination therapy is usually preferred over alternative therapy because it exerts multiple pressures on the virus simultaneously.

[0267] When the compositions of the present invention comprise one or more polymorphs and / or pseudopolymorphs of Compound I as described herein and one or more additional therapeutic or prophylactic agents, Compound I and the additional agent(s) should be present at dose levels of from about 1% to 100%, and more preferably from about 5% to 95% of the dosage administered in a monotherapy regimen. The additional agent(s) can be administered separately from the compounds of the present invention as part of a multi-dose regimen.

[0268] Alternatively, the agents can be part of a single dosage form, combined with the compounds of the present invention into a single composition.

[0269] "Additional therapeutic or prophylactic agents" include, but are not limited to, immunotherapies (e.g., interferons), therapeutic vaccines, anti-fibrotic agents, anti-inflammatory agents (such as corticosteroids or non-steroidal anti-inflammatory drugs), bronchodilators (such as β-2 adrenergic agonists) and xanthines (e.g., theophylline), mucolytics, anti-muscarinic agents, anti-leukotriene agents, cell adhesion inhibitors (e.g., ICAM antagonists), antioxidants (e.g., N-acetylcysteine), cytokine agonists, cytokine antagonists, pulmonary surfactants, and / or antibacterial and antiviral agents (e.g., ribavirin and amantadine). Compositions according to the present invention can also be used in combination with gene replacement therapies.

[0270] Examples

[0271] The compounds and methods of the present invention will be better understood in conjunction with the following examples, which are presented for illustration only and are not intended to limit the scope of the invention. Various changes and modifications to the disclosed embodiments will be apparent to those skilled in the art, and such changes and modifications include, but are not limited to, those relating to the chemical structure, substituents, derivatives, formulations, and / or methods of the present invention, which can be made without departing from the spirit of the invention and the scope of the appended claims.

[0272] Example 1: Preparation of Polymorph 1 of Compound I

[0273] Procedure 1A: Ethyl acetate (700 mL, 10V (volume), where 1 volume represents 1 g of solid in 1 mL of solvent) was added to reactor (R1), and then compound (3R,5'S)-1'-(N-methyl-N-(4,6,7-trifluoro-1H-indole-2-carbonyl)-L-leucyl)-2-oxospiro[indoline-3,3'-pyrrolidine]-5'-carboxamide (70 g) was added. The reaction was cooled to 0 °C (-5 °C to +5 °C), and then triethylamine (89.6 g) was added at 0 °C (-5 °C to 5 °C), followed by trifluoroacetic anhydride (92.4 g). The reaction was stirred at 0 °C (-5 °C to 5 °C) for 1 hour (0.5 to 2 hours). After the reaction was completed, at 0 °C (-5 °C to 5 °C), the reaction mixture was slowly added to 0.2N aqueous hydrochloric acid solution (700 g) within 1 hour. The resulting solution was stirred at 0 °C (-5 °C to 5 °C) for 10 minutes to 30 minutes, and the organic layer was separated. The organic layer was separated and washed 5 times with 10% brine. Then the organic layer was separated and distilled to 280 mL (Note: 280 mL is the total volume of the solution). Seeds of polymorph 1 were added to induce crystallization at 50 °C. Then, by adding approximately 10V of toluene, distilling to 8V, and then adding 8V of toluene again and distilling to 8V (Note: solution volume), this cycle was repeated for toluene-ethyl acetate exchange to control the ethyl acetate level at 1-5% wt / wt (ethyl acetate weight / toluene weight under gas chromatography). The suspension was slowly cooled from 50 °C to 25 °C within 1 hour and stirred at 25 °C (20-30 °C) for 5 hours (3 to 8 hours). The suspension was filtered, and the wet cake was rinsed with toluene (2V). The wet cake was dried at 50 °C (45-55 °C) for 48 hours to provide polymorph 1 of compound I as a white solid with a yield of 80-85%. 1 HNMR (400 MHz, acetone-d 6 ) δ 11.17 (s, 1H), 9.65 (s, 1H), 7.02 (dd, J = 13.7, 7.3 Hz, 2H), 6.94 (dd, J = 6.0, 3.5 Hz, 1H), 6.92–6.85 (m, 2H), 6.81 (t, J = 7.5 Hz, 1H), 5.56 (dd, J = 9.4, 5.6 Hz, 1H), 5.21 (t, J = 8.3 Hz, 1H), 4.25 (d, J = 10.7 Hz, 1H), 3.99 (d, J = 10.6 Hz, 1H), 3.43 (s, 3H), 2.79–2.61 (m, 2H), 1.93 (ddd, J = 14.4, 9.5, 5.1 Hz, 1H), 1.79 (ddd, J = 14.2, 8.7, 5.6 Hz, 1H), 1.64 (dpd, J = 8.7, 6.6, 5.1 Hz, 1H), 0.98 (dd, J = 18.5, 6.6 Hz, 6H).

