WO2023280309A1 - 一种三环四氢异喹啉类衍生物的盐型 - Google Patents

一种三环四氢异喹啉类衍生物的盐型 Download PDF

Info

Publication number
WO2023280309A1
WO2023280309A1 PCT/CN2022/104669 CN2022104669W WO2023280309A1 WO 2023280309 A1 WO2023280309 A1 WO 2023280309A1 CN 2022104669 W CN2022104669 W CN 2022104669W WO 2023280309 A1 WO2023280309 A1 WO 2023280309A1
Authority
WO
WIPO (PCT)
Prior art keywords
formula
compound
compound shown
pharmaceutically acceptable
cancer
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/CN2022/104669
Other languages
English (en)
French (fr)
Inventor
贾丽娜
王林
邵启云
冯君
杨俊然
杜振兴
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Shanghai Hengrui Pharmaceutical Co Ltd
Jiangsu Hengrui Pharmaceutical Co Ltd
Original Assignee
Shanghai Hengrui Pharmaceutical Co Ltd
Jiangsu Hengrui Pharmaceutical Co Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Shanghai Hengrui Pharmaceutical Co Ltd, Jiangsu Hengrui Pharmaceutical Co Ltd filed Critical Shanghai Hengrui Pharmaceutical Co Ltd
Priority to AU2022307207A priority Critical patent/AU2022307207A1/en
Priority to MX2024000486A priority patent/MX2024000486A/es
Priority to KR1020247004295A priority patent/KR20240035515A/ko
Priority to CA3225268A priority patent/CA3225268A1/en
Priority to JP2024500624A priority patent/JP2024525643A/ja
Priority to CN202280048549.0A priority patent/CN117642405B/zh
Priority to US18/576,806 priority patent/US20240374581A1/en
Priority to EP22837054.0A priority patent/EP4368624A4/en
Publication of WO2023280309A1 publication Critical patent/WO2023280309A1/zh
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Images

Classifications

    • 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
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D491/00—Heterocyclic compounds containing in the condensed ring system both one or more rings having oxygen atoms as the only ring hetero atoms and one or more rings having nitrogen atoms as the only ring hetero atoms, not provided for by groups C07D451/00 - C07D459/00, C07D463/00, C07D477/00 or C07D489/00
    • C07D491/02—Heterocyclic compounds containing in the condensed ring system both one or more rings having oxygen atoms as the only ring hetero atoms and one or more rings having nitrogen atoms as the only ring hetero atoms, not provided for by groups C07D451/00 - C07D459/00, C07D463/00, C07D477/00 or C07D489/00 in which the condensed system contains two hetero rings
    • C07D491/04—Ortho-condensed systems
    • C07D491/056—Ortho-condensed systems with two or more oxygen atoms as ring hetero atoms in the oxygen-containing ring
    • 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
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/472—Non-condensed isoquinolines, e.g. papaverine
    • A61K31/4725—Non-condensed isoquinolines, e.g. papaverine containing further heterocyclic rings
    • 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
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/4738—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/4741—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems condensed with ring systems having oxygen as a ring hetero atom, e.g. tubocuraran derivatives, noscapine, bicuculline
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P15/00—Drugs for genital or sexual disorders; Contraceptives
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • 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
    • A61P37/00—Drugs for immunological or allergic disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • 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
    • C07B—GENERAL METHODS OF ORGANIC CHEMISTRY; APPARATUS THEREFOR
    • C07B2200/00—Indexing scheme relating to specific properties of organic compounds
    • C07B2200/13—Crystalline forms, e.g. polymorphs

