WO2021249450A1 - 酪氨酸激酶抑制剂的盐型、晶型、药物组合物及其用途 - Google Patents

酪氨酸激酶抑制剂的盐型、晶型、药物组合物及其用途 Download PDF

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WO2021249450A1
WO2021249450A1 PCT/CN2021/099282 CN2021099282W WO2021249450A1 WO 2021249450 A1 WO2021249450 A1 WO 2021249450A1 CN 2021099282 W CN2021099282 W CN 2021099282W WO 2021249450 A1 WO2021249450 A1 WO 2021249450A1
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phosphate
crystal form
ray powder
compound represented
structural formula
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PCT/CN2021/099282
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French (fr)
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付邦
陈亮
柯永新
吴颢
丁列明
王家炳
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贝达药业股份有限公司
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Priority to CN202180030519.2A priority Critical patent/CN115551863A/zh
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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
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • 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
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems

Definitions

  • This application relates to the salt form of the tyrosine kinase (Protein Tyrosine Kinase, TRK) inhibitor BPI-125, the crystal form of various salt forms, the preparation method of the crystal form, the pharmaceutical composition containing the crystal form, the crystal The use of type and pharmaceutical composition as TRK inhibitors, and the use of treatment of TRK-mediated diseases.
  • TRK Protein Tyrosine Kinase
  • Tropomyosin-related kinases are a class of receptor tyrosine kinases regulated by neurotrophic factors, including three members, TrkA, TrkB, and TrkC, which are encoded by the genes NTRK1, NTRK2, and NTRK3, respectively. Many cell functions, such as cell proliferation, cell differentiation, metabolism and apoptosis, are mediated by Trks through phosphorylation and regulation of its downstream signaling pathway members. Gene fusion involving NTRK genes leads to the continuous activation or overexpression of these kinases, thereby increasing the risk of tumorigenesis.
  • Trk plays an important physiological role in the development of nerves, including the growth and function maintenance of nerve axons, memory development and neuron damage protection.
  • the results indicate that Trk is abnormally expressed in normal tissues or cancer tissues, and fusion can cause abnormally high expression and activation of the Trk kinase domain. Trk fusion is found in a variety of tumor tissues with low fusion rate such as thyroid cancer, lung cancer, colon cancer and melanoma. It is estimated that 1500-5000 patients in the United States suffer from Trk fusion-positive cancer each year.
  • Trk fusion protein has gradually become an effective tumor target.
  • the fastest growing Trk small molecule inhibitor is Loxo Oncology's larotrectinib, which has a strong clinical inhibitory effect on Trk.
  • Previous applications, WO2010048314, WO2011006074, WO2016097869 and WO2018077246 disclosed a series of Trk inhibitors. Accordingly, there is still a need for Trk inhibitors with stronger activity and better metabolic stability of liver microsomes. In addition, in view of the importance of the physiological functions of Trk, there is a great demand for Trk inhibitors.
  • Such inhibitors can not only inhibit Trk A, B, and C, but also inhibit the mutant forms of Trk A, B, and C (such as G595R, G667C). , A608D, F589L and G623R), these mutations have been reported in patients receiving first-generation Trk kinase inhibitors.
  • BPI-125 is an effective small molecule with Trk inhibitor activity, and its structure is shown in structural formula I. The application of this compound in the treatment or prevention of cancer is disclosed in the PCT patent application PCT/CN2019/123719.
  • BPI-125 in the form of free base (FB) has poor solubility in water and low bioavailability, which is not the preferred form of clinical medication.
  • the polymorphs of specific organic drug compounds have different physical properties, such as solubility, moisture absorption, and stability due to their unique three-dimensional structure. However, it is usually impossible to predict whether a specific organic drug compound will form different crystalline forms, let alone the structure and properties of the crystalline form itself. Exploring new crystalline or polymorphic forms of medicinal compounds provides an opportunity to improve the overall performance of pharmaceutical products, and at the same time expands the variety of materials available for formulation scientists to design. The discovery of new crystal forms of useful compounds has expanded the variety of materials for formulation design, which is obviously advantageous.
  • the present invention relates to a compound represented by structural formula I (R)-2-(5-(2-(2,5-difluorophenyl)pyrrolidin-1-yl)pyrazolo[1,5-a]pyrimidine-3 -Yl)-6-methoxy-1H-benzo[d]imidazole-5-carbonitrile salt form, and the crystal form of each salt.
  • the preparation method and structure of the compound represented by structural formula I have been specifically described and disclosed in PCT patent application PCT/CN2019/123719.
  • an acid and BPI-125 free base (hereinafter referred to as BPI-125FB) can form corresponding salts in the corresponding system, and these salt compounds can exist in various physical forms. For example, it may be in the form of a solution, suspension or solid. In certain embodiments, the salt-type compound is in solid form. When in solid form, the compound may be amorphous, crystalline or a mixture thereof.
  • the salt form of BPI-125 is a salt form formed by BPI-125 free base and 27 acids in 5 solvents, wherein the 27 acids are hydrochloric acid, sulfuric acid, phosphoric acid, hydrobromic acid, Maleic acid, tartaric acid, fumaric acid, mucic acid, citric acid, L-malic acid, hippuric acid, L-lactic acid, succinic acid, adipic acid, acetic acid, 1,5-naphthalenedisulfonic acid, 1,2- Ethylenesulfonic acid, p-toluenesulfonic acid, methanesulfonic acid, benzenesulfonic acid, oxalic acid, orotic acid, malonic acid, gentisic acid, D-(+)-camphoric acid, niacin, benzoic acid,
  • the five solvents are acetone, ethyl acetate, tetrahydrofuran, acetonitrile, methanol/water.
  • the salt forms are hydrochloride, sulfate, phosphate, maleate, L-tartrate, fumarate, citrate, L-malate, L-lactate, respectively , Succinate, methanesulfonate, p-toluenesulfonate, benzenesulfonate.
  • the salt form of the Trk inhibitor BPI-125 is phosphate.
  • Exemplary examples of different crystal forms of various salts of Trk inhibitor BPI-125 are as follows:
  • the present invention provides a preferred crystal form of the phosphate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 5.1° ⁇ 0.2°, 8.9° ⁇ 0.2°, 11.5° ⁇ 0.2° And the characteristic peak of 17.9° ⁇ 0.2°.
  • phosphate crystal form A it is referred to as phosphate crystal form A in the present invention.
  • the present invention provides another preferred crystalline form of the phosphate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystalline form has diffraction angles 2 ⁇ of 5.0° ⁇ 0.2°, 13.3° ⁇ 0.2°, 17.7° ⁇ 0.2 ° and 20.1° ⁇ 0.2° characteristic peaks.
  • phosphate crystal form B it is referred to as phosphate crystal form B in the present invention.
  • the present invention provides another preferred crystalline form of the phosphate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystalline form has diffraction angles 2 ⁇ of 5.4° ⁇ 0.2°, 11.7° ⁇ 0.2°, 21.5° ⁇ 0.2 ° characteristic peak. For convenience, it is referred to as phosphate crystal form C in the present invention.
  • the present invention provides another preferred crystal form of the phosphate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 4.8° ⁇ 0.2°, 10.4° ⁇ 0.2°, 15.8° ⁇ 0.2 ° and 20.4° ⁇ 0.2° characteristic peaks.
  • phosphate crystal form D for convenience, it is referred to as phosphate crystal form D in the present invention.
  • the present invention provides another preferred crystal form of the phosphate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 4.7° ⁇ 0.2°, 15.8° ⁇ 0.2°, 18.8° ⁇ 0.2 ° and 20.3° ⁇ 0.2° characteristic peaks.
  • phosphate crystal form E it is referred to as phosphate crystal form E in the present invention.
  • the present invention provides another preferred crystal form of the phosphate of the Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has characteristic peaks with diffraction angles 2 ⁇ of 5.0° ⁇ 0.2° and 8.6° ⁇ 0.2°. For convenience, it is referred to as phosphate crystal form F in the present invention.
  • the present invention provides a preferred crystal form of the hydrochloride salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 5.1° ⁇ 0.2°, 6.4° ⁇ 0.2°, 8.0° ⁇ 0.2 ° and 11.7° ⁇ 0.2° characteristic peaks.
  • hydrochloride crystal form A it is referred to as hydrochloride crystal form A in the present invention.
  • the present invention provides another preferred crystal form of the hydrochloride salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 5.3° ⁇ 0.2°, 9.0° ⁇ 0.2°, 12.0° ⁇
  • the characteristic peaks are 0.2°, 15.4° ⁇ 0.2°, 17.9° ⁇ 0.2°, 21.2° ⁇ 0.2°, 22.0° ⁇ 0.2°, 25.7° ⁇ 0.2° and 26.4° ⁇ 0.2°.
  • hydrochloride crystal form B for convenience, it is referred to as hydrochloride crystal form B in the present invention.
  • the present invention provides a preferred crystal form of the sulfate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 4.8° ⁇ 0.2°, 6.4° ⁇ 0.2°, 7.4° ⁇ 0.2° , 16.0° ⁇ 0.2°, 16.8° ⁇ 0.2°, and 11.7° ⁇ 0.2° characteristic peaks.
  • the sulfate crystal form A it is referred to as the sulfate crystal form A in the present invention.
  • the present invention provides another preferred crystal form of the sulfate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 4.6° ⁇ 0.2°, 15.5° ⁇ 0.2° and 24.7° ⁇ 0.2 ° characteristic peak. For convenience, it is referred to as the sulfate crystal form B in the present invention.
  • the present invention provides another preferred crystal form of the sulfate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 4.7° ⁇ 0.2°, 10.2° ⁇ 0.2°, 18.9° ⁇ 0.2 °, 20.5 and 24.8° ⁇ 0.2° characteristic peaks.
  • the sulfate crystal form C it is referred to as the sulfate crystal form C in the present invention.
  • the present invention provides a preferred crystal form of the L-tartrate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 3.4° ⁇ 0.2°, 5.0° ⁇ 0.2° and 11.5° ⁇ A characteristic peak of 0.2°. For convenience, it is called L-tartrate salt crystal form A in the present invention.
  • the present invention provides another preferred crystal form of the L-tartrate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 7.1° ⁇ 0.2°, 14.1° ⁇ 0.2°, 25.4° The characteristic peaks of ⁇ 0.2° and 25.7° ⁇ 0.2°.
  • L-tartrate salt crystal form B for convenience, it is called L-tartrate salt crystal form B in the present invention.
  • the present invention provides another preferred crystal form of the L-tartrate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has a diffraction angle 2 ⁇ of 5.0° ⁇ 0.2°, 10.9° ⁇ 0.2°, 10.4° The characteristic peaks of ⁇ 0.2° and 26.7° ⁇ 0.2°.
  • L-tartrate salt crystal form C for convenience, it is called L-tartrate salt crystal form C in the present invention.
  • the present invention provides a preferred crystal form of L-malate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 6.8° ⁇ 0.2°, 14.2° ⁇ 0.2°, 26.4° The characteristic peaks of ⁇ 0.2° and 27.2° ⁇ 0.2°. For convenience, it is called L-malate crystal form A in the present invention.
  • the present invention provides another preferred crystal form of L-malate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of this crystal form has diffraction angles 2 ⁇ of 5.0° ⁇ 0.2°, 5.5° ⁇ 0.2°, 5.7 ° ⁇ 0.2°, 6.6° ⁇ 0.2°, 9.0° ⁇ 0.2°, 14.5° ⁇ 0.2°, 21.1° ⁇ 0.2°, 25.5° ⁇ 0.2°, 25.6° ⁇ 0.2° and 26.0° ⁇ 0.2° characteristic peaks.
  • L-malate crystal form B for convenience, it is called L-malate crystal form B in the present invention.
  • the present invention provides a preferred crystal form of L-lactate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has characteristic peaks with diffraction angles 2 ⁇ of 5.0° ⁇ 0.2° and 6.6° ⁇ 0.2° .
  • L-lactate crystal form A for convenience, it is called L-lactate crystal form A in the present invention.
  • the present invention provides a preferred crystal form of the besylate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has a characteristic peak with a diffraction angle 2 ⁇ of 4.6° ⁇ 0.2°. For convenience, it is referred to as benzenesulfonate crystal form A in the present invention.
  • the present invention provides a preferred crystalline form of malonate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystalline form has diffraction angles 2 ⁇ of 5.0° ⁇ 0.2°, 20.2° ⁇ 0.2° and 25.4° ⁇ A characteristic peak of 0.2°. For convenience, it is referred to as malonate crystal form A in the present invention.
  • the present invention provides a preferred crystal form of the oxalate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 5.0° ⁇ 0.2°, 7.9° ⁇ 0.2°, 8.1° ⁇ 0.2 ° and 26.0° ⁇ 0.2° characteristic peaks. For convenience, it is called oxalate crystal form A in the present invention.
  • the present invention provides another preferred crystal form of the oxalate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 6.1° ⁇ 0.2°, 6.5° ⁇ 0.2° and 18.6° ⁇ A characteristic peak of 0.2°. For convenience, it is referred to as oxalate crystal form B in the present invention.