[0274] Procedure 1B: Ethyl acetate (300 mL, 10 volumes, where 1 volume represents 1 g of solid in 1 mL of solvent) was added to reactor (R1), and then compound (3R,5'S)-1'-(N-methyl-N-(4,6,7-trifluoro-1H-indole-2-carbonyl)-L-leucyl)-2-oxospiro[indoline-3,3'-pyrrolidine]-5'-carboxamide (30 g) was added to form a clear solution. The reaction was cooled to 0 °C (-5 °C to 5 °C), and then triethylamine (38.3 g) was added at 0 °C (-5 °C to 5 °C), followed by trifluoroacetic anhydride (39.7 g). The reaction was stirred at 0 °C (-5 °C to 5 °C) for 20 minutes, and then a sample was taken for purity analysis. After the reaction was completed, at 0 °C (-5 °C to 5 °C), the reaction mixture was slowly added to 0.2 N aqueous hydrochloric acid solution (300 g) in reactor R2 within 1 hour. The resulting solution was stirred at 0 °C (-5 °C to 5 °C) for 20 minutes, the organic layer was separated, and washed 5 times with 10% brine (300 mL). Then the organic layer was separated and distilled under vacuum below 50 °C to 60 mL (note: 60 mL is the volume of the solution). Then toluene (300 mL) was added, and the mixture was concentrated to 120 mL. Then additional toluene (300 mL) was added, and the resulting mixture was concentrated to a final volume of 240 mL. The temperature of the mixture was adjusted to 75 °C (70 - 80 °C), and stirred at 75 °C (70 - 80 °C) for 1 hour. Then the mixture was slowly cooled to 20 °C (15 - 25 °C) within 1 hour, and the resulting suspension was stirred at 20 °C (15 - 25 °C) for 1 hour (0.5 - 2 hours). The suspension was filtered, and the wet cake was rinsed with toluene (60 mL). The wet cake was dried at 50 °C (45 - 55 °C) for 16 hours to give crystalline form 1 of compound I as a white solid, with a yield of 80 - 85%. 1 HNMR (400 MHz, acetone-d6) δ 11.17 (s, 1H), 9.65 (s, 1H), 7.02 (dd, J = 13.7, 7.3 Hz, 2H), 6.94 (dd, J = 6.0, 3.5 Hz, 1H), 6.92–6.85 (m, 2H), 6.81 (t, J = 7.5 Hz, 1H), 5.56 (dd, J = 9.4, 5.6 Hz, 1H), 5.21 (t, J = 8.3 Hz, 1H), 4.25 (d, J = 10.7 Hz, 1H), 3.99 (d, J = 10.6 Hz, 1H), 3.43 (s, 3H), 2.79–2.61 (m, 2H), 1.93 (ddd, J = 14.4, 9.5, 5.1 Hz, 1H), 1.79 (ddd, J = 14.2, 8.7, 5.6 Hz, 1H), 1.64 (dpd, J = 8.7, 6.6, 5.1 Hz, 1H), 0.98 (dd, J = 18.5, 6.6 Hz, 6H).