Definitions

  • the disclosure relates to a salt form of a tricyclic tetrahydroisoquinoline derivative, a preparation method thereof, and a medical application, and belongs to the field of pharmacy.
  • fulvestrant was adjusted to a level I recommendation for hormone receptor-positive advanced breast cancer patients without endocrine therapy, and fulvestrant + CDK4/6 inhibitors were added as Class II recommendation; in addition, fulvestrant combined with CDK4/6 inhibitors has become a class I recommendation for patients with non-steroidal AI (NSAI) treatment failure and steroidal AI (SAI) treatment failure.
  • NSAI non-steroidal AI
  • SAI steroidal AI
  • PCT application WO2021139756A discloses that a tricyclic tetrahydroisoquinoline derivative represented by formula (I) is used as an estrogen receptor down-regulator (SERD).
  • SESD estrogen receptor down-regulator
  • the present disclosure provides a pharmaceutically acceptable salt of a compound represented by formula (I), wherein the pharmaceutically acceptable salt is selected from phosphate, sulfate, tartrate, citrate, hydrochloride, hydrobromic acid salt, methanesulfonate, formate, acetate, succinate, maleate, malate or p-toluenesulfonate,
  • the present disclosure provides a pharmaceutically acceptable salt of the compound represented by formula (I), wherein the substance ratio of the compound represented by formula (I) to the acid molecule is selected from 1:5-5:1.
  • the present disclosure provides a pharmaceutically acceptable salt of the compound represented by formula (I), wherein the substance ratio of the compound represented by formula (I) to the acid molecule is 1:1.
  • the present disclosure provides a pharmaceutically acceptable salt of the compound represented by formula (I), wherein the substance ratio of the compound represented by formula (I) to the acid molecule is 1:3.
  • the present disclosure provides a phosphate salt of the compound represented by the formula (I), wherein the substance ratio of the compound represented by the formula (I) to the phosphoric acid molecule is 1:3.
  • the present disclosure provides a crystal form I of a phosphate salt of a compound represented by formula (I), wherein the molar ratio of the compound represented by formula (I) to phosphoric acid molecules is 1:3, and the X-
  • the ray powder diffraction pattern has characteristic peaks at 6.1, 13.5, 19.2, and 22.3.
  • the disclosure provides a method for preparing the phosphate of the compound represented by the formula (I) or the crystal form I of the phosphate of the compound represented by the formula (I), comprising the step of reacting the compound represented by the formula (I) with phosphoric acid.
  • the present disclosure provides a method for preparing crystal form I of a phosphate salt of a compound represented by formula (I), comprising the following steps:
  • Solvent B solution of phosphoric acid is configured, and the solvent B is selected from water or alcohol solvents;
  • a solvent A solution of the compound represented by formula (I) is configured, and the solvent A is selected from acetone, ethyl acetate or methyl tert-butyl ether.
  • a solvent B solution of phosphoric acid is prepared, and the solvent B is selected from methanol, ethanol or water.
  • the present disclosure provides a sulfate salt of the compound represented by the formula (I), wherein the substance ratio of the compound represented by the formula (I) to the sulfuric acid molecule is 1:1.
  • the disclosure provides a crystal form A of the sulfate salt of the compound represented by formula (I), wherein the molar ratio of the compound represented by formula (I) to the sulfuric acid molecule is 1:1, and the X-ray represented by the diffraction angle 2 ⁇ In the powder diffraction pattern, there are characteristic peaks at 6.6, 7.9, 15.1, 20.7, and 22.1.
  • the powder diffraction pattern is shown in Figure 8.
  • the crystal form A of the sulfate salt of the compound shown in formula (I), wherein the molar ratio of the compound shown in formula (I) to the sulfuric acid molecule is 1:1, monoclinic crystal system, axial length Axial angle ⁇ 90°; crystal volume
  • the present disclosure provides a preparation method of the sulfate salt of the compound represented by formula (I) or A crystal form of the sulfate salt of the compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with sulfuric acid.
  • the present disclosure provides a method for crystal form A of a sulfate salt of a compound represented by formula (I), comprising the following steps:
  • a solvent A solution of the compound represented by formula (I) is configured, and the solvent A is selected from acetone, ethyl acetate, acetonitrile, methanol or ethanol.
  • a solvent B solution of sulfuric acid is prepared, and the solvent B is selected from methanol, acetonitrile, ethanol or water.
  • the present disclosure provides an amorphous form of citrate salt of the compound represented by formula (I).
  • the present disclosure provides a method for preparing citrate of the compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with citric acid to form a salt.
  • the present disclosure provides an amorphous form of L-tartrate salt of the compound represented by formula (I).
  • the disclosure provides a method for preparing an L-tartrate salt of a compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with L-tartrate to form a salt.
  • the present disclosure provides a method for preparing the hydrochloride of the compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with hydrochloric acid to form a salt.
  • the disclosure provides a method for preparing a hydrobromide salt of a compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with hydrobromic acid to form a salt.
  • the disclosure provides a method for preparing a methanesulfonic acid salt of a compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with methanesulfonic acid to form a salt.
  • the disclosure provides a method for preparing a formate salt of a compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with formic acid to form a salt.
  • the present disclosure provides a method for preparing the acetate salt of the compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with acetic acid to form a salt.
  • the present disclosure provides a method for preparing a succinate salt of a compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with succinic acid.
  • the disclosure provides a method for preparing a maleate salt of a compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with maleic acid to form a salt.
  • the disclosure provides a method for preparing malate of the compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with malic acid to form a salt.
  • the disclosure provides a method for preparing a p-toluenesulfonic acid salt of a compound represented by formula (I), comprising the step of reacting the compound represented by formula (I) with p-toluenesulfonic acid to form a salt.
  • the crystal form I of the phosphate salt of the compound represented by formula (I) or the crystal form A of the sulfate salt of the compound represented by formula (I) provided by the present disclosure is an X-ray powder represented by a diffraction angle 2 ⁇ angle
  • the error range of the characteristic peak 2 ⁇ angle in the diffraction pattern is ⁇ 0.2.
  • the present disclosure provides a pharmaceutical composition
  • a pharmaceutical composition comprising the following ingredients:
  • the present disclosure provides a preparation method of a pharmaceutical composition, comprising
  • Another aspect of the present disclosure provides the compound phosphate, sulfate, tartrate, citrate, hydrochloride, hydrobromide, methanesulfonate, formate, acetate, succinate represented by the above formula (I) Salt, maleate, malate or p-toluenesulfonate, the I crystal form of the phosphate of the compound shown in the formula (I) or the A crystal form of the sulfate salt of the compound shown in the formula (I), the formula ( I) the amorphous form of the citrate of the compound shown, the amorphous form of the L-tartrate of the compound shown in formula (I), or the use of mixtures and compositions thereof in the preparation of estrogen receptor down-regulators (SERDs).
  • SESDs estrogen receptor down-regulators
  • Another aspect of the present disclosure provides the compound phosphate, sulfate, tartrate, citrate, hydrochloride, hydrobromide, methanesulfonate, formate, acetate, succinate represented by the above formula (I) Salt, maleate, malate or p-toluenesulfonate, the I crystal form of the phosphate of the compound shown in the formula (I) or the A crystal form of the sulfate salt of the compound shown in the formula (I), the formula ( I) the amorphous form of the citrate of the compound shown, the amorphous form of the L-tartrate of the compound shown in formula (I), or the purposes of their mixtures and compositions in the preparation of drugs for the prevention and/or treatment of cancer, wherein Said cancer is preferably selected from breast cancer, endometrial cancer, uterine cancer, cervical cancer, skin cancer, prostate cancer, ovarian cancer, fallopian tube tumors, hemophilia and leukemia.
  • Another aspect of the present disclosure provides the compound phosphate, sulfate, tartrate, citrate, hydrochloride, hydrobromide, methanesulfonate, formate, acetate, succinate represented by the above formula (I) Salt, maleate, malate or p-toluenesulfonate, the I crystal form of the phosphate of the compound shown in the formula (I) or the A crystal form of the sulfate salt of the compound shown in the formula (I), the formula ( I) the amorphous form of the citrate of the compound shown, the amorphous form of the L-tartrate of the compound shown in formula (I), or its mixture, composition in the preparation of prevention and/or treatment of estrogen receptor-mediated or Drugs for dependent diseases or disorders; preferably, wherein said estrogen receptor-mediated or dependent diseases or disorders are selected from cancer, central nervous system defects, cardiovascular system defects, blood system defects , immune and inflammatory diseases, susceptibility to infection, metabolic defects, neurological defects, mental defects and reproductive defects;
  • the "2 ⁇ or 2 ⁇ angle" mentioned in this disclosure refers to the diffraction angle, ⁇ is the Bragg angle, and the unit is ° or degree; the error range of each characteristic peak 2 ⁇ is ⁇ 0.20, specifically -0.20, -0.19, -0.18 , -0.17, -0.16, -0.15, -0.14, -0.13, -0.12, -0.11, -0.10, -0.09, -0.08, -0.07, -0.06, -0.05, -0.04, -0.03, -0.02, - 0.01, 0.00, 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.10, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.20.
  • the "crystallization” mentioned in the present disclosure includes but not limited to stirring crystallization, cooling crystallization, dissolution crystallization and volatilization crystallization.
  • Differential scanning calorimetry or DSC in this disclosure refers to measuring the temperature difference and heat flow difference between the sample and the reference object during the sample heating or constant temperature process, so as to characterize all the physical changes related to thermal effects and Chemical changes, to obtain the phase transition information of the sample.
  • the drying temperature described in the present disclosure is generally 25°C-100°C, preferably 40°C-70°C, and can be dried under normal pressure or under reduced pressure.
  • the values in this disclosure are measured by instruments or calculated by instruments, and there is a certain degree of error. Generally speaking, plus or minus 10% is within a reasonable error range. Of course, the context where the value is used needs to be considered, for example, the content of total impurities. The value is that the error after measurement does not change by more than plus or minus 10%, which can be plus or minus 9%, plus or minus 8%, plus or minus 7%, plus or minus Minus 6%, plus or minus 5%, plus or minus 4%, plus or minus 3%, plus or minus 2% or plus or minus 1%, preferably plus or minus 5%.
  • the starting material used in the preparation method of the disclosed crystal form can be any form of the compound shown in formula I, and the specific form includes but is not limited to: amorphous, any crystal form, hydrate, solvate, etc.
  • crystallization includes but not limited to stirring crystallization, cooling crystallization, beating crystallization and volatilization crystallization.
  • NMR nuclear magnetic resonance
  • MS mass spectroscopy
  • MS was determined with Agilent 1200/1290 DAD-6110/6120 Quadrupole MS liquid mass spectrometer (manufacturer: Agilent, MS model: 6110/6120 Quadrupole MS), waters ACQuity UPLC-QD/SQD (manufacturer: waters, MS Model: waters ACQuity Qda Detector/waters SQ Detector), THERMO Ultimate 3000-Q Exactive (manufacturer: THERMO, MS model: THERMO Q Exactive).
  • HPLC High performance liquid chromatography
  • Chiral HPLC analysis was performed using an Agilent 1260 DAD high performance liquid chromatograph.
  • the CombiFlash rapid preparation instrument uses Combiflash Rf200 (TELEDYNE ISCO).
  • XRPD is X-ray powder diffraction detection: BRUKER D8 DISCOVER X-ray diffractometer is used for the measurement, and the specific collection information: Cu anode (40kV, 40mA), Cu-K ⁇ ray Scanning mode: ⁇ /2 ⁇ , scanning range (2 ⁇ range): 3 ⁇ 48°.
  • DSC is differential scanning calorimetry: the measurement uses METTLER TOLEDO DSC 3+ differential scanning calorimeter, the heating rate is 10°C/min, the specific range of temperature is referred to the corresponding map (25-240°C), and the nitrogen purge rate is 50mL/min.
  • TGA thermogravimetric analysis: METTLER TOLEDO TGA 2 type thermogravimetric analyzer is used for detection, the heating rate is 10°C/min, the specific range of temperature refers to the corresponding map (30-350°C), and the nitrogen purge rate is 20mL/min.
  • DVS dynamic moisture adsorption: SMS DVS Advantage is used for detection. At 25°C, the humidity change is 50%-95%-0%-95%-50%, and the step is 10% (the last step is 5%) (the specific range of humidity Based on the corresponding spectrum, most of the methods are listed here), and the judgment standard is dm/dt ⁇ 0.002%.
  • the thin-layer chromatography silica gel plate uses Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plate.
  • the specification of the silica gel plate used in thin-layer chromatography (TLC) is 0.15mm-0.2mm, and the specification of thin-layer chromatography separation and purification products is 0.4mm. ⁇ 0.5mm.
  • Silica gel column chromatography generally uses Yantai Huanghai silica gel 200-300 mesh silica gel as the carrier.
  • the known starting materials of the present disclosure can be adopted or synthesized according to methods known in the art, or can be purchased from ABCR GmbH & Co. KG, Acros Organics, Aldrich Chemical Company, Shaoyuan Chemical Technology (Accela ChemBio Inc), Darui chemical companies.
  • the reactions can all be carried out under an argon atmosphere or a nitrogen atmosphere.
  • the argon atmosphere or nitrogen atmosphere means that the reaction bottle is connected to an argon or nitrogen balloon with a volume of about 1 L.
  • the hydrogen atmosphere means that the reaction bottle is connected to a hydrogen balloon with a capacity of about 1L.
  • the pressurized hydrogenation reaction uses Parr 3916EKX hydrogenation instrument and Qinglan QL-500 hydrogen generator or HC2-SS hydrogenation instrument.
  • the hydrogenation reaction is usually vacuumized and filled with hydrogen, and the operation is repeated 3 times.
  • the solution refers to an aqueous solution.
  • the temperature of the reaction is room temperature, 20 ⁇ 30 examples.
  • the monitoring of the reaction process in the embodiment adopts thin-layer chromatography (TLC), the developer used for reaction, the eluent system of the column chromatography that purifies compound adopts and the developer system of thin-layer chromatography comprise: A: Dichloromethane/methanol system, B: n-hexane/ethyl acetate system, C: petroleum ether/ethyl acetate system, the volume ratio of the solvent is adjusted according to the polarity of the compound, and a small amount of triethylamine and Alkaline or acidic reagents such as acetic acid for adjustment.
  • TLC thin-layer chromatography
  • Example 1 preparation of the compound shown in formula 1 (priority: the preparation method of Example 13 in the application of 202010680491.4)
  • Test Example 1 Inhibitory Effect of the Disclosed Compound on the Binding of E and ER
  • the disclosed compound has inhibitory effect on the combination of E (estrogen, estrogen) and ER (estrogen receptor, estrogen receptor), thereby blocking the combination of the complex of E and ER and ERE (estrogen response element), and then blocking Downstream expression of luciferase protein.
  • E estrogen, estrogen
  • ER estrogen receptor, estrogen receptor
  • the purpose of this experiment is to test the inhibitory effect of the compound on the combination of E and ER, and evaluate the in vitro activity of the compound according to the IC 50 size.
  • the ERE was cloned upstream of the luciferase gene, and MCF-7/ERE-luciferase (luciferase) monoclonal cells were selected by transfecting MCF-7 (TCHu74, the cell bank of the Type Culture Collection Committee of the Chinese Academy of Sciences).
  • MEM containing 10% charcoal stripped FBS (Moregate, FBSF), 1% sodium pyruvate (sigma, S8636), 1% non-essential amino acids (sigma, M7145) and 500 ⁇ g/ml G418 in a 96-well plate (hyclone, SH30024.01B) medium was used to inoculate MCF-7/ERE-luciferase cells at a seeding density of 30,000 cells/well, and the cells were cultured at 37°C and 5% CO 2 .
  • the drug was formulated as a 20 mM stock solution, diluted with 10-fold gradient with 100% DMSO, and then diluted 20-fold with medium.
  • the medium was removed, and 0.1 nM estradiol (sigma, E2758) and 10 ⁇ L of the drug diluted with the medium were added to each well, and DMSO was added to the control group, and gently oscillated to mix. Place it in a 37°C, 5% CO2 incubator for culture, discard the cell culture medium after 24 hours, add 50 ⁇ L/well of the prepared luciferase substrate (Promega, E6110), place it at room temperature in the dark for 10-15 minutes, and measure Chemiluminescence signal value.
  • the prepared luciferase substrate Promega, E6110
  • the inhibitory effect of the compounds in the present disclosure on the binding of E and ER was determined by the above experiments.
  • Graphpad Prism was used to analyze the chemiluminescent signal value and the logarithmic concentration of the compound to obtain the IC 50 value of the compound. See Table 1 for the results.
  • Test example 2 The inhibitory effect of the disclosed compound on the proliferation of MCF-7 cells
  • the purpose of this experiment is to use the ATP method to test the inhibitory effect of the disclosed compound on the proliferation activity of MCF-7 cells, and evaluate the in vitro activity of the compound according to the IC 50 size.
  • MCF-7 (TCHu74, Cell Bank of Type Culture Collection Committee, Chinese Academy of Sciences) was inoculated in a 96-well plate in a medium containing 10% FBS (Gibco, 10099-141), 1% sodium pyruvate (sigma, S8636), 1 % non-essential amino acids (sigma, M7145) in MEM (hyclone, SH30024.01B) medium, seeded at a density of 4,000 cells/well, and the cells were cultured at 37°C and 5% CO 2 .
  • the compound was formulated as a 20 mM stock solution, diluted with 100% DMSO to a final concentration of 1000X, and then diluted 20 times with medium containing 2% FBS.
  • Graphpad Prism was used to plot the chemiluminescent signal value and the logarithmic concentration of the compound to obtain the IC 50 value of the compound. See Table 2 for the results.
  • Test Example 3 Biological evaluation of the compound of the present disclosure on the inhibition of proliferation of MCF7 cells expressing ER ⁇ mutants
  • the purpose of this experiment is to determine the inhibitory activity of the disclosed compounds on the proliferation of MCF7 cells expressing ER ⁇ mutants.
  • the mutant ER ⁇ Y537S of the human estrogen receptor ⁇ (estrogen receptor ⁇ , ER ⁇ ) protein was obtained by site-directed mutation using the cDNA of the wild-type ESR1 gene (Accession No. NM000125) as a template by double-primer PCR.
  • the primer sequence used for the mutation is as follows (the underlined nucleotide is the mutation site): Y537S: F-AAG AAC GTG GTG CCC CTC TCT GAC CTG CTG CTG GAG ATG (SEQ ID NO: 1); R-CAT CTC CAG CAG CAG GTC AGA GAG GGG CAC CAC GTT CTT (SEQ ID NO: 2).
  • mutant ESR1 was cloned into the target lentiviral vector pCDH-CMV-MCS-EF1-Puro. Then, the lentiviral plasmid and the lentiviral packaging plasmid carrying the mutant ESR1 gene sequence were transfected into HEK-293T cells (ATCC, CRL-3216) by Lipofectamine 3000 Transfection Reagent (ThermoFisher Scientific, Cat#L3000075).
  • MCF7 cells expressing ER ⁇ mutants were cultured with MEM (GE Healthcare, SH30024.01) complete medium containing 10% fetal bovine serum. On the first day of the experiment, cells were seeded in a 96-well plate at a density of 3,000 cells/well using complete medium, 100 ⁇ L of cell suspension per well, and cultured overnight in a cell culture incubator at 37°C and 5% CO 2 .
  • MEM GE Healthcare, SH30024.01
  • Test Example 4 Compounds of the present disclosure have degradative effects on ER ⁇
  • Test Compounds of the Disclosure Cause ERa Degradation This method is used to measure the degradation of ERa by the compounds of the disclosure.
  • ER ⁇ -positive breast cancer cell line MCF-7 was cultured in DMEM/F12 medium (HyClone, SH30023.01) containing 10% fetal bovine serum (Corning, 35-010-CV).
  • DMEM/F12 medium HyClone, SH30023.01
  • fetal bovine serum Corel bovine serum
  • the cells were washed once with phenol red-free DMEM/F12 medium (ThermoFisher, 11039-021) containing 5% activated carbon-treated fetal bovine serum (BIOSUN, BS-0004-500) and re-introduced. Suspend the cells and count them, and adjust the cell density to 1.79 ⁇ 10 5 cells/mL.
  • the capture antibody in the human ER ⁇ /NR3A1 total protein detection kit (R&D, DYC5715-5) to coat the 96-well plate, prepare the capture antibody with PBS to 1 ⁇ g/mL, and mix each in the 96-well plate (Corning, 3590). Add 100 ⁇ L to the wells and place in a 26°C incubator overnight. On the third day of the experiment, the antibody-coated 96-well plate was washed once with PBS, 200 ⁇ L of PBS containing 1% BSA was added to each well, and incubated in a 37° C. incubator for 1.5 hours for blocking.
  • the cell lysate was PBS containing 6M urea, 1 mM EDTA, 0.5% TritonX-100, 1 mM PMSF and protease inhibitors (Roche, 04693159001). Cells were lysed on ice for 15 minutes, and then 300 ⁇ L per well of PBS containing 1 mM EDTA and 0.5% TritonX-100 was added to dilute urea to 1 M.
  • the blocking solution in the blocked 96-well plate was discarded, 100 ⁇ L of diluted cell lysate was added to each well, placed in a 37°C incubator for 2 hours, and the plate was washed five times with PBS. Dilute the biotinylated detection antibody to 0.4 ⁇ g/mL with PBS containing 1% BSA, then add 100 ⁇ L detection antibody to each well, and incubate in a 37°C incubator for 1 hour. Then the well plate was washed five times, and 100 ⁇ L of avidin-HRP diluted 200 times with PBS containing 1% BSA was added to each well, and incubated at 37° C. for 30 minutes.
  • the well plate five times, add 100 ⁇ L of TMB substrate to each well, incubate at room temperature until blue color appears, and add 100 ⁇ L of stop solution to each well.
  • the IC50 value of the inhibitory activity of the compound was calculated with Graphpad Prism software.
  • the maximum degradation rate of the compound is the ratio of the level of remaining ER ⁇ in cells after treatment with 3000 nM compound to the level of remaining ER ⁇ in cells after treatment with 3000 nM fulvestrant.
  • Test Example 5 Inhibitory Effect of Compounds of the Disclosure on Enzyme Activity of CYP3A4 Midazolam Metabolism Site in Human Liver Microsomal
  • the enzyme activity of the disclosed compound on the CYP3A4 midazolam metabolic site of human liver microsomes is determined by the following experimental method:
  • Phosphate buffer solution (20 ⁇ PBS, purchased from Shenggong),
  • CYP probe substrate (midazolam, TRC, M343000/3 ⁇ M) and positive control inhibitor (ketoconazole, SIGMA, Cat No. K1003-100MG).
  • Table 5 shows the IC 50 value of the drug on the CYP3A4 midazolam metabolic site calculated by Graphpad Prism.
  • the compound to be protected in this disclosure has a weak inhibitory effect on the midazolam metabolic site of CYP3A4 in human liver microsomes, showing better safety, suggesting that no metabolism based on the midazolam metabolic site of CYP3A4 metabolism will occur sexual drug interactions.
  • Test Example 6 Inhibitory Effect of Compounds of the Disclosure on Enzyme Activity of CYP3A4 Testosterone Metabolism Site in Human Liver Microsomal
  • the enzyme activity of the disclosed compound on the CYP3A4 testosterone metabolic site of human liver microsomes is determined by the following experimental method:
  • Phosphate buffer solution (20 ⁇ PBS, purchased from Shenggong),
  • CYP probe substrate testosterone, Wokai, CAS No.[58-22-0]/75 ⁇ M
  • positive control inhibitor ketoconazole, SIGMA, Cat No.K1003-100MG
  • Configure the PBS buffer solution of 100mM configure the PBS buffer solution of 100mM, use the buffer solution to prepare the microsomal solution of 0.25mg/ml and the NADPH solution of 7.5mM MgCl 2 and 5mM, and use DMSO to dilute the stock solution with a concentration of 30mM to Concentration is the series solution I of 30mM, 10mM, 3mM, 1mM, 0.3mM, 0.03mM, 0.003mM, 0mM, then use phosphate buffered solution (PBS), the above-mentioned series solution I is diluted 200 times to obtain the series test solution II (150 , 50, 15, 5, 1.5, 0.15, 0.015, 0 ⁇ M). Testosterone working solution diluted to 375 ⁇ M concentration with PBS.
  • PBS phosphate buffered solution
  • Table 6 shows the IC 50 value of the drug on the CYP3A4 testosterone metabolic site calculated by Graphpad Prism.
  • the compounds to be protected in the present disclosure have weaker inhibition on the testosterone metabolic site of CYP3A4 in human liver microsomes, showing better safety.
  • Test Example 7 Time-dependent inhibition of the compound of the present disclosure on the enzymatic activity of the CYP3A4 midazolam metabolic site in human liver microsomes.
  • the time-dependent CYP enzyme activity inhibitory effect of the disclosed compound on the CYP3A4 midazolam metabolic site of human liver microsomal was determined by the following experimental method:
  • Phosphate buffer solution (20 ⁇ PBS, purchased from Shenggong),
  • CYP probe substrate (midazolam, TRC, M343000/3 ⁇ M) and positive control inhibitor (verapamil, Adamas Reagent Co, Ltd, Cat No. 25904A).
  • Table 7 shows the IC 50 value of the drug on the CYP3A4 midazolam metabolic site calculated by Graphpad Prism.
  • the compound to be protected in this disclosure has a weak inhibitory effect on the midazolam metabolic site of CYP3A4 in human liver microsomes, showing better safety, suggesting that no metabolism based on the midazolam metabolic site of CYP3A4 metabolism will occur sexual drug interactions.
  • Test Example 8 Time-dependent inhibitory effect of the disclosed compound on the enzymatic activity of the CYP3A4 testosterone metabolic site in human liver microsomal
  • the time-dependent inhibitory effect of the disclosed compound on the enzyme activity of the CYP3A4 testosterone metabolic site in human liver microsomes is determined by the following experimental method:
  • Phosphate buffer solution (20 ⁇ PBS, purchased from Shenggong),
  • CYP probe substrate testosterone, Wokai, CAS No. [58-22-0]/75 ⁇ M
  • positive control inhibitor verapamil, Adamas Reagent Co Ltd, Cat No. 25904A
  • Table 8 shows the IC 50 value of the drug on the CYP3A4 testosterone metabolic site calculated by Graphpad Prism.
  • the blocking effect of the disclosed compound on hERG potassium current was tested on a stable cell line transfected with hERG potassium channel by automatic patch clamp.
  • HEK293-hERG stable cell lines were passaged in MEM/EBSS medium (10% FBS, 400 ⁇ g/ml G418, 1% MEM non-essential amino acid solution (100 ⁇ ), 1% sodium pyruvate solution) at a density of 1:4 Cultivate and perform automatic patch clamp experiments within 48-72 hours of culture.
  • MEM/EBSS medium 10% FBS, 400 ⁇ g/ml G418, 1% MEM non-essential amino acid solution (100 ⁇ ), 1% sodium pyruvate solution
  • the cells were digested with 0.25% trypsin, collected by centrifugation, and resuspended in extracellular fluid (140mM NaCl, 4mM KCl, 1mM MgCl 2 , 2mM CaCl 2 , 5mMD glucose monohydrate, 10mM Hepes, pH7.4, 298mOsm) Cells were made into cell suspension.
  • the cell suspension is placed on the cell bank of the Patchliner instrument, and the Patchliner instrument uses a negative pressure controller to add cells to the chip (NPC-16), and the negative pressure attracts a single cell to the small hole of the chip.
  • the instrument When the whole-cell mode is formed, the instrument will obtain the hERG current according to the set hERG current and voltage program, and then the instrument will automatically perfuse the compound from low concentration to high concentration. Data collection and analysis were carried out by HEAK Patchmaster, HEAK EPC10 patch clamp amplifier (Nanion), Pathlinersoftware and Pathcontrol HTsoftware, and the current at each concentration of the compound and the blank control current were analyzed.
  • IC 50 ⁇ 30 ⁇ M means no inhibitory activity; 30>IC 50 ⁇ 10 ⁇ M means weak inhibitory activity; 10> IC 50 ⁇ 1 ⁇ M means moderate inhibitory activity; IC 50 ⁇ 1 ⁇ M means strong inhibitory activity.
  • the compound to be protected in the present disclosure has no inhibitory activity on hERG, showing better safety.
  • Embodiment 9 the preparation of citrate amorphous
  • Embodiment 10 the preparation of hydrochloride
  • Embodiment 11 the preparation of hydrobromide
  • Embodiment 12 the preparation of mesylate
  • Embodiment 14 the preparation of acetate
  • Embodiment 15 the preparation of succinate
  • Embodiment 16 the preparation of maleate
  • Embodiment 17 the preparation of malate
  • Embodiment 18 the preparation of p-toluenesulfonate
  • Embodiment 19 research on moisture absorption
  • Embodiment 20 research on the stability of influencing factors
  • the experiment of influencing factors shows that the crystalline form of phosphate I has good physical and chemical stability under the conditions of influencing factors for 30 days.
  • the A crystal form of sulfate has good physical and chemical stability under other conditions except light conditions, and deliquesces under high humidity conditions. Amorphous deliquescent under high humidity conditions, physically stable under other conditions, chemical stability slightly worse under high temperature and light conditions.