  • the present invention provides a preferred crystalline form of p-toluenesulfonate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystalline form has diffraction angles 2 ⁇ of 6.0° ⁇ 0.2°, 7.0° ⁇ 0.2°, 15.6° ⁇ 0.2°, 18.0° ⁇ 0.2°, 18.2° ⁇ 0.2°, 19.3° ⁇ 0.2° and 24.3° ⁇ 0.2° characteristic peaks.
  • p-toluenesulfonate crystal form A in the present invention.
  • the present invention provides a preferred crystal form of the fumarate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 6.8° ⁇ 0.2°, 9.7° ⁇ 0.2°, 26.0° ⁇ 0.2° and 26.9° ⁇ 0.2° characteristic peaks. For convenience, it is referred to as fumarate crystal form A in the present invention.
  • the present invention provides a preferred crystal form of the succinate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has characteristic peaks with diffraction angles 2 ⁇ of 6.5° ⁇ 0.2° and 6.4° ⁇ 0.2°. For convenience, it is called succinate crystal form A in the present invention.
  • the present invention provides another preferred crystal form of the succinate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 6.8° ⁇ 0.2°, 9.7° ⁇ 0.2°, 14.7° ⁇ 0.2°, 25.8° ⁇ 0.2° and 26.7° ⁇ 0.2° characteristic peaks.
  • succinate crystal form B for convenience, it is called succinate crystal form B in the present invention.
  • the present invention provides a preferred crystal form of the adipate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 4.7° ⁇ 0.2°, 6.2° ⁇ 0.2° and 6.6° ⁇ A characteristic peak of 0.2°.
  • the present invention refers to the adipate salt crystal form A.
  • the present invention provides a preferred crystal form of the mesylate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 4.9° ⁇ 0.2°, 6.6° ⁇ 0.2°, 10.1° ⁇ 0.2°, 13.5° ⁇ 0.2° and 17.8° ⁇ 0.2° characteristic peaks.
  • the present invention refers to mesylate salt crystal form A.
  • the present invention provides another preferred crystalline form of the mesylate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystalline form has characteristic peaks with diffraction angles 2 ⁇ of 6.0° ⁇ 0.2° and 18.2° ⁇ 0.2° .
  • mesylate salt crystal form B it is referred to as mesylate salt crystal form B in the present invention.
  • the present invention provides another preferred crystalline form of the mesylate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of this crystalline form has diffraction angles 2 ⁇ of 6.0° ⁇ 0.2°, 18.2° ⁇ 0.2°, 26.4° The characteristic peaks of ⁇ 0.2° and 27.1° ⁇ 0.2°.
  • the present invention refers to mesylate salt crystal form C.
  • the present invention provides a preferred crystalline form of the maleate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystalline form has diffraction angles 2 ⁇ of 4.2° ⁇ 0.2°, 9.0° ⁇ 0.2°, 13.8° ⁇ 0.2°, 14.4° ⁇ 0.2°, 15.4° ⁇ 0.2°, 16.7° ⁇ 0.2°, 18.0° ⁇ 0.2°, 13.8° ⁇ 0.2°, 24.9° ⁇ 0.2°, 25.5° ⁇ 0.2° and 26.3° ⁇ 0.2°
  • the characteristic peak For convenience, the present invention refers to the maleate salt crystal form A.
  • the present invention provides another preferred crystal form of the maleate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 5.0° ⁇ 0.2°, 5.5° ⁇ 0.2°, 5.6°
  • the characteristic peaks are ⁇ 0.2°, 9.0° ⁇ 0.2°, 14.5° ⁇ 0.2°, 21.1° ⁇ 0.2°, 25.5° ⁇ 0.2°, 25.6° ⁇ 0.2° and 26.0° ⁇ 0.2°.
  • the present invention refers to the maleate salt crystal form B.
  • the present invention provides another preferred crystalline form of the maleate salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystalline form has diffraction angles 2 ⁇ of 6.6° ⁇ 0.2°, 26.2° ⁇ 0.2° and 27.0° The characteristic peak of ⁇ 0.2°.
  • the present invention refers to the maleate salt crystal form C.
  • the present invention provides a preferred crystal form of the citrate of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 6.4° ⁇ 0.2°, 26.0° ⁇ 0.2° and 26.8° ⁇ 0.2 ° characteristic peak. For convenience, it is referred to as citrate crystal form A in the present invention.
  • the present invention provides a preferred crystal form of the hydrobromide salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 6.2° ⁇ 0.2°, 7.3° ⁇ 0.2°, 16.1° ⁇ 0.2°, 18.5° ⁇ 0.2°, 19.8° ⁇ 0.2°, 22.6° ⁇ 0.2°, 25.6° ⁇ 0.2° and 26.5° ⁇ 0.2° characteristic peaks.
  • hydrobromide salt crystal form A in the present invention.
  • the present invention provides another preferred crystal form of the hydrobromide salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has diffraction angles 2 ⁇ of 5.4° ⁇ 0.2°, 15.7° ⁇ 0.2°, 21.9° ⁇ 0.2°, 25.4° ⁇ 0.2° and 26.0° ⁇ 0.2° characteristic peaks.
  • hydrobromide salt crystal form B for convenience, it is referred to as hydrobromide salt crystal form B in the present invention.
  • the present invention provides another preferred crystal form of the hydrobromide salt of Trk inhibitor BPI-125.
  • the X-ray powder diffraction pattern of the crystal form has a diffraction angle 2 ⁇ of 6.2° ⁇ 0.2°, 6.5° ⁇ 0.2°, 8.4 ⁇ 0.2°, 18.8° ⁇ 0.2° and 26.1° ⁇ 0.2° characteristic peaks.
  • hydrobromide salt crystal form C for convenience, it is referred to as hydrobromide salt crystal form C in the present invention.
  • the X-ray powder diffraction patterns are all measured using the K ⁇ line of the Cu target.
  • the present invention further provides a preferred embodiment of the Trk inhibitor BPI-125 phosphate crystal form C:
  • the X-ray powder diffraction pattern of the phosphate crystal form C has characteristic peaks with diffraction angles 2 ⁇ of 5.4° ⁇ 0.2°, 11.7° ⁇ 0.2°, and 21.5° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the phosphate crystal form C has diffraction angles 2 ⁇ of 5.4° ⁇ 0.2°, 11.7° ⁇ 0.2°, 15.8° ⁇ 0.2°, 17.8° ⁇ 0.2°, 21.5° ⁇ 0.2° Characteristic peaks.
  • the X-ray powder diffraction pattern of the phosphate crystal form C has diffraction angles 2 ⁇ of 5.4° ⁇ 0.2°, 10.7° ⁇ 0.2°, 11.7° ⁇ 0.2°, 15.8° ⁇ 0.2°, 17.8° ⁇ 0.2°, 21.5° ⁇ 0.2°, 23.4° ⁇ 0.2°, 25.2° ⁇ 0.2°, 31.5° ⁇ 0.2° characteristic peaks.
  • the main data of the X-ray powder diffraction pattern of the phosphate crystal form C is shown in Table 1.
  • the phosphate crystal form C has an X-ray powder diffraction pattern approximately as shown in FIG. 1.
  • the phosphate crystal form C has a differential scanning calorimetry (DSC) spectrum substantially as shown in FIG. 2.
  • DSC differential scanning calorimetry
  • the phosphate crystal form C has a thermogravimetric analysis (TGA) pattern substantially as shown in FIG. 3.
  • TGA thermogravimetric analysis
  • the phosphate crystal form C has a DVS pattern substantially as shown in FIG. 5, and the figure shows that the phosphate crystal form C has a weight gain of about 0.17% at 25° C./80% RH.
  • the present invention further provides a preferred embodiment of the Trk inhibitor BPI-125 phosphate crystal form A:
  • the X-ray powder diffraction pattern of the phosphate crystal form A has diffraction angles 2 ⁇ of 5.1° ⁇ 0.2°, 8.9° ⁇ 0.2°, 11.5° ⁇ 0.2°, 15.4° ⁇ 0.2°, 17.9° ⁇ 0.2° The characteristic peak.
  • the X-ray powder diffraction pattern of the phosphate crystal form A has diffraction angles 2 ⁇ of 5.1° ⁇ 0.2°, 8.9° ⁇ 0.2°, 11.5° ⁇ 0.2°, 15.4° ⁇ 0.2°, 17.9° ⁇ 0.2 °, 20.5° ⁇ 0.2°, 26.7° ⁇ 0.2° characteristic peaks.
  • the main data of the X-ray powder diffraction pattern of the phosphate crystal form A is shown in Table 2.
  • the phosphate crystal form A has an X-ray powder diffraction pattern approximately as shown in FIG. 6.
  • the phosphate crystal form A has a differential scanning calorimetry (DSC) spectrum substantially as shown in FIG. 7.
  • DSC differential scanning calorimetry
  • the phosphate crystal form A has a thermogravimetric analysis (TGA) profile substantially as shown in FIG. 8.
  • TGA thermogravimetric analysis
  • the phosphate crystal form A has a 1 H-NMR spectrum substantially as shown in FIG. 9.
  • the phosphate crystal form A has a DVS pattern substantially as shown in FIG. 10, and the figure shows that the phosphate crystal form A gains about 0.44% by moisture absorption at 25° C./80% RH.
  • the present invention further provides a preferred embodiment of the Trk inhibitor BPI-125 phosphate crystal form B:
  • the X-ray powder diffraction pattern of the phosphate crystal form B has diffraction angles 2 ⁇ of 5.0° ⁇ 0.2°, 13.3° ⁇ 0.2°, 15.3° ⁇ 0.2°, 17.7° ⁇ 0.2°, 20.1° ⁇ 0.2° The characteristic peak.
  • the X-ray powder diffraction pattern of the phosphate crystal form B has a diffraction angle 2 ⁇ of 5.0° ⁇ 0.2°, 8.8° ⁇ 0.2°, 10.0° ⁇ 0.2°, 13.3° ⁇ 0.2°, 15.3° ⁇ 0.2 °, 17.7° ⁇ 0.2°, 19.6° ⁇ 0.2°, 20.1° ⁇ 0.2°, 21.7° ⁇ 0.2°, 22.1° ⁇ 0.2°, 25.6° ⁇ 0.2°.
  • the phosphate crystal form B has an X-ray powder diffraction pattern approximately as shown in FIG. 11.
  • the phosphate crystal form B has a differential scanning calorimetry (DSC) spectrum substantially as shown in FIG. 12.
  • DSC differential scanning calorimetry
  • the phosphate crystal form B has a thermogravimetric analysis (TGA) profile substantially as shown in FIG. 13.
  • TGA thermogravimetric analysis
  • the phosphate crystal form B has a 1 H-NMR spectrum substantially as shown in FIG. 14.
  • the present invention further provides a preferred embodiment of the Trk inhibitor BPI-125 phosphate crystal form D:
  • the X-ray powder diffraction pattern of the phosphate crystal form D has diffraction angles 2 ⁇ of 4.8° ⁇ 0.2°, 10.4° ⁇ 0.2°, 15.8° ⁇ 0.2°, 20.4° ⁇ 0.2°, 24.3° ⁇ 0.2° The characteristic peak.
  • the X-ray powder diffraction pattern of the phosphate crystal form D has diffraction angles 2 ⁇ of 4.8° ⁇ 0.2°, 10.4° ⁇ 0.2°, 12.5° ⁇ 0.2°, 15.4° ⁇ 0.2°, 15.8° ⁇ 0.2 °, 19.3° ⁇ 0.2°, 20.4° ⁇ 0.2°, 23.2° ⁇ 0.2°, 23.4° ⁇ 0.2°, 24.3° ⁇ 0.2°, 30.2° ⁇ 0.2°.
  • the phosphate crystal form D has an X-ray powder diffraction pattern approximately as shown in FIG. 15.
  • the phosphate crystal form D has a differential scanning calorimetry (DSC) spectrum substantially as shown in FIG. 16.
  • DSC differential scanning calorimetry
  • the phosphate crystal form D has a thermogravimetric analysis (TGA) pattern substantially as shown in FIG. 17.
  • TGA thermogravimetric analysis
  • the phosphate crystal form D has a 1 H-NMR spectrum substantially as shown in FIG. 18.
  • the present invention further provides a preferred embodiment of the Trk inhibitor BPI-125 phosphate crystal form E:
  • the X-ray powder diffraction pattern of the phosphate crystal form E has a diffraction angle 2 ⁇ of 4.7° ⁇ 0.2°, 15.2° ⁇ 0.2°, 15.8° ⁇ 0.2°, 18.8° ⁇ 0.2°, 20.3° ⁇ 0.2° , A characteristic peak of 24.3° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the phosphate crystal form E has a diffraction angle 2 ⁇ of 4.7° ⁇ 0.2°, 10.4° ⁇ 0.2°, 11.3° ⁇ 0.2°, 15.2° ⁇ 0.2°, 15.8° ⁇ 0.2 °, 18.8° ⁇ 0.2°, 20.3° ⁇ 0.2°, 23.4° ⁇ 0.2°, 24.3° ⁇ 0.2° characteristic peaks.