[0275] Procedure 2: The following procedure is used to further purify polymorph 1 of Compound I, such as polymorph 1 prepared according to Procedure 1(a) or 1(b) above. At 20 - 30 °C, 20 g of Compound I and ethanol (300 mL, 15V) are added to a reactor. The mixture is stirred at 70 - 80 °C to completely dissolve polymorph 1 of Compound I. The solution is slowly cooled to 60 °C within 1 hour. Seeds of polymorph 1 of Compound I are added to induce crystallization, and the resulting suspension is stirred at 60 °C for 0.5 - 1.5 hours. Then the suspension is slowly cooled to 20 - 30 °C within 1 hour. Water (300 mL, 15V) is slowly added to the suspension within 5 hours. The resulting suspension is stirred at 20 - 30 °C for 0.5 - 2 hours. The slurry is filtered and the wet cake is rinsed with EtOH / H 2 O (30 mL / 30 mL). The wet cake is dried at 50 °C (45 - 55 °C) for 48 hours to obtain solid polymorph 1 of Compound I with a yield of 95 - 100%. 1 HNMR (400 MHz, acetone-d 6 ) δ 11.17 (s, 1H), 9.65 (s, 1H), 7.02 (dd, J = 13.7, 7.3 Hz, 2H), 6.94 (dd, J = 6.0, 3.5 Hz, 1H), 6.92–6.85 (m, 2H), 6.81 (t, J = 7.5 Hz, 1H), 5.56 (dd, J = 9.4, 5.6 Hz, 1H), 5.21 (t, J = 8.3 Hz, 1H), 4.25 (d, J = 10.7 Hz, 1H), 3.99 (d, J = 10.6 Hz, 1H), 3.43 (s, 3H), 2.79–2.61 (m, 2H), 1.93 (ddd, J = 14.4, 9.5, 5.1 Hz, 1H), 1.79 (ddd, J = 14.2, 8.7, 5.6 Hz, 1H), 1.64 (dpd, J = 8.7, 6.6, 5.1 Hz, 1H), 0.98 (dd, J = 18.5, 6.6 Hz, 6H).

[0276] Example 2: Preparation of polymorph 2 of Compound I

[0277] Approximately 15 mg of polymorph J of Compound I is heated to 155 °C at a rate of 10 °C / min and held at 155 °C for 3 minutes to provide polymorph 2 of Compound I.

[0278] Example 3: Preparation of polymorph 3 of Compound I

[0279] In a variable temperature powder X-ray diffraction (VT-XRPD) experiment, approximately 15 mg of polymorph J of Compound I is heated to 80 °C and then cooled under ambient conditions for about 3 hours to provide polymorph 3 of Compound I.

[0280] Example 4: Preparation of Polymorph 4 of Compound I

[0281] In a variable temperature powder X-ray diffraction (VT-XRPD) experiment, approximately 15 mg of polymorph J of Compound I was heated to 60 °C to afford polymorph 4 of Compound I.

[0282] Example 5: Preparation of Polymorph A of Compound I

[0283] 100 mg of amorphous Compound I was stirred in a mixture of benzyl alcohol (BnOH) and toluene (20:80, v / v), and temperature cycling was performed between 5 °C and 50 °C using a heating / cooling rate of 0.1 °C / min. After repeating approximately 4 to 15 cycles, the wet cake was collected by filtration to afford polymorph A of Compound I. As determined by 1 1H-NMR, the wet cake contained approximately 23.1% (1.5 eq) of BnOH and 0.5% (0.04 eq) of toluene by weight.

[0284] Example 6: Preparation of Polymorph B of Compound I

[0285] 100 mg of amorphous Compound I was stirred in anisole, and temperature cycling was performed between 5 °C and 50 °C using a heating / cooling rate of 0.1 °C / min. After repeating approximately 4 to 15 cycles, the wet cake was collected by filtration to afford polymorph B of Compound I. As determined by 1 1H-NMR, the wet cake contained 13.4% (0.8 eq) of anisole by weight.

[0286] Example 7: Preparation of Polymorph C of Compound I

[0287] Approximately 30 mg of amorphous Compound I was dissolved in anisole. The resulting solution was filtered through a 0.45 μm membrane filter. The resulting clear solution was slowly evaporated under ambient conditions to afford polymorph C of Compound I. As determined by 1 1H-NMR, the wet cake contained approximately 60.7% (7.7 eq) of anisole by weight.