Landscapes

  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Animal Behavior & Ethology (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • General Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Epidemiology (AREA)
  • Hematology (AREA)
  • Diabetes (AREA)
  • Oncology (AREA)
  • Biomedical Technology (AREA)
  • Neurology (AREA)
  • Neurosurgery (AREA)
  • Endocrinology (AREA)
  • Reproductive Health (AREA)
  • Cardiology (AREA)
  • Heart & Thoracic Surgery (AREA)
  • Communicable Diseases (AREA)
  • Immunology (AREA)
  • Obesity (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Nitrogen Condensed Heterocyclic Rings (AREA)

Abstract

本公开涉及一种三环四氢异喹啉类衍生物的盐型。具体而言，本公开涉及式(I)所示化合物的不同盐型及其制备方法，本公开提供的式(I)化合物的盐型具备良好的稳定性，可更好地用于临床治疗。

Description

一种三环四氢异喹啉类衍生物的盐型
本申请要求申请日为2021年7月9日的中国专利申请2021107799053的优先权。本申请引用上述中国专利申请的全文。
技术领域
本公开涉及一种三环四氢异喹啉类衍生物的盐型及其制备方法、医药用途，属于制药领域。
背景技术
CSCO乳腺癌诊疗指南(2020版)中氟维司群针对未经内分泌治疗者的激素受体阳性晚期乳腺癌患者，被调整成为Ⅰ级推荐，新增氟维司群+CDK4/6抑制剂为Ⅱ级推荐；另外，氟维司群联合CDK4/6抑制剂已成为非甾体AI(NSAI)治疗失败及甾体AI(SAI)治疗失败患者的I级推荐，氟维司群作为目前唯一获批的SERD，其获批形式为注射剂，为患者用药带来了不便。故寻找新的更有效的SERD不仅具有巨大的市场价值也为乳腺癌患者带来了用药上的可选择性及便利性。目前赛诺菲的SAI439859作为新一代的可口服的SERD抑制剂目前正处于临床III期，另外罗氏的GDC-9545、阿斯利康的AZD-9833及Radius的RAD1901均已进入III期。公开的选择性雌激素受体介导的调节剂专利申请包括WO2014165723、WO2014151899、WO2014141292、WO2014191726、WO2015092634、WO2014135834、WO2014106848和EP1113007。
PCT申请WO2021139756A公开了一种式(I)所示的三环四氢异喹啉类衍生物用于雌激素受体下调剂(SERD)，为满足用药需求，对其盐及其晶型进行研究很有必要，
Figure PCTCN2022104669-appb-000001
发明内容
本公开提供一种式(I)所示化合物的药学上可接受的盐，所述药学上可接受的盐选自磷酸盐、硫酸盐、酒石酸盐、柠檬酸盐、盐酸盐、氢溴酸盐、甲磺酸盐、甲酸盐、乙酸盐、琥珀酸盐、马来酸盐、苹果酸盐或对甲苯磺酸盐，
Figure PCTCN2022104669-appb-000002
一些实施方案中，本公开提供的式(I)所示化合物的药学上可接受的盐，其中式(I)所示化合物与酸分子的物质的量比选自1:5-5:1。
一些实施方案中，本公开提供的式(I)所示化合物的药学上可接受的盐，其中式(I)所示化合物与酸分子的物质的量比为1:1。
一些实施方案中，本公开提供的式(I)所示化合物所的药学上可接受的盐，其中式(I)所示化合物与酸分子的物质的量比为1:3。
一些实施方案中，本公开提供一种式(I)所示化合物的磷酸盐，其中式(I)所示化合物与磷酸分子的物质的量比为1:3。
本公开提供一种式(I)所示化合物的磷酸盐的I晶型，其中式(I)所示化合物与磷酸分子的物质的量比为1:3，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.1、13.5、19.2、22.3处有特征峰。
一些实施方案中，式(I)所示化合物的磷酸盐的I晶型，其中式(I)所示化合物与磷酸分子的物质的量比为1:3，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.1、13.5、13.9、19.2、21.8、22.3、23.4处有特征峰。
一些实施方案中，式(I)所示化合物的磷酸盐的I晶型，其中式(I)所示化合物与磷酸分子的物质的量比为1:3，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.1、12.4、13.5、13.9、15.0、16.3、16.9、19.2、20.8、21.8、22.3、23.4、24.1、24.6、26.2、27.5、30.0处有特征峰。
一些实施方案中，式(I)所示化合物的磷酸盐的I晶型，其中式(I)所示化合物与磷酸分子的物质的量比为1:3，以衍射角2θ角度表示的X-射线粉末衍射图谱如附图2所 示。
一些实施方案中，式(I)所示化合物的磷酸盐的I晶型，其中式(I)所示化合物与磷酸分子的物质的量比为1:3，三斜晶系，轴长
Figure PCTCN2022104669-appb-000003
Figure PCTCN2022104669-appb-000004
轴角α＝89.575(2)°；β＝89.859(2)°；γ＝60.745(2)°；晶胞体积
Figure PCTCN2022104669-appb-000005
Figure PCTCN2022104669-appb-000006
本公开提供一种式(I)所示化合物的磷酸盐或式(I)所示化合物的磷酸盐的I晶型的制备方法，包括式(I)所示化合物与磷酸反应的步骤。
本公开提供一种式(I)所示化合物的磷酸盐的I晶型的制备方法，包括以下步骤：
1)配置式(I)所示化合物的溶剂A溶液，所述溶剂A选自酮类溶剂、酯类溶剂或醚类溶剂；
2)配置磷酸的溶剂B溶液，所述溶剂B选自水或醇类溶剂；
3)混合式(I)所示化合物的溶剂A溶液与磷酸的溶剂B溶液后结晶析出。
一些实施方案中，配置式(I)所示化合物的溶剂A溶液，所述溶剂A选自丙酮、乙酸乙酯或甲基叔丁基醚。
一些实施方案中，配置磷酸的溶剂B溶液，所述溶剂B选自甲醇、乙醇或水。
本公开提供一种式(I)所示化合物的硫酸盐，其中式(I)所示化合物与硫酸分子的物质的量比为1:1。
本公开提供一种式(I)所示化合物的硫酸盐的A晶型，中式(I)所示化合物与硫酸分子的物质的量比为1:1，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.6、7.9、15.1、20.7、22.1处有特征峰。
一些实施方案中，式(I)所示化合物的硫酸盐的A晶型，中式(I)所示化合物与硫酸分子的物质的量比为1:1，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.6、7.9、13.4、14.5、15.1、19.3、19.8、20.7、22.1、24.8、25.4、25.7、27.6处有特征峰。
一些实施方案中，式(I)所示化合物的硫酸盐的A晶型，中式(I)所示化合物与硫酸分子的物质的量比为1:1，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.6、7.9、11.6、13.4、14.5、15.1、15.9、18.1、19.3、19.8、20.7、22.1、24.0、24.8、25.4、25.7、27.2、27.6、28.3、29.2处有特征峰。
一些实施方案中，式(I)所示化合物的硫酸盐的A晶型，中式(I)所示化合物与硫酸分子的物质的量比为1:1，以衍射角2θ角度表示的X-射线粉末衍射图谱如附图8所示。
一些实施方案中，式(I)所示化合物的硫酸盐的A晶型，中式(I)所示化合物与硫酸分子的物质的量比为1:1，单斜晶系，轴长
Figure PCTCN2022104669-appb-000007
Figure PCTCN2022104669-appb-000008
轴角β＝90°；晶体体积
Figure PCTCN2022104669-appb-000009
本公开提供一种(I)所示化合物的硫酸盐或式(I)所示化合物的硫酸盐的A晶型的制备方法，包括式(I)所示化合物与硫酸反应的步骤。
本公开提供一种式(I)所示化合物的硫酸盐的A晶型的方法，包括以下步骤：
1)配置式(I)所示化合物的溶剂A溶液，所述溶剂A选自酮类溶剂、酯类溶剂、腈类溶剂或醇类溶剂；
2)配置硫酸的溶剂B溶液，所述溶剂B选自水、腈类溶剂或醇类溶剂；
3)混合式(I)所示化合物的溶剂A溶液与硫酸的溶剂B溶液后结晶析出。
一些实施方案中，配置式(I)所示化合物的溶剂A溶液，所述溶剂A选自丙酮、乙酸乙酯、乙腈、甲醇或乙醇。
一些实施方案中，配置硫酸的溶剂B溶液，所述溶剂B选自甲醇、乙腈、乙醇或水。
本公开提供一种式(I)所示化合物的柠檬酸盐的无定型。
本公开提供一种制备式(I)所示化合物的柠檬酸盐的方法，包括式(I)所示化合物与柠檬酸反应成盐的步骤。
本公开提供一种式(I)所示化合物的L-酒石酸盐的无定型。
本公开提供一种制备式(I)所示化合物的L-酒石酸盐的方法，包括式(I)所示化合物与L-酒石酸反应成盐的步骤。
本公开提供一种制备式(I)所示化合物的盐酸盐的方法，包括式(I)所示化合物与盐酸反应成盐的步骤。
本公开提供一种制备式(I)所示化合物的氢溴酸盐的方法，包括式(I)所示化合物与氢溴酸反应成盐的步骤。
本公开提供一种制备式(I)所示化合物的甲磺酸盐的方法，包括式(I)所示化合物与甲磺酸反应成盐的步骤。
本公开提供一种制备式(I)所示化合物的甲酸盐的方法，包括式(I)所示化合物与甲酸反应成盐的步骤。
本公开提供一种制备式(I)所示化合物的乙酸盐的方法，包括式(I)所示化合物与乙酸反应成盐的步骤。
本公开提供一种制备式(I)所示化合物的琥珀酸盐的方法，包括式(I)所示化合物与琥珀酸反应的步骤。
本公开提供一种制备式(I)所示化合物的马来酸盐的方法，包括式(I)所示化合物与马来酸反应成盐的步骤。
本公开提供一种制备式(I)所示化合物的苹果酸盐的方法，包括式(I)所示化合物与苹果酸反应成盐的步骤。
本公开提供一种制备式(I)所示化合物的对甲苯磺酸盐的方法，包括式(I)所示化合物与对甲苯磺酸反应成盐的步骤。
一些实施方案中，本公开提供的式(I)所示化合物的磷酸盐的I晶型或式(I)所示化合物的硫酸盐的A晶型，以衍射角2θ角度表示的X-射线粉末衍射图谱中特征峰2θ角的误差范围为±0.2。
另一方面，本公开提供一种药物组合物，包含以下成分：
1)式(I)所示化合物磷酸盐、硫酸盐、酒石酸盐、柠檬酸盐、盐酸盐、氢溴酸盐、甲磺酸盐、甲酸盐、乙酸盐、琥珀酸盐、马来酸盐、苹果酸盐或对甲苯磺酸盐、式(I)所示化合物的磷酸盐的I晶型或式(I)所示化合物的硫酸盐的A晶型、式(I)所示化合物的柠檬酸盐的无定型、式(I)所示化合物的L-酒石酸盐的无定型，或其混合物；和
2)任选自药学上可接受的载体、稀释剂或赋形剂。
另一方面，本公开提供一种药物组合物的制备方法，包括将
1)式(I)所示化合物磷酸盐、硫酸盐、酒石酸盐、柠檬酸盐、盐酸盐、氢溴酸盐、甲磺酸盐、甲酸盐、乙酸盐、琥珀酸盐、马来酸盐、苹果酸盐或对甲苯磺酸盐、式(I)所示化合物的磷酸盐的I晶型或式(I)所示化合物的硫酸盐的A晶型、式(I)所示化合物的柠檬酸盐的无定型、式(I)所示化合物的L-酒石酸盐的无定型，或其混合物；和
2)任选自药学上可接受的载体、稀释剂或赋形剂混合的步骤。
本公开另一方面提供上述式(I)所示化合物磷酸盐、硫酸盐、酒石酸盐、柠檬酸盐、盐酸盐、氢溴酸盐、甲磺酸盐、甲酸盐、乙酸盐、琥珀酸盐、马来酸盐、苹果酸盐或对甲苯磺酸盐、式(I)所示化合物的磷酸盐的I晶型或式(I)所示化合物的硫酸盐的A晶型、式(I)所示化合物的柠檬酸盐的无定型、式(I)所示化合物的L-酒石酸盐的无定型，或其混合物、组合物在制备雌激素受体下调剂(SERD)中的用途。
本公开另一方面提供上述式(I)所示化合物磷酸盐、硫酸盐、酒石酸盐、柠檬酸盐、 盐酸盐、氢溴酸盐、甲磺酸盐、甲酸盐、乙酸盐、琥珀酸盐、马来酸盐、苹果酸盐或对甲苯磺酸盐、式(I)所示化合物的磷酸盐的I晶型或式(I)所示化合物的硫酸盐的A晶型、式(I)所示化合物的柠檬酸盐的无定型、式(I)所示化合物的L-酒石酸盐的无定型，或其混合物、组合物在制备预防和/或治疗癌症的药物中的用途，其中所述的癌症优选选自乳腺癌、子宫内膜癌、子宫癌、宫颈癌、皮肤癌、前列腺癌、卵巢癌、输卵管肿瘤、血友病和白血病。
本公开另一方面提供上述式(I)所示化合物磷酸盐、硫酸盐、酒石酸盐、柠檬酸盐、盐酸盐、氢溴酸盐、甲磺酸盐、甲酸盐、乙酸盐、琥珀酸盐、马来酸盐、苹果酸盐或对甲苯磺酸盐、式(I)所示化合物的磷酸盐的I晶型或式(I)所示化合物的硫酸盐的A晶型、式(I)所示化合物的柠檬酸盐的无定型、式(I)所示化合物的L-酒石酸盐的无定型，或其混合物、组合物在制备预防和/或治疗雌激素受体介导的或依赖性的疾病或病症的药物中的用途；优选地，其中所述的雌激素受体介导的或依赖性的疾病或病症选自癌症、中枢神经系统缺陷、心血管系统缺陷、血液系统缺陷、免疫及炎症疾病、易感性感染、代谢缺陷、神经缺陷、精神缺陷和生殖缺陷；优选地，其中所述的癌症选自乳腺癌、子宫内膜癌、子宫癌、宫颈癌、皮肤癌、前列腺癌、卵巢癌、输卵管肿瘤、血友病和白血病；更优选，其中所述的癌症选自乳腺癌、卵巢癌、子宫内膜癌、前列腺癌和子宫癌。