  • the phosphate crystal form E has an X-ray powder diffraction pattern approximately as shown in FIG. 19.
  • the phosphate crystal form E has a differential scanning calorimetry (DSC) spectrum substantially as shown in FIG. 20.
  • the phosphate crystal form E has a thermogravimetric analysis (TGA) profile substantially as shown in FIG. 21.
  • TGA thermogravimetric analysis
  • the phosphate crystal form E has a 1 H-NMR spectrum substantially as shown in FIG. 22.
  • the present invention further provides a preferred embodiment of the Trk inhibitor BPI-125 phosphate crystal form F:
  • the X-ray powder diffraction pattern of the phosphate crystal form F has a diffraction angle 2 ⁇ of 5.0° ⁇ 0.2°, 8.6° ⁇ 0.2°, 13.3° ⁇ 0.2°, 17.6° ⁇ 0.2°, 20.2° ⁇ 0.2° , A characteristic peak of 24.3° ⁇ 0.2°.
  • the X-ray powder diffraction pattern of the phosphate crystal form F has a diffraction angle 2 ⁇ of 5.0° ⁇ 0.2°, 8.6° ⁇ 0.2°, 13.3° ⁇ 0.2°, 15.3° ⁇ 0.2°, 17.6° ⁇ 0.2 °, 20.2° ⁇ 0.2°, 24.3° ⁇ 0.2° characteristic peaks.
  • the phosphate crystal form F has an X-ray powder diffraction pattern approximately as shown in FIG. 23.
  • the phosphate crystal form F has a differential scanning calorimetry (DSC) spectrum substantially as shown in FIG. 24.
  • the phosphate crystal form F has a thermogravimetric analysis (TGA) pattern substantially as shown in FIG. 25.
  • TGA thermogravimetric analysis
  • the phosphate crystal form F has a 1 H-NMR spectrum substantially as shown in FIG. 26.
  • the purity of the phosphate crystal form C, phosphate crystal form A, phosphate crystal form B, phosphate crystal form D, phosphate crystal form E, and phosphate crystal form F is preferably greater than 50%, such as 85 %. % Or more, 99% or more or 99.5% or more.
  • the present invention further provides a method for preparing Trk inhibitor BPI-125 phosphate crystal form C, phosphate crystal form A, phosphate crystal form B, phosphate crystal form D, phosphate crystal form E, and phosphate crystal form F.
  • the preparation method of phosphate crystal form C is as follows:
  • the preparation method of phosphate crystal form C is as in Example 2.
  • the preparation method of phosphate crystal form A is as follows:
  • Phosphate crystal form A was obtained by suspending and stirring the phosphate crystal form C in acetone solvent at room temperature.
  • the preparation method of phosphate crystal form B is as follows:
  • Phosphate crystal form B is obtained by suspending and stirring the phosphate crystal form C in isopropanol at room temperature.
  • the preparation method of phosphate crystal form D is as follows:
  • Phosphate crystal form D is obtained by suspending and stirring the phosphate crystal form C in a methanol solvent system at room temperature.
  • the preparation method of phosphate crystal form E is as follows:
  • Phosphate crystal form E is obtained by suspending and stirring the phosphate crystal form C in an ethanol solvent system at room temperature.
  • the preparation method of phosphate crystal form F is as follows:
  • Phosphate crystal form F is obtained by suspending and stirring the phosphate crystal form C in a dichloromethane solvent system at room temperature.
  • the present invention further provides a pharmaceutical composition, which contains a therapeutically effective amount of different crystal forms of each salt form of the present invention, and pharmaceutically acceptable excipients, adjuvants or carriers.
  • a pharmaceutical composition which contains a therapeutically effective amount of different crystal forms of each salt form of the present invention, and pharmaceutically acceptable excipients, adjuvants or carriers.
  • the weight ratio of the different crystal forms of the various salt forms to the excipients, adjuvants or carriers is in the range of 0.0001-10.
  • the present invention also provides preferred embodiments of the above-mentioned pharmaceutical composition.
  • the above-mentioned pharmaceutical composition contains a therapeutically effective amount of different crystal forms of each salt form of the present invention, in combination with at least one other active ingredient.
  • the pharmaceutical composition is for oral administration.
  • the pharmaceutical composition is used in tablets or capsules.
  • the pharmaceutical composition contains 0.01% to 99% by weight of the crystal form of the present invention.
  • the pharmaceutical composition contains 0.05% to 50% by weight of the crystal form of the present invention.
  • the pharmaceutical composition contains 0.1% to 30% by weight of the crystal form of the present invention.
  • the present invention further provides the application of the crystal form or the pharmaceutical composition in the preparation of medicines.
  • the application is to treat, prevent, delay or prevent the occurrence or progression of cancer or cancer metastasis.
  • the application is the preparation of drugs for the treatment or prevention of diseases mediated by Trk.
  • the disease is cancer.
  • the cancer is selected from the group consisting of breast-like secretory carcinoma of the salivary glands (MASC), infant fibrosarcoma, Spitz tumor, colon cancer, gastric cancer, thyroid cancer (such as papillary thyroid cancer), lung cancer, leukemia, pancreatic cancer, melanoma Cancer (e.g. multiple melanoma), brain cancer (e.g. pontine glioma), kidney cancer (e.g. congenital mesodermal nephroma), prostate cancer, ovarian cancer, or breast cancer (e.g. secretory breast cancer) .
  • MSC salivary glands
  • infant fibrosarcoma fibrosarcoma
  • Spitz tumor such as papillary thyroid cancer
  • colon cancer gastric cancer
  • thyroid cancer such as papillary thyroid cancer
  • lung cancer leukemia
  • pancreatic cancer melanoma Cancer (e.g. multiple melanoma)
  • brain cancer e.g. pontine glioma
  • kidney cancer e.
  • the application is as a Trk inhibitor.
  • the Trk includes wild-type TrkA, TrkB, TrkC or TrkA G595R, TrkA G667C, TrkA A608D, TrkA F589L or TrkC G623R.
  • the present invention also provides a method for administering a therapeutically effective amount of at least any one crystal form or pharmaceutical composition to a subject to treat and/or prevent diseases mediated by Trk.
  • the Trk includes wild-type TrkA, TrkB, TrkC, or TrkA G595R, TrkA G667C, TrkA A608D, TrkA F589L, or TrkC G623R.
  • the Trk-mediated disease is cancer.
  • the cancer is selected from the group consisting of breast-like secretory carcinoma of the salivary glands (MASC), infant fibrosarcoma, Spitz tumor, colon cancer, gastric cancer, thyroid cancer (such as thyroid papillary carcinoma), lung cancer, leukemia , Pancreatic cancer, melanoma (e.g. multiple melanoma), brain cancer (e.g. pontine glioma), kidney cancer (e.g. congenital mesoderm nephroma), prostate cancer, ovarian cancer or breast cancer (e.g. Secretory breast cancer).
  • MSC salivary glands
  • infant fibrosarcoma fibrosarcoma
  • Spitz tumor such as thyroid papillary carcinoma
  • colon cancer gastric cancer
  • thyroid cancer such as thyroid papillary carcinoma
  • lung cancer leukemia
  • Pancreatic cancer melanoma (e.g. multiple melanoma)
  • brain cancer e.g. pontine glioma
  • kidney cancer e.g. congen
  • the present invention also provides a method for treating cancer, comprising administering a therapeutically effective amount of at least any one crystal form or pharmaceutical composition to a subject, the cancer being breast-like secretory carcinoma of the salivary glands (MASC), infant fibrosarcoma, Spitz tumor, colon cancer, stomach cancer, thyroid cancer (e.g. papillary thyroid carcinoma), lung cancer, leukemia, pancreatic cancer, melanoma (e.g. multiple melanoma), brain cancer (e.g. pontine glioma) , Kidney cancer (such as congenital mesoderm nephroma), prostate cancer, ovarian cancer or breast cancer (such as secretory breast cancer).
  • MSC salivary glands
  • infant fibrosarcoma e.g. papillary thyroid carcinoma
  • lung cancer e.g. papillary thyroid carcinoma
  • lung cancer e.g. papillary thyroid carcinoma
  • lung cancer e.g. papillary thyroid carcinoma
  • lung cancer e.g. papillary
  • the subject to be treated is a human being.
  • substantially pure means that the content of the crystal form is not less than 85% by weight, preferably not less than 95%, and more preferably not less than 99%.
  • the precise positions of the peaks in the drawings should not be interpreted as absolute values.
  • the 2 ⁇ value of the X-ray powder diffraction pattern may have errors due to different measurement conditions (such as the equipment and instruments used) and different samples, and the measurement error of the diffraction angle of the X-ray powder diffraction pattern 5% or less, usually, a difference of ⁇ 0.2° from the given value will be considered appropriate.
  • the relative intensity of the peaks may fluctuate with experimental conditions and sample preparation such as the preferred orientation of the particles in the sample.
  • sample preparation such as the preferred orientation of the particles in the sample.
  • the use of automatic or fixed diverging slits will also affect the calculation of relative strength.
  • the intensities shown in the XRD curves included here are only exemplary and cannot be used as an absolute comparison.
  • Annular tautomerism is a type of proton transfer tautomerism, in which protons can occupy two or more positions in the heterocyclic ring. These two isomers Coexist in a balanced system, changing with each other at a very high rate. For example: 1H- and 3H-imidazole; 1H, 2H- and 4H-1,2,4-triazole; 1H- and 2H-isoindole.
  • BPI-125 has 1H- and 3H-imidazole types of intracyclic tautomerism; due to the coexistence of two isomers, for simplicity of description, the present invention only mentions the structure of one of the isomers, namely Any reference to any structure of the intracyclic tautomer at any place means that the other structure is also mentioned at the same time.
  • the tautomerism of the compound The substance of the compound of formula II is also given at the same time.
  • Figure 1 X-ray powder diffraction pattern of the phosphate crystal form C of the Trk inhibitor BPI-125.
  • Figure 2 Differential scanning calorimetry spectrum of the phosphate crystal form C of the Trk inhibitor BPI-125.
  • Figure 3 Thermogravimetric analysis pattern of the phosphate crystal form C of the Trk inhibitor BPI-125.
  • Figure 4 1 H-NMR spectrum of the phosphate crystal form C of the Trk inhibitor BPI-125.
  • Figure 5 DVS pattern of the phosphate crystal form C of the Trk inhibitor BPI-125.
  • Figure 6 X-ray powder diffraction pattern of the phosphate crystal form A of the Trk inhibitor BPI-125.
  • Figure 7 Differential scanning calorimetry spectrum of the phosphate crystal form A of the Trk inhibitor BPI-125.
  • Figure 8 Thermogravimetric analysis pattern of the phosphate crystal form A of the Trk inhibitor BPI-125.
  • Figure 9 1 H-NMR spectrum of the phosphate crystal form A of the Trk inhibitor BPI-125.
  • Figure 10 DVS pattern of the phosphate crystal form A of the Trk inhibitor BPI-125.
  • Figure 11 X-ray powder diffraction pattern of the phosphate crystal form B of the Trk inhibitor BPI-125.
  • Figure 12 Differential scanning calorimetry spectrum of the phosphate crystalline form B of the Trk inhibitor BPI-125.
  • Figure 13 Thermogravimetric analysis pattern of the phosphate crystal form B of the Trk inhibitor BPI-125.
  • Figure 14 1 H-NMR spectrum of the phosphate crystal form B of the Trk inhibitor BPI-125.
  • Figure 15 X-ray powder diffraction pattern of the phosphate crystal form D of the Trk inhibitor BPI-125.
  • Figure 16 Differential scanning calorimetry spectrum of the phosphate crystal form D of the Trk inhibitor BPI-125.
  • Figure 17 Thermogravimetric analysis pattern of the phosphate crystal form D of the Trk inhibitor BPI-125.
  • Figure 18 1 H-NMR spectrum of the phosphate crystal form D of the Trk inhibitor BPI-125.
  • Figure 19 X-ray powder diffraction pattern of the phosphate crystal form E of the Trk inhibitor BPI-125.
  • Figure 20 Differential scanning calorimetry spectrum of the phosphate crystal form E of the Trk inhibitor BPI-125.
  • Figure 21 Thermogravimetric analysis pattern of the phosphate crystal form E of the Trk inhibitor BPI-125.
  • Figure 22 1 H-NMR spectrum of the phosphate crystal form E of the Trk inhibitor BPI-125.
  • Figure 23 X-ray powder diffraction pattern of the phosphate crystal form F of the Trk inhibitor BPI-125.
  • Figure 24 Differential scanning calorimetry spectrum of the phosphate crystalline form F of the Trk inhibitor BPI-125.
  • Figure 25 Thermogravimetric analysis pattern of the phosphate crystal form F of the Trk inhibitor BPI-125.
  • Figure 26 1 H-NMR spectrum of the phosphate crystal form F of the Trk inhibitor BPI-125.
