WO2019225552A1 - Salts of indazole derivative and crystals thereof - Google Patents

Salts of indazole derivative and crystals thereof Download PDF

Info

Publication number
WO2019225552A1
WO2019225552A1 PCT/JP2019/019947 JP2019019947W WO2019225552A1 WO 2019225552 A1 WO2019225552 A1 WO 2019225552A1 JP 2019019947 W JP2019019947 W JP 2019019947W WO 2019225552 A1 WO2019225552 A1 WO 2019225552A1
Authority
WO
WIPO (PCT)
Prior art keywords
crystal
ppm
powder
diffraction
salt
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/JP2019/019947
Other languages
French (fr)
Inventor
Keiko Onuma
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.)
Eisai R&D Management Co Ltd
Original Assignee
Eisai R&D Management 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 Eisai R&D Management Co Ltd filed Critical Eisai R&D Management Co Ltd
Publication of WO2019225552A1 publication Critical patent/WO2019225552A1/en
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Images

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D231/00—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings
    • C07D231/54—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings condensed with carbocyclic rings or ring systems
    • C07D231/56—Benzopyrazoles; Hydrogenated benzopyrazoles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the present invention relates to salts of an indazole derivative and crystals thereof.
  • tamoxifen has been shown to activate signaling activity in the endometrium leading to an increase in risk of endometrial cancers in the clinic (Fisher et al., (1994) J. Natl. Cancer Inst. Apr 6; 86 (7): 527-37; van Leeuwen et al., (1994) Lancet Feb 19; 343 (8895): 448-52).
  • fulvestrant since fulvestrant is a pure antagonist, it can lead to loss of bone density in post-menopausal women as ER ⁇ activity is critical for bone building.
  • clinical resistance is also beginning to emerge to these classes of ER ⁇ antagonists highlighting the need to develop next-generation compounds.
  • ESR1 gene which encodes ER ⁇ protein
  • ER+ breast cancer patients on average are treated with seven independent therapies including chemotherapies and various anti-estrogen therapies such as tamoxifen, fulvestrant and aromatase inhibitors.
  • chemotherapies such as tamoxifen, fulvestrant and aromatase inhibitors.
  • tamoxifen tamoxifen
  • fulvestrant fulvestrant
  • aromatase inhibitors aromatase inhibitors
  • TDI Time-Dependent Inhibition
  • TDI should be studied in standard in vitro screening protocols by pre-incubating the drug (a potential inhibitor) before the addition of a substrate (Food and Drug Administration (FDA) guidance; Cf. fda.gov/downloads/drugs/guidances/ucm292362.pdf (FDA guidance, In Vitro Metabolism- and Transporter- Mediated Drug-Drug Interaction Studies, Draft Guidance, October 24, 2017.)).
  • FDA Food and Drug Administration
  • TDI parameters i.e., k inact and K I .
  • a compound represented by the formula I namely, (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (hereafter referred to as compound (I)), suppresses the growth of both wild-type (WT) and ER ⁇ -mutant positive tumors.
  • WT wild-type
  • ER ⁇ -mutant positive tumors Generally, the physical properties of a compound, salts thereof, and their crystals used as a pharmaceutical product largely influence on the bioavailability of a drug, the purity of an active pharmaceutical ingredient, prescription of a preparation and the like.
  • An object of the present invention is therefore to provide salts of compound (I) or crystals thereof with a potential to be used as drug substance in pharmaceuticals.
  • the present inventor has found salts of compound (I) or crystals thereof with a potential to be used as drug substance in pharmaceuticals, thereby completing the invention.
  • ⁇ 1> A salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, and an acid selected from the group consisting of hydrobromic acid, phosphoric acid, sulfuric acid, benzenesulfonic acid, p-toluenesulfonic acid, succinic acid, maleic acid, mandelic acid and methanesulfonic acid.
  • ⁇ 3> The crystal according to ⁇ 2> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 19.7° in a powder X-ray diffractometry.
  • ⁇ 4> The crystal according to ⁇ 2> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 16.8°, 18.2° and 19.7° in a powder X-ray diffractometry.
  • ⁇ 9> The crystal according to ⁇ 8> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 18.4° in a powder X-ray diffractometry.
  • ⁇ 10> The crystal according to ⁇ 8> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 18.4°, 19.7° and 25.6° in a powder X-ray diffractometry.
  • ⁇ 15> The crystal according to ⁇ 14> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 6.5° in a powder X-ray diffractometry.
  • ⁇ 16> The crystal according to ⁇ 14> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 6.5°, 16.7° and 19.9° in a powder X-ray diffractometry.
  • ⁇ 21> The crystal according to ⁇ 20> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 19.0° in a powder X-ray diffractometry.
  • ⁇ 22> The crystal according to ⁇ 20> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 7.6°, 19.0° and 19.8° in a powder X-ray diffractometry.
  • ⁇ 27> The crystal according to ⁇ 26> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 10.9° in a powder X-ray diffractometry.
  • ⁇ 28> The crystal according to ⁇ 26> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 10.9°, 17.6° and 24.1° in a powder X-ray diffractometry.
  • ⁇ 33> The crystal according to ⁇ 32> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 8.9° in a powder X-ray diffractometry.
  • ⁇ 34> The crystal according to ⁇ 32> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 8.9°, 21.4° and 22.1° in a powder X-ray diffractometry.
  • ⁇ 39> The crystal according to ⁇ 38> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 18.2° in a powder X-ray diffractometry.
  • ⁇ 40> The crystal according to ⁇ 38> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 17.0°, 18.2° and 19.1° in a powder X-ray diffractometry.
  • ⁇ 45> The crystal according to ⁇ 44> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 18.9° in a powder X-ray diffractometry.
  • ⁇ 46> The crystal according to ⁇ 44> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 16.8°, 18.9° and 20.2° in a powder X-ray diffractometry.
  • ⁇ 51> The crystal according to ⁇ 50> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 19.4° in a powder X-ray diffractometry.
  • ⁇ 52> The crystal according to ⁇ 50> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 15.2°, 19.4° and 20.8° in a powder X-ray diffractometry.
  • ⁇ 57> The crystal according to ⁇ 56> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 18.2° in a powder X-ray diffractometry.
  • ⁇ 58> The crystal according to ⁇ 56> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 5.1°, 10.3° and 18.2° in a powder X-ray diffractometry.
  • ⁇ 63> The crystal according to ⁇ 62> above, characterized by having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 12.7° in a powder X-ray diffractometry.
  • ⁇ 64> The crystal according to ⁇ 62> above, characterized by having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 12.7°, 18.9° and 25.6° in a powder X-ray diffractometry.
  • ⁇ 67> A crystal of methanesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13 C NMR spectrum substantially the same as the solid state 13 C NMR spectrum shown in Figure 22.
  • ⁇ 68> A pharmaceutical composition comprising the salt or the crystal thereof according to any one of ⁇ 1> to ⁇ 67> above.
  • ⁇ 69> The pharmaceutical composition according to ⁇ 68> above, for use in treating cancer.
  • the salts of compound (I) and the crystals thereof provided by the present invention possess properties such as hygroscopicity as shown in the examples described in later and a potential to be used as drug substance in pharmaceuticals.
  • Figure 1 shows a powder X-ray diffraction pattern of the crystal of the compound (I) hydrochloride salt obtained in Example 1.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 2 shows a powder X-ray diffraction pattern of the crystal of the compound (I) hydrobromide salt obtained in Example 2.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 3 shows a powder X-ray diffraction pattern of the crystal of the compound (I) phosphate salt obtained in Example 3.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 4 shows a powder X-ray diffraction pattern of the crystal of the compound (I) sulfate salt obtained in Example 4.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 5 shows a powder X-ray diffraction pattern of the crystal of the compound (I) benzenesulfonate salt obtained in Example 5.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 6 shows a powder X-ray diffraction pattern of the crystal of the compound (I) p-toluenesulfonate salt obtained in Example 6.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 7 shows a powder X-ray diffraction pattern of the crystal of the compound (I) succinate salt obtained in Example 7.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 8 shows a powder X-ray diffraction pattern of the crystal form A of the compound (I) maleate salt obtained in Example 8.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 9 shows a powder X-ray diffraction pattern of the crystal form B of the compound (I) maleate salt obtained in Example 9.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 10 shows a powder X-ray diffraction pattern of the crystal of the compound (I) L-mandelate salt obtained in Example 10.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 11 shows a powder X-ray diffraction pattern of the crystal of the compound (I) methanesulfonate salt obtained in Example 11.
  • the abscissa shows the diffraction angle (2 ⁇ ) and the ordinate shows the peak intensity.
  • Figure 12 shows a solid state 13 C NMR spectrum of the crystal of the compound (I) hydrochloride salt obtained in Example 1.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 13 shows a solid state 13 C NMR spectrum of the crystal of the compound (I) hydrobromide salt obtained in Example 2.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 14 shows a solid state 13 C NMR spectrum of the crystal of the compound (I) phosphate salt obtained in Example 3.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 15 shows a solid state 13 C NMR spectrum of the crystal of the compound (I) sulfate salt obtained in Example 4.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 16 shows a solid state 13 C NMR spectrum of the crystal of the compound (I) benzenesulfonate salt obtained in Example 5.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 17 shows a solid state 13 C NMR spectrum of the crystal of the compound (I) p-toluenesulfonate salt obtained in Example 6.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 18 shows a solid state 13 C NMR spectrum of the crystal of the compound (I) succinate salt obtained in Example 7.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 19 shows a solid state 13 C NMR spectrum of the crystal form A of the compound (I) maleate salt obtained in Example 8.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 20 shows a solid state 13 C NMR spectrum of the crystal form B of the compound (I) maleate salt obtained in Example 9.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 21 shows a solid state 13 C NMR spectrum of the crystal of the compound (I) L-mandelate salt obtained in Example 10.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 22 shows a solid state 13 C NMR spectrum of the crystal of the compound (I) methanesulfonate salt obtained in Example 11.
  • the abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity.
  • Figure 23 is a graph showing hygroscopicity of the crystal of the compound (I) hydrochloride salt obtained in Example 1.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 24 is a graph showing hygroscopicity of the crystal of the compound (I) hydrobromide salt obtained in Example 2.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 25 is a graph showing hygroscopicity of the crystal of the compound (I) phosphate salt obtained in Example 3.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 26 is a graph showing hygroscopicity of the crystal of the compound (I) sulfate salt obtained in Example 4.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 27 is a graph showing hygroscopicity of the crystal of the compound (I) benzenesulfonate salt obtained in Example 5.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 28 is a graph showing hygroscopicity of the crystal of the compound (I) p-toluenesulfonate salt obtained in Example 6.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 29 is a graph showing hygroscopicity of the crystal of the compound (I) succinate salt obtained in Example 7.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 30 is a graph showing hygroscopicity of the crystal form A of the compound (I) maleate salt obtained in Example 8.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 31 is a graph showing hygroscopicity of the crystal form B of the compound (I) maleate salt obtained in Example 9.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 32 is a graph showing hygroscopicity of the crystal of the compound (I) L-mandelate salt obtained in Example 10.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 33 is a graph showing hygroscopicity of the crystal of the compound (I) methanesulfonate salt obtained in Example 11.
  • the abscissa shows the relative humidity and the ordinate shows the weight change.
  • Figure 34 is a graph showing in vitro proliferation effects of wild-type and mutant ER-bearing MCF7 lines to clinical therapies 4-hydroxytamoxifen (4-OHT), raloxifene and fulvestrant, where phenotypic resistance observed in mutant-bearing lines relative to control lines to existing clinical compounds, whereby MCF7 cells engineered to overexpress various ER ⁇ MUT showed partial resistance to various endocrine therapies.
  • Figure 35 is a graph showing antitumor and body weight effects of oral compound (I) as a hydrochloride salt in PDX-Y537S xenograft bearing athymic nude female mice.
  • Figure 36 is a graph showing the anti-tumor and body weight effects of compound (I), prepared as an HCl salt, in the MCF7 tumor model bearing ER ⁇ WT/WT xenograft.
  • Figure 37 is a graph showing the anti-tumor and body weight effects of compound (I), prepared as an HCl salt, in a PDX model bearing ER ⁇ WT/WT xenograft.
  • a “salt” refers to a chemical entity made up of the compound (I) as the basic component and a specific number of equivalents of an acid to the compound (I).
  • salts used herein include salts with organic carboxylic acids, with organic sulfonic acids and with inorganic acids, and in particular, pharmaceutically acceptable salts are preferred.
  • organic carboxylic acids examples include acetic acid, oxalic acid, maleic acid, mandelic acid, tartaric acid, fumaric acid, citric acid, malonic acid, succinic acid, benzoic acid, hippuric acid and malic acid.
  • Preferred examples of organic carboxylic acids include maleic acid, mandelic acid (preferably L-mandelic acid), tartaric acid, malonic acid and succinic acid.
  • organic sulfonic acids examples include methanesulfonic acid, trifluoromethanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid and camphorsulfonic acid.
  • Preferred examples of organic sulfonic acids include methanesulfonic acid, benzenesulfonic acid and p-toluenesulfonic acid.
  • inorganic acids examples include hydrofluoric acid, hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, perchloric acid, phosphoric acid, carbonic acid and bicarbonic acid.
  • Preferred examples of inorganic acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid and phosphoric acid.
  • a salt of the compound (I) may also be a solvate.
  • a solvate of the salt of compound (I) means a solid formed from the salt of the compound (I) together with solvent molecules.
