WO2024165044A1 - 作为蛋白激酶Mek抑制剂的多晶型物、及其制备方法和用途 - Google Patents

作为蛋白激酶Mek抑制剂的多晶型物、及其制备方法和用途 Download PDF

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WO2024165044A1
WO2024165044A1 PCT/CN2024/076655 CN2024076655W WO2024165044A1 WO 2024165044 A1 WO2024165044 A1 WO 2024165044A1 CN 2024076655 W CN2024076655 W CN 2024076655W WO 2024165044 A1 WO2024165044 A1 WO 2024165044A1
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polymorph
ray powder
positions
crystalline form
diffraction pattern
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French (fr)
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田红旗
黄功超
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上海科州药物研发有限公司
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/425—Thiazoles
    • A61K31/428—Thiazoles condensed with carbocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D277/00—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings
    • C07D277/60—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings condensed with carbocyclic rings or ring systems
    • C07D277/62—Benzothiazoles

Definitions

  • the present invention relates to polymorphs of benzothiazole compounds as protein kinase Mek inhibitors, preparation methods thereof, and medical uses thereof.
  • Ras/Raf/Mek/Erk signaling mechanism plays an important role in the proliferation and differentiation of cancer cells; the Ras/Raf/Mek/Erk signaling mechanism has been found to be continuously activated or overactivated in many cancers, such as pancreatic cancer, colon cancer, lung cancer, bladder cancer, kidney cancer, skin cancer, breast cancer, etc. Inhibition of the Ras/Raf/Mek/Erk signaling pathway helps to treat such hyperproliferative diseases, in which Mek, a downstream target of Ras and Raf, plays a key role in this pathway, and the substrate of Mek phosphorylation is MAP kinase Erk.
  • Mek inhibitors can inhibit the growth of cancer cells, especially for cancers caused by overactivation of Ras or Raf. At the same time, Mek is also involved in inflammatory diseases and symptoms, including acute and chronic inflammation.
  • Chinese Patent Application No. 201210190520.4 discloses a number of benzothiazole compounds that exhibit protein kinase Mek inhibitory activity, including compound 4-fluoro-5-(2-fluoro-4-iodophenylamino)-1H-benzo[d]thiazole-6-carboxylic acid (2-hydroxy-ethoxy)-amide (hereinafter referred to as Compound 1).
  • Compound 1 4-fluoro-5-(2-fluoro-4-iodophenylamino)-1H-benzo[d]thiazole-6-carboxylic acid (2-hydroxy-ethoxy)-amide
  • the solid form of Compound 1 is amorphous.
  • the purity of amorphous is usually difficult to control, the physical and chemical stability is usually poor, and the hygroscopicity is poor.
  • Compound 1 is unstable under light and easily degrades to form impurities (A)
  • the present invention provides polymorphs of compound 1, which have the advantages of high purity, good solid-state stability, good powder properties and mechanical stability. More specifically, the present invention provides 7 crystalline forms of compound 1, namely, crystalline form I, crystalline form II, crystalline form IIIA, crystalline form IIIB, crystalline form IV, crystalline form V and VI, wherein crystalline forms I and IIIA show better solid-state stability, mechanical stability, hygroscopicity and light stability than other crystalline forms or amorphous forms, thereby facilitating pharmaceutical processing.
  • the crystalline forms I and IIIA of compound 1 exhibit one or more of the following advantages over the amorphous form:
  • the solid-state stability results showed that Form I did not undergo any crystal transformation or purity reduction after being placed in a closed container at 60°C for 1 day, or in an open container at 25°C/60%RH or 40°C/75%RH for 1 week, indicating that Form I was stable in the evaluation of
  • the crystal form IIIA has good physical and chemical stability under the following conditions; the purity of the crystal form IIIA remains unchanged after one day at 60°C, but the diffraction peak of the crystal form I is observed; after one week at 25°C/60%RH and 40°C/75%RH, no crystal form transformation or purity reduction occurs; the purity of the amorphous form does not change significantly under the three evaluation conditions, but the crystal form transforms into the crystal form I;
  • Form I dissolved slowly in all media (different pH values), and its solubility in 1M HCl was slightly higher than that in other pH buffers, thus ensuring that the dissolution of Form I in the body is more stable, and it is easier to obtain in vivo pharmacokinetics with stable blood drug concentrations, which is conducive to avoiding the risk of drug safety caused by excessive fluctuations in blood drug concentrations.
  • Form I has higher exposure and blood concentration than Form IIIA, indicating the potential to reduce the dosage.
  • Form I exhibited better light stability and no crystal transformation occurred under all evaluation conditions, while the amorphous form transformed into Form I after DVS, solid-state stability, solubility, and mechanical stability tests.
  • the present invention provides a polymorph of formula (I)
  • n 0 or 1
  • X is acetonitrile, water, 1,4-dioxane, ethanol, methanol, dimethylformamide, acetone or a mixture thereof.
  • n is 0. In some embodiments, n is 1; and X is acetonitrile, water, 1,4-dioxane, ethanol, or dimethylformamide.
  • n is 0, and the polymorph is Form I, characterized in that the X-ray powder diffraction pattern of Form I includes the following characteristic diffraction peaks at 2 ⁇ positions: 16.71° ⁇ 0.2°, 21.82° ⁇ 0.2°, and 23.75° ⁇ 0.2°, using Cu-K ⁇ radiation.
  • the X-ray powder diffraction pattern of Form I also includes the following characteristic diffraction peaks at 2 ⁇ positions: 7.48° ⁇ 0.2°, 22.36° ⁇ 0.2°.
  • the Form I The X-ray powder diffraction pattern also includes characteristic diffraction peaks at the following 2 ⁇ positions: 5.3° ⁇ 0.2°, 24.57° ⁇ 0.2°, and 27.08° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form I also includes characteristic diffraction peaks at the following 2 ⁇ positions: 11.81° ⁇ 0.2°, 15.83° ⁇ 0.2°, 17.92° ⁇ 0.2°, 18.95° ⁇ 0.2°, and 19.17° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form I includes the following characteristic diffraction peaks at 2 ⁇ positions: 5.30° ⁇ 0.2°, 7.48° ⁇ 0.2°, 11.81° ⁇ 0.2°, 14.85° ⁇ 0.2°, 15.83° ⁇ 0.2°, 16.71° ⁇ 0.2°, 17.92° ⁇ 0.2°, 18.95° ⁇ 0.2°, 19.17° ⁇ 0.2°, 19.43° ⁇ 0.2°, 21.14° ⁇ 0.2° , 21.82° ⁇ 0.2°, 22.36° ⁇ 0.2°, 23.75° ⁇ 0.2°, 24.57° ⁇ 0.2°, 27.08° ⁇ 0.2°, 27.83° ⁇ 0.2°, 28.88° ⁇ 0.2°, 31.20° ⁇ 0.2°, 31.92° ⁇ 0.2°, 32.40° ⁇ 0.2°, 33.91° ⁇ 0.2°, 35.83° ⁇ 0.2°, 37.51° ⁇ 0.2° and 39.04° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of Form I is substantially as shown in Fig. 3.
  • Form I has a TGA graph and/or DSC graph as shown in Fig. 4.
  • Form I is an anhydrate.
  • n is 1
  • X is acetonitrile
  • the polymorph is Form II, characterized in that the X-ray powder diffraction pattern of Form II includes the following characteristic diffraction peaks at 2 ⁇ positions: 24.99° ⁇ 0.2°, 26.05° ⁇ 0.2°, and 22.6° ⁇ 0.2°, using Cu-K ⁇ radiation.
  • the X-ray powder diffraction pattern of Form II also includes the following characteristic diffraction peaks at 2 ⁇ positions: 6.35° ⁇ 0.2°, 20.34° ⁇ 0.2°, 22.41° ⁇ 0.2°, and 28.71° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form II further includes characteristic diffraction peaks at the following 2 ⁇ positions: 9.18° ⁇ 0.2°, 16.03° ⁇ 0.2°, 18.25° ⁇ 0.2°, 27.07° ⁇ 0.2°, 29.08° ⁇ 0.2°, and 33.93° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form II further includes characteristic diffraction peaks at the following 2 ⁇ positions: 14.44° ⁇ 0.2°, 24.64° ⁇ 0.2°, 26.41° ⁇ 0.2°, 32.27° ⁇ 0.2°, 32.68° ⁇ 0.2°, 37.07° ⁇ 0.2°, and 39.51° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form II includes the following characteristic diffraction peaks at 2 ⁇ positions: 6.35° ⁇ 0.2°, 9.18° ⁇ 0.2°, 9.91° ⁇ 0.2°, 14.44° ⁇ 0.2°, 16.03° ⁇ 0.2°, 18.25° ⁇ 0.2°, 19.77° ⁇ 0.2°, 20.34° ⁇ 0.2°, 21.81° ⁇ 0.2°, 22.41° ⁇ 0.2°, 22.60° ⁇ 0.2°, 23.84° ⁇ 0.2°, 24.64° ⁇ 0.2°, 24.99° ⁇ 0.2°, 25.43 0.2°, 34.19° ⁇ 0.2°, 35.42° ⁇ 0.2°, 37.07° ⁇ 0.2°, 37.56° ⁇ 0.2°, 38.69° ⁇ 0.2° and 39.51° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of Form II is substantially as shown in Fig. 6.
  • the crystalline Form II has a TGA graph and/or a DSC graph as shown in Fig. 7.
  • n is 0, and the polymorph is Form IIIA, characterized in that the X-ray powder diffraction pattern of Form IIIA includes the following characteristic diffraction peaks at 2 ⁇ positions: 6.59° ⁇ 0.2°, 22.69° ⁇ 0.2°, 20.32° ⁇ 0.2°, 23.62° ⁇ 0.2°, 23.91° ⁇ 0.2°, and 24.15° ⁇ 0.2°, using Cu-K ⁇ radiation.
  • the X-ray powder diffraction pattern of Form IIIA also includes the following characteristic diffraction peaks at 2 ⁇ positions: 10.8° ⁇ 0.2°, 17.14° ⁇ 0.2°, 13.75° ⁇ 0.2°, 21.59° ⁇ 0.2°, and 26.01° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form IIIA further includes characteristic diffraction peaks at the following 2 ⁇ positions: 18.71° ⁇ 0.2°, 21.97° ⁇ 0.2°, 25.54° ⁇ 0.2°, 27.13° ⁇ 0.2°, 27.59° ⁇ 0.2° and 30.51° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form IIIA comprises characteristic diffraction peaks at the following 2 ⁇ positions: 6.59° ⁇ 0.2°, 9.9° ⁇ 0.2°, 10.8° ⁇ 0.2°, 13.09° ⁇ 0.2°, 13.75° ⁇ 0.2°, 17.14° ⁇ 0.2°, 17.87° ⁇ 0.2°, 18.71° ⁇ 0.2°, 19.19° ⁇ 0.2°, 20.32° ⁇ 0.2°, 21.59° ⁇ 0.2° °, 21.97° ⁇ 0.2°, 22.69° ⁇ 0.2°, 23.62° ⁇ 0.2°, 23.91° ⁇ 0.2°, 24.15° ⁇ 0.2°, 25.54° ⁇ 0.2°, 26.01° ⁇ 0.2°, 27.13° ⁇ 0.2°, 27.59° ⁇ 0.2°, 28.83° ⁇ 0.2°, 29.24° ⁇ 0.2°, 30.51° ⁇ 0.2°, 31.13° ⁇ 0.2°, 31.79° ⁇ 0.2°, 33.6° ⁇ 0.2°, 34.14° ⁇ 0.2°, 36.08° ⁇ 0.2°, 36
  • the X-ray powder diffraction pattern of the crystalline form IIIA is substantially as shown in Fig.9.
  • the crystalline form IIIA has a TGA graph and/or DSC graph as shown in Fig.10.
  • the crystalline form IIIA is an anhydrate.
  • n is 0, and the polymorph is Form IIIB, characterized in that the X-ray powder diffraction pattern of Form IIIB includes the following characteristic diffraction peaks at 2 ⁇ positions: 6.53° ⁇ 0.2°, 13.69° ⁇ 0.2°, 18.6° ⁇ 0.2°, 20.19° ⁇ 0.2°, 21.52° ⁇ 0.2°, and 22.64° ⁇ 0.2°, using Cu-K ⁇ radiation.
  • the X-ray powder diffraction pattern of Form IIIB also includes the following characteristic diffraction peaks at 2 ⁇ positions: 10.75° ⁇ 0.2°, 17.07° ⁇ 0.2°, 21.93° ⁇ 0.2°, 26.13° ⁇ 0.2°, 23.57° ⁇ 0.2°, and 30.46° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form IIIB also includes characteristic diffraction peaks at the following 2 ⁇ positions: 13.05° ⁇ 0.2°, 16.63° ⁇ 0.2°, 20.82° ⁇ 0.2°, 24.01° ⁇ 0.2°, 27.55° ⁇ 0.2°, and 31.79° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form IIIB includes the following characteristic diffraction peaks at 2 ⁇ positions: 6.53° ⁇ 0.2°, 10.75° ⁇ 0.2°, 12.62° ⁇ 0.2°, 13.05° ⁇ 0.2°, 13.69° ⁇ 0.2°, 16.63° ⁇ 0.2°, 17.07° ⁇ 0.2°, 18.60° ⁇ 0.2°, 19.59° ⁇ 0.2°, 20.19° ⁇ 0.2°, 20.82° ⁇ 0.2°, 21.
  • the X-ray powder diffraction pattern of the crystalline form IIIB is substantially as shown in Fig. 13.
  • the crystalline form IIIB has a TGA graph and/or a DSC graph as shown in Fig. 14.
  • n is 1
  • X is 1,4-dioxane
  • the polymorph is Form IV, characterized in that the X-ray powder diffraction pattern of Form IV includes the following characteristic diffraction peaks at 2 ⁇ positions: 8.56° ⁇ 0.2°, 13.29° ⁇ 0.2°, 17.69° ⁇ 0.2°, 19.75° ⁇ 0.2°, and 22.45° ⁇ 0.2°, using Cu-K ⁇ radiation.
  • the X-ray powder diffraction pattern of Form IV also includes the following characteristic diffraction peaks at 2 ⁇ positions: 5.26° ⁇ 0.2°, 18.29° ⁇ 0.2°, 31.83° ⁇ 0.2°, 25.68° ⁇ 0.2°, 22.86° ⁇ 0.2°, 32.81° ⁇ 0.2°, and 23.44° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form IV further includes characteristic diffraction peaks at the following 2 ⁇ positions: 126.57° ⁇ 0.2°, 27.52° ⁇ 0.2°, 35.69° ⁇ 0.2°, 21.09° ⁇ 0.2°, 20.35° ⁇ 0.2° and 31.43° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form IV comprises characteristic diffraction peaks at the following 2 ⁇ positions: 5.26° ⁇ 0.2°, 8.56° ⁇ 0.2°, 9.85° ⁇ 0.2°, 13.29° ⁇ 0.2°, 17.69° ⁇ 0.2°, 18.29° ⁇ 0.2°, 19.75° ⁇ 0.2°, 20.35° ⁇ 0.2°, 21.09° ⁇ 0.2°, 22.45° ⁇ 0.2°, 22.86
  • the crystalline Form IV has an X-ray powder diffraction pattern substantially as shown in Fig. 17.
  • the crystalline Form IV has a TGA graph and/or a DSC graph as shown in Fig. 18.
  • n is 1
  • X is ethanol
  • the polymorph is Form V, characterized in that the X-ray powder diffraction pattern of Form V includes the following characteristic diffraction peaks at 2 ⁇ positions: 6.21° ⁇ 0.2°, 8.47° ⁇ 0.2°, 15.62° ⁇ 0.2°, 21.73° ⁇ 0.2°, 25.53° ⁇ 0.2°, 25.94° ⁇ 0.2° and 28.05° ⁇ 0.2°, using Cu-K ⁇ radiation.
  • the X-ray powder diffraction pattern of Form V also includes the following characteristic diffraction peaks at 2 ⁇ positions: 9.61° ⁇ 0.2°, 17.55° ⁇ 0.2°, 19.25° ⁇ 0.2°, 22.22° ⁇ 0.2°, 23.12° ⁇ 0.2°, 32.92° ⁇ 0.2° and 34.22° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the crystalline form V further includes characteristic diffraction peaks at the following 2 ⁇ positions: 9.06° ⁇ 0.2°, 20.07° ⁇ 0.2°, 28.49° ⁇ 0.2°, 30.21° ⁇ 0.2°, 31.25° ⁇ 0.2°, In some embodiments, the X-ray powder diffraction pattern of the crystalline form V includes the following characteristic diffraction peaks at 2 ⁇ positions: 6.21° ⁇ 0.2°, 8.47° ⁇ 0.2°, 25.94° ⁇ 0.2°, 15.62° ⁇ 0.2°, 25.53° ⁇ 0.2°, 28.05° ⁇ 0.2°, 21.73° ⁇ 0.2°, 17.55° ⁇ 0.2°, 32.92° ⁇ 0.2°, 23.12° ⁇ 0.2°, 22.22° ⁇ 0.2°, 19.25° ⁇ 0.2°, 34.22° ⁇ 0.2°, 9.61 0.2°, 35.47° ⁇ 0.2°, 28.49° ⁇ 0.2°, 30.21° ⁇ 0.2°, 20.07° ⁇ 0.2°, 39.78°
  • n is 1
  • X is dimethylformamide
  • the polymorph is Form VI.
  • the X-ray powder diffraction pattern of Form VI is substantially as shown in Fig. 23.
  • Form VI has a TGA graph and/or DSC graph as shown in Fig. 24.
  • Form I, Form II, Form IIIA, Form IIIB, Form IV, Form V and Form VI each have a purity of about 85% or more, for example, about 90% or more, for example, about 95% or more, for example, about 97% or more, for example, about 99% or more and including about 99.9% or more, based on the weight of Compound 1, as determined by HPLC (high performance liquid chromatography).
  • the remaining materials may contain a crystalline form of Compound 1 and/or reaction impurities and/or processing impurities produced by its preparation, such as photodegradation impurity compound (A).
  • a mixture of Form I of Compound 1 with other solid forms e.g., other crystalline forms and amorphous is also within the scope of the present disclosure.
  • the polymorph of Compound 1 is substantially free of impurity (A). In some embodiments, the polymorph of Compound 1 contains less than 0.15% by weight impurity (A), such as 0.10%, 0.09%, 0.08%, 0.07%, 0.06%, 0.05%, 0.04%, 0.03%, 0.02% or 0.01% by weight impurity (A) relative to the polymorph.
  • Form I of Compound 1 is substantially free of impurity (A). In some embodiments, Form I of Compound 1 contains less than 0.15% by weight of impurity (A), such as 0.10%, 0.09%, 0.08%, 0.07%, 0.06%, 0.05%, 0.04%, 0.03%, 0.02% or 0.01% by weight of impurity (A) relative to Form I.
  • Form I, Form II, Form IIIA, Form IIIB, Form IV, Form V, and Form VI each have a crystallinity of about 85% or more, such as about 90% or more, such as about 95% or more, such as about 97% or more, such as about 99% or more and including about 99.9% or more.
  • the present invention provides a pharmaceutical composition comprising any polymorph of Form I to Form VI of the present invention, and a pharmaceutically acceptable carrier and/or excipient.
  • the pharmaceutical composition comprises Form I or IIIA of the present invention, and a pharmaceutically acceptable carrier and/or excipient.
  • the pharmaceutical composition comprises Form I of the present invention, and a pharmaceutically acceptable carrier and/or excipient.
  • the present invention provides a method for treating tumors, chronic inflammatory diseases, inflammatory bowel diseases, skin diseases, diabetes, eye diseases, diseases associated with angiogenesis or angiogenesis in mammals, diseases associated with chronic pain, and other diseases modulated by the Mek cascade in mammals, the method comprising administering to the mammal any one of the polymorphs of Form I to Form VI of the present invention; the present invention provides any one of the polymorphs of Form I to Form VI of the present invention, which is used to treat tumors, chronic inflammatory diseases, inflammatory bowel diseases, skin diseases, diabetes, eye diseases, diseases associated with angiogenesis or angiogenesis in mammals, diseases associated with chronic pain, and other diseases modulated by the Mek cascade in mammals; any one of the polymorphs of Form I to Form VI of the present invention is used in the preparation of a method for treating tumors, chronic inflammatory diseases, inflammatory bowel diseases, skin diseases, diabetes, eye diseases, diseases associated with angiogenesis or angiogenesis in mammals, diseases associated with chronic pain, and other diseases modulated
  • the present invention provides a method for treating a mammalian RAS or RAF mutant cancer, the method comprising administering 4-fluoro-5-(2-fluoro-4-iodophenylamino)-1H-benzo[d]thiazole-6-carboxylic acid (2-hydroxy-ethoxy)-amide (Compound 1) or a pharmaceutically acceptable salt thereof to the mammal.
  • the compound 1 is any polymorph of Form I to Form VI.
  • the RAS or RAF mutant cancer is, for example, a KRAS mutant cancer, a NRAS mutant cancer, a HRAS mutant cancer, or a BRAF mutant cancer.
  • the RAS mutant cancer is pancreatic cancer, colorectal cancer, lung cancer, melanoma, acute myeloid leukemia, bladder cancer, or head and neck cancer, etc.
  • the cancer is a NRAS mutant cancer.
  • the NRAS mutant cancer is a melanoma with RNAS mutations.
  • KRAS comprises a mutation at one or more positions selected from codons 12, 13, 59, and 61.
  • the KRAS mutant form has a mutation at one or more amino acid positions selected from G12, G13, S17, P34, A59, and Q61.
  • the KRAS mutant form has one or more amino acid substitutions selected from the group consisting of G12C, G12S, G12R, G12F, G12L, G12N, G12A, G12D, G12V, G13C, G13S, G13D, G13V, G13P, S17G, P34S, A59E, A59G, A59T, Q61K, Q61L, Q61R, and Q61H.
  • the mutant form of KRAS has a mutation at one or more amino acid positions selected from G12, G13, A59, Q61, K117, and A146.
  • the mutant form of KRAS has one or more amino acid substitutions selected from the group consisting of G12C, G12R, G12S, G12A, G12D, G12V, G13C, G13R, G13S, G13A, G13D, G13V, A59E, A59G, A59T, Q61K, Q61L, Q61R, Q61H, K117N, K117R, K117E, A146P, A146T, and A146V.
  • the BRAF mutation is a BRAF V600E mutation.
  • NRAS includes a mutation at one or more positions selected from codons 12, 13, 59, 61, and 146.
  • the NRAS mutant form has a mutation at one or more amino acid positions selected from G12, G13, A59, Q61, K117, and A146.
  • the NRAS mutant form has one or more amino acid substitutions selected from the group consisting of G12C, G12R, G12S, G12A, G12D, G12V, G13C, G13R, G13S, G13A, G13D, G13V, A59D, A59T, Q61K, Q61L, Q61R, Q61H, K117N, K117R, K117E, A146P, A146T, and A146V.
  • the cancer is an early, intermediate or advanced stage cancer.
  • the cancer may be locally advanced or metastatic.
  • the mammal has previously received immunotherapy.
  • the mammal has previously received immunotherapy and has an advanced melanoma with a NRAS mutation.