  • TGA Thermogravimetric Analyzer
  • DSC Differential Scanning Calorimeter
  • the present invention will be further described by the following examples and experimental examples, but the examples and experimental examples do not constitute any limitation to the scope of the present invention.
  • the technique or method is a conventional technique or method in the field.
  • DSC Differential calorimeter scanner
  • RRT relative retention time
  • PK Pharmacokinetics.
  • Step 3 Preparation of ethyl (R)-5-(2-(2-chloro-5-fluorophenyl)pyrrolidin-1-yl)pyrazole[1,5-a]pyrimidine-3-carboxylate
  • Step 2 Preparation of (R)-5-(2-(2,5-difluorophenyl)pyrrolidin-1-yl)pyrazole[1,5-a]pyrimidine-3-carboxylic acid
  • Step 3 (R)-2-(5-(2-(2,5-Difluorophenyl)pyrrolidin-1-yl)pyrazolo[1,5-a]pyrimidin-3-yl)-5 Synthesis of -Methoxy-1H-benzo[d]imidazole-6-nitrile
  • BPI-125 free base can be prepared separately from the above acids in different solvent systems: hydrochloride, sulfate, phosphate, maleate, L-tartrate, fumarate, citrate , L-malate, L-lactate, succinate, methanesulfonate, p-toluenesulfonate, benzenesulfonate and hydrobromide.
  • Aw Water activity, calculated theoretically.
  • Example 7-1 Methanol/water (6:1) Form C
  • Example 7-2 Ethanol/water (6:1) Form C
  • Example 7-3 Isopropyl alcohol/water (6:1) Form C
  • Example 7-4 Acetone/water (6:1) Form C
  • Example 7-5 Tetrahydrofuran/water (6:1) Form C
  • Example 7-6 1,4-Dioxane/water (6:1) Form C
  • Example 7-7 Acetonitrile/water (6:1) Form C
  • Example 7-8 Methanol/water (3:1) Form C
  • Example 7-9 Ethanol/water (3:1) Form C
  • Example 7-10 Isopropyl alcohol/water (3:1) Form C
  • Example 7-11 Acetone/water (3:1) Form C
  • Example 7-12 Tetrahydrofuran/water (3:1) Form C
  • Example 7-13 1,4-Dioxane/water (3:1) Form C
  • Example 7-14 Acetonitrile/water (3:1) Form C
  • Example 7-15 Methanol/water (19:1) Form C
  • the criteria for judging the hygroscopicity are:
  • Deliquescence absorb enough water to form a liquid
  • moisture-absorbing weight gain is not less than 15%
  • Moisture absorption weight gain is less than 15%, but not less than 2%;
  • weight gain by moisture absorption is less than 2%, but not less than 0.2%;
  • the phosphate crystal form C has no or almost no hygroscopicity; the phosphate crystal form A is slightly hygroscopic.
  • ICR mice Eighteen ICR mice were divided into two groups, 9 in each group, all males.
  • the compound crystal form A and compound crystal form C were administered by a single gavage of 24 mg/kg respectively; they passed through the fundus venous plexus at designated time points.
  • the blood was collected, the plasma was separated, and stored in the refrigerator at -80°C.
  • the inhibitory activity of BPI-125 on TrkA kinase was determined by the mobility shift analysis method. The analysis steps are as follows:
  • Stop buffer 100mM HEPES, pH 7.5; 0.015% Brij-35; 0.2% coating reagent #3; 50mM EDTA
  • Inhibition rate (maximum value-conversion value) / (maximum value-minimum value) * 100
  • the “maximum value” is the DMSO control value; the “minimum value” is the value of the control well without kinase.
  • Example 13 BPI-125 inhibits the proliferation of Ba/F3-TPM3-NTRK1, Ba/F3-ETV6-NTRK3 and Ba/F3-TPM3-NTRK1G595R cells
  • Ba/F3 cells with stable expression of TPM3-NTRK or ETV6-NTRK3 fusion mutant genes were named Ba/F3-TPM3-NTRK1 and Ba/F3-ETV6-NTRK3.
  • X logarithm of compound concentration
  • Y luminescence value
  • BPI-125 not only has a strong inhibitory ability on Ba/F3-TPM3-NTRK1 and Ba/F3-ETV6-NTRK3 cells, but also on Ba/F3-TPM3-NTRK1G595R mutant cells. Very strong inhibitory effect.

Abstract

结构式I所示化合物的盐型，以及磷酸盐的多种晶型，其制备方法及其应用。

Description

酪氨酸激酶抑制剂的盐型、晶型、药物组合物及其用途 技术领域
本申请涉及酪氨酸激酶(Protein Tyrosine Kinase,TRK)抑制剂BPI-125的盐型、各种盐型的晶型、晶型的制备方法、含有所述晶型的药物组合物、所述晶型和药物组合物作为TRK抑制剂的用途，以及治疗TRK介导的疾病的用途。
背景技术
原肌球蛋白相关激酶(Trks)是一类受神经营养因子调控的受体酪氨酸激酶，包括TrkA、TrkB和TrkC三个成员，分别由基因NTRK1、NTRK2和NTRK3编码。许多细胞功能，如细胞增殖、细胞分化、代谢和凋亡，都是由Trks通过磷酸化和调控其下游信号通路成员而介导的。涉及NTRK基因的基因融合导致这些激酶的持续激活或过度表达，从而增加肿瘤发生的风险。
Trk在神经的发育过程中起着重要的生理作用，包括神经轴突的生长和功能维持、记忆的发展和神经元的损伤保护等。此外，结果表明，Trk在正常组织或癌组织中表达异常，而融合可引起Trk激酶域的异常高表达和活化。Trk融合见于甲状腺癌、肺癌、结肠癌和黑色素瘤等低融合率的多种肿瘤组织中。据估计，美国每年有1500-5000名患者患有Trk融合阳性癌症。
近年来，Trk融合蛋白正逐渐成为一种有效的肿瘤靶点，其中发展最快的Trk小分子抑制剂是Loxo肿瘤公司的larotrectinib，在临床上对Trk具有很强的抑制作用。先前的申请，WO2010048314、WO2011006074、WO2016097869和WO2018077246公开了一系列Trk抑制剂。相应地，仍然需要具有更强活性和更好的肝微粒体代谢稳定性的Trk抑制剂。另外，鉴于Trk生理学功能的重要性，对Trk抑制剂的需求很大，这种抑制剂不仅可以抑制Trk A、B和C，而且可以抑制Trk A、B和C的突变形式(例如G595R、G667C、A608D、F589L和G623R)，这些突变在接受第一代Trk激酶抑制剂的患者中有报道。
BPI-125是一种具有Trk抑制剂活性的有效小分子，其结构如结构式I所示，在PCT专利申请PCT/CN2019/123719中公开了该化合物在治疗或预防癌症的应用。然而，以游离碱(free base，FB)形式存在的BPI-125在水中溶解性不佳，生物利用度低，不是临床用药的优选形式。
Figure PCTCN2021099282-appb-000001
特定有机药物化合物的多晶型物，由于各自的独特的三维结构，而具有不同的物理性质，如溶解性、引湿性及稳定性等。但是，通常无法预测特定有机药物化合物是否会形成不同的结晶形式，更不可能预测晶型本身的结构和性质。探索可药用化合物的新晶型或多晶型物提供了提高医药产品的整体性能的机会，同时扩大了制剂科学家设计时可用的材料品种。由于发现有用化合物的新晶型而扩大了制剂设计的材料品种，这显然是有利的。
发明内容
本发明涉及结构式I所示化合物(R)-2-(5-(2-(2,5-二氟苯基)吡咯烷-1-基)吡唑并[1,5-a]嘧啶-3-基)-6-甲氧基-1H-苯并[d]咪唑-5-腈的盐型、以及各个盐的晶型。所述结构式I所示的化合物的制备方法和结构均已在PCT专利申请PCT/CN2019/123719中予以具体描述公开。
Trk抑制剂BPI-125的盐型
在一些实施方案中，一种酸和BPI-125游离碱(以下简称为BPI-125FB)可在相应体系中形成相应的盐，这些盐型化合物可以以各种物理形式存在。例如，可以是溶液、悬浮液或固体形式。在某些实施方式中，盐型化合物为固体形式。为固体形式时，所述化合物可以是无定形物，结晶物或其混合物。优选地，BPI-125的盐型为BPI-125游离碱与27种酸在5种溶剂中所形成的盐型，其中，所述的27种酸分别是盐酸、硫酸、磷酸、氢溴酸、马来酸、酒石酸、富马酸、粘酸、柠檬酸、L-苹果酸、马尿酸、L-乳酸、琥珀酸、己二酸、乙酸、1,5-萘二磺酸、1,2-乙二磺酸、对甲苯磺酸、甲磺酸、苯磺酸、草酸、乳清酸、丙二酸、龙胆酸、D-(+)-樟脑酸、烟酸、苯甲酸，所述的5种溶剂分别为丙酮、乙酸乙酯、四氢呋喃、乙腈、甲醇/水。更优选地，所述盐型分别为盐酸盐，硫酸盐，磷酸盐，马来酸盐，L-酒石酸盐，富马酸盐，柠檬酸盐，L-苹果酸盐，L-乳酸盐，琥珀酸盐，甲磺酸盐，对甲苯磺酸盐，苯磺酸盐。最优选地，Trk抑制剂BPI-125的盐型为磷酸盐。
Trk抑制剂BPI-125各种盐的不同晶型示范性举例如下：
本发明提供了Trk抑制剂BPI-125的磷酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.1°±0.2°，8.9°±0.2°，11.5°±0.2°和17.9°±0.2°的特征峰。为方便， 本发明称之为磷酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的磷酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，13.3°±0.2°，17.7°±0.2°和20.1°±0.2°的特征峰。为方便，本发明称之为磷酸盐晶型B。
本发明提供了Trk抑制剂BPI-125的磷酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.4°±0.2°，11.7°±0.2°，21.5°±0.2°的特征峰。为方便，本发明称之为磷酸盐晶型C。
本发明提供了Trk抑制剂BPI-125的磷酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为4.8°±0.2°，10.4°±0.2°，15.8°±0.2°和20.4°±0.2°的特征峰。为方便，本发明称之为磷酸盐晶型D。
本发明提供了Trk抑制剂BPI-125的磷酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为4.7°±0.2°，15.8°±0.2°，18.8°±0.2°和20.3°±0.2°的特征峰。为方便，本发明称之为磷酸盐晶型E。