  • the solvent in the solvate include: a ketone solvent such as acetone, methyl ethyl ketone or cyclohexanone; an ester solvent such as ethyl acetate or methyl acetate; an ether solvent such as 1, 2-dimethoxyethane or methyl-tert-butyl ether; an alcohol solvent such as methanol, ethanol, 1-propanol or isopropanol; a polar solvent such as N-methyl-2-pyrrolidone, N,N-dimethylformamide or dimethyl sulfoxide; and water.
  • a crystal refers to a crystal of the salt of compound (I). Accordingly, a crystal of hydrochloride salt of compound (I), for example, means a crystal of the salt formed between compound (I) and hydrochloric acid.
  • crystals preferred herein include: (a1) a crystal of hydrochloride salt of compound (I), having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 19.7° in a powder X-ray diffractometry; (a2) a crystal of hydrochloride salt of compound (I), having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 16.8°, 18.2° and 19.7° in a powder X-ray diffractometry; (a3) a crystal of hydrochloride salt of compound (I), having diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 12.2°, 16.8°, 18.2°, 19.7° and 23.3° in a powder X-ray diffractometry; (a4) a crystal of hydrochloride salt of compound (I), having one or more diffraction peaks at diffraction angles (2 ⁇ ⁇ 0.2°) of 12.2°, 16.8°, 18.2°,
  • the peaks in a powder X-ray diffractometry, described above, are characteristic for each of the crystal of hydrochloride salt of compound (I), the crystal of hydrobromide salt of compound (I), the crystal of phosphate salt of compound (I), the crystal of sulfate salt of compound (I), the crystal of benzenesulfonate salt of compound (I), the crystal of p-toluenesulfonate salt of compound (I), the crystal of succinate salt of compound (I), the crystal form A of maleate salt of compound (I), the crystal form B of maleate salt of compound (I), the crystal of L-mandelate salt of compound (I), and the crystal of methanesulfonate salt of compound (I).
  • having a diffraction peak at a diffraction angle (2 ⁇ ⁇ 0.2°) of 19.7° means “having a diffraction peak at a diffraction angle (2 ⁇ ) of 19.5° to 19.9°". The same is also applied to other diffraction angles.
  • peak intensities and half-value widths of diffraction angles (2 ⁇ ) in powder X-ray diffraction are different for each measurement because of differences in measurement conditions and dispersions of size and shape of each particle of powder crystal and not always stable even though forms of crystals are same. Therefore, in case of comparing a powder X-ray diffraction pattern, when diffraction angles (2 ⁇ ) are the same but peak intensities, relative peak intensities and half-value widths are different, those differences does not imply that the measured forms of crystals differ from each other.
  • a crystal of salt having a powder X-ray diffraction pattern which has aforementioned differences with respect to characteristic diffraction peaks of a certain crystal of salt according to the present invention, means that the crystal has the same crystal form of the crystal of salt according to the present invention.
  • powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 1 means it includes not only the case of having exactly the same powder X-ray diffraction pattern as shown in Figure 1, but also the case that at least one selected from the group consisting of peak intensities, relative peak intensities and half-value widths are different, or the case of having the characteristic peaks within an error range of approximately ⁇ 0.2° of the diffraction angles.
  • every crystal having such the powder X-ray diffraction pattern means that the crystal is identical to the crystal according to the present invention.
  • having peaks at chemical shifts of 144.5 ppm, 156.8 ppm and 165.8 ppm means “having peaks each substantially equivalent to the peaks at chemical shifts of 144.5 ppm, 156.8 ppm and 165.8 ppm, when solid state 13 C NMR spectrometry is performed under a conventional measurement condition or substantially the same condition as in the present specification".
  • having a solid state 13 C NMR spectrum substantially the same as the solid state 13 C NMR spectrum shown in Figure 12 means it includes not only the case of having exactly the same solid state 13 C NMR spectrum as shown in Figure 12, but also the case that peak intensities are different, or the case of having the characteristic peaks within an error range of approximately ⁇ 0.5 ppm. Thus every crystal having such the solid state 13 C NMR spectrum means that the crystal is identical to the crystal according to the present invention.
  • a salt of the compound (I) can be obtained by a conventional method for producing a salt. Specifically, it can be produced, for example, by suspending or dissolving compound (I) in a solvent, with heating if necessary, then by adding an acid to the obtained suspension or solution and by stirring or leaving the resultant suspension or solution for several minutes to several days at room temperature or with ice-bath cooling.
  • a salt of the compound (I) may be obtained as crystals or amorphous substances according to the production methods.
  • solvents to be used in these methods include alcohol solvents such as ethanol, 1-propanol and isopropanol; acetonitrile; ketone solvents such as acetone and 2-butanone; ester solvents such as ethyl acetate; saturated hydrocarbon solvents such as hexane and heptane; ether solvents such as t-butyl methyl ether or water.
  • alcohol solvents such as ethanol, 1-propanol and isopropanol
  • ketone solvents such as acetone and 2-butanone
  • ester solvents such as ethyl acetate
  • saturated hydrocarbon solvents such as hexane and heptane
  • ether solvents such as t-butyl methyl ether or water.
  • Each of these solvents may be used alone, or two or more may be mixed and used.
  • a crystal of the salt of compound (I) may be produced by the above-mentioned methods for producing a salt of the compound (I), or by heat-dissolving a salt of the compound (I) in a solvent and crystallizing it through cooling with stirring.
  • a salt of the compound (I) to be used in the crystallization may be in any form: it may be a solvate, a hydrate, an anhydrate, an amorphous substance, a crystalline substance (including those consisting of a plurality of crystalline polymorphs) or a combination thereof.
  • solvents to be used in the crystallization examples include alcohol solvents such as methanol, ethanol, isopropanol and 1-propanol; acetonitrile; amide solvents such as N,N-dimethylformamide; ester solvents such as ethyl acetate; saturated hydrocarbon solvents such as hexane and heptane; ketone solvents such as acetone and 2-butanone; ether solvents such as t-butyl methyl ether or water. Furthermore, each of these solvents may be used alone, or two or more may be mixed and used.
  • the amount of the solvent to be used may be suitably selected, provided that the lower limit is the amount with which the free form of compound (I) or the salt thereof is dissolved by heating or the suspension can be stirred, and that the upper limit is the amount with which the yield of the crystal is not significantly reduced.
  • a seed crystal (e.g., the crystal of the desired salt of compound (I)) may be added or may not be added during the crystallization.
  • the temperature at which the seed crystal is added is not particularly limited, but is preferably 0 to 80°C.
  • the temperature to be employed when the salt of compound (I) is dissolved by heating that at which compound (I) dissolves may be suitably selected depending on the solvent, but it is preferably within the range between the temperature at which the recrystallization solvent starts to reflux and 50°C, and more preferably 65 to 55°C.
  • Cooling during the crystallization could give substances containing different forms of crystals (polymorphism) in the case of rapid cooling. It is therefore desirable to perform the cooling while controlling the cooling rate as appropriate based on the consideration of its effect on the quality, grain size and the like of the crystal. Preferred is, for example, cooling at a cooling rate of 40 to 5°C/hour. More preferred is cooling at a cooling rate of, for example, 25 to 5°C/hour.
  • the final crystallization temperature may be selected suitably for the yield, quality and the like of the crystal, but is preferably 30 to -25°C.
  • the target crystal can be obtained by isolating the formed crystal through a conventional filtration procedure, washing the filtered-off crystal with a solvent if necessary, and further drying it.
  • the solvent to be used for washing the crystal the same solvent as in the crystallization can be used.
  • each of these solvents may be used alone, or two or more may be mixed and used.
  • it is, for example, acetone, 2-butanone, ethyl acetate, t-butyl methyl ether, hexane or a mixed solvent of hexane/2-butanone.
  • the crystal isolated through the filtration procedure may be dried appropriately by leaving it in air or under nitrogen flow, or by heating.
  • the time until the amount of residual solvent becomes less than the predefined amount may be selected as appropriate depending on the amount of production, the drying apparatus, the drying temperature and the like. Furthermore, drying may be performed under airflow or under reduced pressure. The degree of pressure reduction may be selected as appropriate depending on the amount of production, the drying apparatus, the drying temperature and the like. The obtained crystal may be left in air as required after drying.
  • a pharmaceutical composition of the present invention could be prepared by mixing pharmaceutically acceptable additives with the salt of compound (I) or the crystal thereof.
  • a pharmaceutical composition of the present invention could be prepared according to the known method such as a method described in the general rules for preparations of the Japanese Pharmacopoeia 17th edition.
  • a pharmaceutical composition of the present invention could be administered to patients appropriately depending on the dosage form.
  • a pharmaceutical composition of the present invention has usability as a therapeutic agent for treating cancers since the salt of compound (I) or the crystal thereof can potently suppress the growth of both wild-type (WT) and ER ⁇ -mutant positive tumors.
  • cancers include breast cancer, uterine endometrial, ovarian carcinoma, sarcoma, thyroid carcinoma, prostate, lung adenocarcinoma, and hepatocellular carcinoma.
  • Preferred example of cancers includes breast cancer. More preferred example of cancers includes ER-positive breast cancer.
  • the dosage of the salt of compound (I) or the crystal thereof varies depending on the extent of the symptom, age, gender, body weight, dosage form, the type of the salt, the specific type of the disease and the like.
  • about 30 ⁇ g to 10 g, preferably 100 ⁇ g to 5 g, and more preferably 100 ⁇ g to 1 g per day is orally administered, or about 30 ⁇ g to 1 g, preferably 100 ⁇ g to 500 mg, and more preferably 100 ⁇ g to 300 mg per day is administered by injection, in each case, in a single dose or in divided doses.
  • ACN Acetonitrile Boc: tert-Butyloxycarbonyl CAN: Ceric ammonium nitrate Conc.: concentrated Cs 2 CO 3 : Cesium carbonate DABCO: 1, 4-Diazabicyclo[2.2.2]octane DCM: Dichloromethane DHP: Dihydropyran DIPEA: N, N-diisopropylethylamine, Hunig’s base DMA: Dimethylacetamide DMF: Dimethylformamide DMSO: Dimethylsulfoxide DPEphos: (Oxydi-2, 1-phenylene)bis(diphenylphosphine) EDCI.HCl: N-(3-Dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride EtOH: Ethanol EtOAc: Ethyl acetate Et 3 N: Triethylamine Ex.: Example h: Hours HATU:
  • splitting patterns are as follows: s: singlet, d: doublet, t: triplet, q: quartet, m: multiplex, bs: broad singlet , br s: broad singlet, dd: doublet of doublets, dt: doublet of triplets, br d: broad doublet, br t: broad triplet Unless indicated otherwise, 1 H NMR spectra were taken on a Bruker 300 MHz or 400 MHz NMR.
  • Powder X-ray diffractometry The powder X-ray diffractometry of the crystal obtained by the method described in the following Examples was analyzed under the following measurement conditions.
  • Solid state 13 C NMR spectroscopy The solid state 13 C NMR spectrum of the crystal obtained by the method described in the following Examples was measured under the following measurement conditions
  • Hygroscopicity The crystal obtained by the method described in the following Examples was weighed into a sampling cup and then the sampling cup was placed inside an isothermal chamber at 25°C.
  • the relative humidity (RH) was controlled from 0% to 95% using a gravimetric vapor sorption system and the weight change of the sample at each stage was measured under the condition described below.
  • the resulting solution was stirred at 80 o C until completion. The reaction progress was monitored by LCMS. The resulting solution was diluted with 1 L of 2-Methyl THF and was washed with 1x1 L of brine. The mixture was dried over anhydrous sodium sulfate and concentrated under vacuum to deliver the title compound in 134 g (crude) as a black oil. The crude product was used directly to the next step.
  • the resulting solution was stirred at 80 o C until completion. The reaction progress was monitored by LCMS.
  • the resulting solution was diluted with 1 L of H 2 O and extracted with 2x1 L of ethyl acetate. The organic layers were combined, dried over anhydrous sodium sulfate, and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (0:100-15:85). The collected fractions were combined and concentrated under vacuum to deliver the title compound in 45 g (47%) as a yellow solid.
  • the resulting solution was stirred at 50 o C until completion.
  • the reaction progress was monitored by LCMS.
  • the solution was diluted with 500 mL of H 2 O, extracted with 2x600 mL of ethyl acetate, then the organic layers were combined, dried over anhydrous sodium sulfate, and concentrated under vacuum.
  • the residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1:2). The collected fractions were combined and concentrated under vacuum to deliver the title compound in 36 g (54%) as yellow oil.
  • the resulting solution was stirred at 80 o C until completion. The reaction progress was monitored by LCMS.
  • the resulting solution was diluted with 200 ml of H 2 O, extracted with 3x500 ml of ethyl acetate, then the organic layers combined, washed with brine (200 ml) and dried over anhydrous sodium sulfate. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1:3). The collected fractions were combined and concentrated under vacuum to deliver the title compound in 16 g (42%) as a yellow solid.
  • Example 8 Preparation of crystal form A of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide maleate To (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.08 mg) was added a solution of maleic acid (84 mg, 1eq) in acetonitrile (3 mL).
  • Cell culture MCF7 BUS cells (Coser, et al., (2003) PNAS 100 (24): 13994-13999) were maintained in Dulbecco’s Modified Eagle Medium supplemented with 10% FBS, 4 mM L-glutamine and 1x non-essential amino acids.
  • Lenti-X 293T cells (Clontech, Cat # 632180) were routinely cultured in Dulbecco’s Modified Eagle Medium supplemented with 10% FBS.