  • the melanoma is selected from: advanced melanoma, unresectable melanoma, metastatic melanoma, melanoma with a BRAF mutation, melanoma with a NRAS mutation, cutaneous melanoma, or intraocular melanoma.
  • compound 1 is in the form of tablets, powders, granules, patches, inhalants, capsules. In some embodiments, compound 1 is in the form of capsules. In some embodiments, compound 1 is administered at a dose of 5-50 mg each time, once or twice a day. In some embodiments, compound 1 is administered at a dose of 12 mg each time, twice a day.
  • the present invention provides a method for preparing the crystalline form I of the present invention, the method comprising any of the following:
  • the heating temperature in method 1) is about 75°C to about 100°C; about 80°C to about 90°C; or about 75°C or about 85°C.
  • the room temperature in method 1) is about 20-about 25°C.
  • the stirring time in method 1) is about 0.5-12 hours, about 1-12 hours, about 1-8 hours, about 1-5 hours or longer, or the stirring time is about 24-about 96 hours.
  • the clear liquid in method 1) is cooled to room temperature within about 2 to about 5 hours or about 2.5 to about 3 hours. In some embodiments, after the room temperature is kept warm and stirred in method 1), the temperature can be further lowered to about 0-about 10°C, and the stirring is kept warm.
  • the stirring is kept warm at about 0-about 10°C for about 1-about 12 hours, about 1-about 8 hours, about 1-about 5 hours, or at about 0-about 10°C for longer stirring as appropriate.
  • the solvent in method 1) is water, methanol, ethanol, isopropanol, acetone, methyl isobutyl ketone, 2-butanone, ethyl acetate, isopropyl acetate, methyl tert-butyl ether, tetrahydrofuran, anisole, 2-methyltetrahydrofuran, cyclopentyl methyl ether, 1,4-dioxane, acetonitrile, dichloromethane, toluene, m-xylene, n-heptane, n-hexane, n-pentane, dimethyl sulfoxide, dimethylacetamide, N-methylpyrrolidone or a mixture thereof.
  • the solvent in method 1) is water, m
  • the good solvent in method 2) is a solvent in which compound 1 is soluble
  • the anti-solvent in method 2) is a solvent in which compound 1 is not soluble.
  • the good solvent in method 2) is MEK, 1,4-dioxane or DMSO.
  • the anti-solvent in method 2) is MTBE, EtOAc, CHCl3, n-heptane, Anisole, EtOAc, H2O , IPAc, CPME, DCM or toluene.
  • the cooling rate of method 3) is about 0.1°C/min.
  • the solvent of method 3) is MIBK, Methyl acetate, 2-MeTHF or acetone/EtOH (1:1).
  • the present invention provides a method for preparing the crystalline form V of the present invention, the method comprising: adding a sample of amorphous compound 1 to a solvent, then heating at a temperature below about 70°C, cooling the resulting clear solution to about 0-about 10°C, standing at about 5°C until solids precipitate, and drying.
  • the method is heated at about 70°C, about 60°C, or about 50°C.
  • the cooling rate is about 0.1-about 0.5°C/min.
  • the cooling rate is about 0.1°C/min.
  • the resulting clear solution is cooled to about 5°C.
  • the present invention provides a method for preparing the crystal form IIIA of the present invention, the method comprising: heating the crystal form II to a first temperature and keeping the temperature constant for about 3 to about 10 minutes, and then cooling to obtain the crystal form IIIA.
  • the first temperature is about 100 to about 140°C; for example, about 110 to about 130°C, for example, about 120°C.
  • the crystal form II is kept constant at the first temperature for about 5 minutes.
  • the crystal form II is cooled to room temperature to about 50°C after being kept constant, for example, cooled to room temperature.
  • the crystalline compound of the present invention has a DSC graph with a characteristic peak position, has substantially the same properties as the DSC graph provided in the accompanying drawings of the present invention, and the measurement value error tolerance is within ⁇ 5°C, generally required to be within ⁇ 3°C.
  • the numerical values described and protected by the present invention are approximate values. Variations in the numerical values may be due to equipment calibration, equipment errors, crystal purity, crystal size, sample size and other factors.
  • crystal forms of the present invention are not limited to characteristic spectra that are completely identical to the characteristic spectra described in the accompanying drawings disclosed in the present invention, such as XRPD, DSC, TGA, DVS, and isothermal adsorption curves. Any crystal forms having characteristic spectra that are substantially the same or essentially the same as those described in the accompanying drawings fall within the scope of the present invention.
  • “Therapeutically effective amount” refers to the amount of a compound that elicits a physiological or medical response in a tissue, system, or subject that is sought, including an amount of a compound that, when administered to a subject, is sufficient to prevent the occurrence of one or more symptoms of the disorder or condition being treated or to alleviate them to some degree.
  • the “therapeutically effective amount” may vary with the compound, the disease, disorder, and/or symptoms of the disease or condition, the severity of the disease or condition, and/or symptoms of the disease or condition, the age of the subject to be treated, and/or the weight of the subject to be treated. The appropriate amount in any given case will be apparent to those skilled in the art or can be determined by routine experimentation.
  • a “therapeutically effective amount” refers to the total amount of the combined subject that is effective in treating the disease, disorder, or condition.
  • Excipient refers to a substance that is not a therapeutic agent itself but is used as a diluent, adjuvant, binder and/or vehicle and is added to a pharmaceutical composition to improve its handling or storage properties or to allow or facilitate the formation of a compound or pharmaceutical composition into a unit dosage form for administration.
  • Crystal or “crystal” or “polymorph” refers to any solid material that exhibits a three-dimensional ordering, in contrast to an amorphous solid material, which produces a characteristic XRPD pattern with well-defined peaks.
  • Amorphous refers to a non-crystalline molecular and/or ionic solid form. Amorphous solids do not exhibit an X-ray diffraction pattern with sharp maxima.
  • Hydrate refers to a crystalline form of a molecule that further comprises water incorporated into the crystalline structure.
  • the water molecules in a hydrate may exist in a regular arrangement and/or a disordered arrangement. Hydrates may contain stoichiometric or non-stoichiometric amounts of water molecules.
  • “Anhydrate” refers to a crystalline form that does not substantially contain any form of water molecules, for example, a crystalline form that does not substantially contain water molecules in the crystal lattice or unit cell.
  • solvate refers to a crystalline form of a molecule, which further comprises one or more solvent molecules incorporated into the crystalline structure.
  • the solvent molecules in the solvate can exist in a regular arrangement and/or a disordered arrangement.
  • the solvate can contain stoichiometric or non-stoichiometric amounts of solvent molecules.
  • Exemplary solvates include, but are not limited to, hydrates, ethanolates, methoxides, and isopropoxides, acetic acid. Methods of solvation are generally known in the art. It is noteworthy that in a solvate, the substance combined with the main molecule (e.g., active pharmaceutical ingredient) is liquid at room temperature, while in a eutectic, the substance is solid at room temperature.
  • the crystal form disclosed in the present application is a substantially pure crystal.
  • substantially pure used in the present application refers to at least 85% by weight, preferably at least 95% by weight, more preferably at least 99% by weight of the crystal form disclosed in the present application, and also includes the like. About 100% by weight of a certain crystalline form.
  • the remaining material includes other one or more forms of the compound and/or reaction impurities and/or processing impurities resulting from its preparation.
  • a crystalline form of Compound 1 can be considered to be substantially pure because it has a purity of greater than 90% by weight, as measured by means known and generally accepted in the art at the time, wherein the remaining less than 10% by weight of the material comprises an amorphous form of Compound 1 and/or other one or more forms and/or reaction impurities and/or processing impurities. .
  • X-ray powder diffraction pattern refers to an experimentally observed diffraction pattern or a parameter, data or value derived therefrom.
  • XRPD patterns are usually characterized by peak positions (abscissa) and/or peak intensities (ordinate).
  • peak positions abcissa
  • peak intensities ordinate
  • main peaks i.e., the most characteristic, significant, unique and/or repeatable peaks
  • main peaks can be repeated within the error limit (within ⁇ 2 of the last decimal place given, or within ⁇ 0.2 of the given value).
  • 2 ⁇ refers to the peak position expressed in degrees (°) based on the setup in the X-ray diffraction experiment, and is usually the unit of the abscissa in the diffraction pattern. If the reflection is diffracted when the incident beam forms an angle ⁇ with a certain lattice plane, the experimental setup requires recording the reflected beam at an angle of 2 ⁇ . It should be understood that the specific 2 ⁇ value of a specific crystal form mentioned in this application is intended to represent the 2 ⁇ value (expressed in degrees) measured using the X-ray diffraction experimental conditions described in this application.
  • the pharmaceutical composition comprising the compound disclosed in the present application can be administered orally, by inhalation, rectally, parenterally or topically to a subject in need thereof.
  • the pharmaceutical composition may be a regular solid preparation such as tablets, powders, granules, capsules, etc., a liquid preparation such as water or oil suspension or other liquid preparations such as syrups, solutions, suspensions, etc.; for parenteral administration, the pharmaceutical composition may be a solution, an aqueous solution, an oil suspension concentrate, a lyophilized powder, etc.
  • the formulation of the pharmaceutical composition is selected from tablets, coated tablets, capsules, suppositories, nasal sprays or injections, more preferably tablets or capsules.
  • the pharmaceutical composition may be administered as a single unit with a precise dose.
  • the pharmaceutical composition may further comprise additional active ingredients.
  • compositions disclosed in the present application can be prepared by conventional methods in the pharmaceutical field.
  • the active ingredient can be mixed with one or more excipients and then the desired preparation can be prepared.
  • “Pharmaceutically acceptable excipients” refer to conventional pharmaceutical carriers suitable for the desired pharmaceutical preparation, for example: diluents, vehicles such as water, various organic solvents, etc., fillers such as starch, sucrose, etc., binders such as cellulose derivatives, alginates, gelatin and polyvinyl pyrrolidone (PVP); wetting agents such as glycerol; disintegrants such as agar, calcium carbonate and sodium bicarbonate; absorption enhancers such as quaternary ammonium compounds; surfactants such as hexadecanol; absorption carriers such as kaolin and soap clay; lubricants such as talc, calcium stearate, magnesium stearate, polyethylene glycol, etc.
  • the pharmaceutical composition further comprises other pharmaceutically acceptable excipient
  • “Pharmaceutical composition” refers to a composition comprising a crystalline form of a compound of the invention and at least one additional pharmaceutically acceptable carrier.
  • “Pharmaceutically acceptable carrier” refers to a medium generally accepted in the art for delivering a biologically active agent to an animal, particularly a mammal, including, i.e., adjuvants, excipients or vehicles, such as diluents, preservatives, fillers, flow regulators, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, fragrances, antibacterial agents, antifungal agents, lubricants and dispensing agents, depending on the nature of the mode of administration and dosage form.
  • Pharmaceutically acceptable carriers are formulated according to a number of factors within the purview of those of ordinary skill in the art. These include, but are not limited to: the type and nature of the active agent being formulated; the subject to whom the composition containing the agent is to be administered; the intended route of administration of the composition; and the therapeutic indication being targeted. Pharmaceutically acceptable carriers include both aqueous and non-aqueous liquid media, as well as a variety of solid and semisolid dosage forms. Such carriers may also include a number of different ingredients and additives in addition to the active agent, such additional ingredients being included in the formulation for a variety of reasons well known to those of ordinary skill in the art (e.g., stabilization of the active agent, binder, etc.).
  • the dosage regimen of the solid form of the present application will vary according to known factors such as the following: the pharmacodynamic characteristics of a particular agent and its mode and route of administration; the species, age, sex, health, medical condition and weight of the recipient; the nature and extent of symptoms; the type of simultaneous treatment; the frequency of treatment; the route of administration, the patient's renal and liver function and the desired effect.
  • the daily oral dosage range of each active ingredient will be between about 0.001 to about 5000 mg/day, preferably between about 0.01 to about 1000 mg/day, and most preferably between 0.1 to about 250 mg/day.
  • Intravenously, during a constant rate infusion the most preferred dosage range will be from about 0.01 to about 10 mg/kg/minute.
  • the compounds of the present invention can be administered in a single daily dose, or the total daily dose can be administered in divided doses twice, three times or four times a day.
  • Dosage forms for administration may contain from about 1 mg to about 2000 mg of active ingredient per dosage unit.
  • the active ingredient will generally be present in an amount of about 0.1%-95% by weight based on the total weight of the composition.
  • Fig. 1 is the XRPD pattern of amorphous compound 1
  • Fig. 2 shows the dynamic water sorption (DVS) of amorphous compound 1
  • Fig. 6 is the XRPD pattern of Form II of Examples 3a and 3b;
  • Fig.7 is the TGA/DSC diagram of Form II of Example 3a;
  • Fig.8 is the 1 H NMR graph of Form II of Example 3a;
  • Fig. 9 is the XRPD diagram of Form IIIA of Example 4a.
  • Fig. 10 is the TGA/DSC diagram of Form IIIA of Example 4a;
  • Fig. 11 is the 1 H NMR graph of Form IIIA of Example 4a;
  • Fig. 12 is an XRPD overlay of Form IIIA of Example 4a before and after heating;
  • Fig. 13 is the XRPD diagram of Form IIIB of Example 4b;
  • Fig. 14 is the TGA/DSC diagram of Form IIIB of Example 4b;
  • Fig. 15 is the 1 H NMR graph of Form IIIB of Example 4b;
  • Fig. 16 is a temperature-dependent XRPD diagram of Form IIIB of Example 4b;
  • Fig. 17 is the XRPD pattern of Form IV of Examples 5a and 5b;
  • Fig. 18 is the TGA/DSC diagram of Form IV of Example 5a;
  • Fig. 19 is the 1 H NMR graph of Form IV of Example 5a;
  • Fig. 20 is the XRPD pattern of Form V of Example 6a;
  • Fig. 21 is the TGA/DSC diagram of Form V of Example 6a;
  • Fig.22 is the 1 H NMR graph of Form V of Example 6a;
  • Fig. 23 is the XRPD pattern of Form VI of Example 7a;
  • Fig. 24 is the DSC/TGA diagram of Form VI of Example 7a;
  • Fig. 25 is the XRPD diagram of Form IV of Example 5a before and after heating;
  • Fig. 26 is an XRPD overlay of Form II of Example 3a before and after heating;
  • Fig. 27 is the XRPD diagram of Form II of Example 3b before and after heating;
  • Fig. 28 is the XRPD diagram of Form V of Example 6a before and after heating
  • Fig. 29 is an XRPD overlay of the competition between Form I and IIIA in EtOAc
  • Fig. 30 is an XRPD overlay of the competition between Form I and IIIA in MIBK;
  • Fig.31 is the XRPD superposition of the competition between Form I and IIIA in acetone/H 2 O (I/II);
  • Fig.32 is the XRPD overlay of the competition between Form I and IIIA in acetone/H 2 O (II/II);
  • Fig. 33 is the DVS diagram of Form I
  • Fig. 34 is the XRPD superposition of Form I before and after DVS testing
  • Fig. 35 is the XRPD superposition of the amorphous sample before and after DVS testing
  • Fig.36 is the DVS diagram of Form IIIA in the moisture induction experiment
  • Fig. 37 is the XRPD superposition of Form IIIA before and after DVS testing
  • Fig. 38 is an XRPD overlay of Example 2a Form I before and after solid state stability evaluation
  • Fig. 39 is the XRPD overlay before and after the amorphous solid state stability evaluation
  • Fig. 40 is an XRPD overlay of Form IIIA before and after solid-state stability evaluation
  • Fig.41 is the solubility diagram of amorphous at different pH values (feeding concentration 0.25 mg/mL);
  • Fig.42 is the solubility diagram of Form I at different pH values (feeding concentration 0.25 mg/mL);
  • Fig.43 is the solubility diagram of amorphous and crystalline form I at different pH values (feeding concentration 0.05 mg/mL);
  • Fig. 44 is an XRPD overlay of Form I before and after grinding
  • Fig.45 is the XRPD superposition of Form I before and after tableting
  • Fig.46 is the XRPD superposition of Form IIIA before and after grinding
  • Fig. 47 is the XRPD superposition of Form IIIA before and after tableting
  • Fig. 48 is the XRPD superposition of amorphous before and after grinding
  • Fig. 49 is the XRPD superposition of amorphous before and after tableting
  • Fig. 50 is an XRPD overlay diagram for evaluating the photostability of Form I;
  • Fig.51 is an XRPD overlay diagram for evaluating the photostability of Form IIIA
  • Fig.52 is an XRPD overlay for evaluating the photostability of amorphous samples
  • Fig.53 is an XRPD overlay diagram for evaluating the light stability of Form V
  • Fig.54 is a HPLC chart for evaluating the light stability of Form I
  • Fig.55 is a HPLC chart for evaluating the photostability of Form IIIA
  • Fig.56 is a HPLC chart for evaluating the photostability of amorphous samples.
  • Fig.57 is the HPLC chart for evaluating the photostability of Form V.
  • X-ray powder diffraction XRPD results were collected on X'Pert 3 and Empyrean X-ray powder diffraction analyzers, and the scanning parameters are shown in Table 1.
  • TGA and DSC were collected on a TA Discovery 5500 thermogravimetric analyzer and a TA Discovery 2500 differential scanning calorimeter, respectively. Table 2 lists the TGA and DSC test parameters.
  • Dynamic moisture sorption (DVS) Dynamic moisture sorption (DVS) curves were collected on the DVS Intrinsic of SMS (Surface Measurement Systems). The relative humidity at 25°C was corrected using the deliquescent point of LiCl, Mg(NO 3 ) 2 and KCl. The DVS test parameters are listed in Table 3.