本公开所述的“2θ或2θ角度”是指衍射角，θ为布拉格角，单位为°或度；每个特征峰2θ的误差范围为±0.20，具体可以为-0.20、-0.19、-0.18、-0.17、-0.16、-0.15、-0.14、-0.13、-0.12、-0.11、-0.10、-0.09、-0.08、-0.07、-0.06、-0.05、-0.04、-0.03、-0.02、-0.01、0.00、0.01、0.02、0.03、0.04、0.05、0.06、0.07、0.08、0.09、0.10、0.11、0.12、0.13、0.14、0.15、0.16、0.17、0.18、0.19、0.20。
本公开所述的“结晶析出”包括但不限于搅拌结晶、降温析晶、溶析结晶和挥发结晶。
本公开中所述的“差示扫描量热分析或DSC”是指在样品升温或恒温过程中，测量样品与参考物之间的温度差、热流差，以表征所有与热效应有关的物理变化和化学变化，得到样品的相变信息。
本公开中所述干燥温度一般为25℃-100℃，优选40℃-70℃，可以常压干燥，也可以减压干燥。
本公开中数值为仪器测量值或仪器测量后计算值，存在一定程度的误差，一般而言，正负10％均属于合理误差范围内。当然需要考虑该数值所用之处的上下文，例如，总杂质的含量，该数值为测量后误差变化不超过正负10％，可以为正负9％、正负8％、正负7％、正负6％、正负5％、正负4％、正负3％、正负2％或正负1％，优选正负5％。
本公开晶型制备方法中所用的起始原料可以是任意形式的式I所示化合物，具体形 式包括但不限于：无定形、任意晶型、水合物、溶剂合物等。
本公开所述的“析晶”包括但不限于搅拌析晶、降温析晶、打浆析晶和挥发析晶。
附图说明
图1、式(I)所示化合物无定型的XRPD谱图；
图2、式(I)所示化合物磷酸盐I晶型的XRPD谱图；
图3、式(I)所示化合物磷酸盐I晶型的TGA谱图；
图4、式(I)所示化合物磷酸盐I晶型的DSC谱图；
图5、式(I)所示化合物磷酸盐I晶型的DVS后的XRPD谱图；
图6、式(I)所示化合物磷酸盐I晶型的DVS谱图；
图7、式(I)所示化合物磷酸盐I晶型的单晶谱图；
图8、式(I)所示化合物硫酸盐的A晶型的XRPD谱图；
图9、式(I)所示化合物硫酸盐的A晶型的TGA谱图；
图10、式(I)所示化合物硫酸盐的A晶型的DSC谱图；
图11、式(I)所示化合物硫酸盐的A晶型的DVS谱图；
图12、式(I)所示化合物硫酸盐的A晶型的单晶谱图；
图13、式(I)所示化合物酒石酸盐无定型的XRPD谱图；
图14、式(I)所示化合物柠檬酸盐无定型的XRPD谱图。
具体实施方式
通过以下实施例和实验例进一步详细说明本发明。这些实施例和实验例仅用于说明性目的，而并不用于限制本发明的范围。
实验所用仪器的测试条件：
化合物的结构是通过核磁共振(NMR)或/和质谱(MS)来确定的。NMR位移(δ)以10 -6(ppm)的单位给出。NMR的测定是用Bruker AVANCE-400核磁仪或Bruker AVANCE NEO 500M，测定溶剂为氘代二甲基亚砜(DMSO-d6)、氘代氯仿(CDCl 3)、氘代甲醇(CD 3OD)，内标为四甲基硅烷(TMS)。
MS的测定用Agilent 1200/1290 DAD-6110/6120 Quadrupole MS液质联用仪(生产商：Agilent，MS型号：6110/6120 Quadrupole MS)、waters ACQuity UPLC-QD/SQD(生产商：waters，MS型号：waters ACQuity Qda Detector/waters SQ Detector)、THERMO Ultimate 3000-Q Exactive(生产商：THERMO，MS型号：THERMO Q Exactive)。
高效液相色谱法(HPLC)分析使用Agilent HPLC 1200DAD、Agilent HPLC 1200VWD和Waters HPLC e2695-2489高压液相色谱仪。
手性HPLC分析测定使用Agilent 1260 DAD高效液相色谱仪。
高效液相制备使用Waters 2767、Waters 2767-SQ Detecor2、Shimadzu LC-20AP和Gilson-281制备型色谱仪。
手性制备使用Shimadzu LC-20AP制备型色谱仪。
CombiFlash快速制备仪使用Combiflash Rf200(TELEDYNE ISCO)。
XRPD为X射线粉末衍射检测：测定使用BRUKER D8 DISCOVER型X射线衍射仪进行，具体采集信息：Cu阳极(40kV，40mA)，Cu-Kα射线
Figure PCTCN2022104669-appb-000010
扫描方式：θ/2θ，扫描范围(2θ范围)：3～48°。
DSC为差示扫描量热：测定采用METTLER TOLEDO DSC 3+差示扫描量热仪，升温速率10℃/min，温度具体范围参照相应图谱(25-240℃)，氮气吹扫速度50mL/min。
TGA为热重分析：检测采用METTLER TOLEDO TGA 2型热重分析仪，升温速率10℃/min，温度具体范围参照相应图谱(30-350℃)，氮气吹扫速度20mL/min。
DVS为动态水分吸附：检测采用SMS DVS Advantage，在25℃，湿度变化为50％-95％-0％-95％-50％，步进为10％(最后一步为5％)(湿度具体范围以相应图谱为准，此处所列为大多使用方法)，判断标准为dm/dt≤0.002％。
薄层层析硅胶板使用烟台黄海HSGF254或青岛GF254硅胶板，薄层色谱法(TLC)使用的硅胶板采用的规格是0.15mm～0.2mm，薄层层析分离纯化产品采用的规格是0.4mm～0.5mm。
硅胶柱色谱法一般使用烟台黄海硅胶200～300目硅胶为载体。
激酶平均抑制率及IC50值的测定用NovoStar酶标仪(德国BMG公司)。
本公开的已知的起始原料可以采用或按照本领域已知的方法来合成，或可购买自ABCR GmbH&Co.KG，Acros Organics，Aldrich Chemical Company，韶远化学科技(Accela ChemBio Inc)、达瑞化学品等公司。
实施例中无特殊说明，反应均能够在氩气氛或氮气氛下进行。
氩气氛或氮气氛是指反应瓶连接一个约1L容积的氩气或氮气气球。
氢气氛是指反应瓶连接一个约1L容积的氢气气球。
加压氢化反应使用Parr 3916EKX型氢化仪和清蓝QL-500型氢气发生器或HC2-SS型氢化仪。
氢化反应通常抽真空，充入氢气，反复操作3次。
微波反应使用CEM Discover-S 908860型微波反应器。
实施例中无特殊说明，溶液是指水溶液。
实施例中无特殊说明，反应的温度为室温，为20施～30例。
实施例中的反应进程的监测采用薄层色谱法(TLC)，反应所使用的展开剂，纯化化合物采用的柱层析的洗脱剂的体系和薄层色谱法的展开剂体系包括：A：二氯甲烷/甲醇体系，B：正己烷/乙酸乙酯体系，C：石油醚/乙酸乙酯体系，溶剂的体积比根据化合物的极性不同而进行调节，也可以加入少量的三乙胺和醋酸等碱性或酸性试剂进行调节。
实施例1、式1所示化合物的制备(优先权为：202010680491.4的申请中实施例13的制备方法)
2,2-二氟-3-((5S,7R)-5-(5-((1-(3-氟丙基)氮杂环丁-3-基)氨基)吡啶-2-基)-7-甲基-7,8-二氢-[1,3]二氧杂戊环并[4,5-g]异喹啉-6(5H)-基-2,2-d 2)丙-1-醇1
Figure PCTCN2022104669-appb-000011
第一步
(R)-N-(1-(3,4-二(苄氧基)苯基)丙-2-基)-3-((叔丁基二苯基硅基)氧基)-2,2-二氟丙-1-胺1b
将化合物1a(1.0g,3.0mmol)溶于二氧六环(20mL)，加入二异丙基乙胺(1.2g,9.0mmol)，3-((叔丁基二苯基硅基)氧基)-2,2-二氟丙基三氟甲磺酸酯(1.0g,2.0mmol，采用公知的方法“Bioorganic&Medicinal Chemistry Letters,2018,28(14),2528-2532”制备而得)， 氩气保护下反应在油浴80℃搅拌20小时。冷却反应液，减压浓缩，加入饱和碳酸氢钠溶液(50mL)，用乙酸乙酯萃取(100mL×2)，合并有机相，无水硫酸钠干燥，过滤，滤液减压浓缩，使用柱层析以展开剂体系B纯化所得残余物，得到标题化合物1b(1.7g)，产率：84％。MS m/z(ESI):680.2[M+1]。
第二步
(R)-4-(2-((3-((叔丁基二苯基硅基)氧基)-2,2-二氟丙基)氨基)丙基)苯-1,2-二酚1c
将化合物1b(1.4g,2.0mmol)溶于甲醇(10mL)，氩气保护，加入氢氧化钯碳(0.2g)，氢气球加氢搅拌3小时，停止搅拌，过滤，滤液减压浓缩，得到标题化合物1c(0.9g)，产率：94％。
第三步
(1S,3R)-1-(5-溴吡啶-2-基)-2-(3-((叔丁基二苯基硅基)氧基)-2,2-二氟丙基)-3-甲基-1,2,3,4-四氢异喹啉-6,7-二酚1d
将化合物1c(0.8g,1.6mmol)溶于甲苯(10mL)，加入乙酸(0.2g,3.2mmol)，5-溴吡啶醛(0.6g,3.2mmol)。油浴80℃搅拌反应16小时，停止反应。冷却反应，减压浓缩，加入水(10mL)，缓慢加入饱和碳酸氢钠溶液(20mL)调节反应液pH至8左右，用乙酸乙酯萃取(10mL×2)，合并有机相，无水硫酸钠干燥，过滤，滤液减压浓缩，使用柱层析以展开剂体系B纯化所得残余物，得到标题化合物1d(256mg)，产率75％。
MS m/z(ESI):667.1[M+1]。
第四步
(5S,7R)-5-(5-溴吡啶-2-基)-6-(3-((叔丁基二苯基硅基)氧基)-2,2-二氟丙基)-7-甲基-5,6,7,8-四氢-[1,3]二氧杂戊环并[4,5-g]异喹啉-2,2-d 2 1e
将化合物1d(350mg,0.5mmol)溶于N,N-二甲基甲酰胺(10mL)，加入二溴二氘甲烷(277mg,1.6mmol)，碳酸铯(512mg,1.6mmol)。油浴70℃搅拌反应16小时，停止反应。冷却反应，减压浓缩，加入水(10mL)，用乙酸乙酯萃取(10mL×2)，合并有机相，无水硫酸钠干燥，过滤，滤液减压浓缩，使用柱层析以展开剂体系B纯化所得残余物，得到标题化合物1e(170mg)，产率：48％。
MS m/z(ESI):681.1[M+1]。
第五步
6-((5S,7R)-6-(3-((叔丁基二苯基硅基)氧基)-2,2-二氟丙基)-7-甲基-5,6,7,8-四氢-[1,3]二氧杂戊环并[4,5-g]异喹啉-5-基-2,2-d 2)-N-(1-(3-氟丙基)氮杂环丁-3-基)吡啶-3-胺1f
将化合物1e(170mg,0.25mmol)溶于二氧六环(10mL)，加入化合物1-(3-氟丙基)氮杂环丁-3-胺(采用专利申请“WO2019228443中说明书第50页实施例1”公开的方法制备而得)(44mg,0.3mmol)，2,2'-双-(二苯膦基)-1,1'-联萘(12mg,0.02mmol)，三(二亚苄基丙酮)二钯(20mg,0.02mmol)，叔丁醇钠(29mg,0.3mmol)，氩气保护。油浴80℃搅拌反应16小时，停止反应。冷却反应液，浓缩，加入饱和碳酸氢钠溶液(10mL)，用乙酸乙酯萃取(10mL×2)，合并有机相，无水硫酸钠干燥，过滤，滤液减压浓缩，使用柱层析以展开剂体系B纯化所得残余物，得到标题化合物1f(47mg)，产率51％。
第六步
2,2-二氟-3-((5S,7R)-5-(5-((1-(3-氟丙基)氮杂环丁-3-基)氨基)吡啶-2-基)-7-甲基-7,8-二氢-[1,3]二氧杂戊环并[4,5-g]异喹啉-6(5H)-基-2,2-d 2)丙-1-醇1
将化合物1f(103mg,0.14mmol)溶于二氯甲烷中(5mL)，冰浴下滴加1M正四丁基氟化铵的四氢呋喃溶液(1mL)，滴毕，室温下搅拌1.5小时，减压浓缩，加入饱和碳酸氢钠溶液(5mL)，用乙酸乙酯萃取(5mL×3)，合并有机相，用饱和氯化钠溶液洗涤(5mL)，无水硫酸钠干燥，过滤，滤液减压浓缩，使用柱层析以展开剂体系B纯化所得残余物，得到标题化合物1(11mg)，产率42％。
MS m/z(ESI):495.2[M+1]。
1H NMR(400MHz,CD 3OD)7.80(d,1H),7.04(d,1H),6.94(dd,1H),6.61(s,1H),6.19(s,1H),4.64(br,2H),4.54-4.52(m,1H),4.45-4.42(m,1H),4.15-4.10(m,1H),3.85-3.83(m,2H),3.76-3.68(m,1H),3.64-3.56(m,1H),3.14-2.97(m,4H),2.78-2.68(m,3H),2.59-2.52(m,1H),1.84-1.74(m,2H),1.05(d,3H)。
经X-射线粉末衍射检测，将该产物为无定形，XRPD谱图如图1。
生物学评价
以下结合测试例进一步描述解释本公开，但这些实施例并非意味着限制本公开的范围。
测试例1本公开化合物对E与ER的结合的抑制作用
本公开化合物对E(estrogen，雌激素)与ER(estrogen receptor，雌激素受体)的结合具有抑制作用，从而阻断E和ER的复合物与ERE(雌激素应答元件)结合，继而阻 断了下游的荧光素酶蛋白的表达。本公开化合物对E与ER结合作用的抑制效果通过以下的方法进行测试。
1、实验目的
本实验的目的是为了测试化合物对E与ER结合的抑制作用，根据IC 50大小评价化合物的体外活性。
2、实验方法
将ERE克隆至荧光素酶基因的上游，通过转染MCF-7(TCHu74，中国科学院典型培养物保藏委员会细胞库)，挑选出MCF-7/ERE-荧光素酶(luciferase)单克隆细胞。在96孔板中用含10％活性炭处理(charcoal stripped)FBS(Moregate，FBSF)，1％丙酮酸钠(sigma，S8636)，1％非必需氨基酸(sigma，M7145)和500μg/ml G418的MEM(hyclone，SH30024.01B)培养基接种MCF-7/ERE-荧光素酶细胞，接种密度为30,000个细胞/孔，细胞在37℃，5％CO 2条件下培养。将药物配置成20mM的储存液，用100％DMSO以10倍梯度稀释，再用培养基稀释20倍。细胞培养24小时后去掉培养基，每孔加入0.1nM雌二醇(sigma，E2758)和10μL用培养基稀释好的药物，对照组加入DMSO，轻轻振荡混匀。放置37℃、5％CO 2培养箱中培养，24小时后弃去细胞培养液，加入50μL/孔配制好的荧光素酶底物(Promega，E6110)，室温避光放置10-15分钟，测定化学发光信号值。