  • NM 000125 was used as a template with the following mutagenesis primers (where the underlined nucleotides represent site mutations); Y537S: F-AAG AAC GTG GTG CCC CTC T C T GAC CTG CTG CTG GAG ATG (SEQ ID NO: 1), R-CAT CTC CAG CAG GTC A G A GAG GGG CAC CAC GTT CTT (SEQ ID NO: 2); Y537N: F-AAG AAC GTG GTG CCC CTC A AT GAC CTG CTG CTG GAG ATG (SEQ ID NO: 3), R-CAT CTC CAG CAG GTC AT T GAG GGG CAC CAC GTT CTT (SEQ ID NO: 4); Y537C: F-AAG AAC GTG GTG CCC CTC T G T GAC CTG CTG CTG GAG ATG (SEQ ID NO: 5), R-CAT CTC CAG CAG CAG GTC A C A GAG GGG
  • WT and mutant ESR1 cDNAs were cloned into the designation lentiviral vector pLenti6.3/V5-Dest (Invitrogen, Cat #V533-06).
  • DNAs WT and mutant ESR1 were co-transfected with packaging plasmids into Lenti-X 293T cells using TransIT (Mirus, Cat #MIR 2700). 48h post-transfection, virus containing media was filtered and added to MCF7 cells in the presence of 8 ⁇ g/ml polybrene overnight. Two days following infection, cells were placed under selection with 10 ⁇ g/ml blasticidin for 2 weeks for stable expression.
  • MCF7-WT and -Y537S cells were seeded at 1500 cells/well in black-walled 96-well plates (assay plates, Costar, Cat #3904). In parallel, cells were also seeded in a separate 96-well plate (8 wells/cell line, control plate) for which a CTG (CellTiter-Glo (registered trademark) Luminescent Viability Assay, Promega, Cat #G7572) was measured the following day (day 0 reading). The day 0 reading was used for the GI 50 calculation at the termination of the experiment. The day following seeding, compounds were added to assay plates.
  • CTG CellTiter-Glo (registered trademark) Luminescent Viability Assay, Promega, Cat #G7572
  • a 1:4 serial dilution was prepared in DMSO at 200x final concentration for a total of 10 concentrations (9 dilutions containing compound and one is DMSO only).
  • Serially diluted compounds were pipetted into medium to prepare a compound-medium mix at 10x final concentration.
  • 10 ⁇ l of compound-medium mix was added to MCF7-WT and -Y537S cells at 3 wells/concentration (triplicate for each concentration).
  • media/compound was removed and replaced with fresh media/compound as described above.
  • CTG was measured and compared to day 0 readings from control plate to assess GI 50 .
  • Results Figure 34 shows that ectopic expression of ER ⁇ Y537S/N/C, D538G in MCF7 cells conferred phenotypic resistance to currently marketed therapies tamoxifen (SERM), raloxifene (SERM) and fulvestrant (SERD). Similar observations were also recently published by several independent labs (Jeselsohn et al., (2014) Clin Cancer Res. Apr 1; 20 (7): 1757-67; Toy et al., (2013) Nat. Genet. 2013 Dec; 45(12):1439-45; Robinson et al., (2013) Nat. Genet. Dec; 45 (12): 1446-51; Merenbakh-Lamin et al., (2013) Cancer Res.
  • SERM tamoxifen
  • SERM raloxifene
  • Results as presented in Table 1 are presented as the average of one or more trials with standard deviations where available. The number of trials for each compound is presented in parentheses following the value.
  • GI50 values may vary depending on the lot of fetal bovine serum (FBS), among other factors, used to supplement the culture media, due to varying concentrations of estrogen between batches.
  • FBS fetal bovine serum
  • results for compound (I) are presented as averages of both free base and HCl salt trials.
  • PDX-Y537S Y537S positive PDX xenograft study
  • PDX-Y537S a Patient-Derived Xenograft (PDX) tumor model representing an ESR1-Y537S mutated human ER+ breast cancer, designated as PDX-Y537S, was propagated subcutaneously in immunocompromised mice. The tumors were excised within 60 days of implantation and processed to mixed tumor fragments. Solid tumor tissues were depleted of necrotic components, cut into 70 mg fragments, mixed with matrigel and subcutaneously implanted into the right flank of 6-12 week old female athymic Nude (Crl:NU(NCr)-Foxn1nu) mice. The precise number of fragments and volume of matrigel were determined on a case by case basis.
  • TGI Tumor Growth Inhibition %
  • FIG. 35 shows the anti-tumor and body weight effects of compound (I), prepared as an HCl salt, in the PDX-Y537S model bearing a heterozygous ER ⁇ Y537/WT xenograft.
  • the ESR1 wild-type human ER+ breast cancer cell line MCF7 was cultured in DMEM media supplemented with 10% FBS at 37°C in a 5% CO 2 atmosphere and kept in the exponential growth phase.
  • the cells were collected in trypsin and re-suspended in a 1:1 mixture of matrigel and HBSS at a final concentration of 5 x10 7 cells/ mL.
  • a 0.2mL aliquot of cells was injected subcutaneously into the 3 rd mammary fat pad of 6-8 week old female Balb/c nude mice, giving 1x 10 7 cells/ mouse. When the average tumor volume reached approximately about 200 mm 3 , animals were randomized prior to treatment.
  • Estrogen was supplemented for the duration of the study.
  • Compound (I) was dosed orally every day at doses ranging from 1 to 10 mg/kg. Each treatment was started on Day 0 and the administration schedule was continued for 28 days.
  • the administration volume was calculated from the individual mouse body weights prior to dose administration.
  • the body weights (BW) were measured daily while the tumor volumes were measured twice a week.
  • Tumor volumes (TV) were calculated based on the above formula.
  • FIG. 36 shows the anti-tumor and body weight effects of compound (I), prepared as an HCl salt, in the MCF7 tumor model bearing ER ⁇ WT/WT xenograft.
  • Compound (I) was given orally once daily for the duration of the study.
  • WHIM20 Xenograft Studies The Patient-Derived Xenograft (PDX) tumor model, WHIM20, representing an ESR1-Y537S mutated human ER+ breast cancer is propagated in mice.
  • the tumors are excised and processed to mixed tumor fragments and the fragments are re-implanted subcutaneously into new recipient mice.
  • Solid tumor tissues are depleted of necrotic components, cut into fragments, mixed with matrigel and subcutaneously implanted into the right flank of 6-8 week old female SCID-bg mice. The precise number of fragments and volume of matrigel are determined on a case by case basis. When the average tumor volume reaches approximately 200 mm 3 , animals are randomized prior to treatment.
  • PDX-WT PDX tumor model representing an ESR1-WT human ER+ breast cancer, designated as PDX-WT
  • the tumors were excised within 60 days of implantation and processed to mixed tumor fragments.
  • Solid tumor tissues were depleted of necrotic components, cut into 70 mg fragments, mixed with matrigel and subcutaneously implanted into the right flank of 6-12 week old female athymic Nude (Crl:NU(NCr)-Foxn1nu) mice. The precise number of fragments and volume of matrigel were determined on a case by case basis.
  • FIG. 37 shows the anti-tumor and body weight effects of compound (I), prepared as an HCl salt, in a PDX model bearing ER ⁇ WT/WT xenograft.
  • the IC 50 shift approach was used to determine whether compounds are time-dependent inhibitors of human CYP3A4 using liver microsomes (0.1 mg/mL).
  • a 30-minute pre-incubation time point was selected, where compounds (9 concentrations, 0 to 30 ⁇ mol/L) are incubated at 37°C in presence and absence of 1 mmol/L NADPH.
  • 5 ⁇ mol/L midazolam (the probe substrate) was added and formation of hydroxymidazolam was measured by high-performance liquid chromatography-mass spectrometry (LC-MS/MS) analysis following a 5 minute incubation period.
  • a TDI assay was also conducted for the following compound, which is reported as Compound 69 of PCT International Application Publication No. WO/2017/196346:
  • the IC 50 shift approach was also used to determine whether Compound 69 of PCT International Application Publication No. WO/2017/196346 is a time-dependent inhibitor of human CYP3A4 using liver microsomes. Similar to the experimental design described above, a 30-minute pre-incubation time point was selected. Minor variations to the method described above were that Compound 69 of PCT International Application Publication No. WO/2017/196346 was tested at 8 concentrations (0 to 10 ⁇ mol/L).
  • IC 50 shift assay was also performed with 15.6 ⁇ mol/L testosterone (0.05 mg/mL liver microsomes, 10 minute incubation) as second probe substrate. Assays for both probe substrates were performed in triplicate, and mifepristone was used as positive control.
  • the CYP3A4 TDI shift result for Compound 69 of PCT International Application Publication No. WO/2017/196346 was greater than 3, indicating it to be a TDI risk.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • General Chemical & Material Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

The present invention provides salts of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I and acids, and crystals thereof, possessing a potential to be used as drug substance in pharmaceuticals. [Chem. 1]

Description

SALTS OF INDAZOLE DERIVATIVE AND CRYSTALS THEREOF
The present invention relates to salts of an indazole derivative and crystals thereof.
Breast cancer is the most commonly diagnosed malignancy among women today with nearly 200,000/1.7 million new cases diagnosed in the US/worldwide each year respectively. Since about 70% of breast tumors are positive for the estrogen receptor alpha (ERα)- a key oncogenic driver in this subset of tumors- several classes of therapies have been developed to antagonize ERα function, including 1) selective estrogen receptor downregulators (SERDs) of which fulvestrant is an example, 2) selective estrogen receptor modulators (SERMs) of which tamoxifen is an example and 3) aromatase inhibitors that reduce systemic levels of estrogen. These therapies have been largely effective in the clinic reducing occurrence and progression of ERα+ breast tumors. However, there are on-target liabilities associated with these different classes of compounds. For example, tamoxifen has been shown to activate signaling activity in the endometrium leading to an increase in risk of endometrial cancers in the clinic (Fisher et al., (1994) J. Natl. Cancer Inst. Apr 6; 86 (7): 527-37; van Leeuwen et al., (1994) Lancet Feb 19; 343 (8895): 448-52). In contrast, since fulvestrant is a pure antagonist, it can lead to loss of bone density in post-menopausal women as ERα activity is critical for bone building. In addition to on-target side effects, clinical resistance is also beginning to emerge to these classes of ERα antagonists highlighting the need to develop next-generation compounds.
Several mechanisms of resistance have been identified using in vitro and in vivo models of resistance to various endocrine therapies. These include increased ERα /HER2 “crosstalk” (Shou et al., (2004) J. Natl. Cancer Inst. Jun 16; 96 (12): 926-35), aberrant expression of ERα coactivators/corepressors (Osborne et al., (2003) J. Natl. Cancer Inst. Mar 5; 95(5): 353-61) or loss of ERα altogether to allow ER-independent growth (Osborne CK, Schiff R (2011) Annu .Rev. Med. 62: 233-47).
In the hopes of identifying clinically relevant mechanisms of resistance, great effort has also recently gone into deeply characterizing the genetics of endocrine-therapy resistant metastases isolated from patients. Several independent labs have recently published the multitude of genetic lesions observed in the resistant vs the primary tumors (Li et al., (2013) Cell Rep. Sep 26; 4(6): 1116-30; Robinson et al., (2013) Nat. Genet. Dec; 45 (12): 1446-51; Toy et al., (2013) Nat. Genet. 2013 Dec; 45(12):1439-45). Among these are the highly recurrent mutations in the ligand-binding domain of ESR1 (gene which encodes ERα protein) found to be significantly enriched in about 20% of resistant tumors relative to endocrine therapy naive tumors (Jeselsohn et al., (2014) Clin. Cancer Res. Apr 1; 20 (7): 1757-67; Toy et al., (2013) Nat. Genet. 2013 Dec; 45 (12): 1439-45; Robinson et al., (2013) Nat. Genet. Dec; 45 (12): 1446-51; Merenbakh-Lamin et al., (2013) Cancer Res. Dec 1; 73 (23): 6856-64; Yu et al., (2014) Science Jul 11; 345 (6193): 216-20; Segal and Dowsett (2014), Clin. Cancer Res. Apr 1; 20 (7): 1724-6), suggesting the potential for these mutations to functionally drive clinical resistance. In contrast to the enrichment in ESR1 mutations observed in therapy-resistant tumors, mutations in other cancer-related genes failed to show such a robust enrichment strongly implying the importance of ERα mutations in promoting resistance (Jeselsohn et al., (2014) Clin. Cancer Res. Apr 1; 20 (7): 1757-67).
ER+ breast cancer patients on average are treated with seven independent therapies including chemotherapies and various anti-estrogen therapies such as tamoxifen, fulvestrant and aromatase inhibitors. Recent genomic profiling has revealed that the ERα pathway remains a critical driver of tumor growth in the resistant setting as activating mutations in ERα have emerged. Thus, it is critical that more potent ER-directed therapies be developed that can overcome resistance in the clinical setting. Hence, there is a need for novel compounds that can potently suppress the growth of both wild-type (WT) and ER α-mutant positive tumors.
Most inhibitory drug interactions with cytochrome (CYP) P450 enzymes are reversible, but in some cases the inhibitory effect increases over time and is not promptly reversible. This effect is due to irreversible covalent binding or quasi-irreversible noncovalent tight binding of a chemically reactive intermediate to the enzyme that catalyzes its formation. This class of inhibitory drug interactions is called Time-Dependent Inhibition (“TDI”). When TDI is the mode of inhibition, the inhibitory interaction will generally be greater over time following multiple dosing and be longer lasting after discontinuation of the inhibitor than in a situation when the inhibitory interaction is reversible. Therefore, TDI should be studied in standard in vitro screening protocols by pre-incubating the drug (a potential inhibitor) before the addition of a substrate (Food and Drug Administration (FDA) guidance; Cf. fda.gov/downloads/drugs/guidances/ucm292362.pdf (FDA guidance, In Vitro Metabolism- and Transporter- Mediated Drug-Drug Interaction Studies, Draft Guidance, October 24, 2017.)). Whether an investigational drug inhibits CYP enzymes is usually investigated in vitro using human liver tissues such as human liver microsomes to determine the inhibition mechanisms (e.g., reversible or TDI) and inhibition potency. Id.