  • Example 1a (Compound 1 is amorphous, repeat Example 1 of Chinese Patent Application No. 201210190520.4): According to the method of Example 1 of Chinese Patent Application No. 201210190520.4, the crude product obtained in step 13 of Example 1 is separated and purified by silica gel column chromatography using dichloromethane/methanol as eluent, and then rotary evaporation is performed to obtain a white solid, which is amorphous by XRPD analysis, and its XRPD pattern is shown in Fig. 1, and the dynamic water sorption (DVS) is shown in Fig. 2.
  • Example 2a Add 4 mL of anhydrous ethanol to about 21 mg of the amorphous sample of Example 1a, heat and stir in an 85°C oil bath for 5 minutes to dissolve; then, cool the resulting clear solution to room temperature (25°C) over 2.5 hours with stirring, continue stirring at room temperature for 24 hours, filter out the resulting solid sample, and perform XRPD analysis.
  • the XRPD results are shown in Fig.3.
  • the TGA/DSC results (Fig.4) show that the sample loses 0.77% of its weight when heated to 150°C, and there is a sharp endothermic peak at 221.3°C (starting temperature).
  • 1H NMR data were collected using DMSO-d6 as solvent, and the results are shown in Fig.5. No obvious solvent residue was detected. According to the characterization results of Form I, it has a relatively The small and gentle weight loss (lower than the theoretical water content of hemihydrate 1.80%) and the single melting endothermic signal indicate that Form I is an anhydrate or anhydrous crystalline form.
  • the XRPD diffraction peak data of Form I are shown in Table 6.
  • Example 2b About 20 mg of each amorphous sample was weighed and added to a 20-mL vial, 0.4-1.0 mL of a good solvent (see Table 7) was added to dissolve, filtered (PTFE filter with a pore size of 0.45 ⁇ m) to obtain a clear solution, and the anti-solvent in Table 7 was added while stirring the clear solution until solids were precipitated, and the precipitated solids were separated by centrifugation and tested by XRPD. The results are shown in Table 7. The anti-solvent was added to obtain a solid, which was determined to be Form I by XRPD.
  • Example 2c Weigh about 20 mg of each amorphous sample of Example 1a into a 5-mL or HPLC vial, add 0.7-4.0 mL of the solvent in Table 8, stir at 50°C for about 2 hours, filter (PTFE filter membrane with a pore size of 0.45 ⁇ m) to take the filtrate, place the obtained filtrate in a biochemical incubator, cool it to 5°C at a cooling rate of 0.1°C/min, collect the precipitated solid and perform XRPD test. The test results are shown in Table 8. The slow cooling test obtained a solid, which was determined to be Form I by XRPD.
  • Example 3a Weigh 71.6 mg of Example 2a Form I sample into a 3 mL vial, add 1.5 mL ACN, and stir magnetically at room temperature for about 1 day. Centrifuge and separate the solid, place it under ambient conditions ( ⁇ 21°C/45% RH) for about 4 hours to dry, and obtain the final product, which is then tested by XRPD and TGA/DSC.
  • the XRPD results are shown in Fig.6.
  • the TGA results are shown in Fig.7.
  • TGA shows that the sample has a step-wise weight loss of 5.90% when heated to 120°C.
  • the DSC results show that there are two endothermic peaks at 94.1°C and 220.9°C (starting temperature) and one exothermic peak at 191.6°C (peak temperature).
  • the 1 H NMR results are shown in Fig.8.
  • the molar ratio of ACN to API in the sample is 0.8 (5.9 wt%, consistent with the TGA weight loss). Based on the characterization and heating experiment results, it is speculated that the crystal Form II is an ACN solvate, which undergoes desolvation and crystallization after heating.
  • the XRPD diffraction peak data of Form II are shown in Table 9.
  • Example 3b Weigh 20.4 mg of Example 2a Form I sample into a 3 mL vial, add 2 mL ACN/acetone (1:1, v/v), and filter (0.45 ⁇ m PTFE filter membrane) after ultrasound to obtain a clear solution. Seal with a sealing film and pierce 4 small holes, place in a fume hood and slowly evaporate for 5 days. Solid precipitation was observed, and after being placed under ambient conditions overnight, it was transferred to room temperature and vacuum dried for about 1 day. The ACN/acetone (1:1, v/v) solution of Example 2a Form I was slowly evaporated, and after the solid was precipitated, it was transferred to room temperature and vacuum dried to obtain the final product, and XRPD test was performed on it. The XRPD results are shown in Fig. 6.
  • Example 3c About 19.7 mg of the amorphous sample obtained in Example 1a was weighed into a HPLC vial, and 0.5 mL of ACN/H 2 O (19:1) solvent was added to obtain a suspension, which was placed at 5° C. and magnetically stirred ( ⁇ 1000 rpm) for about 6 days, and then the solid was separated by centrifugation and subjected to XRPD testing.
  • the XRPD result was Form II.
  • Example 3d About 19.9 mg of the amorphous sample obtained in Example 1a was weighed into an HPLC vial, and 0.5 mL of ACN/acetone (1:1) solvent was added to obtain a suspension. The suspension was placed at 5°C and magnetically stirred ( ⁇ 1000 rpm) for about 6 days, and then the solid was separated by centrifugation and subjected to XRPD testing. The XRPD result was Form II.
  • Example 4a The crystal form II sample of Example 3a was heated to 120°C by DSC and kept at this temperature for 5 minutes before cooling to room temperature to obtain crystal form IIIA.
  • the XRPD results are shown in Fig.9.
  • the TGA/DSC results (Fig.10) show that the sample loses 0.26% of its weight when heated to 200°C, which is lower than the theoretical water content of the hemihydrate (1.80%); the DSC results show that there is an exothermic signal at 200.3°C (peak temperature) and a sharp endothermic signal at 219.6°C (starting temperature).
  • 1 H NMR results (Fig.11) show that no obvious residual solvent is detected.
  • Crystal form IIIA is heated to 205°C, cooled to room temperature and exposed to ambient conditions and then transformed into crystal form I.
  • the XRPD results before and after heating are shown in Fig.12. According to the characterization results of crystal form IIIA, it is inferred that crystal form IIIA is an anhydrate, and the exothermic signal at 198.8°C is a thermal signal for the transformation to crystal form I.
  • the XRPD diffraction peak data of the crystal form IIIA are shown in Table 10.
  • Example 4b Weigh 100.0 mg of the Form I sample of Example 2a into a 20 mL vial, add 10 mL of THF to dissolve, and filter (0.45 ⁇ m PTFE filter) to obtain a clear solution. The filtrate was rotary evaporated at 50°C and the final product was collected. The XRPD results are shown in Fig. 13.
  • the TGA/DSC results show that the sample loses 3.49% weight when heated to 200°C; the DSC results show that there are a weak endothermic signal and an exothermic signal at 121.3°C and 144.6°C (peak temperature), respectively, and a strong endothermic signal at 218.3°C (starting temperature).
  • 1 H NMR results Fig.
  • Example 5a About 20 mg of the amorphous sample obtained in Example 1a was weighed into a HPLC vial, and 0.5 mL of 1,4-dioxane solvent was added. The resulting suspension was placed at room temperature and magnetically stirred ( ⁇ 1000 rpm) for about 6 days, and then the solid was separated by centrifugation and vacuum dried at room temperature overnight to obtain the final product, which was then subjected to XRPD and TGA/DSC tests. The XRPD and TGA/DSC results are shown in Fig. 17 and
  • Example 5b The Form I sample (500 mg) of Example 2a was stirred in 1,4-Dioxane (12.5 mL) at room temperature for 2 days, and the obtained solid was dried under vacuum at room temperature for 1 day.
  • the XRPD results are shown in Fig. 17.
  • Example 6a Weigh 20.8 mg of the Form I sample of Example 2a into a 5 mL vial and add 4 mL of EtOH. Stir at 70°C to dissolve, cool to 5°C within 650 minutes (cooling rate 0.1°C/min) and stand at 5°C. After removing the solution, the solid was left open to dry under ambient conditions (temperature: ⁇ 21°C, humidity: ⁇ 36% RH).
  • XRPD results are shown in Fig. 20.
  • TGA/DSC results (Fig. 21) show that the sample has a step-wise weight loss of 7.64% when heated to 120°C, and has two endothermic peaks at 103.1°C and 223.3°C (starting temperature). 1 H NMR results are shown in Fig.
  • Form V is an EtOH solvate, which is desolvated after heating to transform into anhydrous Form I.
  • the XRPD diffraction peak data of Form V are shown in Table 13.
  • Example 1a The amorphous sample (1 g) of Example 1a was dissolved in DMF (3 mL). After complete dissolution, IPA (6 mL) was added and stirred at room temperature overnight to precipitate a solid, which was filtered and the filter cake was washed with isopropanol and dried in vacuo to obtain a DMF solvate crystalline form, which is Form VI.
  • the XRPD results of Form VI are shown in Fig. 23.
  • the DSC results show that there are two endothermic peaks at 147.39°C and 208.56°C.
  • the TGA results in Fig. 24 show that the weight loss is 0.67% when heated to 180°C.
  • Example 8a Example 5a Form IV was heated to 120°C, cooled to room temperature and exposed to ambient conditions before conversion. It becomes Form I, and the XRPD results are shown in Fig. 25.
  • Example 8b The crystal form II of Example 3a was heated to 120°C and 210°C, respectively, cooled to room temperature and exposed to ambient conditions. The XRPD results are shown in Fig. 26. When heated to 120°C, it was transformed into the crystal form IIIB, and when heated to 210°C, it was transformed into the crystal form I.
  • Example 8c Form II of Example 3b was heated to 120°C, cooled to room temperature and exposed to ambient conditions before transforming into Form I.
  • the XRPD results are shown in Fig. 27. It is speculated that the sample may also transform into Form IIIB first after heating, but transform into Form I more quickly under ambient conditions, and the XRPD test shows that it has transformed into Form I.
  • Example 8d The crystal form IIIB of Example 4a was identified by variable temperature XRPD, and the results are shown in Fig. 16. After N2 purging at 30°C for 20 minutes, the crystal form remained unchanged. When heated to 120°C under N2 protection, the diffraction peak of crystal form I was observed. When heated to 170°C, it was mainly crystal form I, with only a small amount of diffraction peak of crystal form IIIB. It was cooled to 30°C under N2 protection and did not continue to change.
  • Example 8e The Form V of Example 6a was heated to 120°C, cooled to room temperature and exposed to ambient conditions to transform into Form I. The XRPD results are shown in Fig. 28.
  • This example involves suspension competition experiments of Form I and Form IIIA in EtOAc and MIBK at room temperature and 50° C., and suspension competition experiments in acetone/H 2 O of different water activities at room temperature.
  • Example 2a Form I sample was added into an HPLC bottle, add 1 mL of the corresponding solvent, and stir at the corresponding temperature for 4 hours or overnight. Filter (0.45 ⁇ m PTFE filter membrane) to obtain a saturated solution.
  • Example 2 Form I and Example 4a Form IIIA samples about 5 mg each
  • Stir at the corresponding temperature and perform XRPD testing on the solid wet sample cover with Kapton film to avoid potential crystallization due to solvent volatilization during the test).
  • Example 2a Form I The hygroscopicity of Example 2a Form I, Example 1a Amorphous and Example 4a Form IIIA was evaluated by a dynamic moisture sorption (DVS) test at 25°C.
  • DVD dynamic moisture sorption
  • the DVS results of amorphous are shown in Fig. 2.
  • Fig. 35 The XRPD comparison results show that the amorphous sample transforms into crystal form I after the DVS test.
  • the DVS result of Form IIIA is shown in Fig. 36. At 25°C/80%RH, the water absorption increased by 0.060%, indicating that it is almost non-hygroscopic.
  • the XRPD comparison result shows that Form IIIB did not undergo a crystal transformation after the DVS test.
  • Example 2a The crystalline form I of Example 2a and the amorphous form of Example 1a were subjected to different pH conditions (1M HCl and pH The dynamic solubility in 2.0/4.5/6.8/7.4 buffer was tested. The specific steps are as follows:
  • the powder properties of the crystalline form I of Example 2a and the amorphous sample of Example 1a were evaluated, including the angle of repose, bulk density and tap density, in order to understand the powder flow properties of the crystalline form I and the amorphous form.
  • Bulk density and tap density Add a certain mass of the sample to be evaluated into a 5-mL measuring cylinder and record the volume at this time. The bulk density is calculated by dividing the mass of the sample by the volume at this time; tap the measuring cylinder 200 times and record the final volume. The tap density is calculated by dividing the mass of the sample by the final volume. Each parameter is tested three times in parallel.
  • Angle of repose Fix the funnel vertically to the bottom surface and slowly add the material into the funnel.
  • the bottom surface forms a symmetrical cone of the material. Measure the height of the cone and the bottom diameter. Perform the measurement three times in parallel.
  • Carr's index (tapped density - bulk density) / tapped density.
  • Example 1a The crystal form I of Example 2a, the crystal form IIIA of Example 4a and the amorphous form of Example 1a were manually ground and tableted (350MPa pressure), and the samples after grinding and tableting were subjected to XRPD testing to evaluate their mechanical stability.
  • the XRPD results are shown in Fig.44, Fig.45, Fig.46, Fig.47, Fig.48 and Fig.49.
  • crystal form I was selected for hygroscopicity, solid-state stability, solubility, powder properties and mechanical stability evaluation, and the same evaluation was carried out on the amorphous form to compare the properties with crystal form I.
  • the DVS results showed that crystal form I had almost no hygroscopicity and no crystal form transformation occurred after the DVS test; the amorphous form transformed into crystal form I after the DVS test.
  • the solid-state stability results showed that crystal form I did not undergo crystal form transformation or purity reduction after being placed in a closed container at 60°C for 1 day, 25°C/60%RH and 40°C/75%RH for 1 week, indicating that crystal form I has good physical and chemical stability under the evaluation conditions; the purity of the amorphous form did not change significantly under the three evaluation conditions, but it was transformed into crystal form I.
  • the test results of the powder properties of crystal form I showed that crystal form I and amorphous samples had similar fluidity.
  • the mechanical stability results showed that after tableting (350 MPa) and manual grinding (about 3 minutes), Form I did not undergo a crystal form transformation and the crystallinity did not decrease significantly, and the amorphous form transformed into Form I.
  • Form I did not undergo a crystal transformation under all evaluation conditions, while the amorphous form transformed into Form I after DVS, solid-state stability, solubility, and mechanical stability tests, indicating that Form I is superior to amorphous form I.
  • the shape has better physical stability.
  • Example 1a For the crystal form I of Example 2a, the crystal form IIIA of Example 4a, the crystal form V of Example 6a and the amorphous form of Example 1a, some samples were taken out at 6 hours and 24 hours respectively under illumination (white light 5800-5890 Lux + ultraviolet 7.9-8.7 W/m 2 ) for XRPD and HPLC purity tests and stability evaluation.
  • the XRPD results are shown in Fig. 50, Fig. 51, Fig. 52 and Fig.
  • the crystal form I, the crystal form IIIA, the crystal form V and the amorphous form did not undergo crystal transformation after being placed under illumination for 24 hours; under light-proof reference conditions, the crystal form I, the crystal form IIIA and the crystal form V did not undergo crystal transformation after being placed for 24 hours, and the amorphous form was observed to have a weak diffraction peak of the crystal form I after being placed for 6 hours and 24 hours.
  • the collected whole blood was placed in an EDTA-K2 anticoagulant tube and fully mixed, centrifuged (1500-1600 g) for 10 min, and the plasma was separated for biological analysis.
  • concentration of the test substance in the plasma samples was determined by LC-MS/MS analysis (instrument model: Triple Quad 5500; chromatographic column: Agilent ZORBAX XDB-C18; flow rate: 0.50 mL/min; injection volume: 2 ⁇ L; mobile phase A: water [0.1% formic acid + 5 mM ammonium acetate], mobile phase B: acetonitrile [0.1% formic acid]).
  • the corresponding pharmacokinetic parameters (Table 20) were calculated using the non-compartmental model in Pharsight Phoenix 8.3.
  • Form I and Form IIIA are 9820 ng ⁇ h/mL and 8890 ng ⁇ h/mL, respectively, and the C max is 1160 ng/mL and 1030 ng/mL, respectively, indicating that there is sufficient drug plasma exposure in vivo and has good pharmacokinetic properties.
  • Form I has higher exposure and blood drug concentration than Form IIIA, which is reflected in the potential to reduce the dosage.
  • Example 16 Inhibition of proliferation of NRAS mutated melanoma cell lines
  • the human melanoma cell line SK-MEL-2 (NRAS Q61R) was cultured with MEM + 10% FBS + 1% penicillin-streptomycin at 37 degrees Celsius in an incubator with 5% carbon dioxide concentration.
  • the human melanoma cell line HMVII (NRAS Q61K) was cultured with F12K + 10% FBS + 1% penicillin-streptomycin at 37 degrees Celsius in an incubator with 5% carbon dioxide concentration.
  • the adherent cells were digested with trypsin and the cell pellets were collected by centrifugation and counted.
  • 90 ⁇ L of cell suspension was inoculated in a 96-well plate at an appropriate density.
  • a series of gradient dilutions of the compound (crystalline form I) concentration range 0.15nM-10 ⁇ M, 4-fold gradient dilution
  • Three replicate wells were set for each concentration, with a total of 9 concentration points. The wells with the same volume of 5% DMSO were used as controls, and the final concentration of DMSO was 0.5%.
  • Example 17 Inhibition of RAS or RAF mutations
  • the specific method is as follows: for adherent cells, discard the culture medium, add 20 ⁇ L MTS and 100 ⁇ L cell culture medium to each well, and directly add 20 ⁇ L MTS to the suspended cells. Place in an incubator and continue to culture for 1-4 hours, detect OD490, and use OD650 value as a reference. GraphPad Prism software was used to prepare a dose-effect curve and calculate IC 50. The results are shown in Table 23.
  • Embodiment 18 is a diagrammatic representation of Embodiment 18:
  • a multicenter, single-arm Phase II study in patients with advanced melanoma with NRAS mutations evaluated the efficacy and safety of Compound 1 Form I. Patients received 12 mg of this product orally twice daily until intolerable toxicity, disease progression (assessed by the investigator according to RECIST 1.1), withdrawal of consent, death, or at the discretion of the investigator. The trial will be terminated when the risk outweighs the benefit. As of February 19, 2023, a total of 100 subjects were enrolled, of which 95.0% (95/100 subjects) were included in the full analysis set (FAS).
  • FAS full analysis set
  • Compound 1 has a good anti-tumor therapeutic effect on patients with advanced melanoma with NRAS mutations.
  • the ORR evaluated by IRRC is 35.8%, and the efficacy data is significantly better than the clinical research data of similar drugs.