本发明提供了Trk抑制剂BPI-125的磷酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.0°±0.2°和8.6°±0.2°的特征峰。为方便，本发明称之为磷酸盐晶型F。
本发明提供了Trk抑制剂BPI-125的盐酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.1°±0.2°，6.4°±0.2°，8.0°±0.2°和11.7°±0.2°的特征峰。为方便，本发明称之为盐酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的盐酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.3°±0.2°，9.0°±0.2°，12.0°±0.2°，15.4°±0.2°，17.9°±0.2°，21.2°±0.2°，22.0°±0.2°，25.7°±0.2°和26.4°±0.2°的特征峰。为方便，本发明称之为盐酸盐晶型B。
本发明提供了Trk抑制剂BPI-125的硫酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为4.8°±0.2°，6.4°±0.2°，7.4°±0.2°，16.0°±0.2°，16.8°±0.2°，和11.7°±0.2°的特征峰。为方便，本发明称之为硫酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的硫酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为4.6°±0.2°，15.5°±0.2°和24.7°±0.2°的特征峰。为方便，本发明称之为硫酸盐晶型B。
本发明提供了Trk抑制剂BPI-125的硫酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为4.7°±0.2°，10.2°±0.2°，18.9°±0.2°，20.5和24.8°±0.2°的特征峰。 为方便，本发明称之为硫酸盐晶型C。
本发明提供了Trk抑制剂BPI-125的L-酒石酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为3.4°±0.2°，5.0°±0.2°和11.5°±0.2°的特征峰。为方便，本发明称之为L-酒石酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的L-酒石酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为7.1°±0.2°，14.1°±0.2°、25.4°±0.2°和25.7°±0.2°的特征峰。为方便，本发明称之为L-酒石酸盐晶型B。
本发明提供了Trk抑制剂BPI-125的L-酒石酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，10.9°±0.2°、10.4°±0.2°和26.7°±0.2°的特征峰。为方便，本发明称之为L-酒石酸盐晶型C。
本发明提供了Trk抑制剂BPI-125的L-苹果酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.8°±0.2°，14.2°±0.2°，26.4°±0.2°和27.2°±0.2°的特征峰。为方便，本发明称之为L-苹果酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的L-苹果酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，5.5°±0.2°，5.7°±0.2°，6.6°±0.2°，9.0°±0.2°，14.5°±0.2°，21.1°±0.2°，25.5°±0.2°，25.6°±0.2°和26.0°±0.2°的特征峰。为方便，本发明称之为L-苹果酸盐晶型B。
本发明提供了Trk抑制剂BPI-125的L-乳酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.0°±0.2°和6.6°±0.2°的特征峰。为方便，本发明称之为L-乳酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的苯磺酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为4.6°±0.2°的特征峰。为方便，本发明称之为苯磺酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的丙二酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，20.2°±0.2°和25.4°±0.2°的特征峰。为方便，本发明称之为丙二酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的草酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，7.9°±0.2°，8.1°±0.2°和26.0°±0.2°的特征峰。为方便，本发明称之为草酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的草酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.1°±0.2°，6.5°±0.2°和18.6°±0.2°的特征峰。为方便，本发明称 之为草酸盐晶型B。
本发明提供了Trk抑制剂BPI-125的对甲苯磺酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.0°±0.2°，7.0°±0.2°，15.6°±0.2°，18.0°±0.2°，18.2°±0.2°，19.3°±0.2°和24.3°±0.2°的特征峰。为方便，本发明称之为对甲苯磺酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的富马酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.8°±0.2°，9.7°±0.2°，26.0°±0.2°和26.9°±0.2°的特征峰。为方便，本发明称之为富马酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的琥珀酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.5°±0.2°和6.4°±0.2°的特征峰。为方便，本发明称之为琥珀酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的琥珀酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.8°±0.2°，9.7°±0.2°，14.7°±0.2°，25.8°±0.2°和26.7°±0.2°的特征峰。为方便，本发明称之为琥珀酸盐晶型B。
本发明提供了Trk抑制剂BPI-125的己二酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为4.7°±0.2°，6.2°±0.2°和6.6°±0.2°的特征峰。为方便，本发明称之为己二酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的甲磺酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为4.9°±0.2°，6.6°±0.2°，10.1°±0.2°，13.5°±0.2°和17.8°±0.2°的特征峰。为方便，本发明称之为甲磺酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的甲磺酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.0°±0.2°和18.2°±0.2°的特征峰。为方便，本发明称之为甲磺酸盐晶型B。
本发明提供了Trk抑制剂BPI-125的甲磺酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.0°±0.2°，18.2°±0.2°，26.4°±0.2°和27.1°±0.2°的特征峰。为方便，本发明称之为甲磺酸盐晶型C。
本发明提供了Trk抑制剂BPI-125的马来酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为4.2°±0.2°，9.0°±0.2°，13.8°±0.2°，14.4°±0.2°，15.4°±0.2°，16.7°±0.2°，18.0°±0.2°，13.8°±0.2°，24.9°±0.2°，25.5°±0.2°和26.3°±0.2°的特征峰。为方便，本发明称之为马来酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的马来酸盐的另一种优选晶型，该晶型X射线粉 末衍射图具有衍射角2θ为5.0°±0.2°，5.5°±0.2°，5.6°±0.2°，9.0°±0.2°，14.5°±0.2°，21.1°±0.2°，25.5°±0.2°，25.6°±0.2°和26.0°±0.2°的特征峰。为方便，本发明称之为马来酸盐晶型B。
本发明提供了Trk抑制剂BPI-125的马来酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.6°±0.2°，26.2°±0.2°和27.0°±0.2°的特征峰。为方便，本发明称之为马来酸盐晶型C。
本发明提供了Trk抑制剂BPI-125的柠檬酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.4°±0.2°，26.0°±0.2°和26.8°±0.2°的特征峰。为方便，本发明称之为柠檬酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的氢溴酸盐的一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.2°±0.2°，7.3°±0.2°，16.1°±0.2°，18.5°±0.2°，19.8°±0.2°，22.6°±0.2°，25.6°±0.2°和26.5°±0.2°的特征峰。为方便，本发明称之为氢溴酸盐晶型A。
本发明提供了Trk抑制剂BPI-125的氢溴酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为5.4°±0.2°，15.7°±0.2°，21.9°±0.2°，25.4°±0.2°和26.0°±0.2°的特征峰。为方便，本发明称之为氢溴酸盐晶型B。
本发明提供了Trk抑制剂BPI-125的氢溴酸盐的另一种优选晶型，该晶型X射线粉末衍射图具有衍射角2θ为6.2°±0.2°，6.5°±0.2°，8.4±0.2°，18.8°±0.2°和26.1°±0.2°的特征峰。为方便，本发明称之为氢溴酸盐晶型C。
所述X射线粉末衍射图均使用Cu靶的Kα谱线测得。
本发明进一步提供了所述Trk抑制剂BPI-125磷酸盐晶型C的优选实施方式：
作为优选，该磷酸盐晶型C的X射线粉末衍射图具有衍射角2θ为5.4°±0.2°，11.7°±0.2°，21.5°±0.2°的特征峰。
作为优选，该磷酸盐晶型C的X射线粉末衍射图具有衍射角2θ为5.4°±0.2°，11.7°±0.2°，15.8°±0.2°，17.8°±0.2°，21.5°±0.2°的特征峰。
作为优选，该磷酸盐晶型C的X射线粉末衍射图具有衍射角2θ为5.4°±0.2°，10.7°±0.2°，11.7°±0.2°，15.8°±0.2°，17.8°±0.2°，21.5°±0.2°，23.4°±0.2°，25.2°±0.2°，31.5°±0.2°的特征峰。
更优选地，所述磷酸盐晶型C的X射线粉末衍射图的主要数据如表1所示。
表1
Figure PCTCN2021099282-appb-000002
Figure PCTCN2021099282-appb-000003
作为优选，所述磷酸盐晶型C具有约如图1所示的X射线粉末衍射图。
进一步地，所述磷酸盐晶型C具有基本上如图2所示的差示扫描量热(DSC)图谱。
进一步地，所述磷酸盐晶型C具有基本上如图3所示的热重分析(TGA)图谱。
进一步地，所述磷酸盐晶型C具有基本上如图5所示的DVS图谱，图中显示所述磷酸盐晶型C在25℃/80％RH引湿增重约为0.17％。
本发明进一步提供了所述Trk抑制剂BPI-125磷酸盐晶型A的优选实施方式：
作为优选，所述磷酸盐晶型A的X射线粉末衍射图具有衍射角2θ为5.1°±0.2°，8.9°±0.2°，11.5°±0.2°，15.4°±0.2°，17.9°±0.2°的特征峰。
更优选地，所述磷酸盐晶型A的X射线粉末衍射图具有衍射角2θ为5.1°±0.2°，8.9°±0.2°，11.5°±0.2°，15.4°±0.2°，17.9°±0.2°，20.5°±0.2°，26.7°±0.2°的特征峰。
更优选地，所述磷酸盐晶型A的X射线粉末衍射图的主要数据如表2所示。
表2
Figure PCTCN2021099282-appb-000004
作为优选，所述磷酸盐晶型A具有约如图6所示的X射线粉末衍射图。
进一步地，所述磷酸盐晶型A具有基本上如图7所示的差示扫描量热(DSC)图谱。
进一步地，所述磷酸盐晶型A具有基本上如图8所示的热重分析(TGA)图谱。
进一步地，所述磷酸盐晶型A具有基本上如图9所示的 1H-NMR图谱。
进一步地，所述磷酸盐晶型A具有基本上如图10所示的DVS图谱，图中显示所述磷酸盐晶型A在25℃/80％RH引湿增重约为0.44％。
本发明进一步提供了所述Trk抑制剂BPI-125磷酸盐晶型B的优选实施方式：
作为优选，所述磷酸盐晶型B的X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，13.3°±0.2°，15.3°±0.2°，17.7°±0.2°，20.1°±0.2°的特征峰。