Citing to Grimm et al., (“The conduct of in vitro studies to address time-dependent inhibition of drug-metabolizing enzymes: a perspective of the Pharmaceutical Research and Manufacturers of America,” Drug Metab Dispos. 37:1355-1370, 2009), the FDA recently described how pharmaceutical companies should evaluate investigational drugs for TDI potential. In particular, the FDA indicated that pharmaceutical companies “should routinely study TDI in standard in vitro screening protocols by pre-incubating the investigational drug (e.g., for at least 30 min) before adding any substrate. Any significant time-dependent and co-factor-dependent (e.g., NADPH for CYPs) loss of initial product formation may indicate TDI. In these circumstances, the sponsor should conduct definitive in vitro studies to obtain TDI parameters (i.e., kinact and KI).” See FDA guidance, In Vitro Metabolism- and Transporter-Mediated Drug-Drug Interaction Studies Guidance for Industry, Draft Guidance, October 24, 2017, pg. 24, lines 854-858.
Patients frequently use more than one medication at a time. Unanticipated, unrecognized, or mismanaged drug-drug interactions (DDIs) are an important cause of morbidity and mortality associated with prescription drug use and have occasionally caused the withdrawal of approved drugs from the market. Determination of an investigational drug’s potential to inhibit CYPs in both a reversible manner (i.e., reversible inhibition) and time-dependent manner (i.e., TDI) will allow for better characterization of potentially clinically relevant DDI. Hence, there is a need to identify and develop investigational drugs that further mitigate or remove the TDI potential.
A compound represented by the formula I, namely, (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (hereafter referred to as compound (I)), suppresses the growth of both wild-type (WT) and ER α-mutant positive tumors.
Figure JPOXMLDOC01-appb-C000046
 Generally, the physical properties of a compound, salts thereof, and their crystals used as a pharmaceutical product largely influence on the bioavailability of a drug, the purity of an active pharmaceutical ingredient, prescription of a preparation and the like. An object of the present invention is therefore to provide salts of compound (I) or crystals thereof with a potential to be used as drug substance in pharmaceuticals.
The present inventor has found salts of compound (I) or crystals thereof with a potential to be used as drug substance in pharmaceuticals, thereby completing the invention.
Specifically, the present invention provides the following <1> to <69>.
<1> A salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, and an acid selected from the group consisting of hydrobromic acid, phosphoric acid, sulfuric acid, benzenesulfonic acid, p-toluenesulfonic acid, succinic acid, maleic acid, mandelic acid and methanesulfonic acid.
Figure JPOXMLDOC01-appb-C000047
 <2> A crystal of hydrochloride salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 12.2°, 16.8°, 18.2°, 19.7° and 23.3° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000048
 <3> The crystal according to <2> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 19.7° in a powder X-ray diffractometry.
<4> The crystal according to <2> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 16.8°, 18.2° and 19.7° in a powder X-ray diffractometry.
<5> A crystal of hydrochloride salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having peaks at chemical shifts (± 0.5 ppm) of 144.5 ppm, 156.8 ppm and 165.8 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000049
 <6> A crystal of hydrochloride salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 1.
Figure JPOXMLDOC01-appb-C000050
 <7> A crystal of hydrochloride salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 12.
Figure JPOXMLDOC01-appb-C000051
 <8> A crystal of hydrobromide salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 18.4°, 19.7°, 21.6°, 22.9° and 25.6° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000052
 <9> The crystal according to <8> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.4° in a powder X-ray diffractometry.
<10> The crystal according to <8> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 18.4°, 19.7° and 25.6° in a powder X-ray diffractometry.
<11> A crystal of hydrobromide salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 144.2 ppm, 156.7 ppm and 165.2 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000053
 <12> A crystal of hydrobromide salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 2.
Figure JPOXMLDOC01-appb-C000054
 <13> A crystal of hydrobromide salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 13.
Figure JPOXMLDOC01-appb-C000055
 <14> A crystal of phosphate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 6.5°, 13.5°, 16.7°, 19.9° and 20.4° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000056
 <15> The crystal according to <14> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 6.5° in a powder X-ray diffractometry.
<16> The crystal according to <14> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 6.5°, 16.7° and 19.9° in a powder X-ray diffractometry.
<17> A crystal of phosphate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 135.3 ppm, 140.9 ppm and 169.8 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000057
 <18> A crystal of phosphate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 3.
Figure JPOXMLDOC01-appb-C000058
 <19> A crystal of phosphate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 14.
Figure JPOXMLDOC01-appb-C000059
 <20> A crystal of sulfate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 7.6°, 11.3°, 13.0°, 19.0° and 19.8° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000060
 <21> The crystal according to <20> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 19.0° in a powder X-ray diffractometry.
<22> The crystal according to <20> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 7.6°, 19.0° and 19.8° in a powder X-ray diffractometry.
<23> A crystal of sulfate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 117.8 ppm, 157.4 ppm and 165.0 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000061
 <24> A crystal of sulfate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 4.
Figure JPOXMLDOC01-appb-C000062
 <25> A crystal of sulfate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 15.
Figure JPOXMLDOC01-appb-C000063
 <26> A crystal of benzenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 10.9°, 17.6°, 19.9°, 21.9° and 24.1° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000064
 <27> The crystal according to <26> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 10.9° in a powder X-ray diffractometry.
<28> The crystal according to <26> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 10.9°, 17.6° and 24.1° in a powder X-ray diffractometry.
<29> A crystal of benzenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 136.1 ppm, 156.7 ppm and 164.4 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000065
 <30> A crystal of benzenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 5.
Figure JPOXMLDOC01-appb-C000066
 <31> A crystal of benzenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 16.
Figure JPOXMLDOC01-appb-C000067
 <32> A crystal of p-toluenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 8.9°, 12.5°, 21.4°, 22.1° and 23.4° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000068
 <33> The crystal according to <32> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 8.9° in a powder X-ray diffractometry.
<34> The crystal according to <32> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 8.9°, 21.4° and 22.1° in a powder X-ray diffractometry.
<35> A crystal of p-toluenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 140.7 ppm, 156.4 ppm and 166.9 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000069
 <36> A crystal of p-toluenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 6.
Figure JPOXMLDOC01-appb-C000070
 <37> A crystal of p-toluenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 17.
Figure JPOXMLDOC01-appb-C000071
 <38> A crystal of succinate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 14.8°, 17.0°, 17.5°, 18.2° and 19.1° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000072
 <39> The crystal according to <38> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.2° in a powder X-ray diffractometry.
<40> The crystal according to <38> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 17.0°, 18.2° and 19.1° in a powder X-ray diffractometry.
<41> A crystal of succinate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 156.1 ppm, 175.0 ppm and 181.3 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000073
 <42> A crystal of succinate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 7.
Figure JPOXMLDOC01-appb-C000074
 <43> A crystal of succinate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 18.
Figure JPOXMLDOC01-appb-C000075
 <44> A crystal form A of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 5.4°, 11.6°, 16.8°, 18.9° and 20.2° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000076
 <45> The crystal according to <44> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.9° in a powder X-ray diffractometry.
<46> The crystal according to <44> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 16.8°, 18.9° and 20.2° in a powder X-ray diffractometry.
<47> A crystal form A of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 43.4 ppm, 44.8 ppm and 145.0 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000077
 <48> A crystal form A of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 8.
Figure JPOXMLDOC01-appb-C000078
 <49> A crystal form A of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 19.
Figure JPOXMLDOC01-appb-C000079
 <50> A crystal form B of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 10.4°, 15.2°, 19.4°, 20.8° and 22.2° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000080
 <51> The crystal according to <50> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 19.4° in a powder X-ray diffractometry.
<52> The crystal according to <50> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 15.2°, 19.4° and 20.8° in a powder X-ray diffractometry.
<53> A crystal form B of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 25.8 ppm, 143.6 ppm and 166.8 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000081
 <54> A crystal form B of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 9.
Figure JPOXMLDOC01-appb-C000082
 <55> A crystal form B of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 20.
Figure JPOXMLDOC01-appb-C000083
 <56> A crystal of L-mandelate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 5.1°, 8.7°, 10.3°, 18.2° and 20.3° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000084
 <57> The crystal according to <56> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.2° in a powder X-ray diffractometry.
<58> The crystal according to <56> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 5.1°, 10.3° and 18.2° in a powder X-ray diffractometry.
<59> A crystal of L-mandelate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 128.1 ppm, 156.7 ppm and 177.8 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000085
 <60> A crystal of L-mandelate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 10.
Figure JPOXMLDOC01-appb-C000086
 <61> A crystal of L-mandelate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 21.
Figure JPOXMLDOC01-appb-C000087
 <62> A crystal of methanesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 12.7°, 18.9°, 19.7°, 21.8° and 25.6° in a powder X-ray diffractometry.
Figure JPOXMLDOC01-appb-C000088
 <63> The crystal according to <62> above, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 12.7° in a powder X-ray diffractometry.
<64> The crystal according to <62> above, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 12.7°, 18.9° and 25.6° in a powder X-ray diffractometry.
<65> A crystal of methanesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 40.4 ppm, 158.3 ppm and 166.0 ppm in a solid state 13C NMR spectrum.
Figure JPOXMLDOC01-appb-C000089
 <66> A crystal of methanesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 11.
Figure JPOXMLDOC01-appb-C000090
 <67> A crystal of methanesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 22.
Figure JPOXMLDOC01-appb-C000091
 <68> A pharmaceutical composition comprising the salt or the crystal thereof according to any one of <1> to <67> above.
<69> The pharmaceutical composition according to <68> above, for use in treating cancer.
The salts of compound (I) and the crystals thereof provided by the present invention possess properties such as hygroscopicity as shown in the examples described in later and a potential to be used as drug substance in pharmaceuticals.
Figure 1 shows a powder X-ray diffraction pattern of the crystal of the compound (I) hydrochloride salt obtained in Example 1. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 2 shows a powder X-ray diffraction pattern of the crystal of the compound (I) hydrobromide salt obtained in Example 2. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 3 shows a powder X-ray diffraction pattern of the crystal of the compound (I) phosphate salt obtained in Example 3. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 4 shows a powder X-ray diffraction pattern of the crystal of the compound (I) sulfate salt obtained in Example 4. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 5 shows a powder X-ray diffraction pattern of the crystal of the compound (I) benzenesulfonate salt obtained in Example 5. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 6 shows a powder X-ray diffraction pattern of the crystal of the compound (I) p-toluenesulfonate salt obtained in Example 6. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 7 shows a powder X-ray diffraction pattern of the crystal of the compound (I) succinate salt obtained in Example 7. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 8 shows a powder X-ray diffraction pattern of the crystal form A of the compound (I) maleate salt obtained in Example 8. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 9 shows a powder X-ray diffraction pattern of the crystal form B of the compound (I) maleate salt obtained in Example 9. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 10 shows a powder X-ray diffraction pattern of the crystal of the compound (I) L-mandelate salt obtained in Example 10. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 11 shows a powder X-ray diffraction pattern of the crystal of the compound (I) methanesulfonate salt obtained in Example 11. The abscissa shows the diffraction angle (2θ) and the ordinate shows the peak intensity. Figure 12 shows a solid state 13C NMR spectrum of the crystal of the compound (I) hydrochloride salt obtained in Example 1. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 13 shows a solid state 13C NMR spectrum of the crystal of the compound (I) hydrobromide salt obtained in Example 2. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 14 shows a solid state 13C NMR spectrum of the crystal of the compound (I) phosphate salt obtained in Example 3. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 15 shows a solid state 13C NMR spectrum of the crystal of the compound (I) sulfate salt obtained in Example 4. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 16 shows a solid state 13C NMR spectrum of the crystal of the compound (I) benzenesulfonate salt obtained in Example 5. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 17 shows a solid state 13C NMR spectrum of the crystal of the compound (I) p-toluenesulfonate salt obtained in Example 6. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 18 shows a solid state 13C NMR spectrum of the crystal of the compound (I) succinate salt obtained in Example 7. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 19 shows a solid state 13C NMR spectrum of the crystal form A of the compound (I) maleate salt obtained in Example 8. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 20 shows a solid state 13C NMR spectrum of the crystal form B of the compound (I) maleate salt obtained in Example 9. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 21 shows a solid state 13C NMR spectrum of the crystal of the compound (I) L-mandelate salt obtained in Example 10. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 22 shows a solid state 13C NMR spectrum of the crystal of the compound (I) methanesulfonate salt obtained in Example 11. The abscissa shows the chemical shift (ppm) and the ordinate shows the peak intensity. Figure 23 is a graph showing hygroscopicity of the crystal of the compound (I) hydrochloride salt obtained in Example 1. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 24 is a graph showing hygroscopicity of the crystal of the compound (I) hydrobromide salt obtained in Example 2. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 25 is a graph showing hygroscopicity of the crystal of the compound (I) phosphate salt obtained in Example 3. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 26 is a graph showing hygroscopicity of the crystal of the compound (I) sulfate salt obtained in Example 4. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 27 is a graph showing hygroscopicity of the crystal of the compound (I) benzenesulfonate salt obtained in Example 5. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 28 is a graph showing hygroscopicity of the crystal of the compound (I) p-toluenesulfonate salt obtained in Example 6. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 29 is a graph showing hygroscopicity of the crystal of the compound (I) succinate salt obtained in Example 7. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 30 is a graph showing hygroscopicity of the crystal form A of the compound (I) maleate salt obtained in Example 8. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 31 is a graph showing hygroscopicity of the crystal form B of the compound (I) maleate salt obtained in Example 9. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 32 is a graph showing hygroscopicity of the crystal of the compound (I) L-mandelate salt obtained in Example 10. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 33 is a graph showing hygroscopicity of the crystal of the compound (I) methanesulfonate salt obtained in Example 11. The abscissa shows the relative humidity and the ordinate shows the weight change. Figure 34 is a graph showing in vitro proliferation effects of wild-type and mutant ER-bearing MCF7 lines to clinical therapies 4-hydroxytamoxifen (4-OHT), raloxifene and fulvestrant, where phenotypic resistance observed in mutant-bearing lines relative to control lines to existing clinical compounds, whereby MCF7 cells engineered to overexpress various ERαMUT showed partial resistance to various endocrine therapies. Figure 35 is a graph showing antitumor and body weight effects of oral compound (I) as a hydrochloride salt in PDX-Y537S xenograft bearing athymic nude female mice. Figure 36 is a graph showing the anti-tumor and body weight effects of compound (I), prepared as an HCl salt, in the MCF7 tumor model bearing ERαWT/WT xenograft. Figure 37 is a graph showing the anti-tumor and body weight effects of compound (I), prepared as an HCl salt, in a PDX model bearing ERαWT/WT xenograft.