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Abstract

本发明涉及蛋白激酶Mek抑制剂苯并噻唑化合物的多晶型物、其制备方法，以及其医药用途。

Description

作为蛋白激酶Mek抑制剂的多晶型物、及其制备方法和用途 技术领域
本发明涉及蛋白激酶Mek抑制剂苯并噻唑化合物的多晶型物、其制备方法，以及其医药用途。
背景技术
Ras/Raf/Mek/Erk信号传导机制的过度活化在癌细胞的增殖和分化中起了重要作用；已经在多种癌症中发现Ras/Raf/Mek/Erk信号传导机制被连续的活化或过度的活化，如胰腺癌、结肠癌、肺癌、膀胱癌、肾癌、皮肤癌、乳腺癌等等。抑制Ras/Raf/Mek/Erk信号传导途径有助于对这类过度增殖性疾病的治疗，其中位于Ras和Raf下游靶的Mek在该途径中起着关键的作用，Mek磷酸化的底物是MAP激酶Erk。如果Mek被抑制，则Ras/Raf/Mek/Erk信号传导途径就会被关闭，从而癌细胞的增殖就会被抑制。因此，Mek抑制剂可以抑制癌细胞的增长，尤其是对于Ras或Raf过度活化导致的癌症。与此同时Mek也涉及炎症类的疾病和症状，包括急性和慢性炎症。
中国专利申请No.201210190520.4披露了众多苯并噻唑化合物，其表现出蛋白激酶Mek抑制活性，其中包括化合物4-氟-5-(2-氟-4-碘苯基氨基)-1H-苯并[d]噻唑-6-羧酸(2-羟基-乙氧基)-酰胺(以下简称化合物1)。但是，按照中国专利申请No.201210190520.4的制备方法，化合物1的固体形态为无定形。然而，无定形的纯度通常难以控制，物理和化学稳定性通常不佳，引湿性差。而且，化合物1光照不稳定，容易降解生成杂质(A)
从而影响化合物1本身的功效和存储稳定性。
因此有必要开发该化合物的新型晶型，有利于提高工艺生产中的纯度控制，提高化合物的稳定性和耐储存性。
发明内容
本发明提供了化合物1的多晶型物，所述多晶型物具有高纯度、良好的固态稳定性、良好的粉体学性质和机械力稳定性等优点。更具体而言，本发明提供了化合物1的7种晶型，即晶型I、晶型II、晶型IIIA、晶型IIIB、晶型IV、晶型V和VI，其中晶型I和IIIA相对于其它晶型或无定形呈现出了更好的固态稳定性、机械力稳定性、引湿性和光稳定性等性能，从而有利于制药工艺。
具体而言，化合物1的晶型I和IIIA相比于无定形呈现出以下一种或多种优势：
1.混悬竞争结果表明室温至50℃范围内晶型I比晶型IIIA热力学上更稳定；
2.DVS结果显示晶型I和晶型IIIA几乎无引湿性，在DVS测试后均未发生晶型转变；无定形在DVS测试后转变为晶型I；
3.固态稳定性结果显示，晶型I在60℃下闭口放置1天、25℃/60％RH和40℃/75％RH下敞口放置1周后均未发生晶型转变或纯度降低，表明晶型I在评估 条件下具有较好的物理和化学稳定性；晶型IIIA在60℃下1天后纯度不变但观察到晶型I的衍射峰，在25℃/60％RH和40℃/75％RH下1周后未发生晶型转变或纯度降低；无定形在三种评估条件下纯度均未明显变化，但晶型均转变为晶型I；
4.光照稳定性结果表明，晶型I在光照(白光5890Lux+紫外8.7W/m2)条件下24小时后，纯度明显高于晶型IIIA、晶型V和无定形，表现出更好的光照稳定性；
5.晶型I和无定形在室温下1M HCl及pH 1.0/2.0/4.5/7.4缓冲液中的动态溶解度测试结果显示，无定形在10分钟内在所有介质中溶解度均高于晶型I，且均呈现先上升再下降的过程，在1M HCl中测得的最高溶解度高于其它pH缓冲液；相比于无定形，晶型I在各介质(不同pH值)中均缓慢溶解，在1M HCl中溶解度略高于其余pH缓冲液，从而确保晶型I在体内的溶出更稳定，更易得到血药浓度平稳的体内药代动力学，有利于避免因血药浓度波动太大导致的用药安全性风险；
6.粉体学性质测试结果表明，晶型I和无定形样品具有相似的流动性，而且机械力稳定性结果显示，在压片(350MPa)及手动研磨(约3分钟)后，晶型I未发生晶型转变且结晶度无明显降低，无定形转变为晶型I，晶型IIIA未发生晶型转变但结晶度降低；和/或
7.在药代动力学实验中，晶型I比晶型IIIA更高的暴露量和血药浓度，表现在有潜力降低给药剂量。
根据表征数据和评估结果，晶型I表现出更好的光照稳定性，且在所有评估条件下均未发生晶型转变，而无定形在DVS、固态稳定性、溶解度、机械力稳定性测试后转变为晶型I。
第一方面，本发明提供式(I)的多晶型物
其中n为0或1，X为乙腈、水、1,4-二氧六环(1,4-Dioxane)、乙醇、甲醇、二甲基甲酰胺、丙酮或其混合物。
在一些实施方案中，n为0。在一些实施方案中，n为1；X为乙腈、水、1,4-二氧六环、乙醇或二甲基甲酰胺。
在一些实施方案中，n为0，所述多晶型物为晶型I，其特征在于所述晶型I的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：16.71°±0.2°、21.82°±0.2°和23.75°±0.2°，采用Cu-Kα辐射。在一些实施方案中，所述晶型I的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：7.48°±0.2°、22.36°±0.2°。在一些实施方案中，所述晶型I的 X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：5.3°±0.2°、24.57°±0.2°和27.08°±0.2°。在一些实施方案中，所述晶型I的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：11.81°±0.2°、15.83°±0.2°、17.92°±0.2°、18.95°±0.2°和19.17°±0.2°。在一些实施方案中，所述晶型I的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：5.30°±0.2°、7.48°±0.2°、11.81°±0.2°、14.85°±0.2°、15.83°±0.2°、16.71°±0.2°、17.92°±0.2°、18.95°±0.2°、19.17°±0.2°、19.43°±0.2°、21.14°±0.2°、21.82°±0.2°、22.36°±0.2°、23.75°±0.2°、24.57°±0.2°、27.08°±0.2°、27.83°±0.2°、28.88°±0.2°、31.20°±0.2°、31.92°±0.2°、32.40°±0.2°、33.91°±0.2°、35.83°±0.2°、37.51°±0.2°和39.04°±0.2°。在一些实施方案中，所述晶型I的X-射线粉末衍射图谱基本上如Fig.3所示。在一些实施方案中，所述晶型I具有如Fig.4所示的TGA图和/或DSC图。在一些实施方案中，所述晶型I为无水合物。
在一些实施方案中，n为1，X为乙腈，所述多晶型物为晶型II，其特征在于所述晶型II的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：24.99°±0.2°、26.05°±0.2°和22.6°±0.2°，采用Cu-Kα辐射。在一些实施方案中，所述晶型II的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：6.35°±0.2°、20.34°±0.2°、22.41°±0.2°和28.71°±0.2°。在一些实施方案中，所述晶型II的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：9.18°±0.2°、16.03°±0.2°、18.25°±0.2°、27.07°±0.2°、29.08°±0.2°和33.93°±0.2°。在一些实施方案中，所述晶型II的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：14.44°±0.2°、24.64°±0.2°、26.41°±0.2°、32.27°±0.2°、32.68°±0.2°、37.07°±0.2°和39.51°±0.2°。在一些实施方案中，所述晶型II的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.35°±0.2°、9.18°±0.2°、9.91°±0.2°、14.44°±0.2°、16.03°±0.2°、18.25°±0.2°、19.77°±0.2°、20.34°±0.2°、21.81°±0.2°、22.41°±0.2°、22.60°±0.2°、23.84°±0.2°、24.64°±0.2°、24.99°±0.2°、25.43°±0.2°、26.05°±0.2°、26.41°±0.2°、27.07°±0.2°、28.71°±0.2°、29.08°±0.2°、29.81°±0.2°、31.16°±0.2°、31.57°±0.2°、32.27°±0.2°、32.68°±0.2°、33.93°±0.2°、34.19°±0.2°、35.42°±0.2°、37.07°±0.2°、37.56°±0.2°、38.69°±0.2°和39.51°±0.2°。在一些实施方案中，所述晶型II的X-射线粉末衍射图谱基本上如Fig.6所示。在一些实施方案中，所述晶型II具有如Fig.7所示的TGA图和/或DSC图。
在一些实施方案中，n为0，所述多晶型物为晶型IIIA，其特征在于所述晶型IIIA的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.59°±0.2°、22.69°±0.2°、20.32°±0.2°、23.62°±0.2°、23.91°±0.2°和24.15°±0.2°，采用Cu-Kα辐射。在一些实施方案中，所述晶型IIIA的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：10.8°±0.2°、17.14°±0.2°、13.75°±0.2°、21.59°±0.2°和26.01°±0.2°。在一些实施方案中，所述晶型IIIA的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：18.71°±0.2°、21.97°±0.2°、25.54°±0.2°、27.13°±0.2°、27.59°±0.2°和30.51°±0.2°。在一些实施方案中，所述晶型IIIA的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.59°±0.2°、9.9°±0.2°、10.8°±0.2°、13.09°±0.2°、13.75°±0.2°、17.14°±0.2°、17.87°±0.2°、18.71°±0.2°、19.19°±0.2°、20.32°±0.2°、21.59°±0.2°、21.97°±0.2°、22.69°±0.2°、23.62°±0.2°、23.91°±0.2°、24.15°±0.2°、25.54°±0.2°、26.01°±0.2°、27.13°±0.2°、27.59°±0.2°、28.83°±0.2°、29.24°±0.2°、30.51°±0.2°、31.13°±0.2°、31.79°±0.2°、 33.6°±0.2°、34.14°±0.2°、36.08°±0.2°、36.67°±0.2°和37.26°±0.2°。在一些实施方案中，所述晶型IIIA的X-射线粉末衍射图谱基本上如Fig.9所示。在一些实施方案中，所述晶型IIIA具有如Fig.10所示的TGA图和/或DSC图。在一些实施方案中，所述晶型IIIA为无水合物。
在一些实施方案中，n为0，所述多晶型物为晶型IIIB，其特征在于所述晶型IIIB的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.53°±0.2°、13.69°±0.2°、18.6°±0.2°、20.19°±0.2°、21.52°±0.2°和22.64°±0.2°，采用Cu-Kα辐射。在一些实施方案中，所述晶型IIIB的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：10.75°±0.2°、17.07°±0.2°、21.93°±0.2°、26.13°±0.2°、23.57°±0.2°和30.46°±0.2°。在一些实施方案中，所述晶型IIIB的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：13.05°±0.2°、16.63°±0.2°、20.82°±0.2°、24.01°±0.2°、27.55°±0.2°、31.79°±0.2°。在一些实施方案中，所述晶型IIIB的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.53°±0.2°、10.75°±0.2°、12.62°±0.2°、13.05°±0.2°、13.69°±0.2°、16.63°±0.2°、17.07°±0.2°、18.60°±0.2°、19.59°±0.2°、20.19°±0.2°、20.82°±0.2°、21.52°±0.2°、21.93°±0.2°、22.64°±0.2°、23.57°±0.2°、24.01°±0.2°、25.46°±0.2°、26.13°±0.2°、27.55°±0.2°、30.46°±0.2°、31.04°±0.2°、31.79°±0.2°、32.81°±0.2°、33.54°±0.2°、34.06°±0.2°和34.46°±0.2°。在一些实施方案中，所述晶型IIIB的X-射线粉末衍射图谱基本上如Fig.13所示。在一些实施方案中，所述晶型IIIB具有如Fig.14所示的TGA图和/或DSC图。
在一些实施方案中，n为1，X为1,4-二氧六环，所述多晶型物为晶型IV，其特征在于所述晶型IV的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：8.56°±0.2°、13.29°±0.2°、17.69°±0.2°、19.75°±0.2°和22.45°±0.2°，采用Cu-Kα辐射。在一些实施方案中，所述晶型IV的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：5.26°±0.2°、18.29°±0.2°、31.83°±0.2°、25.68°±0.2°、22.86°±0.2°、32.81°±0.2°和23.44°±0.2°。在一些实施方案中，所述晶型IV的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：126.57°±0.2°、27.52°±0.2°、35.69°±0.2°、21.09°±0.2°、20.35°±0.2°和31.43°±0.2°。在一些实施方案中，所述晶型IV的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：5.26°±0.2°、8.56°±0.2°、9.85°±0.2°、13.29°±0.2°、17.69°±0.2°、18.29°±0.2°、19.75°±0.2°、20.35°±0.2°、21.09°±0.2°、22.45°±0.2°、22.86°±0.2°、23.44°±0.2°、24.44°±0.2°、25.68°±0.2°、26.57°±0.2°、27.52°±0.2°、28.40°±0.2°、29.78°±0.2°、31.43°±0.2°、31.83°±0.2°、32.81°±0.2°、34.29°±0.2°、35.69°±0.2°和37.72°±0.2°。在一些实施方案中，所述晶型IV的X-射线粉末衍射图谱基本上如Fig.17所示。在一些实施方案中，所述晶型IV具有如Fig.18所示的TGA图和/或DSC图。
在一些实施方案中，n为1，X为乙醇，所述多晶型物为晶型V，其特征在于所述晶型V的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.21°±0.2°、8.47°±0.2°、15.62°±0.2°、21.73°±0.2°、25.53°±0.2°、25.94°±0.2°和28.05°±0.2°，采用Cu-Kα辐射。在一些实施方案中，所述晶型V的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：9.61°±0.2°、17.55°±0.2°、19.25°±0.2°、22.22°±0.2°、23.12°±0.2°、32.92°±0.2°和34.22°±0.2°。在一些实施方案中，所述晶型V的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：9.06°±0.2°、20.07°±0.2°、28.49°±0.2°、30.21°±0.2°、31.25°±0.2°、 35.47°±0.2°和38.94°±0.2°。在一些实施方案中，所述晶型V的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.21°±0.2°、8.47°±0.2°、25.94°±0.2°、15.62°±0.2°、25.53°±0.2°、28.05°±0.2°、21.73°±0.2°、17.55°±0.2°、32.92°±0.2°、23.12°±0.2°、22.22°±0.2°、19.25°±0.2°、34.22°±0.2°、9.61°±0.2°、9.06°±0.2°、38.94°±0.2°、31.25°±0.2°、35.47°±0.2°、28.49°±0.2°、30.21°±0.2°、20.07°±0.2°、39.78°±0.2°、14.26°±0.2°、24.32°±0.2°、16.95°±0.2°、32.33°±0.2°、36.43°±0.2°、24.94°±0.2°、12.42°±0.2°和37.86°±0.2°。在一些实施方案中，所述晶型V的X-射线粉末衍射图谱基本上如Fig.20所示。在一些实施方案中，所述晶型V具有如Fig.21所示的TGA图和/或DSC图。
在一些实施方案中，n为1，X为二甲基甲酰胺，所述多晶型物为晶型VI，在一些实施方案中，所述晶型VI的X-射线粉末衍射图谱基本上如Fig.23所示。在一些实施方案中，所述晶型VI具有如Fig.24所示的TGA图和/或DSC图。
在一些实施方案中，晶型I、晶型II、晶型IIIA、晶型IIIB、晶型IV、晶型V和VI各自具有基于化合物1重量的约85％或更高，例如约90％或更高，例如约95％或更高，例如约97％或更高，例如约99％或更高并且包括约99.9％或更高的纯度，如通过HPLC(高效液相色谱法)确定的。其余材料可以包含化合物1的晶型和/或由其制备产生的反应杂质和/或加工杂质，如光降解杂质化合物(A)。化合物1的晶型I与其它固体形式(例如其它晶型和无定形)的混合物也在本申请公开的范围内。
在一些实施方案中，化合物1的多晶型物基本上不含有杂质(A)。在一些实施方案中，化合物1的多晶型物含有相对于多晶型物少于0.15重量％杂质(A)，例如0.10％，0.09％。0.08％，0.07％，0.06％，0.05％，0.04％，0.03％，0.02％或0.01％重量％杂质(A)。
在一些实施方案中，化合物1的晶型I基本上不含有杂质(A)。在一些实施方案中，化合物1的晶型I含有相对于晶型I少于0.15重量％杂质(A)，例如0.10％，0.09％。0.08％，0.07％，0.06％，0.05％，0.04％，0.03％，0.02％或0.01％重量％杂质(A)。
在一些实施方案中，晶型I、晶型II、晶型IIIA、晶型IIIB、晶型IV、晶型V和VI各自具有约85％或更高，例如约90％或更高，例如约95％或更高，例如约97％或更高，例如约99％或更高并且包括约99.9％或更高的结晶度。
第二方面，本发明提供一种药物组合物，其含有本发明晶型I至晶型VI中的任意一种多晶型物，以及药学上可接受的载体和/或赋形剂。在一些实施方案中，所述药物组合物包含本发明晶型I或IIIA，以及药学上可接受的载体和/或赋形剂。在一些实施方案中，所述药物组合物包含本发明晶型I，以及药学上可接受的载体和/或赋形剂。