3、测试结果
本公开中化合物对E与ER的结合的抑制作用通过以上的实验进行测定，用Graphpad Prism对化学发光信号值与化合物的对数浓度分析，得出化合物的IC 50值，结果参见表1。
表1.本公开中化合物对E与ER的结合的抑制作用
化合物编号 IC 50(nM)
1 0.69
结论：本公开所要保护的化合物对E与ER的结合有明显的抑制作用。
测试例2本公开化合物对MCF-7细胞增殖的抑制效应
1、实验目的
本实验的目的是采用ATP法测试本公开化合物对MCF-7细胞增殖活性的抑制作用，根据IC 50大小评价化合物的体外活性。
2、实验方法
在96孔板中接种MCF-7(TCHu74，中国科学院典型培养物保藏委员会细胞库)，培养基为含10％FBS(Gibco，10099-141)，1％丙酮酸钠(sigma，S8636)，1％非必需氨基酸(sigma，M7145)的MEM(hyclone，SH30024.01B)培养基，接种密度为4,000个细胞/孔，细胞在37℃，5％CO 2条件下培养。将化合物配置成20mM的储存液，用 100％DMSO梯度稀释成1000X终浓度，再用含2％FBS的培养基稀释20倍。培养24小时后去掉培养基，每孔加入90μL含2％FBS的培养基和10μL药物，对照组加入10μL DMSO，轻轻振荡混匀，空白组只含100μL 2％FBS培养基，放置37℃、5％CO 2培养箱中培养，72小时后每孔加入50μL混合后的Cell Titer-Glo(Promega，G7571)，振荡混匀，室温放置10分钟，测定化学发光信号值。
3、数据分析
用Graphpad Prism对化学发光信号值与化合物的对数浓度作图，得出化合物的IC 50值，结果参见表2。
表2.本公开化合物对MCF-7细胞增殖的抑制效应
化合物编号 IC 50(nM)
1 0.29
测试例3本公开化合物对表达ERα突变体MCF7细胞增殖抑制实验生物学评价
1、实验目的
本实验的目的是测定本公开化合物对表达ERα突变体MCF7细胞增殖的抑制活性。
2、实验方法
定点突变和细胞系构建
人雌激素受体α(estrogen receptorα，ERα)蛋白的突变体ERαY537S使用双引物PCR的方式以野生型ESR1基因的cDNA(Accession No.NM000125)为模板进行定点突变获得。突变所使用的引物序列如下(下划线标出的核苷酸为突变的位点)：Y537S:F-AAG AAC GTG GTG CCC CTC TCT GAC CTG CTG CTG GAG ATG(SEQ ID NO:1)；R-CAT CTC CAG CAG CAG GTC AGA GAG GGG CAC CAC GTT CTT(SEQ ID NO:2)。将突变体ESR1的cDNA克隆至目标慢病毒载体pCDH-CMV-MCS-EF1-Puro上。然后将带有突变体ESR1基因序列的慢病毒质粒以及慢病毒包装质粒通过Lipofectamine 3000 Transfection Reagent(ThermoFisher Scientific，Cat#L3000075)转染到HEK-293T细胞(ATCC，CRL-3216)中。转染后48小时，将带有病毒的培养基上清过滤、超速离心获得病毒沉淀，用适量的培养基重悬溶解后，加入到MCF7细胞(ATCC，HTB-22)中，并加入终浓度为8μg/ml的聚凝胺(polybrene)孵育过夜。转染两天后，在细胞培养液中加入1μg/ml的嘌呤霉素进行抗性筛选，约两周后得到能够稳定表达ERαY537S突变体的MCF7细胞系。
细胞增殖抑制实验
将表达ERα突变体的MCF7细胞用含有10％胎牛血清的MEM(GE Healthcare，SH30024.01)完全培养基进行培养。实验第一天，使用完全培养基将细胞以3,000个/孔的密度种于96孔板，每孔100μL细胞悬液，放置37℃，5％CO 2的细胞培养箱培养过夜。第 二天吸掉培养基，每孔更换为135μL含有2％胎牛血清的MEM不完全培养基，同时每孔加入15μL用不完全培养基配制的不同浓度的待测化合物，化合物的终浓度是从100nM开始进行4倍梯度稀释的9个浓度点，设置含有0.5％DMSO的空白对照，放置37℃，5％CO 2的细胞培养箱培养144小时。第八天，取出96孔细胞培养板，每孔加入150μL CellTiter-
Figure PCTCN2022104669-appb-000012
Luminescent Cell Viability Assay(Promega，G7573)，室温放置10分钟后，使用多标记微孔板酶标仪(PerkinElmer，VICTOR 3)读取发光信号值，用Graphpad Prism软件根据化合物的浓度和发光信号值计算化合物抑制活性的IC 50值见表3。
3、测试结果
表3.本公开化合物对表达ERα突变体MCF7细胞增殖的抑制效应的IC 50值
化合物编号 IC 50(nM)
1 1.36
结论：本公开所要保护的化合物对表达ERα突变体的MCF7细胞增殖具有明显的抑制作用。
测试例4本公开化合物对ERα降解作用
1、实验目的
测试本公开化合物引起ERα降解作用，该方法用来测定本公开化合物对ERα的降解作用。
2、实验方法
ERα阳性乳腺癌细胞系MCF-7细胞用含有10％胎牛血清(Corning,35-010-CV)的DMEM/F12培养基(HyClone,SH30023.01)进行培养。实验第一天，消化细胞后，用含有5％活性炭处理的胎牛血清(BIOSUN,BS-0004-500)的无酚红DMEM/F12培养基(ThermoFisher,11039-021)将细胞洗一遍后重悬细胞并计数，将细胞密度调整为1.79×10 5个/mL。在48孔板(Corning，3548)中每孔加入280μL细胞悬液，将细胞放置于37℃，5％CO 2培养箱中培养过夜。实验第二天将化合物用DMSO进行梯度稀释，并用含有5％活性炭处理胎牛血清的无酚红DMEM/F12培养基进一步稀释。在48孔板细胞中每孔加入20μL稀释后的化合物，终浓度分别是3000，300，30，3，0.3，0.03和0.003nM。将48孔板在培养箱中放置16至18小时。使用人ERα/NR3A1总蛋白检测试剂盒(R&D,DYC5715-5)中的俘获抗体包被96孔板，将俘获抗体用PBS配制成1μg/mL，并在96孔板(Corning,3590)中每孔加入100μL，置于26℃孵箱中放置过夜。实验第三天，将包被抗体的96孔板用PBS洗一遍，每孔加入200μL含有1％BSA的PBS，置于37℃孵 箱中孵育1.5小时进行封闭。弃去细胞培养基上清，用PBS将细胞洗一遍，每孔中加入60μL细胞裂解液。细胞裂解液为含有6M尿素、1mM EDTA、0.5％TritonX-100、1mM PMSF以及蛋白酶抑制剂(Roche,04693159001)的PBS。将细胞放置在冰上裂解15分钟，然后加入300μL每孔的含有1mM EDTA和0.5％TritonX-100的PBS将尿素稀释至1M。将封闭后的96孔板中的封闭液弃去，每孔加入100μL稀释后的细胞裂解液，放置于37℃孵箱孵育2小时，用PBS洗板五遍。用含有1％BSA的PBS将生物素化的检测抗体稀释至0.4μg/mL，然后每孔加入100μL检测抗体，置于37℃孵箱孵育1小时。然后再将孔板洗五遍，每孔加入100μL用含有1％BSA的PBS稀释200倍的亲和素-HRP，置于37℃孵育30分钟。再将孔板洗五遍，每孔加入100μL的TMB底物，室温孵育至出现蓝色，每孔加入100μL终止液。使用PHERAstar多功能酶标仪读取OD450信号值。用Graphpad Prism软件计算化合物抑制活性的IC 50值。化合物最大降解率为3000nM化合物处理后细胞剩余ERα的水平与3000nM氟维司群(Fulvestrant)处理细胞后细胞剩余ERα水平的比值。
3、测试结果
本公开化合物对ERα降解作用所测得的EC 50值见表4。
表4.本公开化合物对ERα降解作用
实施例编号 EC 50(nM) Emax降解(％)
氟维司群 0.06 100
1 0.38 107
结论：本公开所要保护的化合物对ERα有明显的降解作用。
测试例5、本公开化合物对人肝微粒体CYP3A4咪达唑仑代谢位点的酶活性的抑制作用
本公开化合物对人肝微粒体CYP3A4咪达唑仑代谢位点的酶活性采用如下实验方法测定：
一、实验材料及仪器
1.磷酸缓冲液(20×PBS，购买自生工),
2.NADPH(ACROS，A2646-71-1),
3.人肝微粒体(Corning Gentest，Cat No，452161，Lot No.9050002,Donor,36)，
4.ABI QTrap 4000液质两用仪(AB Sciex),
5.ZORBAX Extend-C18,3×50mm,3.5μm(美国安捷伦公司)，
6.CYP探针底物(咪达唑仑,TRC,M343000/3μM)和阳性对照抑制剂(酮康唑,SIGMA,Cat No.K1003-100MG)。
二、实验步骤
配置100mM的PBS缓冲液，用该缓冲液配制0.25mg/mL的微粒体溶液和7.5mM的MgCl 2和5mM的NADPH溶液，用DMSO将浓度为30mM的储备液稀释成浓度为30mM、10mM、3mM、1mM、0.3mM、0.03mM、0.003mM、0mM的系列溶液I，再用磷酸缓冲液(PBS)，将上述系列溶液I稀释200倍得到系列待测溶液II(150、50、15、5、1.5、0.15、0.015、0μM)。用PBS稀释至15μM浓度的咪达唑仑工作液。
分别取配制在7.5mM MgCl 2中的0.25mg/mL的微粒体溶液40μL，再分别取15μM的咪达唑仑工作液和化合物工作液(150、50、15、5、1.5、0.15、0.015、0μM)各20μL，混合均匀。阳性对照组用相同浓度的酮康唑代替化合物。同时将5mM的NADPH溶液一起在37℃预孵育5分钟。5分钟之后每个孔中加入20μL NADPH，启动反应，孵育30分钟。所有孵育样品设双样本。30分钟后向所有样本中加入250μL含内标的乙腈，混匀，800rpm摇10分钟，然后3700rpm离心10分钟。取100μL的上清液与80μL的超纯水混匀，转移至LC-MS/MS分析。
数值经Graphpad Prism计算得到药物对CYP3A4咪达唑仑代谢位点的IC 50值见表5。
表5.本公开化合物对人肝微粒体CYP3A4的咪达唑仑代谢位点的IC 50值
化合物编号 IC 50(μM)
1 15.38
结论：本公开所要保护的化合物对人肝微粒体CYP3A4的咪达唑仑代谢位点抑制作用弱，表现出更好的安全性，提示不会发生基于CYP3A4代谢咪达唑仑代谢位点的代谢性药物相互作用。
测试例6、本公开化合物对人肝微粒体CYP3A4睾酮代谢位点的酶活性的抑制作用
本公开化合物对人肝微粒体CYP3A4睾酮代谢位点的酶活性采用如下实验方法测定：
一、实验材料及仪器
1.磷酸缓冲液(20×PBS，购买自生工),
2.NADPH(ACROS，A2646-71-1),
3.人肝微粒体(Corning Gentest，Cat No，452161，Lot No.905002,Donor36),
4.ABI QTrap 4000液质两用仪(AB Sciex),
5.ZORBAX Extend-C18,3×50mm,3.5μm(美国安捷伦公司),
6.CYP探针底物(睾酮,沃凯，CAS No.[58-22-0]/75μM),和阳性对照抑制剂(酮康唑,SIGMA,Cat No.K1003-100MG)。
二、实验步骤
配置100mM的PBS缓冲液，配置100mM的PBS缓冲液，用该缓冲液配制0.25mg/ml 的微粒体溶液和7.5mM的MgCl 2和5mM的NADPH溶液，用DMSO将浓度为30mM的储备液稀释成浓度为30mM、10mM、3mM、1mM、0.3mM、0.03mM、0.003mM、0mM的系列溶液I，再用磷酸缓冲液(PBS)，将上述系列溶液I稀释200倍得到系列待测溶液II(150、50、15、5、1.5、0.15、0.015、0μM)。用PBS稀释至375μM浓度的睾酮工作液。
取配制在7.5mM MgCl 2中的0.25mg/mL的微粒体溶液40μL，再分别取375μM的睾酮工作液和化合物工作液(150、50、15、5、1.5、0.15、0.015、0μM)各20μL，混合均匀。阳性对照组用相同浓度的酮康唑代替化合物。同时将5mM的NADPH溶液一起在37℃预孵育5分钟。5分钟之后每个孔中加入20μL NADPH，启动反应，孵育30分钟。30分钟后向所有样本中加入250μL含内标的乙腈，混匀，800rpm摇10分钟，然后3700rpm离心10分钟。取100μL的上清液与80μL的超纯水混匀，转移至LC-MS/MS分析。
数值经Graphpad Prism计算得到药物对CYP3A4睾酮代谢位点的IC 50值见表6。
表6.本公开化合物对人肝微粒体CYP3A4的睾酮代谢位点的IC 50值
化合物编号 IC 50(μM)
1 >30
结论：本公开所要保护的化合物对人肝微粒体CYP3A4的睾酮代谢位点的抑制较弱，表现出更好的安全性。
测试例7、本公开化合物对人肝微粒体CYP3A4咪达唑仑代谢位点的酶活性的时间依赖性抑制作用(Time-dependent inhibition)。
本公开化合物对人肝微粒体CYP3A4咪达唑仑代谢位点的时间依赖性CYP酶活性抑制作用采用如下实验方法测定：
一、实验材料及仪器
1.磷酸缓冲液(20×PBS，购买自生工),
2.NADPH(ACROS，A2646-71-1),
3.人肝微粒体(Corning Gentest，Cat No，452161，Lot No.9050002,Donor,36)，
4.ABI QTrap 4000液质两用仪(AB Sciex),
5.ZORBAX Extend-C18,3×50mm,3.5μm(美国安捷伦公司)，
6.CYP探针底物(咪达唑仑,TRC,M343000/3μM)和阳性对照抑制剂(维拉帕米,Adamas Reagent Co,Ltd,Cat No.25904A)。
二、实验步骤