A salt of the compound (I) of the present invention, a crystal thereof, and production methods thereof will be described in detail.
As used herein, a "salt" refers to a chemical entity made up of the compound (I) as the basic component and a specific number of equivalents of an acid to the compound (I). Here, the term " a salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, and an acid selected from the group consisting of hydrobromic acid, phosphoric acid, sulfuric acid, benzenesulfonic acid, p-toluenesulfonic acid, succinic acid, maleic acid, mandelic acid and methanesulfonic acid" is used for the same meaning as " a salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I formed with an acid selected from the group consisting of hydrobromic acid, phosphoric acid, sulfuric acid, benzenesulfonic acid, p-toluenesulfonic acid, succinic acid, maleic acid, mandelic acid and methanesulfonic acid".
Examples of a "salt" used herein include salts with organic carboxylic acids, with organic sulfonic acids and with inorganic acids, and in particular, pharmaceutically acceptable salts are preferred.
Examples of organic carboxylic acids include acetic acid, oxalic acid, maleic acid, mandelic acid, tartaric acid, fumaric acid, citric acid, malonic acid, succinic acid, benzoic acid, hippuric acid and malic acid. Preferred examples of organic carboxylic acids include maleic acid, mandelic acid (preferably L-mandelic acid), tartaric acid, malonic acid and succinic acid.
Examples of organic sulfonic acids include methanesulfonic acid, trifluoromethanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid and camphorsulfonic acid. Preferred examples of organic sulfonic acids include methanesulfonic acid, benzenesulfonic acid and p-toluenesulfonic acid.
Examples of inorganic acids include hydrofluoric acid, hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, perchloric acid, phosphoric acid, carbonic acid and bicarbonic acid. Preferred examples of inorganic acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid and phosphoric acid.
A salt of the compound (I) may also be a solvate. As used herein, a solvate of the salt of compound (I) means a solid formed from the salt of the compound (I) together with solvent molecules. Examples of the solvent in the solvate include: a ketone solvent such as acetone, methyl ethyl ketone or cyclohexanone; an ester solvent such as ethyl acetate or methyl acetate; an ether solvent such as 1, 2-dimethoxyethane or methyl-tert-butyl ether; an alcohol solvent such as methanol, ethanol, 1-propanol or isopropanol; a polar solvent such as N-methyl-2-pyrrolidone, N,N-dimethylformamide or dimethyl sulfoxide; and water.
As used herein, a "crystal" refers to a crystal of the salt of compound (I). Accordingly, a crystal of hydrochloride salt of compound (I), for example, means a crystal of the salt formed between compound (I) and hydrochloric acid.
Examples of crystals preferred herein include:
(a1) a crystal of hydrochloride salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 19.7° in a powder X-ray diffractometry;
(a2) a crystal of hydrochloride salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 16.8°, 18.2° and 19.7° in a powder X-ray diffractometry;
(a3) a crystal of hydrochloride salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 12.2°, 16.8°, 18.2°, 19.7° and 23.3° in a powder X-ray diffractometry;
(a4) a crystal of hydrochloride salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 12.2°, 16.8°, 18.2°, 19.7°, 21.3°, 21.8°, 23.3°, 24.5°, 25.5° and 29.4° in a powder X-ray diffractometry;
(b1) a crystal of hydrobromide salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.4° in a powder X-ray diffractometry;
(b2) a crystal of hydrobromide salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 18.4°, 19.7° and 25.6° in a powder X-ray diffractometry;
(b3) a crystal of hydrobromide salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 18.4°, 19.7°, 21.6°, 22.9° and 25.6° in a powder X-ray diffractometry;
(b4) a crystal of hydrobromide salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 6.1°, 12.0°, 16.6°, 18.4°, 19.7°, 21.6°, 22.9°, 24.1°, 25.6° and 29.8° in a powder X-ray diffractometry;
(c1) a crystal of phosphate salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 6.5° in a powder X-ray diffractometry;
(c2) a crystal of phosphate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 6.5°, 16.7° and 19.9° in a powder X-ray diffractometry;
(c3) a crystal of phosphate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 6.5°, 13.5°, 16.7°, 19.9° and 20.4° in a powder X-ray diffractometry;
(c4) a crystal of phosphate salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 6.5°, 13.5°, 14.0°, 16.7°, 19.9°, 20.4°, 21.6° and 23.2° in a powder X-ray diffractometry;
(d1) a crystal of sulfate salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 19.0° in a powder X-ray diffractometry;
(d2) a crystal of sulfate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 7.6°, 19.0° and 19.8° in a powder X-ray diffractometry;
(d3) a crystal of sulfate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 7.6°, 11.3°, 13.0°, 19.0° and 19.8° in a powder X-ray diffractometry;
(d4) a crystal of sulfate salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 3.7°, 7.6°, 11.3°, 13.0°, 16.9°, 19.0° and 19.8° in a powder X-ray diffractometry;
(e1) a crystal of benzenesulfonate salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 10.9° in a powder X-ray diffractometry;
(e2) a crystal of benzenesulfonate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 10.9°, 17.6° and 24.1° in a powder X-ray diffractometry;
(e3) a crystal of benzenesulfonate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 10.9°, 17.6°, 19.9°, 21.9° and 24.1° in a powder X-ray diffractometry;
(e4) a crystal of benzenesulfonate salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 5.5°, 9.8°, 10.9°, 17.6°, 19.2°, 19.9°, 21.9°, 24.1° and 26.5° in a powder X-ray diffractometry;
(f1) a crystal of p-toluenesulfonate salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 8.9° in a powder X-ray diffractometry;
(f2) a crystal of p-toluenesulfonate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 8.9°, 21.4° and 22.1° in a powder X-ray diffractometry;
(f3) a crystal of p-toluenesulfonate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 8.9°, 12.5°, 21.4°, 22.1° and 23.4° in a powder X-ray diffractometry;
(f4) a crystal of p-toluenesulfonate salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 8.9°, 11.5°, 12.5°, 16.2°, 17.5°, 20.0°, 20.9°, 21.4°, 22.1° and 23.4° in a powder X-ray diffractometry;
(g1) a crystal of succinate salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.2° in a powder X-ray diffractometry;
(g2) a crystal of succinate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 17.0°, 18.2° and 19.1° in a powder X-ray diffractometry;
(g3) a crystal of succinate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 14.8°, 17.0°, 17.5°, 18.2° and 19.1° in a powder X-ray diffractometry;
(g4) a crystal of succinate salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 5.2°, 10.4°, 12.9°, 14.8°, 17.0°, 17.5°, 18.2°, 19.1°, 21.8° and 23.6° in a powder X-ray diffractometry;
(h1) a crystal form A of maleate salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.9° in a powder X-ray diffractometry;
(h2) a crystal form A of maleate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 16.8°, 18.9° and 20.2° in a powder X-ray diffractometry;
(h3) a crystal form A of maleate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 5.4°, 11.6°, 16.8°, 18.9° and 20.2° in a powder X-ray diffractometry;
(h4) a crystal form A of maleate salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 5.4°, 11.6°, 13.3°, 14.3°, 16.8°, 18.9°, 20.2°, and 21.6° in a powder X-ray diffractometry;
(i1) a crystal form B of maleate salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 19.4° in a powder X-ray diffractometry;
(i2) a crystal form B of maleate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 15.2°, 19.4° and 20.8° in a powder X-ray diffractometry;
(i3) a crystal form B of maleate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 10.4°, 15.2°, 19.4°, 20.8° and 22.2° in a powder X-ray diffractometry;
(i4) a crystal form B of maleate salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 10.4°, 15.2°, 19.4°, 20.8°, 22.2°, 23.7°, 25.7° and 28.6° in a powder X-ray diffractometry;
(j1) a crystal of L-mandelate salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.2° in a powder X-ray diffractometry;
(j2) a crystal of L-mandelate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 5.1°, 10.3° and 18.2° in a powder X-ray diffractometry;
(j3) a crystal of L-mandelate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 5.1°, 8.7°, 10.3°, 18.2° and 20.3° in a powder X-ray diffractometry;
(j4) a crystal of L-mandelate salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 5.1°, 8.7°, 10.3°, 13.9°, 18.2° and 20.3° in a powder X-ray diffractometry;
(k1) a crystal of methanesulfonate salt of compound (I), having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 12.7° in a powder X-ray diffractometry;
(k2) a crystal of methanesulfonate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 12.7°, 18.9° and 25.6° in a powder X-ray diffractometry;
(k3) a crystal of methanesulfonate salt of compound (I), having diffraction peaks at diffraction angles (2θ ± 0.2°) of 12.7°, 18.9°, 19.7°, 21.8° and 25.6° in a powder X-ray diffractometry;
(k4) a crystal of methanesulfonate salt of compound (I), having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) of 7.5°, 12.1°, 12.7°, 17.1°, 18.9°, 19.7°, 20.9°, 21.8° and 25.6° in a powder X-ray diffractometry;
(l1) a crystal of hydrochloride salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 144.5 ppm, 156.8 ppm and 165.8 ppm in a solid state 13C NMR spectrum;
(l2) a crystal of hydrochloride salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 139.0 ppm, 141.4 ppm, 144.5 ppm, 156.8 ppm and 165.8 ppm in a solid state 13C NMR spectrum;
(l3) a crystal of hydrochloride salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 25.8 ppm, 45.9 ppm, 117.3 ppm, 128.5 ppm, 133.9 ppm, 139.0 ppm, 141.4 ppm, 144.5 ppm, 156.8 ppm and 165.8 ppm in a solid state 13C NMR spectrum;
(m1) a crystal of hydrobromide salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 144.2 ppm, 156.7 ppm and 165.2 ppm in a solid state 13C NMR spectrum;
(m2) a crystal of hydrobromide salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 139.4 ppm, 141.6 ppm, 144.2 ppm, 156.7 ppm and 165.2 ppm in a solid state 13C NMR spectrum;
(m3) a crystal of hydrobromide salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 26.4 ppm, 45.2 ppm, 47.3 ppm, 116.9 ppm, 129.3 ppm, 139.4 ppm, 141.6 ppm, 144.2 ppm, 156.7 ppm and 165.2 ppm in a solid state 13C NMR spectrum;
(n1) a crystal of phosphate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 135.3 ppm, 140.9 ppm and 169.8 ppm in a solid state 13C NMR spectrum;
(n2) a crystal of phosphate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 126.6 ppm, 135.3 ppm, 140.9 ppm, 146.6 ppm and 169.8 ppm in a solid state 13C NMR spectrum;
(n3) a crystal of phosphate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 26.9 ppm, 45.5 ppm, 49.2 ppm, 126.6 ppm, 135.3 ppm, 138.3 ppm, 140.9 ppm, 145.5 ppm, 146.6 ppm and 169.8 ppm in a solid state 13C NMR spectrum;
(o1) a crystal of sulfate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 117.8 ppm, 157.4 ppm and 165.0 ppm in a solid state 13C NMR spectrum;
(o2) a crystal of sulfate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 47.8 ppm, 49.9 ppm, 117.8 ppm, 157.4 ppm and 165.0 ppm in a solid state 13C NMR spectrum;
(o3) a crystal of sulfate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 25.0 ppm, 47.8 ppm, 49.9 ppm, 113.6 ppm, 117.8 ppm, 124.7 ppm, 141.3 ppm, 146.5 ppm, 157.4 ppm and 165.0 ppm in a solid state 13C NMR spectrum;
(p1) a crystal of benzenesulfonate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 136.1 ppm, 156.7 ppm and 164.4 ppm in a solid state 13C NMR spectrum;
(p2) a crystal of benzenesulfonate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 116.8 ppm, 132.3 ppm, 136.1 ppm, 156.7 ppm and 164.4 ppm in a solid state 13C NMR spectrum;
(p3) a crystal of benzenesulfonate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 27.6 ppm, 44.9 ppm, 47.1 ppm, 116.8 ppm, 132.3 ppm, 136.1 ppm, 156.7 ppm and 164.4 ppm in a solid state 13C NMR spectrum;
(q1) a crystal of p-toluenesulfonate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 140.7 ppm, 156.4 ppm and 166.9 ppm in a solid state 13C NMR spectrum;
(q2) a crystal of p-toluenesulfonate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 125.9 ppm, 140.7 ppm, 144.2 ppm, 156.4 ppm and 166.9 ppm in a solid state 13C NMR spectrum;
(q3) a crystal of p-toluenesulfonate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 19.2 ppm, 27.5 ppm, 46.4 ppm, 112.3 ppm, 117.0 ppm, 125.9 ppm, 140.7 ppm, 144.2 ppm, 156.4 ppm and 166.9 ppm in a solid state 13C NMR spectrum;
(r1) a crystal of succinate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 156.1 ppm, 175.0 ppm and 181.3 ppm in a solid state 13C NMR spectrum;
(r2) a crystal of succinate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 145.8 ppm, 156.1 ppm, 165.5 ppm, 175.0 ppm and 181.3 ppm in a solid state 13C NMR spectrum;
(r3) a crystal of succinate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 26.0 ppm, 35.1 ppm, 116.0 ppm, 125.3 ppm, 139.2 ppm, 145.8 ppm, 156.1 ppm, 165.5 ppm, 175.0 ppm and 181.3 ppm in a solid state 13C NMR spectrum;
(s1) a crystal form A of maleate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 43.4 ppm, 44.8 ppm and 145.0 ppm in a solid state 13C NMR spectrum;
(s2) a crystal form A of maleate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 43.4 ppm, 44.8 ppm, 63.0 ppm, 133.8 ppm and 145.0 ppm in a solid state 13C NMR spectrum;
(s3) a crystal form A of maleate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 43.4 ppm, 44.8 ppm, 47.8 ppm, 63.0 ppm, 66.0 ppm, 125.2 ppm, 133.8 ppm, 140.6 ppm, 145.0 ppm and 165.5 ppm in a solid state 13C NMR spectrum;
(t1) a crystal form B of maleate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 25.8 ppm, 143.6 ppm and 166.8 ppm in a solid state 13C NMR spectrum;
(t2) a crystal form B of maleate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 25.8 ppm, 48.8 ppm, 135.1 ppm, 143.6 ppm and 166.8 ppm in a solid state 13C NMR spectrum;
(t3) a crystal form B of maleate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 25.8 ppm, 48.8 ppm, 110.7 ppm, 115.7 ppm, 135.1ppm, 136.6 ppm, 143.6 ppm, 156.3 ppm, 166.8 ppm and 172.6 ppm in a solid state 13C NMR spectrum;
(u1) a crystal of L-mandelate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 128.1 ppm, 156.7 ppm and 177.8 ppm in a solid state 13C NMR spectrum;
(u2) a crystal of L-mandelate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 128.1 ppm, 141.9 ppm, 156.7 ppm, 164.6 ppm and 177.8 ppm in a solid state 13C NMR spectrum;
(u3) a crystal of L-mandelate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 62.9 ppm, 76.5 ppm, 116.2 ppm, 123.6 ppm, 128.1 ppm, 141.9 ppm, 146.3 ppm, 156.7 ppm, 164.6 ppm and 177.8 ppm in a solid state 13C NMR spectrum;
(v1) a crystal of methanesulfonate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 40.4 ppm, 158.3 ppm and 166.0 ppm in a solid state 13C NMR spectrum;
(v2) a crystal of methanesulfonate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 40.4 ppm, 136.0 ppm, 146.7 ppm, 158.3 ppm and 166.0 ppm in a solid state 13C NMR spectrum; and
(v3) a crystal of methanesulfonate salt of compound (I), having peaks at chemical shifts (± 0.5 ppm) of 24.8 ppm, 40.4 ppm, 47.2 ppm, 117.8 ppm, 133.9 ppm, 136.0 ppm, 140.4 ppm, 146.7 ppm, 158.3 ppm and 166.0 ppm in a solid state 13C NMR spectrum.