[0288] Example 8: Preparation of Polymorph D of Compound I

[0289] 10 g of amorphous Compound I was dissolved in 100 ml of ethyl acetate. Solvent exchange was carried out at no more than 50 °C to replace ethyl acetate with toluene under vacuum to a final volume of approximately 30 - 40 ml. Additional toluene (50 mL) was added, and the slurry was stirred at 50 °C for 2 hours. The suspension was cooled to 20 °C within 1 - 2 hours and then stirred at 20 °C (15 °C to 25 °C) for 10 hours. The suspension was filtered, rinsed with toluene (20 mL), and then dried to afford polymorph D of Compound I.

[0290] Example 9: Preparation of Polymorph E of Compound I

[0291] Add approximately 200 mg of polymorph D of Compound I into a 2 mL glass vial. Add 1.2 mL of DMSO / water (1:1, v / v) into the vial. Stir the mixture at 25 °C for approximately 5 minutes to obtain a suspension. Add approximately 10 mg of polymorph E seed crystals into the suspension. Stir the suspension and perform a temperature cycle between 5 °C and 50 °C 20 times using a heating / cooling rate of 0.1 °C / min. Centrifugally filter the resulting suspension through a 0.45 μm membrane filter. Collect the wet cake to provide polymorph E of Compound I.

[0292] Example 10: Preparation of Polymorph F of Compound I

[0293] Stir approximately 50 mg of polymorph D of Compound I in 0.1 - 0.2 mL of PEG / water (1:1, v / v) and perform a temperature cycle between 5 °C and 50 °C at least 10 times using a heating / cooling rate of 0.1 °C / min. Centrifugally filter the resulting suspension through a 0.45 μm membrane filter at a rotational speed of 14000 rpm. Collect the wet cake to provide polymorph F of Compound I.

[0294] Example 11: Preparation of Polymorph G of Compound I

[0295] Add approximately 200 mg of polymorph D of Compound I into a 2 mL glass vial. Add 0.4 mL of MEK / heptane (1:1, v / v) into the vial. Stir the mixture at 25 °C for approximately 5 minutes to obtain a suspension. Add approximately 10 mg of polymorph G seed crystals into the suspension. Stir the suspension and perform a temperature cycle between 5 °C and 50 °C 10 times using a heating / cooling rate of 0.1 °C / min. Centrifugally filter the obtained suspension through a 0.45 μm membrane filter at a rotational speed of 14000 rpm. Collect the wet cake to provide polymorph G of Compound I.

[0296] Example 12: Preparation of Polymorph H of Compound I

[0297] Add approximately 200 mg of polymorph D of Compound I into a 2 mL glass vial. Add 0.4 mL of IPA / water (1:1, v / v) into the vial. Stir the resulting mixture at 25 °C for approximately 5 minutes to obtain a suspension. Add approximately 10 mg of polymorph H seed crystals into the above suspension. Stir the suspension at 25 °C for approximately 10 days. Collect the solid by centrifugation and then dry it under vacuum at 50 °C for approximately 1 hour. Obtain approximately 180 mg of polymorph H of Compound I as an off-white solid with a yield of 90%.

[0298] Example 13: Preparation of Polymorph H1 of Compound I

[0299] About 10 mg of crystalline form H of Compound I was stored at 4 °C for 6 months to provide crystalline form H1 of Compound I.

[0300] Example 14: Preparation of crystalline form J of Compound I

[0301] At about 22 - 25 °C, about 130 mg of crystalline form 1 of Compound I was dissolved in 0.3 mL of THF. The resulting solution was filtered through a 0.45 μm membrane filter. Seeds of crystalline form J were added to the obtained clear solution, and then 0.4 mL of heptane was slowly added to form a suspension. The solid was collected by centrifugal filtration at 14000 rpm through a 0.45 μm membrane filter to provide crystalline form J of Compound I.

[0302] Abbreviation

[0303] DSC: Differential Scanning Calorimetry

[0304] DVS: Dynamic Vapor Sorption

[0305] FT-IR: Fourier Transform Infrared

[0306] RH: Relative Humidity

[0307] PEG: Polyethylene Glycol

[0308] XRPD: X-Ray Powder Diffraction

[0309] 1 H-NMR: Proton Nuclear Magnetic Resonance

[0310] T3P: Propylphosphonic Anhydride

[0311] TFAA: Trifluoroacetic Anhydride

[0312] P 2 O 5 : Phosphorus Pentoxide

[0313] MEK: Methyl Ethyl Ketone

[0314] EtOH: Ethanol

[0315] H 2 O: Water

[0316] DMSO: Dimethyl Sulfoxide

[0317] EtOAc: Ethyl Acetate

[0318] HCl: Hydrogen Chloride

[0319] ACN: Acetonitrile

[0320] While the invention has been particularly shown and described with reference to its preferred embodiments, those skilled in the art will understand that various changes in form and detail may be made therein without departing from the scope of the invention as defined by the appended claims.