第三方面，本发明提供治疗哺乳动物的肿瘤、慢性炎症疾病、炎症性肠道疾病、皮肤病、糖尿病、眼部疾病、与哺乳动物的血管发生或血管再生相关的疾病、与慢性疼痛相关的疾病和其它由Mek级联调制的疾病的方法，所述方法包括对所述哺乳动物给药本发明晶型I至晶型VI中的任一种多晶型物；本发明提供本发明晶型I至晶型VI中的任一种多晶型物，其用于治疗哺乳动物的肿瘤、慢性炎症疾病、炎症性肠道疾病、皮肤病、糖尿病、眼部疾病、与哺乳动物的血管发生或血管再生相关的疾病、与慢性疼痛相关的疾病和其它由Mek级联调制的疾病；本发明晶型I至晶型VI中的任一种多晶型物在制备用于治疗哺乳动物的肿瘤、慢性炎症疾病、炎症性肠道疾病、皮肤病、糖尿病、眼部疾病、与哺乳动物的血管发生或血管再生相关的疾病、与慢性疼痛相关的疾病和其它由 Mek级联调制的疾病中的用途。在一些实施方案中，所述哺乳动物为人类。
第四方面，本发明提供治疗哺乳动物RAS或RAF突变型癌症的方法，所述方法包括对所述哺乳动物给药4-氟-5-(2-氟-4-碘苯基氨基)-1H-苯并[d]噻唑-6-羧酸(2-羟基-乙氧基)-酰胺(化合物1)或其可药用盐。在一些实施方案中，所述化合物1为晶型I至晶型VI中的任一种多晶型物。在一些实施方案中，所述RAS或RAF突变型癌症为例如KRAS突变型癌症、NRAS突变型癌症、HRAS突变型癌症或BRAF突变癌症。在一些实施方案中，所述RAS突变型癌症是胰腺癌、结直肠癌、肺癌、黑色素瘤、急性骨髓性白血病、膀胱癌或头颈癌等。在优选的实施方案中，所述癌症是NRAS突变型癌症。在一些实施方案中，所述NRAS突变型癌症是RNAS突变的黑色素瘤。
在一实施方案中，KRAS包括在选自密码子12、13、59和61的一个或多个位置处的突变。在一实施方案中，KRAS突变形式在选自G12、G13、S17、P34、A59和Q61的一个或多个氨基酸位置处具有突变。在一实施方案中，所述KRAS突变形式具有选自以下的一个或多个氨基酸取代：G12C、G12S、G12R、G12F、G12L、G12N、G12A、G12D、G12V、G13C、G13S、G13D、G13V、G13P、S17G、P34S、A59E、A59G、A59T、Q61K、Q61L、Q61R和Q61H。在一实施方案中，所述KRAS突变形式在选自G12、G13、A59、Q61、K117和A146的一个或多个氨基酸位置处具有突变。在一实施方案中，所述KRAS突变形式具有选自以下的一个或多个氨基酸取代：G12C、G12R、G12S、G12A、G12D、G12V、G13C、G13R、G13S、G13A、G13D、G13V、A59E、A59G、A59T、Q61K、Q61L、Q61R、Q61H、K117N、K117R、K117E、A146P、A146T和A146V。在一实施方案中，所述BRAF突变为BRAF V600E突变。
在一实施方案中，NRAS包括在选自密码子12、13、59、61和146的一个或多个位置处的突变。在一些实施方案中，所述NRAS突变形式在选自G12、G13、A59、Q61、K117和A146的一个或多个氨基酸位置处具有突变。在一些实施方案中，所述NRAS突变形式具有选自以下的一个或多个氨基酸取代：G12C、G12R、G12S、G12A、G12D、G12V、G13C、G13R、G13S、G13A、G13D、G13V、A59D、A59T、Q61K、Q61L、Q61R、Q61H、K117N、K117R、K117E、A146P、A146T和A146V。
在一些实施方案中，所述癌症为早期、中期或晚期阶段癌症。癌症可以是局部晚期的或转移性的。在一些实施方案中，所述哺乳动物之前接受过免疫治疗。在一些实施方案中，所述哺乳动物之前接受过免疫治疗且患有NRAS突变的晚期黑色素瘤。在一些实施方案中，所述黑色素瘤选自：晚期黑色素瘤、不可切除的黑色素瘤、转移性黑色素瘤、具有BRAF突变的黑色素瘤、具有NRAS突变的黑色素瘤、皮肤黑色素瘤、或眼内黑色素瘤。
在一些实施方案中，化合物1为片剂、粉剂、颗粒剂、贴剂、吸入剂、胶囊剂的形式。在一些实施方案中，化合物1为胶囊剂的形式。在一些实施方案中，化合物1以每次5-50mg，每日一次或两次的剂量给药。在一些实施方案中，化合物1以每次12mg，每日两次的剂量给药。
第五方面，本发明提供制备本发明晶型I的方法，所述方法包括以下任一种：
a)将化合物1无定形样品加至溶剂中，然后在约70℃以上的温度加热，所得清液降温至室温，室温保温继续搅拌，固体析出，过滤，干燥；或
b)将化合物1无定形样品溶于良溶剂中，过滤得到澄清溶液，边搅拌该澄清溶液边 加入反溶剂至有固体析出；或
c)将化合物1无定形样品溶于溶剂中，在约50℃搅拌，然后过滤取滤液，将所得滤液降温至约5℃，收集析出的固体。
在一些实施方案中，方法1)中加热温度为约75℃至约100℃；约80℃至约90℃；或约75℃或约85℃。在一些实施方案中，方法1)中室温为约20-约25℃。在一些实施方案中，方法1)中继续搅拌的时间为约0.5-12小时、约1-12小时、约1-8小时、约1-5小时或更长，或者，继续搅拌的时间为约24-约96小时。在一些实施方案中，方法1)中清液在约2至约5小时内或约2.5-约3小时内降温至室温。在一些实施方案中，方法1)中室温保温继续搅拌后可任选地将温度进一步降低至约0-约10℃，保温搅拌。在一些实施方案中，在约0-约10℃保温搅拌约1-约12小时、约1-约8小时、约1-约5小时或者在约0-约10℃视情况保温搅拌更长。在一些实施方案中，方法1)中所述溶剂为水、甲醇、乙醇、异丙醇、丙酮、甲基异丁基酮、2-丁酮、乙酸乙酯、乙酸异丙酯、甲基叔丁基醚、四氢呋喃、苯甲醚、2-甲基四氢呋喃、环戊基甲醚、1,4-二氧六环、乙腈、二氯甲烷、甲苯、间二甲苯、正庚烷、正己烷、正戊烷、二甲亚砜、二甲基乙酰胺、N-甲基吡咯烷酮或其混合物。在一些实施方案中，方法1)中所述溶剂为乙醇。
在一些实施方案中，方法2)中的良溶剂为化合物1能溶于其中的溶剂，方法2)中的反溶剂为化合物1不能溶于其中的溶剂。在一些实施方案中，方法2)中的良溶剂为MEK、1,4-二氧六环或DMSO。在一些实施方案中，方法2)中的反溶剂为MTBE、EtOAc、CHCl3、正庚烷、Anisole、EtOAc、H2O、IPAc、CPME、DCM或甲苯。
在一些实施方案中，方法3)所述降温速度为约0.1℃/分钟。在一些实施方案中，方法3)的溶剂为MIBK、Methyl acetate、2-MeTHF或丙酮/EtOH(1:1)。
本发明提供制备本发明晶型V的方法，所述方法包括：将化合物1无定形样品加至溶剂中，然后在约70℃以下的温度加热，所得清液降温至约0-约10℃，在约5℃静置直至固体析出，干燥。在一些实施方案中，所述方法在约70℃、约60℃或约50℃加热。在一些实施方案中，所述降温速度为约0.1-约0.5℃/分。在一些实施方案中，所述降温速度为约0.1℃/分。在一些实施方案中，所得清液降温至约5℃。
本发明提供制备本发明晶型IIIA的方法，所述方法包括：将晶型II加热至第一温度并恒温约约3-约10分钟，然后冷却，得到晶型IIIA。在一些实施方案中，所述第一温度为约100-约140℃；例如约110-约130℃，例如约120℃。在一些实施方案中，将所述晶型II在第一温度恒温约5分钟。在一些实施方案中，将所述晶型II恒温后冷却至室温至约50℃，例如冷却至室温。
差示扫描量热(DSC)为本领域中所熟知的，DSC曲线的熔融峰高取决于与样品制备和仪器几何形状有关的许多因素，而峰位置对实验细节相对不敏感。因此，在一些实施方案中，本发明的结晶化合物具有特征峰位置的DSC图，具有与本发明附图中提供的DSC图实质上相同的性质，测量值误差容限为±5℃内，一般要求在±3℃内。
本发明描述的和保护的数值为近似值。数值内的变化可能归因于设备的校准、设备误差、晶体的纯度、晶体大小、样本大小以及其它因素。
本发明的晶型不限于与本发明公开的附图中描述的特征图谱完全相同的特征图谱，比如XRPD、DSC、TGA、DVS、等温吸附曲线图，具有与附图中描述的那些图谱基本上相同或本质上相同的特征图谱的任何晶型均落入本发明的范围内。
术语
除非在本文件中另外具体指定，否则本申请所使用的所有其它技术和科学术语具有本发明所属领域的技术人员通常理解的含义。
本申请包括权利要求使用的，单数形式的词语诸如“一”、“一种”和“所述”，包括它们对应的复数指代，除非上下文清楚地表明相反情况。因此，例如，提及“晶型”包括一个或多个这样不同的晶型并且提及“所述方法”包括提及本领域普通技术人员已知的等同步骤和方法，其可经改良或代替本申请所述的方法。
在以下整个说明书和权利要求书中，除非上下文有其它要求，否则术语“包含”及变体诸如“含有”和“包括”，将会被理解为隐含包括所述整数或步骤或者整数或步骤的集合，但是不排除任何其它整数或步骤或者整数或步骤的集合。当本申请使用时，术语"包含"可替换为术语“含有”，或有时在使用时可替换为术语“具有”。
术语“约”指指定项加上或减去值的10％或者5％或2％。
“治疗有效量”指引起组织、系统或受试者生理或医学反应的化合物的量，此量是所寻求的，包括在受治疗者身上施用时足以预防受治疗的疾患或病症的一种或几种症状发生或使其减轻至某种程度的化合物的量。“治疗有效量”可随化合物，疾病，病症，和/或疾病或病症的症状，疾病或病症的严重程度，和/或疾病或病症的症状，待治疗受试者的年龄，和/或待治疗受试者的重量而变化。在任何给定情况中的适当量对于本领域技术人员是显而易见的或者可通过常规实验确定。在联合疗法的情况中，“治疗有效量”是指有效治疗疾病、病症或病况的组合对象的总量。
“赋形剂”是指：其本身并非治疗剂，用作稀释剂、辅料、粘合剂和/或媒介物，用于添加至药物组合物中以改善其处置或储存性质或允许或促进化合物或药物组合物形成用于给药的单位剂型。
“晶型”或“晶体”或“多晶型物”是指呈现三维排序的任意固体物质，与无定型固体物质相反，其产生具有边界清楚的峰的特征性XRPD图谱。
“无定形”是指非结晶的分子和/或离子固体形式。无定形固体不显示具有尖锐最大值的X射线衍射图。
“水合物”是指分子的结晶形式，其进一步包含掺入结晶结构中的水。在水合物中的水分子可以以规则排列和/或无序排列存在。水合物可以包含化学计量量或非化学计量量的水分子。
“无水合物”是指晶体中基本上不含有任何形式的水分子，例如在晶体的晶格或晶胞中基本不不含有水分子的结晶形式。
术语“溶剂合物”是指分子的结晶形式，其进一步包含掺入结晶结构的一个或多个溶剂分子。在溶剂化物中的溶剂分子可以以规则排列和/或无序排列存在。溶剂化物可以包含化学计量量或非化学计量量的溶剂分子。示例性溶剂化物包括但不限于水合物、乙醇盐、甲醇盐、和异丙醇盐、乙酸。溶剂化的方法在本领域通常是已知的。值得注意的是，在溶剂化物中，与主要分子(例如，活性药物成分)结合的物质在室温下是液体，而在共晶中，所述物质在室温下是固体。
本申请公开的晶型为基本上纯的结晶。本申请使用的术语“基本上纯的”是指至少85重量％、优选至少95重量％、更优选至少99重量％的本申请公开的晶型，并且还包括等 于某种晶型的约100重量％。其余材料包括化合物的其它一种或多种形式和/或由其制备产生的反应杂质和/或加工杂质。例如，化合物1的结晶形式可以被认为是基本上纯的，因为它具有大于90重量％的纯度，如通过当时在本领域中已知和普遍接受的手段所测量的，其中其余的小于10重量％的材料包含化合物1的无定形和/或其它一种或多种形式和/或反应杂质和/或加工杂质。。
“X射线粉末衍射图谱(XRPD图谱)”是指实验观察的衍射图或源于其的参数、数据或值。XRPD图谱通常由峰位(横坐标)和/或峰强度(纵坐标)表征。对于本申请公开的晶型，仅总结了主峰(即，最具有特征性的、显著的、独特的和/或可重复的峰)；其它峰可通过常规方法从衍射图谱获得。上述主峰可在误差界限的范围内重复(在最后给出的小数位±2，或者在所给值±0.2)。
“2θ”是指基于X射线衍射实验中设置的以度数(°)表示的峰位，并且通常是在衍射图谱中的横坐标单位。如果入射束与某晶格面形成θ角时反射被衍射，则实验设置需要以2θ角记录反射束。应当理解，在本申请中提到的特定晶型的特定2θ值意图表示使用本申请所述的X射线衍射实验条件所测量的2θ值(以度数表示)。
对于X射线衍射峰的术语“基本上相同”或“基本如图XX所示”意指将代表性峰位和强度变化考虑在内。例如，本领域技术人员会理解峰位(2θ)会显示一些变化，通常多达0.1-0.2°，并且用于测量衍射的仪器也会导致一些变化。另外，本领域技术人员会理解相对峰强度会因仪器间的差异以及结晶性程度、择优取向、制备的样品表面以及本领域技术人员已知的其它因素而出现变化，并应将其看作仅为定性测量。
包含本申请公开的化合物的药物组合物可经口服、吸入、直肠、肠胃外或局部给予有此需要的受试者。对于口服给予，药物组合物可为规则固体制剂诸如片剂、粉剂、颗粒剂、胶囊剂等，液体制剂诸如水或油悬浮液或其它液体制剂诸如糖浆、溶液、悬浮液等；对于肠胃外给予，药物组合物可为溶液、水溶液、油悬浮液浓缩物、冻干粉末等。优选地，药物组合物的制剂选自片剂、包衣片剂、胶囊、栓剂、鼻喷雾或注射剂，更优选片剂或胶囊。药物组合物可为具有精确剂量的单个单元给予。此外，药物组合物可进一步包含额外的活性成分。
本申请公开的药物组合物的所有制剂都可通过制药领域的常规方法制备。例如，活性成份可与一种或多种赋形剂混合然后制备所需的制剂。“药学上可接受的赋形剂”是指适于所需药物制剂的常规药物载体，例如：稀释剂，媒介物诸如水、各种有机溶剂等，填充剂诸如淀粉、蔗糖等，粘合剂诸如纤维素衍生物、藻酸盐、明胶和聚乙烯吡咯烷酮(PVP)；润湿剂诸如甘油；崩解剂诸如琼脂、碳酸钙和碳酸氢钠；吸收增强剂诸如季铵化合物；表面活性剂诸如十六醇；吸收载体诸如高岭土和皂粘土；润滑剂诸如滑石粉、硬脂酸钙、硬脂酸镁、聚乙二醇等。此外，药物组合物进一步包含其它药学上可接受的赋形剂诸如分散剂(decentralized agent)、稳定剂、增稠剂、络合剂、缓冲剂、渗透促进剂、聚合物、芳香化合物、增甜剂和染料。
“药物组合物”是指包含本发明化合物的晶型与至少一种另外的药学上可接受的载体的组合物。“药学上可接受的载体”是指本领域中通常接受的用于将生物活性剂递送至动物、特别是哺乳动物的介质，包括，即，佐剂，赋形剂或媒介物，如稀释剂、防腐剂、填充剂、流动调节剂、崩解剂、润湿剂、乳化剂、混悬剂、甜味剂、调味剂、芳香剂、抗细菌剂、抗真菌剂、润滑剂和分配剂，取决于施用方式和剂型的性质。
药学上可接受的载体根据本领域普通技术人员认知范围内的许多因素来配制。这些包括但不限于：正在配制的活性剂的类型和性质；含有药剂的组合物待给予的受试者；组合物施用的预期途径；和正在靶向的治疗适应症。药学上可接受的载体包括水性和非水性液体介质两者，以及多种固体和半固体剂型。此类载体还可以包括除活性剂之外的许多不同的成分和添加剂，此类另外的成分出于本领域普通技术人员熟知的多种原因(例如，活性剂、粘合剂等的稳定化)被包括在配制品中。合适的药学上可接受的载体以及其选择中涉及的因素的描述可在多种可易获得的来源例如像Allen,Jr.,L.V.等人,Remington:The Science and Practice of Pharmacy(2卷),第22版,Pharmaceutical Press(2012)中找到。
当然，本申请的固体形式的剂量方案将根据诸如以下的已知因素而变化：特定药剂的药效学特征及其施用方式和途径；接受者的物种、年龄、性别、健康、医疗状况和体重；症状的性质和程度；同时治疗的种类；治疗频率；施用途径、患者的肾和肝功能和所希望的效果。通常每种活性成分的每日口服剂量范围将为在约0.001至约5000mg/天之间、优选在约0.01至约1000mg/天之间、并且最优选在0.1至约250mg/天之间。经静脉内，在恒定速率输注期间，最优选的剂量的范围将是从约0.01至约10mg/kg/分钟。可以将本发明化合物以单一每日剂量施用，或者可以将每日总剂量以每日两次、三次或四次的分剂量施用。
用于施用的剂型(药物组合物)可以含有每剂量单位约1mg至约2000mg的活性成分。在这些药物组合物中，活性成分将通常以基于组合物的总重量的按重量计约0.1％-95％的量存在。
附图说明
Fig.1为化合物1无定形的XRPD图；
Fig.2为化合物1无定形的动态水分吸附(DVS)；
Fig.3实施例2a晶型I的XRPD图；
Fig.4实施例2a晶型I的TGA/DSC图；
Fig.5实施例2a晶型I的1H NMR图；
Fig.6为实施例3a和3b晶型II的XRPD图；
Fig.7为实施例3a晶型II的TGA/DSC图；
Fig.8为实施例3a晶型II的1H NMR图；
Fig.9为实施例4a晶型IIIA的XRPD图；
Fig.10为实施例4a晶型IIIA的TGA/DSC图；
Fig.11为实施例4a晶型IIIA的1H NMR图；
Fig.12为实施例4a晶型IIIA加热前后的XRPD叠图；
Fig.13为实施例4b晶型IIIB的XRPD图；
Fig.14为实施例4b晶型IIIB的TGA/DSC图；
Fig.15为实施例4b晶型IIIB的1H NMR图；
Fig.16为实施例4b晶型IIIB的变温XRPD图；
Fig.17为实施例5a和5b的晶型IV的XRPD图；
Fig.18为实施例5a晶型IV的TGA/DSC图；
Fig.19为实施例5a晶型IV的1H NMR图；
Fig.20为实施例6a晶型V的XRPD图；
Fig.21为实施例6a晶型V的TGA/DSC图；
Fig.22为实施例6a晶型V的1H NMR图；
Fig.23为实施例7a晶型VI的XRPD图；
Fig.24为实施例7a晶型VI的DSC/TGA图；
Fig.25为实施例5a晶型IV加热前后的XRPD图；
Fig.26为实施例3a晶型II加热前后的XRPD叠图；
Fig.27为实施例3b晶型II加热前后的XRPD图；
Fig.28为实施例6a晶型V加热前后的XRPD图；
Fig.29为晶型I和IIIA在EtOAc中混悬竞争的XRPD叠图；
Fig.30为晶型I和IIIA在MIBK中混悬竞争的XRPD叠图；
Fig.31为晶型I和IIIA在丙酮/H2O中混悬竞争的XRPD叠图(I/II)；
Fig.32为晶型I和IIIA在丙酮/H2O中混悬竞争的XRPD叠图(II/II)；
Fig.33为晶型I的DVS图；
Fig.34为晶型I在DVS测试前后的XRPD叠图；
Fig.35为无定形样品在DVS测试前后的XRPD叠图；
Fig.36为晶型IIIA在引湿实验中的DVS图；
Fig.37为晶型IIIA在DVS测试前后的XRPD叠图；
Fig.38为实施例2a晶型I固态稳定性评估前后的XRPD叠图；
Fig.39为无定形固态稳定性评估前后的XRPD叠图；
Fig.40为晶型IIIA固态稳定性评估前后的XRPD叠图；
Fig.41为无定形在不同pH值的溶解度图(投料浓度0.25mg/mL)；
Fig.42为晶型I在不同pH值的溶解度图(投料浓度0.25mg/mL)；
Fig.43为无定形和晶型I在不同pH值的溶解度图(投料浓度0.05mg/mL)；
Fig.44为晶型I研磨前后的XRPD叠图；
Fig.45为晶型I压片前后的XRPD叠图；
Fig.46为晶型IIIA研磨前后的XRPD叠图；
Fig.47为晶型IIIA压片前后的XRPD叠图；
Fig.48为无定形研磨前后的XRPD叠图；
Fig.49为无定形压片前后的XRPD叠图；
Fig.50为评价晶型I的光照稳定性的XRPD叠图；
Fig.51为评价晶型IIIA光照稳定性的XRPD叠图；
Fig.52为评价无定形样品光照稳定性的XRPD叠图；
Fig.53为评价晶型V光照稳定性的XRPD叠图；
Fig.54为评价晶型I光照稳定性的HPLC图；
Fig.55为评价晶型IIIA光照稳定性的HPLC图；
Fig.56为评价无定形样品光照稳定性的HPLC图；以及
Fig.57为评价晶型V光照稳定性的HPLC图。
实施例
以下实施例仅用于示例本发明，而非以任何方式对本发明进行限制。
溶剂名称对应表
在本发明中采用以下仪器和方法：
仪器和方法
X射线粉末衍射(XRPD)：XRPD结果是在X’Pert3和Empyrean X射线粉末衍射分析仪上采集，扫描参数如表1所示。
表1：XRPD测试参数
热重分析(TGA)和差示扫描量热(DSC):TGA和DSC分别在TA Discovery 5500热重分析仪和TA Discovery 2500差示扫描量热仪上采集，表2列出了TGA及DSC测试参数。
表2:TGA和DSC测试参数