The peaks in a powder X-ray diffractometry, described above, are characteristic for each of the crystal of hydrochloride salt of compound (I), the crystal of hydrobromide salt of compound (I), the crystal of phosphate salt of compound (I), the crystal of sulfate salt of compound (I), the crystal of benzenesulfonate salt of compound (I), the crystal of p-toluenesulfonate salt of compound (I), the crystal of succinate salt of compound (I), the crystal form A of maleate salt of compound (I), the crystal form B of maleate salt of compound (I), the crystal of L-mandelate salt of compound (I), and the crystal of methanesulfonate salt of compound (I).
Generally, errors in diffraction angles (2θ) within the range of ± 0.2° may arise in powder X-ray diffractometry, and thus the above-described values of diffraction angles need to be considered to include values within the range of approximately ± 0.2°. Included in the present invention are, therefore, not only crystals with peaks at exactly the same diffraction angles in powder X-ray diffractometry, but also crystals with peaks within an error range of approximately ± 0.2° of the diffraction angles. Hence, "having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 19.7°" as used herein, for example, means "having a diffraction peak at a diffraction angle (2θ) of 19.5° to 19.9°". The same is also applied to other diffraction angles.
Generally, peak intensities and half-value widths of diffraction angles (2θ) in powder X-ray diffraction are different for each measurement because of differences in measurement conditions and dispersions of size and shape of each particle of powder crystal and not always stable even though forms of crystals are same. Therefore, in case of comparing a powder X-ray diffraction pattern, when diffraction angles (2θ) are the same but peak intensities, relative peak intensities and half-value widths are different, those differences does not imply that the measured forms of crystals differ from each other. Thus, a crystal of salt having a powder X-ray diffraction pattern, which has aforementioned differences with respect to characteristic diffraction peaks of a certain crystal of salt according to the present invention, means that the crystal has the same crystal form of the crystal of salt according to the present invention.
As used herein, "having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 1" means it includes not only the case of having exactly the same powder X-ray diffraction pattern as shown in Figure 1, but also the case that at least one selected from the group consisting of peak intensities, relative peak intensities and half-value widths are different, or the case of having the characteristic peaks within an error range of approximately ± 0.2° of the diffraction angles. Thus every crystal having such the powder X-ray diffraction pattern means that the crystal is identical to the crystal according to the present invention.
The peaks in solid state 13C NMR spectrum, described above, are characteristic for each of the crystal of hydrochloride salt of compound (I), the crystal of hydrobromide salt of compound (I), the crystal of phosphate salt of compound (I), the crystal of sulfate salt of compound (I), the crystal of benzenesulfonate salt of compound (I), the crystal of p-toluenesulfonate salt of compound (I), the crystal of succinate salt of compound (I), the crystal form A of maleate salt of compound (I), the crystal form B of maleate salt of compound (I), the crystal of L-mandelate salt of compound (I), and the crystal of methanesulfonate salt of compound (I).
As used herein, "having peaks at chemical shifts of 144.5 ppm, 156.8 ppm and 165.8 ppm" means "having peaks each substantially equivalent to the peaks at chemical shifts of 144.5 ppm, 156.8 ppm and 165.8 ppm, when solid state 13C NMR spectrometry is performed under a conventional measurement condition or substantially the same condition as in the present specification".
When determining whether "having peaks substantially equivalent to" or not, the above-described values of the chemical shifts need to be considered to include values within the range of approximately ± 0.5 ppm since generally errors in chemical shifts (ppm) within the range of ± 0.5 ppm may arise in a solid state 13C NMR spectrum. Included in the present invention are, therefore, not only crystals with exactly the same chemical shifts in a solid state 13C NMR spectrum, but also crystals with chemical shifts within an error range of approximately ± 0.5 ppm. Hence, "having a peak at chemical shift of 144.5 ppm" as used herein, for examples, means "having a peak at a chemical shift of 144.0 ppm to 145.0 ppm". The same is also applied to other chemical shifts in solid state 13C NMR spectra.
As used herein, " having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 12" means it includes not only the case of having exactly the same solid state 13C NMR spectrum as shown in Figure 12, but also the case that peak intensities are different, or the case of having the characteristic peaks within an error range of approximately ± 0.5 ppm. Thus every crystal having such the solid state 13C NMR spectrum means that the crystal is identical to the crystal according to the present invention.
Methods for producing a salt of the compound (I) and a crystal thereof will be described in detail.
(Production of compound (I))
Compound (I) can be synthesized as described specifically in Production Example 1 below.
(Method for producing a salt of the compound (I))
A salt of the compound (I) can be obtained by a conventional method for producing a salt. Specifically, it can be produced, for example, by suspending or dissolving compound (I) in a solvent, with heating if necessary, then by adding an acid to the obtained suspension or solution and by stirring or leaving the resultant suspension or solution for several minutes to several days at room temperature or with ice-bath cooling. A salt of the compound (I) may be obtained as crystals or amorphous substances according to the production methods. Examples of the solvents to be used in these methods include alcohol solvents such as ethanol, 1-propanol and isopropanol; acetonitrile; ketone solvents such as acetone and 2-butanone; ester solvents such as ethyl acetate; saturated hydrocarbon solvents such as hexane and heptane; ether solvents such as t-butyl methyl ether or water. Each of these solvents may be used alone, or two or more may be mixed and used.
(Method for producing a crystal of the salt of compound (I))
A crystal of the salt of compound (I) may be produced by the above-mentioned methods for producing a salt of the compound (I), or by heat-dissolving a salt of the compound (I) in a solvent and crystallizing it through cooling with stirring.
A salt of the compound (I) to be used in the crystallization may be in any form: it may be a solvate, a hydrate, an anhydrate, an amorphous substance, a crystalline substance (including those consisting of a plurality of crystalline polymorphs) or a combination thereof.
Examples of the solvents to be used in the crystallization include alcohol solvents such as methanol, ethanol, isopropanol and 1-propanol; acetonitrile; amide solvents such as N,N-dimethylformamide; ester solvents such as ethyl acetate; saturated hydrocarbon solvents such as hexane and heptane; ketone solvents such as acetone and 2-butanone; ether solvents such as t-butyl methyl ether or water. Furthermore, each of these solvents may be used alone, or two or more may be mixed and used.
The amount of the solvent to be used may be suitably selected, provided that the lower limit is the amount with which the free form of compound (I) or the salt thereof is dissolved by heating or the suspension can be stirred, and that the upper limit is the amount with which the yield of the crystal is not significantly reduced.
A seed crystal (e.g., the crystal of the desired salt of compound (I)) may be added or may not be added during the crystallization. The temperature at which the seed crystal is added is not particularly limited, but is preferably 0 to 80°C.
As the temperature to be employed when the salt of compound (I) is dissolved by heating, that at which compound (I) dissolves may be suitably selected depending on the solvent, but it is preferably within the range between the temperature at which the recrystallization solvent starts to reflux and 50°C, and more preferably 65 to 55°C.
Cooling during the crystallization could give substances containing different forms of crystals (polymorphism) in the case of rapid cooling. It is therefore desirable to perform the cooling while controlling the cooling rate as appropriate based on the consideration of its effect on the quality, grain size and the like of the crystal. Preferred is, for example, cooling at a cooling rate of 40 to 5°C/hour. More preferred is cooling at a cooling rate of, for example, 25 to 5°C/hour.
Furthermore, the final crystallization temperature may be selected suitably for the yield, quality and the like of the crystal, but is preferably 30 to -25°C.
The target crystal can be obtained by isolating the formed crystal through a conventional filtration procedure, washing the filtered-off crystal with a solvent if necessary, and further drying it. As the solvent to be used for washing the crystal, the same solvent as in the crystallization can be used. Furthermore, each of these solvents may be used alone, or two or more may be mixed and used. Preferably, it is, for example, acetone, 2-butanone, ethyl acetate, t-butyl methyl ether, hexane or a mixed solvent of hexane/2-butanone.
The crystal isolated through the filtration procedure may be dried appropriately by leaving it in air or under nitrogen flow, or by heating.
As the drying time, the time until the amount of residual solvent becomes less than the predefined amount may be selected as appropriate depending on the amount of production, the drying apparatus, the drying temperature and the like. Furthermore, drying may be performed under airflow or under reduced pressure. The degree of pressure reduction may be selected as appropriate depending on the amount of production, the drying apparatus, the drying temperature and the like. The obtained crystal may be left in air as required after drying.
A pharmaceutical composition of the present invention could be prepared by mixing pharmaceutically acceptable additives with the salt of compound (I) or the crystal thereof. A pharmaceutical composition of the present invention could be prepared according to the known method such as a method described in the general rules for preparations of the Japanese Pharmacopoeia 17th edition.
A pharmaceutical composition of the present invention could be administered to patients appropriately depending on the dosage form.
A pharmaceutical composition of the present invention has usability as a therapeutic agent for treating cancers since the salt of compound (I) or the crystal thereof can potently suppress the growth of both wild-type (WT) and ER α-mutant positive tumors. Examples of cancers include breast cancer, uterine endometrial, ovarian carcinoma, sarcoma, thyroid carcinoma, prostate, lung adenocarcinoma, and hepatocellular carcinoma. Preferred example of cancers includes breast cancer. More preferred example of cancers includes ER-positive breast cancer.
The dosage of the salt of compound (I) or the crystal thereof varies depending on the extent of the symptom, age, gender, body weight, dosage form, the type of the salt, the specific type of the disease and the like. In the case of adults, typically, about 30 μg to 10 g, preferably 100 μg to 5 g, and more preferably 100 μg to 1 g per day is orally administered, or about 30 μg to 1 g, preferably 100 μg to 500 mg, and more preferably 100 μg to 300 mg per day is administered by injection, in each case, in a single dose or in divided doses.
Example
Hereinafter, the present invention will be described in detail with the production examples and examples. However, the present invention is not intended to be limited by these examples.