更优选地，所述磷酸盐晶型B的X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，8.8°±0.2°，10.0°±0.2°，13.3°±0.2°，15.3°±0.2°，17.7°±0.2°，19.6°±0.2°，20.1°±0.2°，21.7°±0.2°，22.1°±0.2°，25.6°±0.2°的特征峰。
作为优选，所述磷酸盐晶型B具有约如图11所示的X射线粉末衍射图。
进一步地，所述磷酸盐晶型B具有基本上如图12所示的差示扫描量热(DSC)图谱。
进一步地，所述磷酸盐晶型B具有基本上如图13所示的热重分析(TGA)图谱。
进一步地，所述磷酸盐晶型B具有基本上如图14所示的 1H-NMR图谱。
本发明进一步提供了所述Trk抑制剂BPI-125磷酸盐晶型D的优选实施方式：
作为优选，所述磷酸盐晶型D的X射线粉末衍射图具有衍射角2θ为4.8°±0.2°，10.4°±0.2°，15.8°±0.2°，20.4°±0.2°，24.3°±0.2°的特征峰。
更优选地，所述磷酸盐晶型D的X射线粉末衍射图具有衍射角2θ为4.8°±0.2°，10.4°±0.2°，12.5°±0.2°，15.4°±0.2°，15.8°±0.2°，19.3°±0.2°，20.4°±0.2°，23.2°±0.2°，23.4°±0.2°，24.3°±0.2°，30.2°±0.2°的特征峰。
作为优选，所述磷酸盐晶型D具有约如图15所示的X射线粉末衍射图。
进一步地，所述磷酸盐晶型D具有基本上如图16所示的差示扫描量热(DSC)图谱。
进一步地，所述磷酸盐晶型D具有基本上如图17所示的热重分析(TGA)图谱。
进一步地，所述磷酸盐晶型D具有基本上如图18所示的 1H-NMR图谱。
本发明进一步提供了所述Trk抑制剂BPI-125磷酸盐晶型E的优选实施方式：
作为优选，所述磷酸盐晶型E的X射线粉末衍射图具有衍射角2θ为4.7°±0.2°，15.2°±0.2°，15.8°±0.2°，18.8°±0.2°，20.3°±0.2°，24.3°±0.2°的特征峰。
更优选地，所述磷酸盐晶型E的X射线粉末衍射图具有衍射角2θ为4.7°±0.2°，10.4°±0.2°，11.3°±0.2°，15.2°±0.2°，15.8°±0.2°，18.8°±0.2°，20.3°±0.2°，23.4°±0.2°，24.3°±0.2°的特征峰。
作为优选，所述磷酸盐晶型E具有约如图19所示的X射线粉末衍射图。
进一步地，所述磷酸盐晶型E具有基本上如图20所示的差示扫描量热(DSC)图谱。
进一步地，所述磷酸盐晶型E具有基本上如图21所示的热重分析(TGA)图谱。
进一步的，所述磷酸盐晶型E具有基本上如图22所示的 1H-NMR图谱。
本发明进一步提供了所述Trk抑制剂BPI-125磷酸盐晶型F的优选实施方式：
作为优选，所述磷酸盐晶型F的X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，8.6°±0.2°，13.3°±0.2°，17.6°±0.2°，20.2°±0.2°，24.3°±0.2°的特征峰。
更优选地，所述磷酸盐晶型F的X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，8.6°±0.2°，13.3°±0.2°，15.3°±0.2°，17.6°±0.2°，20.2°±0.2°，24.3°±0.2°的特征峰。
作为优选，所述磷酸盐晶型F具有约如图23所示的X射线粉末衍射图。
进一步地，所述磷酸盐晶型F具有基本上如图24所示的差示扫描量热(DSC)图谱。
进一步地，所述磷酸盐晶型F具有基本上如图25所示的热重分析(TGA)图谱。
进一步地，所述磷酸盐晶型F具有基本上如图26所示的 1H-NMR图谱。
根据本发明，所述磷酸盐晶型C、磷酸盐晶型A、磷酸盐晶型B、磷酸盐晶型D、磷酸盐晶型E、磷酸盐晶型F的纯度优选大于50％，例如85％以上、99％以上或99.5％以上。
本发明进一步提供了制备Trk抑制剂BPI-125磷酸盐晶型C、磷酸盐晶型A、磷酸盐晶型B、磷酸盐晶型D、磷酸盐晶型E、磷酸盐晶型F的方法。
其中，磷酸盐晶型C的制备方法如下：
反应容器中加入BPI-125游离碱样品、四氢呋喃和水，搅拌升温至50度，溶清后热滤，滤液中滴加磷酸溶液，室温搅拌，过滤、滤饼用四氢呋喃和水混合溶剂打浆，抽滤，淋洗。滤饼在60℃下真空干燥制得磷酸盐晶型C。
优选地，磷酸盐晶型C的制备方法如实施例2。
其中，磷酸盐晶型A的制备方法如下：
由磷酸盐晶型C在丙酮溶剂中室温悬浮搅拌得到磷酸盐晶型A。
其中，磷酸盐晶型B的制备方法如下：
由磷酸盐晶型C在异丙醇中通过室温悬浮搅拌得到磷酸盐晶型B。
其中，磷酸盐晶型D的制备方法如下：
由磷酸盐晶型C在甲醇溶剂体系中通过室温悬浮搅拌得到磷酸盐晶型D。
其中，磷酸盐晶型E的制备方法如下：
由磷酸盐晶型C在乙醇溶剂体系中通过室温悬浮搅拌得到磷酸盐晶型E。
其中，磷酸盐晶型F的制备方法如下：
由磷酸盐晶型C在二氯甲烷溶剂体系中通过室温悬浮搅拌得到磷酸盐晶型F。
本发明进一步提供了一种药物组合物，其含有治疗有效量的本发明所述各盐型的不同晶型，和药学上可接受的辅料、辅助剂或载体。在上述药物组合物中，所述各盐型的不同晶型和所述辅料、辅助剂或载体的重量比范围是0.0001～10。
其次，本发明还提供了上述药物组合物的优选实施方式。
作为优选，上述药物组合物含有治疗有效量的本发明的各盐型的不同晶型，联用至少一种其他的活性成分。
作为优选，所述药物组合物用于口服给药。
作为优选，所述药物组合物用于片剂或胶囊。
作为优选，所述药物组合物含有0.01重量％-99重量％的本发明的晶型。
作为优选，所述药物组合物含有0.05重量％-50重量％的本发明的晶型。
作为优选，所述药物组合物含有0.1重量％-30重量％的本发明的晶型。
本发明进一步提供了所述晶型或药物组合物在制备药物中的应用。
本发明进一步提供了所述应用的优选技术方案：
作为优选，所述应用为治疗、预防、延迟或阻止癌症或癌症转移的发生或进展。
作为优选，所述应用为制备治疗或预防由Trk介导的疾病的药物。
作为优选，所述疾病是癌症。
作为优选，所述癌症选自唾液腺的乳腺类似分泌癌(MASC)、婴儿纤维肉瘤、斯皮茨瘤、结肠癌、胃癌、甲状腺癌(例如甲状腺乳头状癌)、肺癌、白血病、胰腺癌、黑素瘤(例如多发性黑素瘤)、脑癌(例如桥脑神经胶质瘤)、肾癌(例如先天性中胚层肾瘤)、前列腺癌、卵巢癌或乳腺癌(例如分泌型乳腺癌)。
作为优选，所述应用为用作Trk抑制剂。
作为优选，在上述应用中，所述Trk包括野生型TrkA、TrkB、TrkC或TrkA G595R、TrkA G667C、TrkA A608D、TrkA F589L或TrkC G623R。
本发明还提供了一种在治疗对象上施用治疗有效量的至少任意一种晶型或药物组合物治疗和/或预防由Trk介导的疾病的方法。
作为优选，在上述方法中，所述Trk包括野生型TrkA、TrkB、TrkC或TrkA G595R、TrkA G667C、TrkA A608D、TrkA F589L或TrkC G623R。
作为优选，在上述方法中，所述Trk介导的疾病是癌症。
作为优选，在上述方法中，所述癌症选自唾液腺的乳腺类似分泌癌(MASC)、婴儿 纤维肉瘤、斯皮茨瘤、结肠癌、胃癌、甲状腺癌(例如甲状腺乳头状癌)、肺癌、白血病、胰腺癌、黑素瘤(例如多发性黑素瘤)、脑癌(例如桥脑神经胶质瘤)、肾癌(例如先天性中胚层肾瘤)、前列腺癌、卵巢癌或乳腺癌(例如分泌型乳腺癌)。
本发明还提供了一种治疗癌症的方法，包括向治疗对象施用治疗有效量的至少任意一种晶型或药物组合物，所述癌症是唾液腺的乳腺类似分泌癌(MASC)、婴儿纤维肉瘤、斯皮茨瘤、结肠癌、胃癌、甲状腺癌(例如甲状腺乳头状癌)、肺癌、白血病、胰腺癌、黑素瘤(例如多发性黑素瘤)、脑癌(例如桥脑神经胶质瘤)、肾癌(例如先天性中胚层肾瘤)、前列腺癌、卵巢癌或乳腺癌(例如分泌型乳腺癌)。
作为优选，在上述方法中，所述的治疗对象为人类。
本发明的所有晶型都是基本上纯的。
本文所用的术语“基本上纯的”是指所述晶型的含量以重量计，不小于85％，优选不小于95％，更优选不小于99％。
本发明中，“具有约如图1所示的X射线粉末衍射图”或“其X射线粉末衍射图基本上如图1所示”中所使用的术语“约”和“基本上”是表示附图中的峰的精确位置不应当被解释为绝对值。因为本领域技术人员可知，X射线粉末衍射图的2θ值可能会由于不同的测量条件(如所使用的设备和仪器)和不同的样品而产生误差，X射线粉末衍射图的衍射角的测量误差为5％或更小，通常，给定的值的±0.2°的差别会被认为是恰当的。还应理解，峰值的相对强度可能随实验条件和样品制备诸如颗粒在样品中的优选的取向而波动。自动或固定的发散狭缝的使用也将会影响相对强度的计算。在这里所包括的XRD曲线所示强度只是示例性的，不能被用作绝对比较。
本领域的技术人员应当理解，环内互变异构(Annular tautomerism)是质子转移互变异构的一种，其中质子可以占杂环中的两个或多个位置，这两个异构体共存于一个平衡体系中，以相当高的速率互相变换着。例如：1H-和3H-咪唑；1H,2H-和4H-1,2,4-三唑；1H-和2H-异吲哚。本发明中BPI-125即存在1H-和3H-咪唑类型的环内互变异构；由于两个异构体共存，为叙述简便，本发明只提及其中一种异构体的结构，即在任何处提及环内互变异构体其中任何一种结构，则表示也同时提及另一种结构，如虽然本发明中只给出结构式I的化合物，但该化合物的互变异构体结构式II化合物实质也是同时给出的。
Figure PCTCN2021099282-appb-000005
本领域的技术人员也会理解，由于样品纯度、样品制备以及测量条件(例如加热速率)的变化，由DSC测量的数据可能会发生小的变化。应当理解，通过其它种类的仪器或通过使用不同于那些在下文中描述的条件，可能会给出可替换的熔点的读数。因此，本申请所引用的吸热图并不作为绝对值，且当解释DSC数据时将考虑这样的测量误差。
附图说明
图1：Trk抑制剂BPI-125的磷酸盐晶型C的X射线粉末衍射图。
图2：Trk抑制剂BPI-125的磷酸盐晶型C的差示扫描量热图谱。
图3：Trk抑制剂BPI-125的磷酸盐晶型C的热重分析图谱。
图4：Trk抑制剂BPI-125的磷酸盐晶型C的 1H-NMR图谱。
图5：Trk抑制剂BPI-125的磷酸盐晶型C的DVS图谱。
图6：Trk抑制剂BPI-125的磷酸盐晶型A的X射线粉末衍射图。
图7：Trk抑制剂BPI-125的磷酸盐晶型A的差示扫描量热图谱。
图8：Trk抑制剂BPI-125的磷酸盐晶型A的热重分析图谱。
图9：Trk抑制剂BPI-125的磷酸盐晶型A的 1H-NMR图谱。
图10：Trk抑制剂BPI-125的磷酸盐晶型A的DVS图谱。
图11：Trk抑制剂BPI-125的磷酸盐晶型B的X射线粉末衍射图。
图12：Trk抑制剂BPI-125的磷酸盐晶型B的差示扫描量热图谱。
图13：Trk抑制剂BPI-125的磷酸盐晶型B的热重分析图谱。
图14：Trk抑制剂BPI-125的磷酸盐晶型B的 1H-NMR图谱。
图15：Trk抑制剂BPI-125的磷酸盐晶型D的X射线粉末衍射图。
图16：Trk抑制剂BPI-125的磷酸盐晶型D的差示扫描量热图谱。
图17：Trk抑制剂BPI-125的磷酸盐晶型D的热重分析图谱。
图18：Trk抑制剂BPI-125的磷酸盐晶型D的 1H-NMR图谱。
图19：Trk抑制剂BPI-125的磷酸盐晶型E的X射线粉末衍射图。
图20：Trk抑制剂BPI-125的磷酸盐晶型E的差示扫描量热图谱。
图21：Trk抑制剂BPI-125的磷酸盐晶型E的热重分析图谱。
图22：Trk抑制剂BPI-125的磷酸盐晶型E的 1H-NMR图谱。
图23：Trk抑制剂BPI-125的磷酸盐晶型F的X射线粉末衍射图。
图24：Trk抑制剂BPI-125的磷酸盐晶型F的差示扫描量热图谱。
图25：Trk抑制剂BPI-125的磷酸盐晶型F的热重分析图谱。
图26：Trk抑制剂BPI-125的磷酸盐晶型F的 1H-NMR图谱。
除非另有说明，本发明所用到的检测仪器信息和检测方法参数如表3-表7所示：
表3
Figure PCTCN2021099282-appb-000006
表4
设备名称 热重分析仪(TGA)
设备型号 Discovery TGA 550
样品盘 Al 2O 3坩埚
保护气体 氮气
气体流速 40mL/min
检测方法 Ramp 10℃/min
表5
设备名称 差示扫描量热仪(DSC)
设备型号 Discovery DSC 2500
样品盘 铝坩埚
保护气体 氮气
气体流速 50mL/min
检测方法 Ramp 10℃/min
表6
设备名称 动态蒸汽吸附仪
厂家 Surface Measurement Systems
设备型号 DVS Resolution 1
样品盘 铝坩埚
检测样品量 30-50mg
保护气体 氮气
气体流速 200sccm
表7
Figure PCTCN2021099282-appb-000007
具体实施方式
下面通过给出的各实施例和实验例对本发明作出进一步说明，但所述实施例和实验例并不能对本发明要求保护的范围构成任何限制。在本发明的具体实施例中，除非特别说明，所述技术或方法为本领域的常规技术或方法等。
缩略语：
DSC：差示热量扫描仪；
DVS：动态蒸汽吸附仪
h：小时；
min：分钟；
RRT：相对保留时间；
ND：未检出；
XRPD：X射线粉末衍射；
1H-NMR：核磁共振氢谱；
TGA：热重分析；
PK：药代动力学。
实施例1 BPI-125游离碱的制备方法
步骤1：4-氨基-2-甲氧基-5-硝基苯甲腈的制备
Figure PCTCN2021099282-appb-000008
在15℃下，向CH 3ONa(14.6g)的MeOH(300mL)溶液中，加入4-氨基-2-氟-5-硝基苯甲腈(9.8g)。然后将溶液升温至室温并搅拌8h。LCMS显示反应完成，减压浓缩以除去MeOH，向残余物中加入1L水，并用2N HCl水溶液调节pH至4-5。过滤，得到固体用水洗涤。在50℃下减压干燥10小时，得到产物4-氨基-2-甲氧基-5-硝基苯甲腈(9.6g)。
步骤2：4,5-二氨基-2-甲氧基苯甲腈的制备
Figure PCTCN2021099282-appb-000009
向4-氨基-2-甲氧基-5-硝基苯甲腈(9.6g)的DCM/MeOH(1:1,60mL)溶液中加入饱和的NH 4Cl(aq)(60mL)溶液。添加Zn粉至混合物(32.5g)，然后将混合物在室温搅拌2h。LCMS显示反应完成。将反应混合物过滤并将滤液用DCM(3*100mL)萃取，合并有机层，用盐水洗涤，减压浓缩，残余物用combi flash纯化(PE:EA＝50％:50％)得到产物4,5-二氨基-2-甲氧基苯甲腈(7.3g)，为红色固体。