配置100mM的PBS缓冲液，用该缓冲液配制0.25mg/ml的微粒体溶液和7.5mM的MgCl 2和5mM的NADPH溶液，用DMSO将浓度为30mM的储备液稀释成浓度为30mM、10mM、3mM、1mM、0.3mM、0.1mM、0.03mM、0mM的系列溶液I，再用磷酸缓冲液(PBS)，将上述系列溶液I稀释200倍得到系列待测溶液II(150、50、15、5、1.5、0.5、0.15、0μM)。用PBS稀释至15μM浓度的咪达唑仑工作液。
将上述制备好的待测系列溶液摇匀，分装20μL到对应的反应板(设+NADPH、T0、 -NADPH组)中，设3个平行，再往每块96孔板中加入40μL肝微粒体工作液，T0板中加入20μL对应的底物溶液，T0板和+NADPH组分别加入20μLNADPH，记时开始，放入水浴锅37℃孵育，孵育30min后取出T0板，用含内标的ACN的溶液250μL终止反应；+NADPH组补上20μL对应的底物溶液，-NADPH组补上20μL对应的底物溶液和20μLNADPH，记时开始，放入水浴锅37℃孵育，孵育30min后取出，用含内标的ACN的溶液250μL终止反应。然后摇床800rpm摇10min，离心机4000rpm离心15min。取上清100μL与80μL超纯水混匀后转移至LC-MS/MS分析。
数值经Graphpad Prism计算得到药物对CYP3A4咪达唑仑代谢位点的IC 50值见表7。
表7.本公开化合物对人肝微粒体CYP3A4的咪达唑仑代谢位点的IC 50值和IC 50Shift倍数
Figure PCTCN2022104669-appb-000013
结论：本公开所要保护的化合物对人肝微粒体CYP3A4的咪达唑仑代谢位点抑制作用弱，表现出更好的安全性，提示不会发生基于CYP3A4代谢咪达唑仑代谢位点的代谢性药物相互作用。
测试例8、本公开化合物对人肝微粒体CYP3A4睾酮代谢位点的酶活性的时间依赖性抑制作用
本公开化合物对人肝微粒体CYP3A4睾酮代谢位点的酶活性的时间依赖性抑制作用采用如下实验方法测定：
一、实验材料及仪器
1.磷酸缓冲液(20×PBS，购买自生工),
2.NADPH(ACROS，A2646-71-1),
3.人肝微粒体(Corning Gentest，Cat No，452161，Lot No.905002,Donor36),
4.ABI QTrap 4000液质两用仪(AB Sciex),
5.ZORBAX Extend-C18,3×50mm,3.5μm(美国安捷伦公司),
6.CYP探针底物(睾酮,沃凯，CAS No.[58-22-0]/75μM),和阳性对照抑制剂(维拉帕米,Adamas Reagent Co Ltd,Cat No.25904A)。
二、实验步骤
配置100mM的PBS缓冲液，用该缓冲液配制0.25mg/mL的微粒体溶液和7.5mM的MgCl 2和5mM的NADPH溶液，用DMSO将浓度为30mM的储备液稀释成浓度为30mM、10mM、3mM、1mM、0.3mM、0.1mM、0.03mM、0mM的系列溶液I，再用磷酸缓冲液(PBS)，将上述系列溶液I稀释200倍得到系列待测溶液II(150、50、15、5、1.5、0.5、0.15、0μM)。用PBS稀释至15μM浓度的咪达唑仑工作液。
将上述制备好的待测系列溶液摇匀，分装20μL到对应的反应板(设+NADPH、T0、 -NADPH组)中，设3个平行，再往每块96孔板中加入40μL肝微粒体工作液，T0板中加入20μL对应的底物溶液，T0板和+NADPH组分别加入20μLNADPH，记时开始，放入水浴锅37℃孵育，孵育30min后取出T0板，用含内标的ACN的溶液250μL终止反应；+NADPH组补上20μL对应的底物溶液，-NADPH组补上20μL对应的底物溶液和20μLNADPH，记时开始，放入水浴锅37℃孵育，孵育30min后取出，用含内标的ACN的溶液250μL终止反应。然后摇床800rpm摇10min,离心机4000rpm离心15min。取上清100μL与80μL超纯水混匀后转移至LC-MS/MS分析。
数值经Graphpad Prism计算得到药物对CYP3A4睾酮代谢位点的IC 50值见表8。
表8.本公开化合物对人肝微粒体CYP3A4的睾酮代谢位点的IC 50值和IC 50shift倍数
Figure PCTCN2022104669-appb-000014
结论：本公开所要保护的化合物对人肝微粒体CYP3A4的睾酮代谢位点的抑制较弱，表现出更好的安全性，提示不会发生基于CYP3A4发生代谢性药物相互作用。
测试例9
1、实验目的
应用全自动膜片钳在转染hERG钾通道的稳定细胞株上测试本公开化合物对hERG钾电流的阻断作用。
2、实验方法
2.1实验材料与仪器
2.1.1实验材料：
Figure PCTCN2022104669-appb-000015
2.1.2实验仪器：
Figure PCTCN2022104669-appb-000016
Figure PCTCN2022104669-appb-000017
2.2全自动膜片钳实验步骤
HEK293-hERG稳定细胞株按照1:4的密度在MEM/EBSS培养基(10％FBS，400μg/ml G418,1％MEM非必需氨基酸溶液(100×),1％丙酮酸钠溶液)中进行传代培养，培养48-72小时之内进行全自动膜片钳实验。实验当天将细胞用0.25％胰酶消化后，离心收集细胞，用细胞外液(140mM NaCl，4mM KCl，1mM MgCl 2，2mM CaCl 2，5mMD一水葡萄糖，10mM Hepes,pH7.4,298mOsm)重悬细胞制成细胞悬液。将细胞悬液放置在Patchliner仪器的细胞库上，Patchliner仪器利用负压控制器将细胞加到芯片(NPC-16)上，负压将单个细胞吸引在芯片的小孔上。当形成全细胞模式后，仪器将按照设定的hERG电流电压程序得到hERG电流，然后仪器自动的由低浓度到高浓度，进行化合物灌流。通过HEAK Patchmaster,HEAK EPC10膜片钳放大器(Nanion)和Pathlinersoftware以及Pathcontrol HTsoftware进行数据收集和分析，对化合物各浓度下的电流以及空白对照电流进行分析。
2.3测试结果
本公开化合物对hERG钾电流的阻断作用通过以上的试验进行测定，测得的IC 50值见表9。
表9.本公开化合物对hERG钾电流的阻断作用的IC 50
化合物编号 IC 50(μM)
1 >30
注：IC 50≥30μM为无抑制活性；30>IC 50≥10μM为有微弱抑制活性；10>IC 50≥1μM为有中等的抑制活性；IC 50<1μM为有强的抑制活性。
结论：本公开所要保护的化合物对hERG没有抑制活性，表现为更好的安全性。
实施例2、式(I)所示化合物磷酸盐I晶型的制备
称取约450mg式(I)所示化合物溶于10mL丙酮，加入1.25mL 1.46M磷酸乙醇溶液，升降温24h后样品离心，40℃真空干燥得固体，经X-射线粉末衍射检测，将该固体定义为I晶型，XRPD谱图如图2所示，其特征峰位置如表10所示，TGA谱图(附图3)显示：I晶型在30-115℃之间失重0.38％；DSC谱图(附图4)显示I晶型具有一个吸热峰，峰值为188.98℃；DVS后的XRPD谱图(附图5)显示晶型未发生变化；DVS谱图见附图6；单晶谱图见附图7，其中游离碱与磷酸根的摩尔比值为1：3；轴长
Figure PCTCN2022104669-appb-000018
Figure PCTCN2022104669-appb-000019
轴角α＝89.575(2)°；β＝89.859(2)°；γ＝60.745 (2)°；晶体体积
Figure PCTCN2022104669-appb-000020
表10.I晶型特征峰
Figure PCTCN2022104669-appb-000021
实施例3、式(I)所示化合物磷酸盐I晶型的制备
称取约15mg式(I)所示化合物溶于0.2mL乙酸乙酯，加入41μL 1.46M的磷酸乙醇溶液，升降温24h后离心干燥得固体，经X-射线粉末衍射检测为I晶型。
实施例4、式(I)所示化合物硫酸盐的A晶型的制备
称取约15mg式(I)所示化合物溶于0.2mL乙醇，加入16.3μL 1.84M硫酸乙醇溶液，升降温24h后离心干燥，得固体，经X-射线粉末衍射检测，将该产物定义为晶型A，XRPD谱图如图8；其特征峰位置如表11所示，TGA谱图(附图9)显示：A晶型在30-90℃之间失重6.84％；DSC谱图(附图10)显示A晶型具有两个吸热峰，峰值为68.13℃和146.66℃；DVS后的样品潮解；DVS谱图见图11；单晶谱图见附图12，其中游离碱与硫酸根的摩尔比值为1：1；轴长
Figure PCTCN2022104669-appb-000022
Figure PCTCN2022104669-appb-000023
轴角β＝90°；晶体体积
Figure PCTCN2022104669-appb-000024
表11.A晶型特征峰
Figure PCTCN2022104669-appb-000025
Figure PCTCN2022104669-appb-000026
实施例5、式(I)所示化合物硫酸盐的A晶型的制备
称取约15mg式(I)所示化合物溶于0.2mL乙酸乙酯，加入16.3μL 1.84M硫酸乙醇溶液，升降温24h后离心干燥的固体，经X-射线粉末衍射检测为A晶型。
实施例6、式(I)所示化合物硫酸盐的A晶型的制备
称取约148mg式(I)所示化合物溶于1mL丙酮，加入0.163mL 1.84M硫酸乙醇溶液，升降温40h后离心干燥后的固体，经X-射线粉末衍射检测为A晶型。
实施例7、式(I)所示化合物硫酸盐的A晶型的制备
称取约296mg式(I)所示化合物溶于1mL甲醇，加入0.33mL 1.84M硫酸甲醇溶液，升降温24h后离心干燥后的固体，经X-射线粉末衍射检测为A晶型。
实施例8、式(I)所示化合物L-酒石酸盐无定型的制备
将式(I)所示化合物25mg加入到0.5mL异丙醇中，搅拌溶清，L-酒石酸8mg用异丙醇0.5mL稀释，滴加入以上反应液中，室温下氮气保护搅拌形成白色浑浊液，继续搅拌16小时后过滤，收集滤饼，真空干燥，得到白色固体，经X-射线粉末衍射检测，该产物为无定型，XRPD谱图如图13。
实施例9、柠檬酸盐无定型的制备
将式(I)所示化合物25mg加入到0.5mL异丙醇中，搅拌溶清，柠檬酸10mg用异丙醇0.5mL稀释，滴加入以上反应液中，室温下氮气保护搅拌形成白色浑浊液，继续搅拌16小时后过滤，收集滤饼，真空干燥，得到白色固体，经X-射线粉末衍射检测，该产物为无定型，XRPD谱图如图14。
实施例10、盐酸盐的制备
称取约15mg式(I)所示化合物溶于0.2mL乙醇，加入50μL 1.2M的盐酸乙醇溶液，升降温24h并搅拌，室温挥发得油胶状固体。
实施例11、氢溴酸盐的制备
称取约15mg式(I)所示化合物溶于0.2mL乙酸乙酯，加入80μL 0.75M的氢溴酸乙醇溶液，升降温24h并搅拌，室温挥发得油胶状固体。
实施例12、甲磺酸盐的制备
称取约15mg式(I)所示化合物溶于0.2mL乙酸乙酯，加入39μL 1.53M的甲磺酸水溶液，升降温24h并搅拌，室温挥发得油胶状固体。
实施例13、甲酸盐的制备
称取约15mg式(I)所示化合物溶于0.2mL丙酮，加入22.6μL 2.65M的甲酸水溶液，升降温24h并搅拌，室温挥发得油胶状固体。
实施例14、乙酸盐的制备
称取约15mg式(I)所示化合物溶于0.2mL乙醇，加入34.2μL 1.75M的乙酸水溶液，升降温24h并搅拌，室温挥发得油胶状固体。
实施例15、琥珀酸盐的制备
称取约15mg式(I)所示化合物溶于0.2mL丙酮，加入36μL 0.83M的琥珀酸乙醇溶液，升降温24h并搅拌，室温挥发得油胶状固体。
实施例16、马来酸盐的制备
称取约15mg式(I)所示化合物溶于0.2mL乙酸乙酯，加入30μL 1M的马来酸水溶液，升降温24h并搅拌，室温挥发得油胶状固体。
实施例17、苹果酸盐的制备
称取约15mg式(I)所示化合物溶于0.2mL乙酸乙酯，加入30μL 1M的苹果酸水溶液，升降温24h并搅拌，室温挥发得油胶状固体。
实施例18、对甲苯磺酸盐的制备
称取约15mg式(I)所示化合物溶于0.2mL乙醇，加入10.28mg的对甲苯磺酸，
加入60μL纯化水，升降温24h并搅拌，室温挥发得油胶状固体。
实施例19、引湿性研究
采用Surface Measurement Systems intrinsic，在25℃，湿度从50％起，考察湿度范围为0％-95％，步进为10％，判断标准为每个梯度质量变化dM/dT≤0.002％，TMAX 360min，循环两圈。
表12.引湿性研究数据
Figure PCTCN2022104669-appb-000027
实施例20、影响因素稳定性研究
将游离态无定形、磷酸盐I晶型、硫酸盐A晶型敞口平摊放置，分别考察在光照(4500Lux)、高温(40℃、60℃)、高湿(RH 75％、RH 92.5％)条件下样品的稳定性，取样考察期为30天。
表13.影响因素30天试验1个月结果
Figure PCTCN2022104669-appb-000028
Figure PCTCN2022104669-appb-000029
结论：影响因素实验表明：磷酸盐I晶型在影响因素条件下放置30天物理和化学稳定性良好。硫酸盐的A晶型除光照条件下，其他条件下物理和化学稳定性良好，高湿条件下潮解。无定形高湿条件下潮解，其他条件下物理稳定，化学稳定性高温和光照条件下略差。
实验例21、长期/加速稳定性
将游离态无定形、磷酸盐I晶型、硫酸盐A晶型分别放置25℃/60％RH和40℃/75％RH条件考察其稳定性：
表14.游离碱无定形长期/加速稳定性
Figure PCTCN2022104669-appb-000030
Figure PCTCN2022104669-appb-000031
表15.磷酸盐I晶型长期/加速稳定性
Figure PCTCN2022104669-appb-000032
表16.硫酸盐A晶型长期/加速稳定性
Figure PCTCN2022104669-appb-000033
结论：长期加速3个月的稳定性结果显示，磷酸盐I晶型、硫酸盐A晶型物理化学稳定性良好，略优于游离态无定形。