The following abbreviations may be used herein:
ACN: Acetonitrile
Boc: tert-Butyloxycarbonyl
CAN: Ceric ammonium nitrate
Conc.: concentrated
Cs2CO3: Cesium carbonate
DABCO: 1, 4-Diazabicyclo[2.2.2]octane
DCM: Dichloromethane
DHP: Dihydropyran
DIPEA: N, N-diisopropylethylamine, Hunig’s base
DMA: Dimethylacetamide
DMF: Dimethylformamide
DMSO: Dimethylsulfoxide
DPEphos: (Oxydi-2, 1-phenylene)bis(diphenylphosphine)
EDCI.HCl: N-(3-Dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride
EtOH: Ethanol
EtOAc: Ethyl acetate
Et3N: Triethylamine
Ex.: Example
h: Hours
HATU: 1-[Bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxid hexafluorophosphate
HCl: Hydrochloric acid
HMPA: Hexamethylphosphoramide
HPLC: High-performance liquid chromatography
H2SO4: Sulfuric acid
IPA: Isopropyl alcohol
K2CO3: Potassium carbonate
KOH: Potassium hydroxide
LCMS: Liquid chromatography - mass spectrometry
MeOH: Methanol
Na2CO3: Sodium carbonate
NBS: n-Bromosuccinimide
nBuLi: n-Butyllithium
NH4Cl: Ammonium chloride
NH4OH: Ammonium hydroxide
NMR: nuclear magnetic resonance
on or o.n.: overnight
Pd/C: Palladium (0) on carbon
Pd2(dba)3: Tris(dibenzylideneacetone)dipalladium(0)
PPTS: Pyridinium p-toluenesulfonate
PTSA: p-Toluenesulfonic acid
RT or r.t.: room temperature
TBAF: Tetrabutylammonium fluoride
TEA: Triethylamine
TFA: Trifluoroacetic acid
THF: Tetrahydrofuran
TLC: Thin-layer chromatography
Pt/C: Platinum (0) on carbon
1H NMR: proton nuclear magnetic resonance
The coupling constant is recorded in hertz (Hz). The abbreviations of splitting patterns are as follows:
s: singlet, d: doublet, t: triplet, q: quartet, m: multiplex, bs: broad singlet , br s: broad singlet, dd: doublet of doublets, dt: doublet of triplets, br d: broad doublet, br t: broad triplet
Unless indicated otherwise, 1H NMR spectra were taken on a Bruker 300 MHz or 400 MHz NMR.
Powder X-ray diffractometry
The powder X-ray diffractometry of the crystal obtained by the method described in the following Examples was analyzed under the following measurement conditions.
Measurement conditions
Apparatus: RINT TTR-III (Rigaku)
Sample pan: aluminum
X-ray: Cu K alpha
Detection: scintillation counter
Tube voltage: 50 kV
Tube current: 300 mA
Slit: divergence slit 0.5 mm (Height limiting slit 2 mm), scattering slit open, receiving slit open
Scan speed: 5°/minute
Step size: 2θ = 0.02°
Scan range: 2θ = 3° to 35°
Solid state 13C NMR spectroscopy
The solid state 13C NMR spectrum of the crystal obtained by the method described in the following Examples was measured under the following measurement conditions
Measurement conditions
Apparatus: AVANCE400MHz (Bruker)
Measurement temperature: room temperature (23°C)
Reference material: glycine (external standard: 176.03 ppm)
Measured nucleus: 13C (100.6248425 MHz)
Number of transients: 5000
Pulse repetition time: 4 seconds
Contact time: 1 m second
Rotational speed: 5000 Hz
Pulse mode: TOSS measurement
Hygroscopicity
The crystal obtained by the method described in the following Examples was weighed into a sampling cup and then the sampling cup was placed inside an isothermal chamber at 25°C. The relative humidity (RH) was controlled from 0% to 95% using a gravimetric vapor sorption system and the weight change of the sample at each stage was measured under the condition described below.
Measurement conditions
Sample temperature: 25°C
First stage RH: 0%
Maximum stage RH: 95%
Step number: 39 (0, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 90, 85, 80, 75, 70, 65, 60, 55, 50, 45, 40, 35, 30, 25, 20, 15, 10, 5, 0)
Equilibrium criterion: 0.002 wt% in 1 minute
Max equilibrium time: 360 minutes
Production Example 1
Preparation of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (compound (I))
Figure JPOXMLDOC01-appb-C000092
Production Example 1a
Synthesis of 5-bromo-3-fluoro-1H-indazole
Figure JPOXMLDOC01-appb-C000093
 Into a 5-L 3-necked round-bottom flask purged and maintained with an inert atmosphere of nitrogen was placed 5-bromo-1H-indazole (200 g, 1.0204mol, 1.00 equiv), CH3CN (3.5L), acetic acid (120 mL), and selectfluoro (544 g, 1.5367mol, 1.51 equiv). The resulting solution was stirred at 80 oC until completion. The reaction progress was monitored by LCMS. The resulting solution was diluted with 8 L of ethyl acetate and washed with 3x4000 mL of H2O. The organic layer was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (0:100-15:85). The collected fractions were combined and concentrated under vacuum to deliver the title compound in 72 g (33%) as a yellow solid.
1H NMR (400 MHz, DMSO-d6) δ 12.77 (s, 1H), 8.03-7.90 (m, 1H), 7.59-7.48 (m, 2H). LCMS: 215 [M+H]+.
Production Example 1b
Synthesis of 5-bromo-3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-1H-indazole
Figure JPOXMLDOC01-appb-C000094
 Into a 2-L 3-necked round-bottom flask was placed 5-bromo-3-fluoro-1H-indazole (70 g, 325.55 mmol, 1.00 equiv), DCM (700 mL), and TsOH (5.6 g, 32.52 mmol, 0.10 equiv). This was followed by the drop-wise addition of DHP (82.4 g, 979.55mmol, 3.01 equiv) while stirring at 0 oC. The resulting solution was stirred at 0 oC until completion. The reaction was monitored by LCMS. The resulting mixture was washed with 2x500 mL of H2O, and the organic layer was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (0:100-10:90). The collected fractions were combined and concentrated under vacuum to deliver the title compound in 96.3 g (99%) as yellow oil.
1H NMR (300 MHz, DMSO-d6) δ 8.02 (d, J = 1.8 Hz, 1H), 7.78-7.74 (m, 1H), 7.67-7.63 (m, 1H), 5.88-5.71 (m, 1H), 3.95-3.79 (m, 1H), 3.75-3.71 (m, 1H), 2.31-2.13 (m, 1H), 2.11-1.86 (m, 2H), 1.74-1.70 (m, 1H), 1.58-1.50 (m, 2H). LCMS: 299 [M+H]+.
Production Example 1c
Synthesis of 3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-5-((trimethylsilyl)ethynyl)-1H-indazole
Figure JPOXMLDOC01-appb-C000095
 Into a 2-L round-bottom flask purged and maintained with an inert atmosphere of nitrogen was placed 5-bromo-3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-1H-indazole (94.3 g, 315.24 mmol, 1.00 equiv), 2-Methyl THF (950mL), TEA (95.6 g, 944.76 mmol, 3.00 equiv), ethynyltrimethylsilane (154.5 g, 1.57 mol, 4.99 equiv), PdCl2 (5.6 g, 31.64 mmol, 0.10 equiv), Xantphos (36.5 g, 63.08 mmol, 0.20 equiv), and CuI (12 g, 63.01mmol, 0.20 equiv). The resulting solution was stirred at 80 oC until completion. The reaction progress was monitored by LCMS. The resulting solution was diluted with 1 L of 2-Methyl THF and was washed with 1x1 L of brine. The mixture was dried over anhydrous sodium sulfate and concentrated under vacuum to deliver the title compound in 134 g (crude) as a black oil. The crude product was used directly to the next step.
Production Example 1d
Synthesis of 5-ethynyl-3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-1H-indazole
Figure JPOXMLDOC01-appb-C000096
 Into a 2-L round-bottom flask was placed 3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-5-((trimethylsilyl)ethynyl)-1H-indazole (131.3 g, 414.92mmol, 1.00 equiv), methanol (950 mL), and potassium carbonate (114.7 g, 829.90mmol, 2.00 equiv). The resulting solution was stirred at 0 oC until completion. The reaction progress was monitored by LCMS. The resulting mixture was concentrated under vacuum and then diluted with 1 L of H2O. The solution was extracted with 3x1 L of ethyl acetate and the organic layers combined, dried over anhydrous sodium sulfate, and concentrated under vacuum to deliver the title compound in 77 g (76%) as black oil.
LCMS: 245 [M+H]+.
Production Example 1e
Synthesis of 3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-5-(4,4,4-trifluorobut-1-yn-1-yl)-1H-indazole
Figure JPOXMLDOC01-appb-C000097
 Into a 2-L round-bottom flask purged and maintained with an inert atmosphere of nitrogen was placed 5-ethynyl-3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-1H-indazole (72 g, 294.76mmol, 1.00 equiv), toluene (900 mL), 1,1,1-trifluoro-2-iodoethane (186 g, 885.98mmol, 3.01 equiv), DABCO (99 g, 883.93mmol, 3.00equiv), DPEPhos (31.8 g, 59.00mmol, 0.20 equiv), and Pd2(dba)3CHCl3 (15.3 g, 14.78mmol, 0.05 equiv). The resulting solution was stirred at 80 oC until completion. The reaction progress was monitored by LCMS. The resulting solution was diluted with 1 L of H2O and extracted with 2x1 L of ethyl acetate. The organic layers were combined, dried over anhydrous sodium sulfate, and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (0:100-15:85). The collected fractions were combined and concentrated under vacuum to deliver the title compound in 45 g (47%) as a yellow solid.
1H NMR (300 MHz, DMSO-d6) δ 7.89 (d, J = 1.4 Hz, 1H), 7.81-7.77 (m, 1H), 7.58-7.54 (m, 1H), 5.84-5.80 (m, 1H), 3.94-3.60 (m, 4H), 2.35-2.12 (m, 1H), 2.06-1.89 (m, 2H), 1.86-1.64 (m, 1H), 1.58-1.54 (m,2H). LCMS: 327 [M+H]+
Production Example 1f
Synthesis of (Z)-3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-5-(4,4,4-trifluoro-1,2-bis(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)but-1-en-1-yl)-1H-indazole
Figure JPOXMLDOC01-appb-C000098
 Into a 500-mL round-bottom flask purged and maintained with an inert atmosphere of nitrogen was placed 4,4,5,5-tetramethyl-2-(tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3,2-dioxaborolane (57 g, 219.74 mmol, 2.00 equiv), 3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-5-(4,4,4-trifluorobut-1-en-1-yl)-1H-indazole (36 g, 110.34 mmol, 1.00 equiv), Pt(PPh3)4 (6.84 g, 0.05 equiv), and 2-Methyl THF (450 mL). The solution was stirred at 90 °C until completion to deliver the title compound (crude) that was used directly to the next step.
Production Example 1g
Synthesis of (E)-4-bromobut-2-enoyl chloride
Figure JPOXMLDOC01-appb-C000099
 Into a 500-mL round-bottom flask, was placed (E)-4-bromobut-2-enoic acid (10 g, 60.61mmol, 1.00 equiv), DCM (200 mL), and N, N-dimethylformamide (0.5 mL). Oxalyl dichloride (7.7 g, 1.00 equiv) was added at 0 oC in dropwise. The resulting solution was stirred until completion at 0 °C. The mixture was used directly to the next step without isolation of the product.
Production Example 1h
Synthesis of (E)-4-bromo-N-methylbut-2-enamide
Figure JPOXMLDOC01-appb-C000100
 Into a 250-mL round-bottom flask was placed CH3NH2.HCl (1.005g, 1.00 equiv), sodium carbonate (3.18 g, 30.00 mmol, 2.00 equiv), and DCM (100 mL). Then, (E)-4-bromobut-2-enoyl chloride (15.00 mmol, 1.00 equiv) was added dropwise at 0 oC. The resulting solution was stirred at 0 oC in a water/ice bath until completion. The mixture was then washed with 2x100 mL of water. The organic layer was concentrated under vacuum to deliver the title compound in 3 g (62%) as a yellow solid.
LCMS: 178, 180[M+H]+.
Production Example 1i
Synthesis of tert-butyl (2-(4-iodophenoxy)ethyl)carbamate
Figure JPOXMLDOC01-appb-C000101
 To a stirred solution of 4-iodophenol (50 g, 0.227 mol) in DMF (750 mL) was added Cs2CO3 (493 g, 1.363 mol). The mixture was stirred for 30 min at room temperature and then tert-butyl (2-bromoethyl)carbamate (71.27 g, 0.318 mol) was added. The solution was stirred at 80 oC until completion. The reaction mixture was then poured onto ice water, solid separated was filtered and dried under reduced pressure to deliver the title compound in 80 g (97%) as an off-white solid.
LCMS: 264 [M-Boc+H]+.
Production Example 1j
Synthesis of 2-(4-iodophenoxy)ethan-1-amine hydrochloride
Figure JPOXMLDOC01-appb-C000102
 To a stirred solution of tert-butyl (2-(4-iodophenoxy)ethyl)carbamate (25 g, 68.6 mmol) in dioxane (50 mL) at 0 oC was added 4M HCl in dioxane (250 mL). The reaction mixture was stirred at room temperature until completion. The reaction mixture was then concentrated under reduced pressure to deliver the title compound in 16 g (88%) as crude material, used in next step without further purification.