步骤3：乙基(R)-5-(2-(2-氯-5-氟苯基)吡咯烷-1-基)吡唑[1,5-a]嘧啶-3-羧酸酯的制备
Figure PCTCN2021099282-appb-000010
向(R)-2-(2,5-二氟苯基)吡咯烷盐酸盐(76g)的1-BuOH(1L)溶液中加入乙基5-氯吡唑[1,5-a]嘧啶-3-羧酸酯(78g)和DIEA(89g)。将混合物加热至120℃反应14h。通过LCMS监测直至反应完成。
将混合物在减压下浓缩以除去1-BuOH，将残余物倒入冰水中，并用EA(300mL×3)萃取，合并有机层，用盐水洗涤并经Na2SO4干燥。真空浓缩，残余物用己烷(500mL)洗涤，得到终产物乙基(R)-5-(2-(2-氯-5-氟苯基)吡咯烷-1-基)吡唑[1,5-a]嘧啶-3-羧酸酯(122g，95％)，为白色固体。
步骤2：(R)-5-(2-(2,5-二氟苯基)吡咯烷-1-基)吡唑[1,5-a]嘧啶-3-羧酸的制备
Figure PCTCN2021099282-appb-000011
向乙基(R)-5-(2-(2-氯-5-氟苯基)吡咯烷-1-基)吡唑[1,5-a]嘧啶-3-羧酸酯(122g)的EtOH(1L)溶液中加入LiOH(1M,1L)水溶液。将反应混合物加热至80℃，反应8小时。通过LCMS监测直至反应完成。将混合物真空浓缩以除去EtOH，向残余物中加入水(1L)，并用HCl(1M)酸化至pH＝4～5，过滤，将固体用水洗涤，真空干燥，得到终产物(R)-5-(2-(2,5-二氟苯基)吡咯烷-1-基)吡唑[1,5-a]嘧啶-3-羧酸(110g,98％)，为白色固体。
步骤3：(R)-2-(5-(2-(2,5-二氟苯基)吡咯烷-1-基)吡唑并[1,5-a]嘧啶-3-基)-5-甲氧基-1H-苯并[d]咪唑-6-腈的合成
Figure PCTCN2021099282-appb-000012
向(R)-5-(2-(2,5-二氟苯基)吡咯烷-1-基)吡唑并[1,5-a]嘧啶-3-羧酸(3.44g)的POCl 3(30mL)溶液中加入4,5-二氨基-2-甲氧基苯甲腈(1.96g)。将混合物加热到90℃并搅拌3h。LCMS显示反应完成。冷却至室温并减压浓缩以除去POCl 3，将残留物倒入水(300mL)中析出固体，过滤，滤饼加入到1N NaOH水溶液(100mL)中搅拌过夜，过滤，滤饼水洗，在 60℃下真空干燥10h得到终产物BPI-125游离碱。
实施例2-1 BPI-125磷酸盐晶型C的制备方法
Figure PCTCN2021099282-appb-000013
搅拌下，向反应釜中加入BPI-125游离碱(1.58Kg)、四氢呋喃(31.43Kg)和纯化水(3.92Kg)，升温至50.8℃，待体系溶清后热过滤，滤液中滴加磷酸溶液(0.85Kg 85wt.％磷酸溶于6.29Kg四氢呋喃和0.79Kg纯化水中)，5小时滴加完毕，室温搅拌12小时，过滤，滤饼用四氢呋喃(8Kg)纯化水(1Kg)的混合液淋洗，滤饼加入到四氢呋喃(13Kg)和水(1.63Kg)的混合溶剂中，室温打浆12小时，抽滤，滤饼用四氢呋喃(4Kg)和水(0.5Kg)的混合溶剂淋洗。滤饼真空60℃干燥12小时，得BPI-125磷酸盐晶型C的固体(1.85Kg)。
实施例2-2 BPI-125盐酸盐晶型A的制备方法
称取约500mg BPI-125游离碱样品和对应1.2摩尔比的盐酸加入20mL玻璃瓶中，加入10mL乙酸乙酯，25℃下磁力搅拌48h，反应液离心后的固体，50℃真空干燥12h。
实施例2-3 BPI-125盐酸盐晶型B的制备方法
称取约20mg BPI-125和对应1.2摩尔比的盐酸加入2mL玻璃瓶中，加入0.5mL四氢呋喃，25℃下磁力搅拌48h，反应液离心后的固体，50℃真空干燥12h。
实施例2-4 BPI-125L-酒石酸盐晶型A的制备方法
称取约500mg BPI-125游离碱样品和对应1.2摩尔比的L-酒石酸加入20mL玻璃瓶中，加入10mL四氢呋喃，25℃下磁力搅拌48h，反应液离心后的固体，50℃真空干燥12h。
实施例2-5 BPI-125L-酒石酸盐晶型B的制备方法
称取约20mg BPI-125游离碱样品和1.2摩尔比的L-酒石酸加入2mL玻璃瓶中，加入0.5mL丙酮，25℃下磁力搅拌48h，反应液离心后的固体，50℃真空干燥12h。
实施例2-6 BPI-125L-酒石酸盐晶型C的制备方法
称取约500mg BPI-125游离碱样品和对应1.2摩尔比的L-酒石酸加入20mL玻璃瓶中，加入10mL甲醇/水(19:1,v/v)，25℃下磁力搅拌48h，反应液离心后的固体，50℃真空干燥12h。
实施例3离子色谱测定
取BPI-125磷酸盐晶型C约25mg配成浓度为0.1mg/ml的溶液，以无水磷酸钠为对照品，通过离子色谱测定BPI-125磷酸盐晶型C中磷酸的含量，结果如表8所示。
表8
Figure PCTCN2021099282-appb-000014
结果表明，单磷酸盐中磷酸的理论含量与磷酸盐晶型C中磷酸的实际测得含量的比为1:1.07，证明BPI-125磷酸盐晶型C为单磷酸盐。
实施例4成盐筛选试验
根据BPI-125游离碱的pKa值和溶解度，选择了将其与27种酸在5种溶剂(丙酮、乙酸乙酯、四氢呋喃、乙腈、甲醇/水)体系中进行成盐/共晶筛选，具体操作步骤如下：
称取约20mg的BPI-125游离碱样品和对应摩尔比的酸加入HPLC小瓶中。加入0.5ml对应的溶剂，室温下磁力搅拌，观察现象，离心分离干燥得到固体进行XRPD表征。具体结果见下表9：
表9
Figure PCTCN2021099282-appb-000015
Figure PCTCN2021099282-appb-000016
由表征可知，BPI-125游离碱可与上述酸在不同溶剂体系中分别制备得到：盐酸盐，硫酸盐，磷酸盐，马来酸盐，L-酒石酸盐，富马酸盐，柠檬酸盐，L-苹果酸盐，L-乳酸盐，琥珀酸盐，甲磺酸盐，对甲苯磺酸盐，苯磺酸盐和氢溴酸盐。
实施例5稳定性试验
测定BPI-125的盐酸盐晶型A、L-酒石酸盐晶型C、磷酸盐晶型C在0天和5天高湿92.5％、高温60℃、光照4500lx影响因素下的稳定性。实验数据如表10所示。
表10
Figure PCTCN2021099282-appb-000017
Figure PCTCN2021099282-appb-000018
上述结果显示：BPI-125盐酸盐晶型A和BPI-125L-酒石酸盐晶型C在高温、高湿、光照等影响因素条件(尤其是高温和光照)下有明显杂质增长；而磷酸盐晶型C在相同影响条件下则显现出优异的稳定性。
实施例6不同水活度体系室温悬浮搅拌试验
取约40mg-55mg BPI-125磷酸盐晶型C样品，加入1.0毫升相应溶剂得到悬浊液，在室温条件下磁力搅拌1周后，均得到固体。并对得到的固体进行XRPD测试，结果如下表11所示：
表11
Figure PCTCN2021099282-appb-000019
Aw：水活度，理论计算得到。
实验结果显示，Aw≥0.4时，BPI-125磷酸盐晶型C在水或甲醇/水溶剂体系的不同水活度中均可得到磷酸盐晶型C，说明磷酸盐晶型C在水或甲醇/水溶剂体系的不同水活度中均具有较好的稳定性。
实施例7混合体系室温悬浮搅拌试验
取约20mg-50mg磷酸盐晶型C样品，加入0.5-1.0毫升相应溶剂得到悬浊液，在室温条件下磁力搅拌1周后，均得到固体。并对得到的固体进行XRPD测试，结果如下表12所示：
表12
实验编号 溶剂(v:v) 晶型结果
实施例7-1 甲醇/水(6:1) 晶型C
实施例7-2 乙醇/水(6:1) 晶型C
实施例7-3 异丙醇/水(6:1) 晶型C
实施例7-4 丙酮/水(6:1) 晶型C
实施例7-5 四氢呋喃/水(6:1) 晶型C
实施例7-6 1,4-二氧六环/水(6:1) 晶型C
实施例7-7 乙腈/水(6:1) 晶型C
实施例7-8 甲醇/水(3:1) 晶型C
实施例7-9 乙醇/水(3:1) 晶型C
实施例7-10 异丙醇/水(3:1) 晶型C
实施例7-11 丙酮/水(3:1) 晶型C
实施例7-12 四氢呋喃/水(3:1) 晶型C
实施例7-13 1,4-二氧六环/水(3:1) 晶型C
实施例7-14 乙腈/水(3:1) 晶型C
实施例7-15 甲醇/水(19:1) 晶型C
实施例7-16 乙醇/水(19:1) 晶型C
实施例7-17 异丙醇/水(19:1) 晶型C
实施例7-18 正丁醇/水(19:1) 晶型C
实施例7-19 丙酮/水(19:1) 晶型C
实施例7-20 丁酮/水(19:1) 晶型C
实施例7-21 1,4-二氧六环/水(19:1) 晶型C
实施例7-22 乙腈/水(19:1) 晶型C
实施例7-23 甲醇/水(12:1) 晶型C
实施例7-24 乙醇/水(12:1) 晶型C
实施例7-25 异丙醇/水(12:1) 晶型C
实施例7-26 正丁醇/水(12:1) 晶型C
实施例7-27 丙酮/水(12:1) 晶型C
实施例7-28 丁酮/水(12:1) 晶型C
实施例7-29 1,4-二氧六环/水(12:1) 晶型C
实施例7-30 乙腈/水(12:1) 晶型C
结果显示：磷酸盐晶型C在不同混合溶剂体系均具有良好的稳定性。
实施例8磷酸盐晶型转化关系
设置不同温度和不同水活度体系的混悬竞争试验，具体方法如下：
(1)使用丙酮，四氢呋喃，乙腈配置不同水活度的混合溶剂体系，其中水活度值为理论计算得到。
(2)称取适量的磷酸盐晶型A样品加入对应的溶剂体系，分别在5℃，室温(25±3℃)和55℃平衡2h得到饱和溶液，然后用0.22微米的滤膜过滤得到澄清溶液。
(3)加入磷酸盐晶型A/B/C/D/E/F晶型各5毫克，置于5℃、室温(25±3℃)和55℃条件下磁力搅拌。离心分离，得到的固体进行XRPD测试。总结固体晶型结果见表13。
表13
Figure PCTCN2021099282-appb-000020
Figure PCTCN2021099282-appb-000021
*：水活度通过理论计算得到。
混悬竞争结果显示：温度5-55℃/水活度≥0.4时，磷酸盐晶型C热力学上较磷酸盐晶型A/B/D/E/F更稳定。
实施例9晶型引湿性测定
使用动态水分吸附仪(DVS)对BPI-125磷酸盐晶型A和晶型C进行引湿性测定，并进行引湿增重变化检测，检测结果见图5、图10以及表14。
表14样品0％RH至80％RH范围内重量变化
化合物 引湿增重
磷酸盐晶型C 0.17％
磷酸盐晶型A 0.44％
引湿性的判断标准为：
潮解：吸收足量水分形成液体；
极具引湿性：引湿增重不小于15％；
有引湿性：引湿增重小于15％，但不小于2％；
略有引湿性：引湿增重小于2％，但不小于0.2％；
无或几乎无引湿性：引湿增重小于0.2％。
由检测结果可知，磷酸盐晶型C为无或几乎无引湿性；磷酸盐晶型A为略有引湿性。
实施例10 BPI-125磷酸盐与BPI-125的体内药代动力学(PK)对比实验
实验方法：使用比格犬8只，分为两组，每组4只，雌雄各半，分别单次灌胃100mg/kg Trk抑制剂BPI-125磷酸盐以及Trk抑制剂BPI-125游离碱；分别在指定的时间点通过前 肢静脉采血，分离血浆，放入-80℃冰箱保存。
通过乙腈沉淀上述血浆样品中的蛋白，取上清用水稀释2倍，取10μL至LC-MS/MS检测，试验数据如表15所示：
表15
Figure PCTCN2021099282-appb-000022
由以上结果提示，相较于Trk抑制剂BPI-125游离碱，BPI-125的磷酸盐晶型C在比格犬体内吸收更好。
实施例11 BPI-125磷酸盐不同晶型的体内药代动力学实验：
ICR小鼠18只，分为两组，每组9只，均为雄性，分别单次灌胃24mg/kg给药化合晶型A和化合物晶型C；分别在指定的时间点通过眼底静脉丛采血，分离血浆，放入-80℃冰箱保存。
上述血浆样品，乙腈沉淀蛋白，取上清用水稀释2倍，取10μL至LC-MS/MS检测，试验数据如表16所示：
表16
Figure PCTCN2021099282-appb-000023
由以上结果提示，BPI-125磷酸盐晶型A与BPI-125磷酸盐晶型C在体内吸收基本无差异。
BPI-125磷酸盐的PK试验结果显示：生物利用度满足开发需求。
实施例12 BPI-125的TrkA激酶测定
用迁移率变动分析法测定BPI-125对TrkA激酶的抑制活性。分析步骤如下：
1.反应缓冲液
1x激酶基础缓冲液(50mM HEPES,pH 7.5；0.0015％Brij-35)
终止缓冲液(100mM HEPES,pH 7.5；0.015％Brij-35；0.2％涂层试剂#3；50mM EDTA)
2.制备BPI-125溶液:
1)用100％DMSO将BPI-125稀释至反应中最终所需的最高抑制剂浓度的50倍。将100μl BPI-125稀释液转移至96孔板的孔中。
2)将管中的BPI-125转移到96孔存储板上的一个孔中，并通过在下一个孔中将30μl转移到60μl的100％DMSO中来依次稀释BPI-125，依此类推，总共10种浓度。
3)在相同的96孔板中，将100μl 100％DMSO加至无BPI-125对照和无酶对照两个空孔中。将该板标记为源板。
4)制备中间板
从源板转移10μl BPI-125到新的96孔板作为中间板
在中间板的每个孔中加入90μl 1x激酶缓冲液。
在振荡器上将BPI-125在中间板上混合10分钟。
3.制备测定板
1)将每孔5μl从96孔中间板转移到384孔板中，一式两份。例如，将96孔板的A1转移到384孔板的A1和A2中。将96孔板转的A2移到384孔板的A3和A4，依此类推。
4.激酶反应
1)制备2.5倍酶溶液
在1x激酶基本缓冲液中添加激酶。
2)制备2.5倍肽溶液
在1x激酶基本缓冲液中添加FAM标记的肽和ATP。
3)分析板中已经包含5μl化合物的10％DMSO溶液。
4)将2.5倍酶溶液转移到测定板上
在384孔测定板的每个孔中加入10μl 2.5x酶溶液。
5)在室温下孵育10分钟。
6)将2.5倍肽溶液转移至测定板
在384孔测定板的每个孔中加入10μl 2.5x肽溶液。
激酶反应条件如表17所示：
表17
名称 酶(nM) ATP(μM) 肽 肽浓度(μM)
TRKA 5 415 P22 3
7)激酶反应并停止
在28℃下孵育指定的时间。
加入25μl终止缓冲液以终止反应。
5.Caliper读数
收集Caliper数据。
6.曲线拟合