Claims (17)

  1. 一种式(I)所示化合物的药学上可接受的盐，所述药学上可接受的盐选自磷酸盐、硫酸盐、酒石酸盐、柠檬酸盐、盐酸盐、氢溴酸盐、甲磺酸盐、甲酸盐、乙酸盐、琥珀酸盐、马来酸盐、苹果酸盐和对甲苯磺酸盐，
    Figure PCTCN2022104669-appb-100001
  2. 根据权利要求1所述的式(I)所示化合物的药学上可接受的盐，其中式(I)所示化合物与酸分子的物质的量比选自5:1-1:5；优选1:1或1:3。
  3. 根据权利要求1-2任一项所述的式(I)所示化合物的药学上可接受的盐，其为磷酸盐，其中式(I)所示化合物与磷酸分子的物质的量比为1:3。
  4. 一种式(I)所示化合物的磷酸盐的I晶型，其中式(I)所示化合物与磷酸分子的物质的量比为1:3，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.1、13.5、19.2、22.3处有特征峰，
    Figure PCTCN2022104669-appb-100002
    优选地，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.1、13.5、13.9、19.2、21.8、22.3、23.4处有特征峰；
    最优选地，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.1、12.4、13.5、13.9、15.0、16.3、16.9、19.2、20.8、21.8、22.3、23.4、24.1、24.6、26.2、27.5、30.0处有特征峰。
  5. 根据权利要求1-2任一项所述的式(I)所示化合物的药学上可接受的盐，其为硫酸盐，其中式(I)所示化合物与硫酸分子的物质的量比为1:1。
  6. 一种式(I)所示化合物的硫酸盐的A晶型，其中式(I)所示化合物与硫酸分子的物质的量比为1:1，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.6、7.9、15.1、20.7、22.1处有特征峰，
    Figure PCTCN2022104669-appb-100003
    优选地，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.6、7.9、13.4、14.5、15.1、19.3、19.8、20.7、22.1、24.8、25.4、25.7、27.6处有特征峰；
    最优选地，以衍射角2θ角度表示的X-射线粉末衍射图谱，在6.6、7.9、11.6、13.4、14.5、15.1、15.9、18.1、19.3、19.8、20.7、22.1、24.0、24.8、25.4、25.7、27.2、27.6、28.3、29.2处有特征峰。
  7. 根据权利要求4或6任一项所述的式(I)所示化合物的药学上可接受的盐的晶型，所述2θ角的误差范围为±0.2。
  8. 一种权利要求1-3任一项所述的式(I)所示化合物的药学上可接受的盐，所述药学上可接受的盐为磷酸盐，或权利要求4所述的式(I)所示化合物的磷酸盐的I晶型的制备方法，包括式(I)所示化合物与磷酸反应的步骤。
  9. 一种权利要求4所述的式(I)所示化合物的磷酸盐的I晶型的制备方法，包括以下步骤：
    1)配置式(I)所示化合物的溶剂A溶液，所述溶剂A选自酮类溶剂、酯类溶剂或醚类溶剂；优选丙酮、乙酸乙酯或甲基叔丁基醚；
    2)配置磷酸的溶剂B溶液，所述溶剂B选自水或醇类溶剂；优选甲醇、乙醇或水；
    3)混合式(I)所示化合物的溶剂A溶液与磷酸的溶剂B溶液后结晶析出。
  10. 一种权利要求1-2或5任一项所述的式(I)所示化合物的药学上可接受的盐，所述药学上可接受的盐为硫酸盐，或根据权利要求6所述的式(I)所示化合物的硫酸盐 的A晶型的制备方法，包括式(I)所示化合物与硫酸反应的步骤。
  11. 一种权利要求6所述的式(I)所示化合物的硫酸酸盐的A晶型的制备方法，包括以下步骤：
    1)配置式(I)所示化合物的溶剂A溶液，所述溶剂A选自酮类溶剂、酯类溶剂、腈类溶剂或醇类溶剂；所述溶剂A优选丙酮、乙酸乙酯、乙腈、甲醇或乙醇；
    2)配置硫酸的溶剂B溶液，所述溶剂B选自水、腈类溶剂或醇类溶剂；所述溶剂B优选甲醇、乙醇、水、乙腈；
    3)混合式(I)所示化合物的溶剂A溶液与硫酸的溶剂B溶液后结晶析出。
  12. 一种权利要求1所述的式(I)所示化合物的药学上可接受的盐的制备方法，包括式(I)所示化合物与酒石酸、柠檬酸、盐酸、氢溴酸、甲磺酸、甲酸、乙酸、琥珀酸、马来酸、苹果酸或对甲苯磺酸反应成盐的步骤。
  13. 一种药物组合物，包含以下成分：
    1)根据权利要求1-3、5任一项所述的式(I)所示化合物的药学上可接受的盐或根据权利要求4、6-7任一项所述的式(I)所示化合物的药学上可接受的盐的晶型，或其混合物；和
    2)任选自药学上可接受的载体、稀释剂或赋形剂。
  14. 一种药物组合物的制备方法，包括将
    1)根据权利要求1-3、5任一项所述的式(I)所示化合物的药学上可接受的盐或根据权利要求4、6-7任一项所述的式(I)所示化合物的药学上可接受的盐的晶型，或其混合物；和
    2)任选自药学上可接受的载体、稀释剂或赋形剂混合的步骤。
  15. 权利要求1-3、5任一项所述的式(I)所示化合物的药学上可接受的盐或根据权利要求4、6-7任一项所述的式(I)所示化合物的药学上可接受的盐的晶型，或其混合物，或权利要求13所述的药物组合物在制备雌激素受体调节剂中的用途；优选为在制备雌激素受体下调剂中的用途。
  16. 权利要求1-3、5任一项所述的式(I)所示化合物的药学上可接受的盐或根据权利要求4、6-7任一项所述的式(I)所示化合物的药学上可接受的盐的晶型，或其混合物，或权利要求13所述的药物组合物在制备预防和/或治疗癌症的药物中的用途，其中所述的癌症优选选自乳腺癌、子宫内膜癌、子宫癌、宫颈癌、皮肤癌、前列腺癌、卵巢癌、输 卵管肿瘤、血友病和白血病。
  17. 权利要求1-3、5任一项所述的式(I)所示化合物的药学上可接受的盐或根据权利要求4、6-7任一项所述的式(I)所示化合物的药学上可接受的盐的晶型，或其混合物，或权利要求13所述的药物组合物在制备预防和/或治疗雌激素受体介导的或依赖性的疾病或病症的药物中的用途；优选地，其中所述的雌激素受体介导的或依赖性的疾病或病症选自癌症、中枢神经系统缺陷、心血管系统缺陷、血液系统缺陷、免疫及炎症疾病、易感性感染、代谢缺陷、神经缺陷、精神缺陷和生殖缺陷；优选地，其中所述的癌症选自乳腺癌、子宫内膜癌、子宫癌、宫颈癌、皮肤癌、前列腺癌、卵巢癌、输卵管肿瘤、血友病和白血病；更优选，其中所述的癌症选自乳腺癌、卵巢癌、子宫内膜癌、前列腺癌和子宫癌。
PCT/CN2022/104669 2021-07-09 2022-07-08 一种三环四氢异喹啉类衍生物的盐型 Ceased WO2023280309A1 (zh)