氢谱液态核磁(1H Solution NMR):氢谱液态核磁谱图在Bruker 400M核磁共振仪上采集，DMSO-d6作为溶剂。
动态水分吸附(DVS):动态水分吸附(DVS)曲线在SMS(Surface Measurement Systems)的DVS Intrinsic上采集。在25℃时的相对湿度用LiCl、Mg(NO3)2和KCl的潮解点校正。DVS测试参数列于表3。
表3:DVS测试参数
高效液相色谱(UPLC):实验中纯度和溶解度由Agilent 1290超高效液相色谱仪测试，分析条件如表4和表5所示。
表4:纯度测试的超高效液相色谱测试条件

表5:溶解度测试的超高效液相色谱测试条件
对于在“仪器和方法”部分没有明确指出的仪器和方法，可以采用本领域已知的仪器和方法。
实施例1a(化合物1无定形，重复中国专利申请No.201210190520.4实施例1)：按照中国专利申请No.201210190520.4实施例1的方法，即将实施例1步骤13所获得的粗产品通过硅胶柱色谱分离纯化，以二氯甲烷/甲醇作为洗脱剂，然后旋蒸得到白色固体，经XRPD分析为无定形，其XRPD图如Fig.1所示，以及动态水分吸附(DVS)如Fig.2所示。
实施例2a：将约21mg实施例1a的无定形样品中加入4mL无水乙醇，在85℃油浴中加热搅拌5分钟溶清；随后，所得清液在2.5h内伴随着搅拌降温至室温25℃，室温继续搅拌24h，将所得固体样品滤出，进行XRPD测定。
XRPD结果如Fig.3所示。TGA/DSC结果(Fig.4)显示，样品加热至150℃失重0.77％，在221.3℃(起始温度)处有1个尖锐的吸热峰。1H NMR数据以DMSO-d6为溶剂采集，结果如Fig.5所示，未检测到明显的溶剂残留。根据晶型I的表征结果，其具有较 小且平缓的失重(低于半水合物的理论含水量1.80％)及单一的熔化吸热信号，推测晶型I为无水合物或无水晶型。该晶型I的XRPD衍射峰数据参见表6。
表6：晶型I的XRPD衍射峰数据
实施例2b：分别称取约20mg每份的无定形样品加至20-mL的小瓶内，加入0.4～1.0mL的良溶剂(见表7)溶解，过滤(孔径0.45μm的PTFE滤膜)得到澄清溶液，边搅拌该澄清溶液边加入表7中的反溶剂至有固体析出，离心分离析出固体并进行XRPD测试。结果如表7所示，反溶剂添加得到固体，经XRPD确定为晶型I。
表7：反溶剂添加试验小结

[1]：反溶剂添加后澄清，转移至5℃搅拌析出固体。
[2]：反溶剂添加后澄清，转至5℃搅拌后仍澄清，转至-20℃搅拌析出固体。
[3]：反溶剂添加后澄清，转至5℃及-20℃搅拌后仍澄清，转至室温挥发。
[4]：反溶剂添加后成油，转至5～50℃温度循环悬浮搅拌后仍成油，转至室温挥发。
实施例2c：称取约20mg每份的实施例1a的无定形样品于5-mL或HPLC小瓶中，加入0.7～4.0mL表8中的溶剂，在50℃下搅拌约2小时后过滤(孔径0.45μm的PTFE滤膜)取滤液，将所得滤液放置在生化培养箱中，以0.1℃/分钟的降温速度降温至5℃，收集析出的固体并进行XRPD测试。试验结果见表8。缓慢降温试验得到固体，经XRPD确定为晶型I。
表8 5℃缓慢降温试验小结
[1]：缓慢降温至5℃后澄清，转移至-20℃静置后出固体。
[2]：缓慢降温至5℃后澄清，转移至-20℃静置后仍无固体析出，转至室温挥发。
实施例3晶型II
实施例3a：称取71.6mg实施例2a晶型I样品于3mL小瓶中，加入1.5mL ACN，室温下磁力搅拌约1天。离心分离固体，环境条件(～21℃/45％RH)下放置约4小时干燥，得到最终产品，并且对其进行XRPD和TGA/DSC测试。
XRPD结果如Fig.6所示。TGA结果如Fig.7所示，TGA显示，样品加热至120℃有5.90％的台阶式失重；DSC结果表明，在94.1℃和220.9℃(起始温度)处有2个吸热峰，在191.6℃(峰值温度)处有1个放热峰。1H NMR结果如Fig.8所示，样品中ACN与API的摩尔比为0.8(5.9重量％，与TGA失重一致)。根据表征及加热实验结果，推测晶 型II为ACN溶剂合物，其加热后脱溶剂发生转晶。该晶型II的XRPD衍射峰数据参见表9。
表9：晶型II的XRPD衍射峰数据
实施例3b：称取20.4mg实施例2a晶型I样品于3mL小瓶中，加入2mL ACN/丙酮(1:1,v/v)，超声后过滤(0.45μm PTFE滤膜)，得澄清溶液。用封口膜密封后扎4个小孔，放置在通风橱中缓慢挥发5天。观察到固体析出，放置在环境条件下过夜后，转至室温真空干燥约1天。将实施例2a晶型I的ACN/丙酮(1:1,v/v)溶液缓慢挥发，析出固体后转至室温真空干燥，得到最终产品，并且对其进行XRPD测试。XRPD结果如Fig.6所示。
实施例3c：称量约19.7mg的实施例1a得到的无定形样品至HPLC小瓶中，分别加入0.5mL ACN/H2O(19:1)溶剂，得到悬浊液置于5℃下磁力搅拌(～1000rpm)约6天后，离心分离固体并进行XRPD测试。XRPD结果为晶型II。
实施例3d：称量约19.9mg的实施例1a得到的无定形样品至HPLC小瓶中，分别加入0.5mL ACN/丙酮(1:1)溶剂，得到悬浊液置于5℃下磁力搅拌(～1000rpm)约6天后，离心分离固体并进行XRPD测试。XRPD结果为晶型II。
实施例4(晶型IIIA和IIIB)
实施例4a：将实施例3a的晶型II样品用DSC加热至120℃并恒温5分钟后冷却至室温，得到晶型IIIA。XRPD结果如Fig.9所示。TGA/DSC结果(Fig.10)显示，样品加热至200℃失重0.26％，低于半水合物的理论含水量(1.80％)；DSC结果显示，在200.3℃(峰值温度)处分别有1个放热信号，在219.6℃(起始温度)处有1个尖锐的吸热信号。1H NMR结果(Fig.11)显示，未检测到明显的溶剂残留。将晶型IIIA加热至205℃、冷却至室温并暴露在环境条件下后转变为晶型I，加热前后XRPD结果如Fig.12所示。根据晶型IIIA的表征结果，推测晶型IIIA为无水合物，在198.8℃处的放热信号为向晶型I转变的热信号。该晶型IIIA的XRPD衍射峰数据参见表10。
表10晶型IIIA的XRPD衍射峰数据

实施例4b：称取100.0mg实施例2a晶型I样品于20mL小瓶中，加入10mL THF溶解，过滤(0.45μm PTFE滤膜)得澄清溶液。滤液在50℃旋蒸，收集得到最终产品。XRPD结果如Fig.13所示。TGA/DSC结果(Fig.14)显示，样品加热至200℃失重3.49％；DSC结果显示，在121.3℃和144.6℃(峰值温度)处分别有1个微弱的吸热信号和放热信号，在218.3℃(起始温度)处有1个较强的吸热信号。1H NMR结果(Fig.15)显示，残留溶剂THF与API的摩尔比为0.09(1.3重量％)，推测为表面吸附的溶剂。晶型IIIB在121.3℃和144.6℃处的热信号为向晶型I转变的热信号。该晶型IIIB的XRPD衍射峰数据参见表11。
表11：晶型IIIB的XRPD衍射峰数据

实施例5(晶型IV)
实施例5a：称量约20mg的实施例1a得到的无定形样品至HPLC小瓶中，加入0.5mL1,4-二氧六环(1,4-dioxane)溶剂，将得到的悬浊液置于室温下磁力搅拌(～1000rpm)约6天后，离心分离固体，在室温真空干燥过夜，得到最终产品，并且对其进行XRPD和TGA/DSC测试。XRPD和TGA/DSC结果如Fig.17和
Fig.18所示。TGA结果显示，样品加热至120℃有16.80％的台阶式失重；DSC结果表明，在95.6℃和220.5℃(起始温度)处有2个吸热峰。1H NMR结果如Fig.19所示，样品中1,4-Dioxane与API的摩尔比为1.2(17.8重量％，与TGA失重一致)。根据以上表征结果及变温XRPD结果，推测晶型IV为1,4-Dioxane溶剂合物。该晶型IV的XRPD衍射峰数据参见表12。
表12：晶型IV的XRPD衍射峰数据