1)从Caliper程序中复制转换数据。
2)将转换值转换为抑制值。
抑制率＝(最大值-转换值)/(最大值-最小值)*100
“最大值”为DMSO对照值；“最小值”为无激酶对照孔值。
3)使用XLFit excel插件版本5.4.0.8中的数据拟合来获得IC 50值。
方程是：Y＝最小抑制率+(最大抑制率-最小抑制率)/(1+(IC 50/X)^斜率)。
试验数据如表18所示：
表18
样品 Trk A激酶IC 50(nM)
BPI-125 1.1
实施例13 BPI-125对Ba/F3-TPM3-NTRK1、Ba/F3-ETV6-NTRK3和Ba/F3-TPM3-NTRK1G595R细胞增殖抑制试验
1.细胞培养
细胞系：具有TPM3-NTRK或ETV6-NTRK3融合突变基因稳定表达的Ba/F3细胞命名为Ba/F3-TPM3-NTRK1和Ba/F3-ETV6-NTRK3。
A.培养基
RPMI 1640和10％FBS和1％PS和2ug/mL嘌呤霉素。
B.细胞复苏
a)预先在37℃水浴中预热培养基。
b)从液氮罐中取出冻存管，迅速将其放入37℃的水浴中，并在1分钟内完全融化。
c)将细胞悬液转移至装有8mL培养基的15mL离心管中，以1000rpm离心5分钟。
d)弃去上清液，将细胞重悬于1mL培养基中，转移至装有15mL培养基的75cm 2烧瓶中，在37℃，5％CO 2的培养箱中培养细胞。
C.细胞传代
a)预先在37℃水浴中预热培养基。
b)将细胞收集到15mL离心管中，以1000rpm离心5分钟。弃去上清液，计数，使细胞密度达到1.49x10 4细胞/mL，然后将其置于37℃，5％CO 2的培养箱中。
2.BPI-125的制备
a)将BPI-125(20mM储备溶液)以100％DMSO稀释至60μM作为起始浓度，然后以"9+0"浓度进行3倍系列稀释。在96孔稀释板中(Thermo，Cat.No.249944)；
b)用培养基将BPI-125溶液稀释1:20倍以制备10倍的工作溶液；
3.细胞铺板
a)取处于对数生长期的细胞，以1000rpm离心5分钟，然后用培养基重悬细胞，然后计数细胞；
b)将细胞以2000个细胞/孔的密度接种到96孔细胞培养板上；
4.BPI-125处理
a)将第2步制备的BPI-125溶液以每孔15μL的量添加到细胞板中，终浓度为300、100、33.33、11.11、3.70、1.23、0.41、0.14、0.05和0nM，DMSO终浓度为0.5％。空白对照孔是培养基(0.5％DMSO)；
c)将细胞在培养箱中再培养72小时。
5.检测
a)移开96孔细胞培养板，并添加50μl CTG试剂(CellTiter Glo试剂盒，promega，Cat#G7573)。
b)将板摇动2分钟，然后在室温下冷却30分钟。
c)使用PerkinElmer读取器读取发光信号值。
实验数据分析
用GraphPad Prism 6.0软件分析数据，得到BPI-125活性的拟合曲线。
根据非线性回归方程拟合BPI-125IC 50：
方程是：Y＝最小抑制率+(最大抑制率-最小抑制率)/(1+(IC 50/X)^斜率)。
X：化合物浓度的对数；Y：发光值。
试验数据如表19所示：
表19
Figure PCTCN2021099282-appb-000024
Figure PCTCN2021099282-appb-000025
由表19可以看出，BPI-125不但对Ba/F3-TPM3-NTRK1和Ba/F3-ETV6-NTRK3细胞具有极强的抑制能力，对Ba/F3-TPM3-NTRK1G595R突变型细胞也可以起到非常强的抑制作用。

Claims (32)

  1. 结构式I所示化合物的盐型：
    Figure PCTCN2021099282-appb-100001
    其特征在于，所述盐型选自磷酸盐、L-酒石酸盐、L-苹果酸盐、L-乳酸盐、丙二酸盐、草酸盐、对甲苯磺酸盐、富马酸盐、琥珀酸盐、己二酸盐、甲磺酸盐、硫酸盐、马来酸盐、柠檬酸盐、乳清酸盐、氢溴酸盐、烟酸盐、盐酸盐、乙酸盐或樟脑酸盐。
  2. 权利要求1所述的结构式I所示化合物的盐型，其特征在于，所述盐型为晶型。
  3. 权利要求1或2所述的结构式I所示化合物的盐型，其特征在于，所述盐型为磷酸盐。
  4. 权利要求1-3所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐晶型C，其X射线粉末衍射图具有衍射角2θ为5.4°±0.2°，11.7°±0.2°，21.5°±0.2°的特征峰。
  5. 权利要求1-4所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐晶型C，其X射线粉末衍射图具有衍射角2θ为5.4°±0.2°，11.7°±0.2°，15.8°±0.2°，17.8°±0.2°，21.5°±0.2°的特征峰。
  6. 权利要求1-5中任意一项权利要求所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐晶型C，其X射线粉末衍射图具有衍射角2θ为5.4°±0.2°，8.9°±0.2°，10.7°±0.2°，11.7°±0.2°，15.8°±0.2°，17.8°±0.2°，21.5°±0.2°，23.4°±0.2°，25.2°±0.2°的特征峰。
  7. 权利要求1-6中任意一项权利要求所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐晶型C，其X射线粉末衍射图基本上如表1所示。
  8. 权利要求1-7中任意一项权利要求所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐晶型C，其X射线粉末衍射图基本上如图1所示。
  9. 权利要求1-3中任意一项权利要求所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型A，其X射线粉末衍射图具有衍射角2θ为5.1°±0.2°， 8.9°±0.2°，11.5°±0.2°，15.4°±0.2°，17.9°±0.2°的特征峰。
  10. 权利要求1-3或9所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型A，其X射线粉末衍射图具有衍射角2θ为5.1°±0.2°，8.9°±0.2°，11.5°±0.2°，15.4°±0.2°，17.9°±0.2°，20.5°±0.2°，26.7°±0.2°的特征峰。
  11. 权利要求1-3或9-10所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型A，其X射线粉末衍射图基本上如图6所示。
  12. 权利要求1-3所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型B，其X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，13.3°±0.2°，15.3°±0.2°，17.7°±0.2°，20.1°±0.2°的特征峰。
  13. 权利要求1-3或12所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型B，其X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，8.8°±0.2°，10.0°±0.2°，13.3°±0.2°，15.3°±0.2°，17.7°±0.2°，19.6°±0.2°，20.1°±0.2°21.7°±0.2°，22.1°±0.2°，25.6°±0.2°的特征峰。
  14. 权利要求1-3或12-13所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型B，其X射线粉末衍射图基本上如图11所示。
  15. 权利要求1-3所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型D，其X射线粉末衍射图具有衍射角2θ为4.8°±0.2°，10.4°±0.2°，15.8°±0.2°，20.4°±0.2°，24.3°±0.2°的特征峰。
  16. 权利要求1-3或15所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型D，其X射线粉末衍射图具有衍射角2θ为4.8°±0.2°，10.4°±0.2°，12.5°±0.2°，15.4°±0.2°，15.8°±0.2°，19.3°±0.2°，20.4°±0.2°，23.2°±0.2°，23.4°±0.2°，24.3°±0.2°，30.2°±0.2°的特征峰。
  17. 权利要求1-3或15-16所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐的晶型D，其X射线粉末衍射图基本上如图15所示。
  18. 权利要求1-3所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型E，其X射线粉末衍射图具有衍射角2θ为4.7°±0.2°，15.2°±0.2°，15.8°±0.2°，18.8°±0.2°，20.3°±0.2°，24.3°±0.2°的特征峰。
  19. 权利要求1-3或18所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型E，其X射线粉末衍射图具有衍射角2θ为4.7°±0.2°，10.4°±0.2°，11.3°±0.2°，15.2°±0.2°，15.8°±0.2°，18.8°±0.2°，20.3°±0.2°，23.4°±0.2°，24.3°±0.2°的特 征峰。
  20. 权利要求1-3或18-19所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型E，其X射线粉末衍射图基本上如图19所示。
  21. 权利要求1-3所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型F，其X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，8.6°±0.2°，13.3°±0.2°，17.6°±0.2°，20.2°±0.2°，24.3°±0.2°的特征峰。
  22. 权利要求1-3或21所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型F，其X射线粉末衍射图具有衍射角2θ为5.0°±0.2°，8.6°±0.2°，13.3°±0.2°，15.3°±0.2°，17.6°±0.2°，20.2°±0.2°，24.3°±0.2°的特征峰。
  23. 权利要求1-3或21-22所述的结构式I所示化合物的盐型，其特征在于，所述磷酸盐为磷酸盐的晶型F，其X射线粉末衍射图基本上如图23所示。
  24. 一种结构式I所示化合物的磷酸盐晶型C的制备方法，其特征在于，由以下步骤制得：
    1)反应容器中加入结构式I所示化合物、四氢呋喃和水，搅拌升温至50度；
    2)溶清后热滤，滤液中滴加磷酸溶液，室温搅拌；
    3)过滤、滤饼用四氢呋喃和水混合溶剂打浆，抽滤，淋洗；
    4)滤饼真空干燥。
  25. 一种药物组合物，其特征在于：含有治疗有效量的权利要求1-23任一项所述的盐型，和药学上可接受的辅料，辅助剂和/或载体。
  26. 如权利要求25所述的药物组合物，其特征在于，所述组合物用于口服给药。
  27. 权利要求1-23任一项所述的盐型或权利要求25或26所述的药物组合物在制备药物中的应用，其特征在于，所述药物用于治疗由Trk介导的疾病。
  28. 如权利要求27所述的应用，其特征在于，所述由Trk介导的疾病为癌症。
  29. 如权利要求28所述的应用，其特征在于，所述的Trk包括野生型TrkA、TrkB、TrkC或TrkA G595R、TrkA G667C、TrkA A608D、TrkA F589L或TrkC G623R。
  30. 如权利要求28所述的应用，其特征在于，所述癌症选自唾液腺的乳腺类似分泌癌、婴儿纤维肉瘤、斯皮茨瘤、结肠癌、胃癌、甲状腺癌、肺癌、白血病、胰腺癌、黑素瘤、脑癌、肾癌、前列腺癌、卵巢癌或乳腺癌。
  31. 权利要求27-30任一项所述的应用，其特征在于，向治疗对象施用治疗有效量的权利要求1-23任一项所述的盐型或权利要求25或26所述的药物组合物。
  32. 如权利要求31所述的应用，其特征在于，所述治疗对象为人类。
PCT/CN2021/099282 2020-06-11 2021-06-10 酪氨酸激酶抑制剂的盐型、晶型、药物组合物及其用途 WO2021249450A1 (zh)

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CN102596957A (zh) * 2009-07-09 2012-07-18 阵列生物制药公司 作为TRK激酶抑制剂的被取代的吡唑并[1,5-a]嘧啶化合物
CN107207514A (zh) * 2014-12-15 2017-09-26 康联制药有限公司 稠环杂芳基化合物及其作为trk抑制剂的用途
WO2018077246A1 (zh) * 2016-10-28 2018-05-03 正大天晴药业集团股份有限公司 用作神经营养因子酪氨酸激酶受体抑制剂的氨基吡唑并嘧啶化合物
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Publication number Priority date Publication date Assignee Title
CN102596957A (zh) * 2009-07-09 2012-07-18 阵列生物制药公司 作为TRK激酶抑制剂的被取代的吡唑并[1,5-a]嘧啶化合物
CN107207514A (zh) * 2014-12-15 2017-09-26 康联制药有限公司 稠环杂芳基化合物及其作为trk抑制剂的用途
WO2018077246A1 (zh) * 2016-10-28 2018-05-03 正大天晴药业集团股份有限公司 用作神经营养因子酪氨酸激酶受体抑制剂的氨基吡唑并嘧啶化合物
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