Priority Applications (8)

Application Number Priority Date Filing Date Title
AU2022307207A AU2022307207A1 (en) 2021-07-09 2022-07-08 Salt types of tricyclic tetrahydro isoquinoline derivative
MX2024000486A MX2024000486A (es) 2021-07-09 2022-07-08 Tipos de sal de derivado de tetrahidroisoquinolina triciclico.
KR1020247004295A KR20240035515A (ko) 2021-07-09 2022-07-08 삼중 고리 테트라하이드로 이소퀴놀린계 유도체의 염의 형태
CA3225268A CA3225268A1 (en) 2021-07-09 2022-07-08 Salt types of tricyclic tetrahydro isoquinoline derivative
JP2024500624A JP2024525643A (ja) 2021-07-09 2022-07-08 三環式テトラヒドロイソキノリン系誘導体の塩の形態
CN202280048549.0A CN117642405B (zh) 2021-07-09 2022-07-08 一种三环四氢异喹啉类衍生物的盐型
US18/576,806 US20240374581A1 (en) 2021-07-09 2022-07-08 Salt types of tricyclic tetrahydro isoquinoline derivative
EP22837054.0A EP4368624A4 (en) 2021-07-09 2022-07-08 SALT TYPES OF A TRICYCLIC TETRAHYDROISOQUINOLINE DERIVATIVE

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CN202110779905.3 2021-07-09
CN202110779905 2021-07-09

Publications (1)

Publication Number Publication Date
WO2023280309A1 true WO2023280309A1 (zh) 2023-01-12

Family

ID=84801329

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2022/104669 Ceased WO2023280309A1 (zh) 2021-07-09 2022-07-08 一种三环四氢异喹啉类衍生物的盐型

Country Status (10)

Country Link
US (1) US20240374581A1 (zh)
EP (1) EP4368624A4 (zh)
JP (1) JP2024525643A (zh)
KR (1) KR20240035515A (zh)
CN (1) CN117642405B (zh)
AU (1) AU2022307207A1 (zh)
CA (1) CA3225268A1 (zh)
MX (1) MX2024000486A (zh)
TW (1) TW202317582A (zh)
WO (1) WO2023280309A1 (zh)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2025157258A1 (zh) * 2024-01-25 2025-07-31 山东盛迪医药有限公司 一种包含选择性雌激素受体降解剂的药物组合物及其制备方法
WO2025209455A1 (zh) * 2024-04-01 2025-10-09 山东盛迪医药有限公司 一种选择性雌激素受体降解剂的制备方法

Citations (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1113007A1 (en) 1999-12-24 2001-07-04 Pfizer Inc. Tetrahydroisoquinoline compounds as estrogen agonists/antagonists
WO2014106848A1 (en) 2013-01-07 2014-07-10 B.G. Negev Technologies And Applications Ltd. Coumestan, coumestrol, coumestan derivatives and processes of making the same and uses of same
WO2014135834A1 (en) 2013-03-05 2014-09-12 Cipla House Oxalic acid addition salts and/or solvates of a selective estrogen receptor modulator
WO2014141292A2 (en) 2013-03-04 2014-09-18 Intas Pharmaceuticals Limited Endoxifen citrate polymorph and process for preparing the same
WO2014151899A1 (en) 2013-03-14 2014-09-25 Seragon Pharmaceuticals, Inc. Polycyclic estrogen receptor modulators and uses thereof
WO2014165723A2 (en) 2013-04-04 2014-10-09 The Administrators Of The Tulane Educational Fund Enhanced osteogenic activity of daidzein analogs on human mesenchymal stem cells
WO2014191726A1 (en) 2013-05-28 2014-12-04 Astrazeneca Ab Chemical compounds
WO2015092634A1 (en) 2013-12-16 2015-06-25 Novartis Ag 1,2,3,4-tetrahydroisoquinoline compounds and compositions as selective estrogen receptor antagonists and degraders
WO2016202161A1 (zh) * 2015-06-16 2016-12-22 江苏恒瑞医药股份有限公司 哌啶类衍生物、其制备方法及其在医药上的应用
CN109219604A (zh) * 2016-04-08 2019-01-15 豪夫迈·罗氏有限公司 四氢异喹啉雌激素受体调节剂及其用途
CN109776523A (zh) * 2017-11-14 2019-05-21 杭州安霖药业有限公司 杂环化合物及其在医药上的应用
WO2019228443A1 (en) 2018-05-30 2019-12-05 Dizal (Jiangsu) Pharmaceutical Co., Ltd. Selective estrogen receptor downregulators and uses thereof
WO2021139756A1 (zh) 2020-01-10 2021-07-15 江苏恒瑞医药股份有限公司 三环四氢异喹啉类衍生物、其制备方法及其在医药上的应用

Patent Citations (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1113007A1 (en) 1999-12-24 2001-07-04 Pfizer Inc. Tetrahydroisoquinoline compounds as estrogen agonists/antagonists
WO2014106848A1 (en) 2013-01-07 2014-07-10 B.G. Negev Technologies And Applications Ltd. Coumestan, coumestrol, coumestan derivatives and processes of making the same and uses of same
WO2014141292A2 (en) 2013-03-04 2014-09-18 Intas Pharmaceuticals Limited Endoxifen citrate polymorph and process for preparing the same
WO2014135834A1 (en) 2013-03-05 2014-09-12 Cipla House Oxalic acid addition salts and/or solvates of a selective estrogen receptor modulator
WO2014151899A1 (en) 2013-03-14 2014-09-25 Seragon Pharmaceuticals, Inc. Polycyclic estrogen receptor modulators and uses thereof
WO2014165723A2 (en) 2013-04-04 2014-10-09 The Administrators Of The Tulane Educational Fund Enhanced osteogenic activity of daidzein analogs on human mesenchymal stem cells
WO2014191726A1 (en) 2013-05-28 2014-12-04 Astrazeneca Ab Chemical compounds
WO2015092634A1 (en) 2013-12-16 2015-06-25 Novartis Ag 1,2,3,4-tetrahydroisoquinoline compounds and compositions as selective estrogen receptor antagonists and degraders
WO2016202161A1 (zh) * 2015-06-16 2016-12-22 江苏恒瑞医药股份有限公司 哌啶类衍生物、其制备方法及其在医药上的应用
CN109219604A (zh) * 2016-04-08 2019-01-15 豪夫迈·罗氏有限公司 四氢异喹啉雌激素受体调节剂及其用途
CN109776523A (zh) * 2017-11-14 2019-05-21 杭州安霖药业有限公司 杂环化合物及其在医药上的应用
WO2019228443A1 (en) 2018-05-30 2019-12-05 Dizal (Jiangsu) Pharmaceutical Co., Ltd. Selective estrogen receptor downregulators and uses thereof
WO2021139756A1 (zh) 2020-01-10 2021-07-15 江苏恒瑞医药股份有限公司 三环四氢异喹啉类衍生物、其制备方法及其在医药上的应用

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
BIOORGANIC & MEDICINAL CHEMISTRY LETTERS, vol. 28, no. 14, 2018, pages 2528 - 2532
See also references of EP4368624A4

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2025157258A1 (zh) * 2024-01-25 2025-07-31 山东盛迪医药有限公司 一种包含选择性雌激素受体降解剂的药物组合物及其制备方法
WO2025209455A1 (zh) * 2024-04-01 2025-10-09 山东盛迪医药有限公司 一种选择性雌激素受体降解剂的制备方法

Also Published As

Publication number Publication date
EP4368624A1 (en) 2024-05-15
EP4368624A4 (en) 2024-12-18
CN117642405A (zh) 2024-03-01
CA3225268A1 (en) 2023-01-12
KR20240035515A (ko) 2024-03-15
MX2024000486A (es) 2024-01-30
AU2022307207A1 (en) 2024-02-15
JP2024525643A (ja) 2024-07-12
US20240374581A1 (en) 2024-11-14
CN117642405B (zh) 2026-05-12
TW202317582A (zh) 2023-05-01

Similar Documents

Publication Publication Date Title
US20050159451A1 (en) Amide derivatives as NMDA receptor antagonists
CN114746424B (zh) 三环四氢异喹啉类衍生物、其制备方法及其在医药上的应用
WO2020244349A1 (zh) 呋喃并咪唑并吡啶类化合物的合成方法、多晶型物、及盐的多晶型物
CA3219940A1 (en) Heterocyclic compounds and methods of preparation thereof
WO2023116882A1 (zh) 一种glp-1受体激动剂的可药用盐、结晶形式及其制备方法
KR20100031578A (ko) 5-ｈｔ6 수용체 친화성을 갖는 4&#39;-치환된 화합물
CN117642405B (zh) 一种三环四氢异喹啉类衍生物的盐型
WO2023116879A1 (zh) 一种glp-1受体激动剂的结晶形式及其制备方法
US9120764B2 (en) Chromene derivatives
WO2019149089A1 (zh) 含有羧酸基团的苯并氮杂环类化合物及其制备方法和用途
JP2020502188A (ja) Ａｒ−ｖ７インヒビター
CN115650902A (zh) 1′,2′-二氢-3′h-螺[环丁烷1,4′-异喹啉]-3′-酮衍生物的盐及其应用
TW202446373A (zh) 二氧代哌啶化合物或其可藥用鹽的無定形物、結晶、製備方法和用途
JP2025528074A (ja) 縮合三環系誘導体の結晶型及び調製方法
CA3219804A1 (en) Fused heterocyclic derivatives as negative allosteric modulators of mglu7 receptor
CN116891465A (zh) 一类n-取代的喹啉酮化合物、其制备方法和用途
CN111094273B (zh) 催产素受体抑制剂的晶型及其制备方法
CN111094272B (zh) 一种otr抑制剂的可药用盐、晶型及制备方法
EA052236B1 (ru) Типы солей трициклического производного тетрагидроизохинолина
HU226059B1 (en) Novel disubstituted trans-3,4,4a,5,6,10b-hexahydro-2h-naphto[1,2-b][1,4]oxazine derivatives, process for producing them and pharmaceutical compositions containing them
HK40076301B (zh) 三环四氢异喹啉类衍生物、其制备方法及其在医药上的应用
CN117510490A (zh) 一种稠合氮杂萘烷类衍生物的晶型及其制备方法
WO2023174300A1 (zh) Btk抑制剂的晶型及其酸式盐和其酸式盐的晶型
TW202436295A (zh) 喹啉胺類化合物的可藥用鹽、晶型及其製備方法
TW202436296A (zh) 喹啉胺類化合物晶型及其製備方法

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 22837054

Country of ref document: EP

Kind code of ref document: A1

WWE Wipo information: entry into national phase

Ref document number: 18576806

Country of ref document: US

WWE Wipo information: entry into national phase

Ref document number: 202280048549.0

Country of ref document: CN

Ref document number: MX/A/2024/000486

Country of ref document: MX

Ref document number: 3225268

Country of ref document: CA

Ref document number: 2401000122

Country of ref document: TH

ENP Entry into the national phase

Ref document number: 2024500624

Country of ref document: JP

Kind code of ref document: A

REG Reference to national code

Ref country code: BR

Ref legal event code: B01A

Ref document number: 112024000404

Country of ref document: BR

WWE Wipo information: entry into national phase

Ref document number: AU2022307207

Country of ref document: AU

ENP Entry into the national phase

Ref document number: 20247004295

Country of ref document: KR

Kind code of ref document: A

WWE Wipo information: entry into national phase

Ref document number: 1020247004295

Country of ref document: KR

WWE Wipo information: entry into national phase

Ref document number: 202490188

Country of ref document: EA

WWE Wipo information: entry into national phase

Ref document number: 2022837054

Country of ref document: EP

NENP Non-entry into the national phase

Ref country code: DE

ENP Entry into the national phase

Ref document number: 2022307207

Country of ref document: AU

Date of ref document: 20220708

Kind code of ref document: A

ENP Entry into the national phase

Ref document number: 2022837054

Country of ref document: EP

Effective date: 20240209

WWE Wipo information: entry into national phase

Ref document number: 11202400155Y

Country of ref document: SG

ENP Entry into the national phase

Ref document number: 112024000404

Country of ref document: BR

Kind code of ref document: A2

Effective date: 20240109