实施例5b：将实施例2a的晶型I样品(500mg)在1,4-Dioxane(12.5mL)中室温搅拌2天，所得固体室温真空干燥1天得到。XRPD结果如Fig.17所示。
实施例6(晶型V)
实施例6a:称取20.8mg实施例2a晶型I样品于5mL小瓶中，加入4mL EtOH。置于70℃下搅拌后溶清，在650分钟内降温(降温速率0.1℃/分)至5℃并在5℃下静置。去除溶液后，将固体在环境条件(温度：～21℃，湿度：～36％RH)下敞口放置干燥。XRPD结果如Fig.20所示。TGA/DSC结果(Fig.21)显示，样品加热至120℃有7.64％的台阶式失重，在103.1℃和223.3℃(起始温度)处有2个吸热峰。1H NMR结果如Fig.22所示，样品中EtOH与API的摩尔比为0.7(6.2重量％，与TGA失重较一致)。根据以上表征结果及变温XRPD结果，推测晶型V为EtOH溶剂合物，其加热后脱溶剂转变为无水晶型I。该晶型V的XRPD衍射峰数据参见表13。
表13：晶型V的XRPD衍射峰数据

实施例7(晶型VI)
将实施例1a的无定形样品(1g)溶解于DMF(3mL)中，完全溶解后，加入IPA(6mL),室温搅拌过夜，析出固体，过滤，滤饼用异丙醇洗，真空干燥得到DMF的溶剂化物晶型，为晶型VI。
晶型VI的XRPD结果如Fig.23所示，DSC结果显示，在147.39℃、208.56℃处有2个吸热峰；Fig.24的TGA结果显示，加热至180℃失重0.67％。
实施例8(晶体转换)
实施例8a:将实施例5a晶型IV加热至120℃、冷却至室温并暴露在环境条件下后转 变为晶型I，XRPD结果如Fig.25所示。
实施例8b:将实施例3a晶型II分别加热至120℃和210℃、冷却至室温并暴露在环境条件下，XRPD结果如Fig.26显示，加热至120℃转变为晶型IIIB，加热至210℃转变为晶型I。
实施例8c:将实施例3b晶型II加热至120℃、冷却至室温并暴露在环境条件下后转变为晶型I，XRPD结果如Fig.27所示。推测样品加热后可能同样先转变为晶型IIIB，但在环境条件下又较快的转变为晶型I，在XRPD测试时显示转变为晶型I。
实施例8d:通过变温XRPD对实施例4a晶型IIIB进行鉴定，结果如Fig.16所示。在30℃下N2吹扫20分钟后晶型不变，在N2保护下加热至120℃时观察到晶型I的衍射峰，继续加热至170℃时主要为晶型I，仅有少量晶型IIIB的衍射峰，在N2保护下冷却至30℃未继续转变。
实施例8e:将实施例6a晶型V加热至120℃、冷却至室温并暴露在环境条件下后转变为晶型I，XRPD结果如Fig.28所示。
实施例8f(晶型I和IIIA混悬竞争实验)
该实施例涉及晶型I和晶型IIIA在室温和50℃下EtOAc和MIBK中的混悬竞争实验，以及室温下不同水活度的丙酮/H2O中的混悬竞争实验。
称取约15mg实施例2a晶型I样品与HPLC瓶中，加入1mL相应溶剂，在对应温度下搅拌4小时或过夜。过滤(0.45μm PTFE滤膜)得到饱和溶液。称取等质量的实施例2晶型I和实施例4a晶型IIIA样品(各约5mg)于新的HPLC瓶中，加入步骤1得到的饱和溶液。在对应温度下搅拌，对固体湿样进行XRPD测试(用Kapton膜覆盖，避免在测试过程中由于溶剂挥发导致的潜在转晶)。
混悬竞争的结果汇总在表14。XRPD结果见Fig.29、Fig.30、Fig.31、Fig.32。
表14晶型I和IIIA混悬竞争实验结果
其中aw：理论水活度。
根据XRPD比对结果，在所有条件下晶型I和IIIA的物理混合物在混悬竞争后均转变为晶型I，表明在室温至50℃范围内无水条件下以及室温下水活度0～1条件下，无水晶型I比晶型IIIA热力学上更稳定。
实施例9(晶型I的引湿性)
通过25℃下的动态水分吸附(DVS)试验对实施例2a晶型I、实施例1a无定形和实施例4a的晶型IIIA的引湿性进行了评估。
晶型I的DVS结果如Fig.33所示，在25℃/80％RH时吸水增重0.047％，表明其几乎无引湿性。XRPD对比结果(Fig.34)显示，晶型I在DVS测试后未发生晶型转变。
无定形的DVS结果如Fig.2所示，当湿度由50％RH增加至95％RH过程中，吸水增重不断减小，推测无定形样品在湿度升高过程中发生转晶，原样品中吸附或包裹的水或有机溶剂在N2吹扫作用下而去除。XRPD对比结果(Fig.35)显示，无定形样品在DVS测试后转变为晶型I。
晶型IIIA的DVS结果如Fig.36所示，在25℃/80％RH时吸水增重0.060％，表明其几乎无引湿性。XRPD对比结果(Fig.37)显示，晶型IIIB在DVS测试后未发生晶型转变。
实施例10(晶型I的固态稳定性)
为了评估晶型I、晶型IIIA和无定形的固态稳定性，分别称取适量实施例2a晶型I、实施例1a的无定形样品和实施例4a的晶型IIIA，设置在60℃/闭口/1天、25℃/60％RH/敞口/1周和40℃/75％RH/敞口/1周条件下的稳定性实验。将不同条件下的稳定性样品，通过XRPD测试晶型评估物理稳定性，通过HPLC测试纯度评估化学稳定性。评估结果汇总于表15中，XRPD结果见Fig.38、Fig.39和Fig.40(Fig.40中星标处观察到微弱的晶型I的衍射峰)。稳定性结果显示：晶型I在三种评估条件下均未发生晶型转变或纯度降低，表明晶型I在评估条件下具有较好的物理和化学稳定性；无定形在三种评估条件下纯度均无明显变化，但均转变为晶型I；晶型IIIA在60℃下闭口放置1天后观察到微弱的晶型I的衍射峰，在25℃/60％RH和40℃/75％RH下敞口放置1周后晶型不变。
表15实施例2a晶型I的固态稳定性评估结果汇总

*：无定形样品在稳定性评估后均转变为晶型I；
#：观察到微弱的晶型I的衍射峰。
实施例11(溶解度)
对实施例2a的晶型I和实施例1a的无定形在室温下不同pH条件下(1M HCl以及pH 2.0/4.5/6.8/7.4缓冲液)中的动态溶解度进行了测试。具体步骤如下：
(1)称取约2.5mg晶型I或无定形样品于20mL玻璃瓶中，加入10mL不同pH缓冲液，或称取约2.0mg晶型I或无定形样品于20mL玻璃瓶中，加入8mL盐酸(1M)。
(2)室温下震荡(～500rpm)3min，用注射器吸取0.8～1mL样品，过滤(0.45μm PTFE滤膜)后进行HPLC测试。(3)对于以无定形为原料得到的溶液，取出100μL并用对应缓冲液稀释10倍备用。若稀释前的溶液有固体析出的现象，则对稀释后样品进行测试，若无固体析出则对稀释前的样品进行测试；对于以晶型I为原料得到的溶液，因预估其溶解度较低，因此未进行稀释，直接对过滤后的澄清溶液进行HPLC测试。
晶型I和无定形在不同PH条件下的溶解度见表16。结果表明，晶型I的溶解度在不同的pH下几乎无变化，而无定形随着pH变化，呈现较大的变化(如Fig.41)。而晶型I在体内的溶出更稳定(如Fig.42)，更易得到血药浓度平稳的体内药代动力学结果，有利于避免因血药浓度波动太大导致的用药安全性风险。当投料浓度为0.05mg/mL时也观察到了类似结果(如Fig.43)。
表16晶型I和无定形在不同介质中的3min溶解度结果(投料浓度0.25mg/mL)
LOQ＝0.28μg/mL。
实施例12(粉末学性质)
对实施例2a的晶型I和实施例1a的无定形样品开展粉体学性质的评估，包括休止角、松密度和振实密度，以了解晶型I和无定形的粉体流动性能。
松密度与振实密度：将一定质量的待评估样品加入一个5-mL量筒中，记录此时体积。松密度由样品的质量除以此时的体积计算得到；将量筒轻叩200次，记录最终体积，振实密度由样品质量除以最终体积得到。每个参数平行测试三次。
休止角：将漏斗与底面垂直固定，将物料缓慢加入到漏斗中。底面形成一个匀称的物料的椎体。测量椎体的高度和底面直径。平行测定三次。
松密度/振实密度的结果汇总于表17。结果显示，晶型I的平均松密度和振实密度分别为0.34g/cm3和0.46g/cm3，计算得到卡尔指数为26％；无定形的平均松密度和振实密度分别为0.31g/cm3和0.46g/cm3，计算得到卡尔指数为33％。
休止角的结果汇总于表18。结果显示，晶型I和无定形样品的休止角分别为27.7°和26.7°。综合评估的结果，晶型I和无定形样品具有较为接近的卡尔指数和休止角，表明晶型I和无定形有相似的流动性。
表17松密度/振实密度测试结果
卡尔指数＝(振实密度-松密度)/振实密度。
表18休止角测试结果
休止角α计算公式：α＝tan-1(h/D)，h为椎体高度，D为椎体直径。
实施例13(机械力稳定性)
对实施例2a的晶型I、实施例4a的晶型IIIA和实施例1a的无定形进行手动研磨和压片机压片(350MPa压力)，并对研磨及压片后的样品进行XRPD测试以评估其机械力稳定性。XRPD结果如Fig.44、Fig.45、Fig.46、Fig.47、Fig.48和Fig.49所示。评估结果显示：在研磨及压片后，晶型I未发生晶型转变且结晶度无明显降低；晶型III在研磨后晶型不变但结晶度略有降低，在压片后晶型不变但观察到结晶度明显降低；无定形在研磨及压片后转变为晶型I。
根据晶型表征及鉴定结果，选择晶型I开展引湿性、固态稳定性、溶解度、粉体学性质及机械力稳定性评估，同时对无定形开展相同的评估以与晶型I进行性质比较。DVS结果显示晶型I几乎无引湿性，在DVS测试后未发生晶型转变；无定形在DVS测试后转变为晶型I。固态稳定性结果显示，晶型I在60℃下闭口放置1天、25℃/60％RH和40℃/75％RH下敞口放置1周后均未发生晶型转变或纯度降低，表明晶型I在评估条件下具有较好的物理和化学稳定性；无定形在三种评估条件下纯度均无明显变化，但均转变为晶型I。晶型I粉体学性质测试结果表明，晶型I和无定形样品具有相似的流动性。机械力稳定性结果显示，在压片(350MPa)及手动研磨(约3分钟)后，晶型I未发生晶型转变且结晶度无明显降低，无定形转变为晶型I。
根据表征数据和评估结果，晶型I在所有评估条件下均未发生晶型转变，而无定形在DVS、固态稳定性、溶解度、机械力稳定性测试后均转变为晶型I，表明晶型I相比无定 形具有更好的物理稳定性。
实施例14(光照稳定性)
对实施例2a的晶型I、实施例4a晶型IIIA、实施例6a晶型V和实施例1a的无定形分别在光照(白光5800～5890Lux+紫外7.9～8.7W/m2)下，分别在6小时和24小时时取出部分样品，用于XRPD和HPLC纯度测试，进行稳定性评估。XRPD结果如Fig.50、Fig.51、Fig.52、Fig.53所示：晶型I、晶型IIIA、晶型V和无定形在光照条件下放置24小时后均未发生晶型转变；在避光参照条件下，晶型I、晶型IIIA和晶型V放置24小时后未发生转晶，无定形放置6小时和24小时后观察到微弱的晶型I的衍射峰。HPLC结果如Fig.54、Fig.55、Fig.56、Fig.57和表19所示：光照24h后，晶型I在光照条件下纯度由100.00面积％略降低至99.14面积％(杂质A含量0.79％)，晶型IIIA、晶型V和无定形在光照条件下纯度由100.00面积％分别降低至98.46面积％(杂质A含量1.34％)、92.05面积％(杂质A含量7.95％)和90.42面积％(杂质A含量8.83％)；四种样品在避光参照条件下纯度均无明显变化(在Fig.54、Fig.55和Fig.56中光照6小时的样品与光降解杂质的测试是在两个HPLC测试序列中分别进行，因此杂质的保留时间可能略有偏移)。根据光照稳定性的结果，晶型I相比于晶型IIIA、晶型V和无定形表现出更好的光照稳定性。
表19晶型I/IIIA/V与无定形的光照稳定性结果
实施例15(SD大鼠药代动力学实验)
分别将2.39mg实施例2a的晶型I、2.38mg实施例4a的晶型IIIA、2.46mg实施例1的无定形作为受试物，分别加入7.967mL、7.906mL、8.193mL的0.5％CMC-Na水溶液，配制成终浓度为0.3mg/mL的灌胃给药制剂。3只雄性SD大鼠，分别以3mg/kg动物体重的剂量，灌胃给予上述三种受试物溶液。按时间点0,0.25h,0.5h,1h,2h,4h,8h,10h,24h由颈静脉窦采血0.15mL。将采集的全血置于EDTA-K2的抗凝管中并充分混匀，离心(1500～1600g)10min，分离血浆，用于生物分析。采用LC-MS/MS分析方法(仪器型号：Triple Quad 5500；色谱柱：Agilent ZORBAX XDB-C18；流速：0.50mL/min；进样量：2μL；流动相A:水[0.1％甲酸+5mM乙酸铵]，流动相B:乙腈[0.1％甲酸])，测定血浆样品中受试物的浓度。采用Pharsight Phoenix 8.3中的非房室模型计算相应的药代动力学参数(表20)。由数据可知，晶型I和晶型IIIA的AUC0-t分别9820ng·h/mL和8890ng·h/mL，Cmax分别为1160ng/mL和1030ng/mL，表明在体内有充分的药物血浆暴露，具有较好的药代动力学性质，并且晶型I比晶型IIIA更高的暴露量和血药浓度，表现在有潜力降低给药剂量。
表20晶型I、晶型IIIA及无定形的药代动力参数
实施例16：对NRAS突变的黑色素瘤细胞株增殖的抑制
所有细胞由北京肿瘤医院提供，人类黑色素瘤细胞株SK-MEL-2(NRAS Q61R)用MEM+10％FBS+1％青链霉素培养于37摄氏度，5％二氧化碳浓度培养箱中，人类黑色素瘤细胞株HMVII(NRAS Q61K)用F12K+10％FBS+1％青链霉素培养于37摄氏度，5％二氧化碳浓度培养箱中。
当培养的细胞汇合度达到80％以上时，用胰酶消化贴壁细胞并且离心收集细胞沉淀，计数，按适当密度接种90μL细胞悬液于96孔板，24小时后，每孔按照10μl的体积加入一系列梯度稀释的化合物(晶型I)(浓度范围0.15nM-10μM，4倍梯度稀释)。每个浓度设三个复孔，一共9个浓度点，以加入同样体积的5％DMSO的孔作为对照，DMSO终浓度为0.5％。药物处理3天后，MTT检测细胞活力，每孔加入10μl MTT放入培养箱继续培养4小时后，弃上清并加入150μl DMSO溶解结晶甲瓒，利用酶标仪检测490nM吸光度。以上实验重复3次，GraphPad Prism 8软件制作量效曲线并计算IC50。结果以三次实验的IC50平均值±SD值进行表示(表21和表22)。
表21.化合物1在SK-MEL-2肿瘤细胞上的IC50值

表22.化合物1在HMVII肿瘤细胞上的IC50值
实施例17：对RAS或RAF突变的抑制
采用四唑盐(MTS)方法检测化合物1对RAS突变或RAF突变的肿瘤细胞株和RAS/RAF野生型肿瘤细胞株，以及正常人源细胞株的体外抗增殖活性。
胰酶消化对数生长期的贴壁细胞或者离心收集悬浮细胞，计数，将细胞接种于150μL于96孔板，24小时后加入培养基稀释的4倍终浓度化合物(晶型I)50μL/孔(浓度范围0.15nM-1000nM，3倍梯度稀释)。以加入同样体积的2％DMSO的孔作为对照，DMSO终浓度为0.5％。细胞继续培养72小时后，MTS检测细胞活力。具体方法如下：贴壁细胞，弃去培养基，每孔加入20μL MTS和100μL细胞培养基，悬浮细胞直接加入20μL MTS。放入培养箱继续培养1-4小时后检测OD490，以OD650值作为参考。GraphPad Prism软件制作量效曲线并计算IC50。结果见表23。
表23.化合物1对RAS和RAF突变的肿瘤细胞和人胚肺细胞MRC-5的体外抗增殖IC50值
实施例18：
一项在NRAS突变的晚期黑色素瘤患者中开展的多中心、单臂Ⅱ期研究，评价化合物1晶型I的有效性和安全性。患者接受本品每次12mg，每日口服2次，直至出现不可耐受的毒性、疾病进展(研究者根据RECIST 1.1进行评估)、撤回知情、死亡或经研究者判 断风险大于获益时终止。截止2023年2月19日，共入组100例受试者，其中95.0％(95/100例)受试者纳入全分析集(FAS)。
主要疗效指标：
FAS人群中：IRRC(独立影像评审委员会)评估的ORR(客观缓解率)为35.8％(34/95例)(95％CI：26.2％，46.3％)。
FAS人群中，IRRC评估结果：中位PFS(无进展生存期)为4.2个月(95％CI：3.5，5.6)，DCR为72.6％(69/95例)(95％CI：62.5％，81.3％)，中位DoR为6.1个月(95％CI：3.9，8.9)；
化合物1对NRAS突变的晚期黑色素瘤患者具有良好的抗肿瘤治疗效果，IRRC评估的ORR为35.8％，疗效数据显著优于同类药物临床研究数据。

Claims (85)