Claims

1. Polymorph 1 of Compound I, wherein, the polymorph is characterized by having characteristic peaks in the X-ray powder diffraction pattern at the following 2θ (°2θ) values: 6.9, 8.3, 10.8, 11.9, 13.2, 13.7, 14.5, 15.4, 16.1, 18.2, 19.2, 20.0, 20.6, 21.7, 22.3, 23.9, 24.8, 25.3, 26.1, 26.7, 27.6, 27.9, 28.9, 29.3, 30.1, 31.3, 31.3, 31.7, 32.5, 32.9, 34.7, 35.0, 35.8, 36.1, 36.8, 37.1, 37.8, 38.8 and 39.

7.

2. The polymorph according to claim 1, further characterized in that the differential scanning calorimetry thermogram has an endothermic peak at 251 °C with an onset temperature of 250 °C.

3. The polymorph according to claim 1 or 2, characterized in that, the X-ray powder diffraction pattern of Figure 3.

4. The polymorph according to any one of claims 1 to 3, wherein, the polymorph is characterized in that the differential scanning calorimetry thermogram has an endothermic peak at 175 °C with an onset temperature of 175 °C and an endothermic peak at 251 °C with an onset temperature of 250 °C.

5. Polymorph 2 of Compound I, wherein, the polymorph is characterized by having characteristic peaks in the X-ray powder diffraction pattern at the following 2θ (°2θ) values: 6.9, 10.8, 11.9, 13.7, 14.5, 16.1, 18.1, 20.0, 20.6, 21.7, 22.3, 23.9, 24.8, 25.2, 26.1, 26.7, 27.9, 28.8, 29.2, 30.1, 31.3, 32.8, 34.3, 34.7, 35.77, 36.1, 37.1 and 39.

9.

6. The polymorph according to claim 4, wherein, the polymorph is further characterized in that the differential scanning calorimetry thermogram has an endothermic peak at 175 °C with an onset temperature of 175 °C and an endothermic peak at 251 °C with an onset temperature of 250 °C.

7. The polymorph according to claim 5 or 6, wherein, the polymorph is characterized by the XRPD pattern shown in Figure 7.

8. Polymorph 3 of Compound I, wherein, The characteristics of the crystal form are: (i) the differential scanning calorimeter thermogram has an endothermic peak at 150 °C with an onset temperature of 142 °C, an exothermic peak at 176 °C with an onset temperature of 171 °C, and an endothermic peak at 252 °C with an onset temperature of 251 °C; (ii) the X-ray powder diffraction pattern has main peaks (2θ) at the following: 7.10, 8.71, 8.92, 11.03, 11.70, 13.07, 14.06, 15.23, 15.55, 16.17, 17.14, 17.45, 17.96, 19.16, 19.56, 20.43, 20.68, 21.89, 22.56, 23.45, 23.83, 24.74, 25.79, 26.26, 26.95, 28.12, 28.64, 29.68, 30.44, 31.28, 31.76, 32.34, 32.79, 33.30, 34.67, 35.56, 36.11, 36.47, 37.07, 37.85 and 38.

22.

9. Crystal form 3 of compound I wherein, the characteristics of the crystal form are that the XRPD pattern has characteristic peaks at the following 2θ (°2θ) values: 7.1, 8.7, 8.9, 11.0, 11.7, 13.1, 14.1, 15.2, 15.6, 16.2, 17.1, 17.5, 18.0, 19.2, 19.6, 20.4, 20.7, 21.9, 22.6, 23.5, 23.8, 24.7, 25.8, 26.3, 27.0, 28.1, 28.6, 29.7, 30.4, 31.3, 31.8, 32.3, 32.8, 33.3, 34.7, 35.6, 36.1, 36.5, 37.1, 37.9 and 38.

2.

10. The crystal form according to claim 8, wherein, the characteristics of the crystal form lie in the XRPD pattern of Figure 9.

11. Crystal form 4 of compound I wherein, the characteristics of the crystal form are that the X-ray powder diffraction pattern has characteristic peaks at the following 2θ (°2θ) values of (2θ): 4.39, 5.14, 7.06, 7.50, 8.47, 8.92, 9.75, 10.08, 10.33, 11.03, 11.71, 12.45, 13.09, 13.83, 14.06, 14.62, 14.84, 15.38, 15.78, 15.95, 16.91, 17.45, 17.93, 18.54, 19.23, 19.66, 20.07, 20.96, 21.84, 22.50, 23.42, 24.19, 24.91, 25.13, 26.18, 26.54, 27.81, 28.54, 29.41 and 30.

99.

12. Crystal form 4 of compound I wherein, The crystal form is characterized in that the XRPD pattern has characteristic peaks at the following 2θ (°2θ) values: 4.4, 5.1, 7.1, 7.5, 8.5, 8.9, 9.8, 10.1, 10.3, 11.0, 11.7, 12.5, 13.1, 13.8, 14.1, 14.6, 14.8, 15.4, 15.8, 16.0, 16.9, 17.5, 17.9, 18.5, 19.2, 19.7, 20.1, 21.0, 21.8, 22.5, 23.4, 24.2, 24.9, 25.1, 26.2, 26.5, 27.8, 28.5, 29.4 and 31.

0.

13. The crystal form according to claim 1, wherein, the XRPD pattern is shown in Figure 12.

14. A composition comprising the crystal form according to any one of claims 1 to 12.

15. A method for manufacturing a pharmaceutical composition comprising Compound I, comprising dissolving the crystal form according to any one of claims 1 to 12 in a solvent.

16. A method for preparing Crystal Form 1 of Compound I, the method comprising the following steps: 1) Prepare a solution of Compound I by dehydrating (3R,5'S)-1'-(N-methyl-N-(4,6,7-trifluoro-1H-indole-2-carbonyl)-L-leucyl)-2-oxospiro[indoline-3,3'-pyrrolidine]-5'-carboxamide (Compound n) in the presence of one or more suitable dehydrating agents, a suitable base and a suitable solvent; 2) Concentrate the solution of Compound I in step 1 at a temperature of about 20 °C to about 80 °C to a volume (milliliters) / weight (grams) ratio of solvent / Compound n of about 2:1 to about 6:1; 3) Optionally add Crystal Form 1 of Compound I in an amount of about 0.0001X (X = the weight of Compound n in step 1) to about 0.1X by weight as a seed crystal to the concentrated solution in step 2 to induce crystallization of Crystal Form 1 of Compound I; 4) At a temperature of about 20 °C to about 80 °C, exchange the solvent to a suitable solvent to a volume (milliliters) / weight (grams) ratio of solvent / Compound n of about 6:1 to about 10:1, the suitable solvents including but not limited to anisole, toluene, xylene, ethyl acetate, isopropyl acetate and mixtures of two or more thereof; 5) Cool to a temperature of about 0 °C to about 50 °C; and 6) Filter to provide Crystal Form 1 of Compound I.

17. The method according to claim 15, wherein, the solvent is selected from anisole, toluene, xylene, ethyl acetate, isopropyl acetate and mixtures of two or more thereof.

18. A method for preparing Crystal Form 1 of Compound I, the method comprising the following steps: I. Add Compound I to a first solvent to form a solution; Compound I for forming the solution can be in the solid form of the compound or a mixture of two or more solid forms; II. Remove a portion of the first solvent from the solution, for example by distillation; and then add more of the first solvent; III. Repeatedly remove a portion of the first solvent from the solution and add more of the first solvent until a solution of Compound I is produced, with the residual solvent controlled to be no greater than a predetermined value, such as no greater than 0.1% (wt / wt); IV. Add a second solvent to the solution to precipitate polymorph 1 of Compound I; and V. Separate polymorph 1 of Compound I by filtration.

19. The method according to claim 18, wherein, the first solvent is EtOH, isopropanol, acetone, EtOAc, IPAc or MTBE.

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