  1. 式(I)的多晶型物
    其中n为0或1，X为乙腈、水、1,4-二氧六环(1,4-Dioxane)、乙醇、甲醇、二甲基甲酰胺、丙酮或其混合物。
  2. 权利要求1的多晶型物，其中n为0。
  3. 权利要求1的多晶型物，其中n为1；X为乙腈、水、1,4-二氧六环或乙醇。
  4. 权利要求1的多晶型物，其中n为0，所述多晶型物为晶型I，其特征在于所述晶型I的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：16.71°±0.2°、21.82°±0.2°和23.75°±0.2°，采用Cu-Kα辐射。
  5. 权利要求4的多晶型物，其中所述晶型I的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：7.48°±0.2°和22.36°±0.2°。
  6. 权利要求5的多晶型物，其中所述晶型I的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：5.3°±0.2°、24.57°±0.2°和27.08°±0.2°。
  7. 权利要求6的多晶型物，其中所述晶型I的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：11.81°±0.2°、15.83°±0.2°、17.92°±0.2°、18.95°±0.2°和19.17°±0.2°。
  8. 权利要求4的多晶型物，其中所述晶型I的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：5.30°±0.2°、7.48°±0.2°、11.81°±0.2°、14.85°±0.2°、15.83°±0.2°、16.71°±0.2°、17.92°±0.2°、18.95°±0.2°、19.17°±0.2°、19.43°±0.2°、21.14°±0.2°、21.82°±0.2°、22.36°±0.2°、23.75°±0.2°、24.57°±0.2°、27.08°±0.2°、27.83°±0.2°、28.88°±0.2°、31.20°±0.2°、31.92°±0.2°、32.40°±0.2°、33.91°±0.2°、35.83°±0.2°、37.51°±0.2°和39.04°±0.2°。
  9. 权利要求4的多晶型物，其中所述晶型I的X-射线粉末衍射图谱基本上如Fig.3所示。
  10. 权利要求4的多晶型物，其中所述晶型I具有如Fig.4所示的TGA图和/或DSC图。
  11. 权利要求1的多晶型物，其中n为1，X为乙腈，所述多晶型物为晶型II，其特征在于所述晶型II的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.35° ±0.2°、20.34°±0.2°、22.41°±0.2°、22.6°±0.2°、24.99°±0.2°、26.05°±0.2°和28.71°±0.2°，采用Cu-Kα辐射。
  12. 权利要求11的多晶型物，其中所述晶型II的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：9.18°±0.2°、16.03°±0.2°、18.25°±0.2°、27.07°±0.2°、29.08°±0.2°和33.93°±0.2°。
  13. 权利要求12的多晶型物，其中所述晶型II的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：14.44°±0.2°、24.64°±0.2°、26.41°±0.2°、32.27°±0.2°、32.68°±0.2°、37.07°±0.2°和39.51°±0.2°。
  14. 权利要求11的多晶型物，其中所述晶型II的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.35°±0.2°、9.18°±0.2°、9.91°±0.2°、14.44°±0.2°、16.03°±0.2°、18.25°±0.2°、19.77°±0.2°、20.34°±0.2°、21.81°±0.2°、22.41°±0.2°、22.60°±0.2°、23.84°±0.2°、24.64°±0.2°、24.99°±0.2°、25.43°±0.2°、26.05°±0.2°、26.41°±0.2°、27.07°±0.2°、28.71°±0.2°、29.08°±0.2°、29.81°±0.2°、31.16°±0.2°、31.57°±0.2°、32.27°±0.2°、32.68°±0.2°、33.93°±0.2°、34.19°±0.2°、35.42°±0.2°、37.07°±0.2°、37.56°±0.2°、38.69°±0.2°和39.51°±0.2°。
  15. 权利要求11的多晶型物，其中所述晶型II的X-射线粉末衍射图谱基本上如Fig.6所示。
  16. 权利要求11的多晶型物，其中所述晶型II具有如Fig.7所示的TGA图和/或DSC图。
  17. 权利要求1的多晶型物，其中n为0，所述多晶型物为晶型IIIA，其特征在于所述晶型IIIA的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.59°±0.2°、22.69°±0.2°、20.32°±0.2°、23.62°±0.2°、23.91°±0.2°和24.15°±0.2°，采用Cu-Kα辐射。
  18. 权利要求17的多晶型物，其中所述晶型IIIA的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：10.8°±0.2°、17.14°±0.2°、13.75°±0.2°、21.59°±0.2°和26.01°±0.2°。
  19. 权利要求18的多晶型物，其中所述晶型IIIA的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：18.71°±0.2°、21.97°±0.2°、25.54°±0.2°、27.13°±0.2°、27.59°±0.2°和30.51°±0.2°。
  20. 权利要求17的多晶型物，其中所述晶型IIIA的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.59°±0.2°、9.9°±0.2°、10.8°±0.2°、13.09°±0.2°、13.75°±0.2°、17.14°±0.2°、17.87°±0.2°、18.71°±0.2°、19.19°±0.2°、20.32°±0.2°、21.59°±0.2°、21.97°±0.2°、22.69°±0.2°、23.62°±0.2°、23.91°±0.2°、24.15°±0.2°、25.54°±0.2°、26.01°±0.2°、27.13°±0.2°、27.59°±0.2°、28.83°±0.2°、29.24°±0.2°、30.51°±0.2°、31.13°±0.2°、31.79°±0.2°、33.6°±0.2°、34.14°±0.2°、36.08°±0.2°、36.67°±0.2°和37.26°±0.2°。
  21. 权利要求17的多晶型物，其中所述晶型IIIA的X-射线粉末衍射图谱基本上如Fig.9所示。
  22. 权利要求17的多晶型物，其中所述晶型IIIA具有如Fig.10所示的TGA图和/或DSC图。
  23. 权利要求1的多晶型物，其中n为0，所述多晶型物为晶型IIIB，其特征在于所述晶型IIIB的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.53°±0.2°、13.69°±0.2°、18.6°±0.2°、20.19°±0.2°、21.52°±0.2°和22.64°±0.2°，采用Cu-Kα辐射。
  24. 权利要求23的多晶型物，其中所述晶型IIIB的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：10.75°±0.2°、17.07°±0.2°、21.93°±0.2°、26.13°±0.2°、23.57°±0.2°和30.46°±0.2°。
  25. 权利要求24的多晶型物，其中所述晶型IIIB的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：13.05°±0.2°、16.63°±0.2°、20.82°±0.2°、24.01°±0.2°、27.55°±0.2°、31.79°±0.2°。
  26. 权利要求23的多晶型物，其中所述晶型IIIB的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.53°±0.2°、10.75°±0.2°、12.62°±0.2°、13.05°±0.2°、13.69°±0.2°、16.63°±0.2°、17.07°±0.2°、18.60°±0.2°、19.59°±0.2°、20.19°±0.2°、20.82°±0.2°、21.52°±0.2°、21.93°±0.2°、22.64°±0.2°、23.57°±0.2°、24.01°±0.2°、25.46°±0.2°、26.13°±0.2°、27.55°±0.2°、30.46°±0.2°、31.04°±0.2°、31.79°±0.2°、32.81°±0.2°、33.54°±0.2°、34.06°±0.2°和34.46°±0.2°。
  27. 权利要求23的多晶型物，其中所述晶型IIIB的X-射线粉末衍射图谱基本上如Fig.13所示。
  28. 权利要求23的多晶型物，其中所述晶型IIIB具有如Fig.14所示的TGA图和/或DSC图。
  29. 权利要求1的多晶型物，其中n为1，X为1,4-二氧六环，所述多晶型物为晶型IV，其特征在于所述晶型IV的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：8.56°±0.2°、13.29°±0.2°、17.69°±0.2°、19.75°±0.2°和22.45°±0.2°，采用Cu-Kα辐射。
  30. 权利要求29的多晶型物，其中所述晶型IV的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：5.26°±0.2°、18.29°±0.2°、31.83°±0.2°、25.68°±0.2°、22.86°±0.2°、32.81°±0.2°和23.44°±0.2°。
  31. 权利要求30的多晶型物，其中所述晶型IV的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：126.57°±0.2°、27.52°±0.2°、35.69°±0.2°、21.09°±0.2°、20.35°±0.2°和31.43°±0.2°。
  32. 权利要求29的多晶型物，其中所述晶型IV的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：5.26°±0.2°、8.56°±0.2°、9.85°±0.2°、13.29°±0.2°、17.69°±0.2°、18.29°±0.2°、19.75°±0.2°、20.35°±0.2°、21.09°±0.2°、22.45°±0.2°、22.86°±0.2°、23.44°±0.2°、24.44°±0.2°、25.68°±0.2°、26.57°±0.2°、27.52°±0.2°、28.40°±0.2°、29.78°±0.2°、31.43°±0.2°、31.83°±0.2°、32.81°±0.2°、34.29°±0.2°、35.69°±0.2°和37.72°±0.2°。
  33. 权利要求29的多晶型物，其中所述晶型IV的X-射线粉末衍射图谱基本上如Fig.17所示。
  34. 权利要求29的多晶型物，其中所述晶型IV具有如Fig.18所示的TGA图和/或DSC图。
  35. 权利要求1的多晶型物，其中n为1，X为乙醇，所述多晶型物为晶型V，其特征在于所述晶型V的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.21°±0.2°、8.47°±0.2°、15.62°±0.2°、21.73°±0.2°、25.53°±0.2°、25.94°±0.2°和28.05°±0.2°，采用Cu-Kα辐射。
  36. 权利要求35的多晶型物，其中所述晶型V的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：9.61°±0.2°、17.55°±0.2°、19.25°±0.2°、22.22°±0.2°、23.12°±0.2°、32.92°±0.2°和34.22°±0.2°。
  37. 权利要求36的多晶型物，其中所述晶型V的X-射线粉末衍射图谱还包括以下2θ位置的特征衍射峰：9.06°±0.2°、20.07°±0.2°、28.49°±0.2°、30.21°±0.2°、31.25°±0.2°、35.47°±0.2°和38.94°±0.2°。
  38. 权利要求35的多晶型物，其中所述晶型V的X-射线粉末衍射图谱包括以下2θ位置的特征衍射峰：6.21°±0.2°、8.47°±0.2°、25.94°±0.2°、15.62°±0.2°、25.53°±0.2°、28.05°±0.2°、21.73°±0.2°、17.55°±0.2°、32.92°±0.2°、23.12°±0.2°、22.22°±0.2°、19.25°±0.2°、34.22°±0.2°、9.61°±0.2°、9.06°±0.2°、38.94°±0.2°、31.25°±0.2°、35.47°±0.2°、28.49°±0.2°、30.21°±0.2°、20.07°±0.2°、39.78°±0.2°、14.26°±0.2°、24.32°±0.2°、16.95°±0.2°、32.33°±0.2°、36.43°±0.2°、24.94°±0.2°、12.42°±0.2°和37.86°±0.2°。
  39. 权利要求35的多晶型物，其中所述晶型V的X-射线粉末衍射图谱基本上如Fig.20所示。
  40. 权利要求35的多晶型物，其中所述晶型V具有如Fig.21所示的TGA图和/或DSC图。
  41. 权利要求1-40中任一项的多晶型物，其中所述多晶型物基本上不含有杂质(A)。
  42. 权利要求40的多晶型物，其中化合物1的多晶型物含有相对于多晶型物少于0.15重量％杂质(A)。
  43. 权利要求4的多晶型物，其中所述晶型I基本上不含有杂质(A)。
  44. 权利要求43的多晶型物，其中晶型I含有相对于晶型I少于0.15重量％杂质(A)。
  45. 一种药物组合物，其含有权利要求1-44中任一项的多晶型物，以及药学上可接受的载体和/或赋形剂。
  46. 权利要求45的药物组合物，其中所属多晶型物为晶型I。
  47. 一种治疗哺乳动物的肿瘤、慢性炎症疾病、炎症性肠道疾病、皮肤病、糖尿病、眼部疾病、与哺乳动物的血管发生或血管再生相关的疾病、与慢性疼痛相关的疾病和其它由Mek级联调制的疾病的方法，所述方法包括对所述哺乳动物给药权利要求1-44中任一项的多晶型物。
  48. 权利要求1-44中任一项的多晶型物，其用于治疗哺乳动物的肿瘤、慢性炎症疾病、炎症性肠道疾病、皮肤病、糖尿病、眼部疾病、与哺乳动物的血管发生或血管再生相关的疾病、与慢性疼痛相关的疾病和其它由Mek级联调制的疾病。
  49. 权利要求1-44中任一项的多晶型物在制备用于治疗哺乳动物的肿瘤、慢性炎症疾病、炎症性肠道疾病、皮肤病、糖尿病、眼部疾病、与哺乳动物的血管发生或血管再生相关的疾病、与慢性疼痛相关的疾病和其它由Mek级联调制的疾病中的用途。
  50. 一种制备权利要求4晶型I的方法，所述方法包括以下任一种：
    a)将化合物1无定形样品加至溶剂中，然后在约70℃以上的温度加热，所得清液降温至室温，室温保温继续搅拌，固体析出，过滤，干燥；或
    b)将化合物1无定形样品溶于良溶剂中，过滤得到澄清溶液，边搅拌该澄清溶液边加入反溶剂至有固体析出；或
    c)将化合物1无定形样品溶于溶剂中，在约50℃搅拌，然后过滤取滤液，将所得滤液降温至约5℃，收集析出的固体。
  51. 权利要求50的方法，其中方法1)中加热温度为约75℃至约100℃；约80℃至约90℃；或约75℃或约85℃。
  52. 权利要求50的方法，其中方法1)中室温为约20-约25℃。
  53. 权利要求50的方法，其中方法1)中继续搅拌的时间为约1-约12小时、约1-约8小时、约1-约5小时或更长，或者约24-约96小时或更长。
  54. 权利要求50的方法，其中方法1)中清液在约2至约5小时内或约2.5-约3小时内降温至室温。
  55. 权利要求50的方法，其中方法1)中室温保温继续搅拌后可任选地将温度进一步降低至约0-约10℃，保温搅拌。
  56. 权利要求50的方法，其中在约0-约10℃保温搅拌约1-约12小时、约1-约8小时、约1-约5小时或更长。
  57. 权利要求50的方法，其中方法1)中所述溶剂为水、甲醇、乙醇、异丙醇、丙酮、甲基异丁基酮、2-丁酮、乙酸乙酯、乙酸异丙酯、甲基叔丁基醚、四氢呋喃、苯甲醚、2-甲基四氢呋喃、环戊基甲醚、1,4-二氧六环、乙腈、二氯甲烷、甲苯、间二甲苯、正庚烷、正己烷、正戊烷、二甲亚砜、二甲基乙酰胺、N-甲基吡咯烷酮或其混合物。
  58. 权利要求50的方法，其中方法1)中所述溶剂为乙醇。
  59. 权利要求50的方法，其中方法2)中的良溶剂为化合物1能溶于其中的溶剂，方法2)中的反溶剂为化合物1不能溶于其中的溶剂。
  60. 权利要求50的方法，其中方法2)中的良溶剂为MEK、1,4-二氧六环或DMSO；方法2)中的反溶剂为MTBE、EtOAc、CHCl3、正庚烷、Anisole、EtOAc、H2O、IPAc、CPME、DCM或甲苯。
  61. 权利要求50的方法，其中方法3)所述降温速度为0.1℃/分钟。
  62. 权利要求50的方法，其中方法3)的溶剂为MIBK、Methyl acetate、2-MeTHF或 丙酮/EtOH(1:1)。
  63. 一种制备权利要求35晶型V的方法，所述方法包括：将化合物1无定形样品加至溶剂中，然后在约70℃以下的温度加热，所得清液降温至约0-约10℃，在该温度静置直至固体析出，干燥。
  64. 权利要求63的方法，其中所述方法在约70℃、约60℃或约50℃加热。
  65. 权利要求63的方法，其中所述降温速度为约0.1-约0.5℃/分。
  66. 权利要求63的方法，其中所述降温速度为约0.1℃/分。
  67. 权利要求63的方法，其中所得清液降温至约5℃。
  68. 一种治疗哺乳动物RAS或RAF突变型癌症的方法，所述方法包括对所述哺乳动物给药4-氟-5-(2-氟-4-碘苯基氨基)-1H-苯并[d]噻唑-6-羧酸(2-羟基-乙氧基)-酰胺(化合物1)或其可药用盐。
  69. 权利要求68的方法，其中所述RAS或RAF突变型癌症为例如KRAS突变型癌症、NRAS突变型癌症、HRAS突变型癌症或BRAF突变癌症。
  70. 权利要求68或69的方法，其中所述RAS突变型癌症是胰腺癌、结直肠癌、肺癌、黑色素瘤、急性骨髓性白血病、膀胱癌或头颈癌等。
  71. 权利要求68至70的方法，其中所述癌症是NRAS突变型癌症。
  72. 权利要求71的方法，其中所述NRAS突变型癌症是NRAS突变的黑色素瘤。
  73. 权利要求72的方法，其中KRAS包括在选自密码子12、13、59和61的一个或多个位置处的突变。
  74. 权利要求68至70的方法，其中NRAS包括在选自密码子12、13、59、61和146的一个或多个位置处的突变。
  75. 权利要求68至70的方法，其中所述NRAS突变形式在选自G12、G13、A59、Q61、K117和A146的一个或多个氨基酸位置处具有突变。
  76. 权利要求75的方法，其中所述NRAS突变形式具有选自以下的一个或多个氨基酸取代：G12C、G12R、G12S、G12A、G12D、G12V、G13C、G13R、G13S、G13A、G13D、G13V、A59D、A59T、Q61K、Q61L、Q61R、Q61H、K117N、K117R、K117E、A146P、A146T和A146V。
  77. 权利要求68至70的方法，其中所述癌症为早期、中期或晚期阶段癌症。癌症可以是局部晚期的或转移性的。
  78. 权利要求68至70的方法，其中所述哺乳动物之前接受过免疫治疗。
  79. 权利要求78的方法，其中所述哺乳动物之前接受过免疫治疗且患有NRAS突变的晚期黑色素瘤。
  80. 权利要求68至70的方法，其中所述黑色素瘤选自：晚期黑色素瘤、不可切除的黑色素瘤、转移性黑色素瘤、具有BRAF突变的黑色素瘤、具有NRAS突变的黑色素瘤、皮肤黑色素瘤、或眼内黑色素瘤。
  81. 权利要求68至80的方法，其中化合物1为胶囊剂的形式。
  82. 权利要求81的方法，其中化合物1以每次5-50mg，每日一次或两次的剂量给药。
  83. 权利要求82的方法，其中化合物1以每次12mg，每日两次的剂量给药。
  84. 权利要求68至83的方法，其中所述化合物1为晶型I至晶型VI中的任一种多晶型物。
  85. 权利要求68至83的方法，其中所述化合物1为晶型I。
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