Production Example 1k
Synthesis of tert-butyl (E)-2-(4-iodophenoxy)ethyl(4-(methylamino)-4-oxobut-2-enyl)carbamate
Figure JPOXMLDOC01-appb-C000103
 Into a 40-mL vial was placed 2-(4-iodophenoxy)ethan-1-amine hydrochloride (2.26 g, 7.57 mmol, 1.00 equiv), DIEA (1.9 g, 14.70mmol, 2.00 equiv), and N,N-dimethylformamide (20 mL). (E)-4-bromo-N-methylbut-2-enamide (1.08 g, 6.07mmol, 0.80 equiv) was then added to the solution at 0 oC, which was then stirred at R.T. until completion. Boc2O (2.62 g, 12mmol) was then added and the resulting mixture was stirred at room temperature until completion. Upon completion by TLC, the reaction mixture was cooled to 0 oC, quenched with ice cold water (100 mL) and extracted with 3x250 mL of DCM. The combined organic extracts were washed with brine (250 mL), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude material was purified by column chromatography over 100-200 mesh silica using 50-80% ethyl acetate in n-hexane as an eluent, then further purified with C18 column (MeOH/H2O=7:3) to deliver the title compound in 690 mg (20%).
1H NMR (400 MHz, DMSO-d6) δ 7.97-7.91 (m, 1H), 7.63-7.55 (m, 2H), 6.84-6.76 (m, 2H), 6.52 (d, J = 12.9 Hz, 1H), 5.89 (d, J = 15.5 Hz, 1H), 4.08-4.02 (m, 2H), 4.01-3.94 (m, 2H), 3.49 (d, J = 4.8 Hz, 2H), 2.63 (d, J = 4.6 Hz, 3H), 1.37 (s, 9H). LCMS: 461 [M+H]+.
Production Example 1l
Synthesis of ((Z)-1-(4-(2-((tert-butoxycarbonyl)((E)-4-(methylamino)-4-oxobut-2-en-1-yl)amino)ethoxy)phenyl)-4,4,4-trifluoro-1-(3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-1H-indazol-5-yl)but-1-en-2-yl)boronic acid
Figure JPOXMLDOC01-appb-C000104
 Into a 1000-mL round-bottom flask purged and maintained with an inert atmosphere of nitrogen, was placed (Z)-3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-5-(4,4,4-trifluoro-1,2-bis(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)but-1-en-1-yl)-1H-indazole (53 g, 91.94 mmol, 1.00 equiv), 2-Methyl THF (500 mL), tert-butyl (E)-(2-(4-iodophenoxy)ethyl)(4-(methylamino)-4-oxobut-2-en-1-yl)carbamate (42.3 g, 91.90 mmol, 1.00 equiv), Cs2CO3 (90 g, 276.23 mmol, 3.00 equiv), Pd(PPh3)2Cl2 (6.46 g, 9.20 mmol, 0.10 equiv), and water (100 mL) were added. The resulting solution was stirred at 50 oC until completion. The reaction progress was monitored by LCMS. The solution was diluted with 500 mL of H2O, extracted with 2x600 mL of ethyl acetate, then the organic layers were combined, dried over anhydrous sodium sulfate, and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1:2). The collected fractions were combined and concentrated under vacuum to deliver the title compound in 36 g (54%) as yellow oil.
Production Example 1m
Synthesis of tert-butyl ((E)-4-(methylamino)-4-oxobut-2-en-1-yl)(2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)carbamate
Figure JPOXMLDOC01-appb-C000105
 Into a 1-L round-bottom flask purged and maintained with an inert atmosphere of nitrogen, was placed ((Z)-1-(4-(2-((tert-butoxycarbonyl)((E)-4-(methylamino)-4-oxobut-2-en-1-yl)amino)ethoxy)phenyl)-4,4,4-trifluoro-1-(3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-1H-indazol-5-yl)but-1-en-2-yl)boronic acid (36 g, 51.10 mmol, 1.00 equiv), Pd(PPh3)2Cl2 (3.5 g, 4.99 mmol, 0.10 equiv), potassium hydroxide (8.4 g, 149.71 mmol, 3.00 equiv), dioxane (200 mL), water (40 mL), and bromobenzene (8.4 g, 53.50 mmol, 1.00 equiv). The resulting solution was stirred at 80 oC until completion. The reaction progress was monitored by LCMS. The resulting solution was diluted with 200 ml of H2O, extracted with 3x500 ml of ethyl acetate, then the organic layers combined, washed with brine (200 ml) and dried over anhydrous sodium sulfate. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1:3). The collected fractions were combined and concentrated under vacuum to deliver the title compound in 16 g (42%) as a yellow solid.
Production Example 1n
Synthesis of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (compound (I))
Figure JPOXMLDOC01-appb-C000106
 Into a 250-mL round-bottom flask was placed tert-butyl ((E)-4-(methylamino)-4-oxobut-2-en-1-yl)(2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1-(tetrahydro-2H-pyran-2-yl)-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)carbamate (16 g, 21.72 mmol, 1.00 equiv) and TFA (100 mL). The resulting solution was stirred at 25 oC until completion. The reaction progress was monitored by LCMS. The resulting solution was concentrated under vacuum and the crude product was purified by Prep-HPLC with the following conditions: Column: X-Bridge Prep OBD C18 Column 30×150mm 5um; Mobile Phase A: Water(10mmol/L NH4HCO3), Mobile Phase B: ACN; Flow rate: 60 mL/min; Gradient: 40% B to 55% B in 60 min; 254,220 nm, to deliver the title compound as a free base in 4.05 g as a yellow solid.
1H NMR (400 MHz, Methanol-d4) δ 7.60 (s, 1H), 7.52-7.42 (m, 1H), 7.25-7.11 (m, 6H), 6.83-6.81 (m, 2H), 6.78-6.71 (m, 1H), 6.63-6.61 (m, 2H), 6.05-6.01 (m, 1H), 3.97-3.94 (t, J = 5.3 Hz, 2H), 3.41-3.33 (m, 4H), 2.98-2.88 (t, J = 5.2 Hz, 2H), 2.76 (s, 3H).
The solid was then dissolved in 100 mL CH3CN and acidified with 8.07 mL HCl (1N) (1 mL 12N HCl(aq) dissolved in 11 mL CH3CN) at 0 oC, and stirred for 30 min at R.T., then evaporated at 30 oC to remove the excess HCl. Then the product was dissolved in 150 mL H2O and lyophilized for 48h to deliver the title compound in 4.4 g, 0.96% overall yield, as a yellow solid.
1H NMR (400 MHz, Methanol-d4) δ 7.59 (s, 1H), 7.46-7.44 (m, 1H), 7.27 (m, 1H) 7.25-7.12 (m, 5H), 6.91-6.87 (m, 2H), 6.72-6.65 (m, 3H), 6.30-6.26 (m, 1H), 4.16-4.14 (t, J = 4.9 Hz, 2H), 3.86-3.84 (m, 2H), 3.42-3.34 (m, 4H), 2.79 (s, 3H). LCMS: 553 [M+H]+.
Example 1
Preparation of crystal of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide hydrochloride
To a solution of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.01 mg) in acetonitrile (3 mL) was added a solution of hydrochloric acid (62.1 μL, 1eq) in acetonitrile (2 mL). Then, 3 mL of acetonitrile was added to the mixture and it was stirred at 50 °C for an hour, at room temperature overnight, at 50 °C for an hour, at room temperature for 6 hours, at 50 °C for an hour. Then, 2 mL of acetonitrile was added to the mixture again and it was stirred at room temperature for 4 days. The obtained solid was filtered off, washed with acetonitrile and dried for 10 days to give the titled crystal (358 mg, 84%). A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 1 are shown in Figures 1, 12 and 23, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.64 (d, J=4.6 Hz, 3H), 3.22 (t, J=4.6 Hz, 2H), 3.44 (q, J=10.7 Hz, 2H), 3.74 (d, J=6.5 Hz, 2H), 4.10 (t, J=4.7 Hz, 2H), 6.16 (d, J=15.4 Hz, 1H), 6.57 (dt, J=15.4, 6.8 Hz, 1H), 6.68 (d, J=8.9 Hz, 2H), 6.85 (d, J=8.8 Hz, 2H), 7.15 (dddd, J=6.9, 6.9, 1.3, 1.3 Hz, 1H), 7.17-7.26 (m, 5H), 7.52 (dd, J=8.7, 1.5 Hz, 1H), 7.57 (br s, 1H), 8.15 (q, J=4.6 Hz, 1H), 9.09 (br s, 2H), 12.69 (s, 1H).
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 1 are shown below.
13C NMR (100MHz, solid state): δ 25.8, 45.9, 117.3, 128.5, 133.9, 139.0, 141.4, 144.5, 156.8, 165.8.
Powder X-ray diffraction angle (2θ ± 0.2°): 12.2°, 16.8°, 18.2°, 19.7°, 21.3°, 21.8°, 23.3°, 24.5°, 25.5°, 29.4°.
Example 2
Preparation of crystal of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide hydrobromide
To a solution of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.44 mg) in acetonitrile (2 mL) was added a solution of hydrobromic acid (119.6 μL, 1eq) in acetonitrile (2 mL). Then, 5 mL of tert-butyl methyl ether was added to the solution and the mixture was stirred at room temperature for 4 days. The obtained solid was filtered off, washed with tert-butyl methyl ether and dried for 10 days to give the titled crystal (322 mg, 70%). A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 2 are shown in Figures 2, 13 and 24, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.64 (d, J=4.7 Hz, 3H), 3.25 (t, J=4.6 Hz, 2H), 3.44 (q, J=10.8 Hz, 2H), 3.77 (d, J=6.7 Hz, 2H), 4.08 (t, J=4.7 Hz, 2H), 6.16 (d, J=15.5 Hz, 1H), 6.55 (dt, J=15.4, 6.8 Hz, 1H), 6.69 (d, J=8.8 Hz, 2H), 6.85 (d, J=8.8 Hz, 2H), 7.15 (dddd, J=6.9, 6.9, 1.5, 1.5 Hz, 1H), 7.18-7.25 (m, 5H), 7.52 (dd, J=8.8, 1.5 Hz, 1H), 7.56 (br s, 1H), 8.13 (q, J=4.6 Hz, 1H), 8.83 (br s, 2H), 12.66 (s, 1H),
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 2 are shown below.
13C NMR (100MHz, solid state): δ 26.4, 45.2, 47.3, 116.9, 129.3, 139.4, 141.6, 144.2, 156.7, 165.2.
Powder X-ray diffraction angle (2θ ± 0.2°): 6.1°, 12.0°, 16.6°, 18.4°, 19.7°, 21.6°, 22.9°, 24.1°, 25.6°, 29.8°.
Example 3
Preparation of crystal of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide phosphate
To a solution of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.25 mg) in acetonitrile (4 mL) was added a solution of phosphoric acid (49.4 μL, 1eq) in acetonitrile (1 mL). The mixture was stirred at 50 °C for 3 hours and at room temperature overnight. Again, it was stirred at 50 °C for an hour, 3 mL of acetonitrile was added to the mixture and it was stirred at room temperature for 3 days. The obtained solid was filtered off, washed with acetonitrile and dried for 10 days to give the titled crystal (416 mg, 88%). A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 3 are shown in Figures 3, 14 and 25, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.61 (d, J=4.7 Hz, 3H), 2.87 (m, 2H), 3.39 (m, 2H), 3.43 (q, J=10.8 Hz, 2H), 3.92 (t, J=5.1 Hz, 2H), 6.01 (d, J=15.4 Hz, 1H), 6.56 (dt, J=15.4, 5.7 Hz, 1H), 6.64 (d, J=8.7 Hz, 2H), 6.81 (d, J=8.7 Hz, 2H), 7.14 (m, 1H), 7.17-7.25 (m, 5H), 7.52 (dd, J=8.7, 1.2 Hz, 1H), 7.56 (br s, 1H), 7.92 (m, 1H), 12.66 (s, 1H).
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 3 are shown below.
13C NMR (100MHz, solid state): δ 26.9, 45.5, 49.2, 126.6, 135.3, 138.3, 140.9, 145.5, 146.6, 169.8.
Powder X-ray diffraction angle (2θ ± 0.2°): 6.5°, 13.5°, 14.0°, 16.7°, 19.9°, 20.4°, 21.6°, 23.2°.
Example 4
Preparation of crystal of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide sulfate
To a solution of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.08 mg) in acetonitrile (2 mL) was added a solution of sulfuric acid (40.6 μL, 1eq) in acetonitrile (2 mL). Then, 1 mL of acetonitrile was added to the solution, the mixture was stirred at 50 °C for an hour and at room temperature overnight. Again, it was stirred at 50 °C for 2 hours, 3 mL of acetonitrile was added to the mixture and it was stirred at room temperature for 3 days. The obtained solid was filtered off, and dried for 10 days to give the titled crystal (428 mg, 91%). A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 4 are shown in Figures 4, 15 and 26, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.65 (d, J=4.7 Hz, 3H), 3.24 (t, J=4.7 Hz, 2H), 3.44 (q, J=10.8 Hz, 2H), 3.76 (d, J=6.6 Hz, 2H), 4.07 (t, J=4.7 Hz, 2H), 6.15 (d, J=15.5 Hz, 1H), 6.55 (dt, J=15.5, 6.8 Hz, 1H), 6.69 (d, J=8.8 Hz, 2H), 6.85 (d, J=8.8 Hz, 2H), 7.15 (dddd, J=7.1, 7.1, 1.5, 1.5 Hz, 1H), 7.18-7.26 (m, 5H), 7.52 (dd, J=8.8, 1.3 Hz, 1H), 7.56 (br s, 1H), 8.11 (q, J=4.6 Hz, 1H), 8.83 (br s, 2H), 12.66 (s, 1H).
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 4 are shown below.
13C NMR (100MHz, solid state): δ 25.0, 47.8, 49.9, 113.6, 117.8, 124.7, 141.3, 146.5, 157.4, 165.0.
Powder X-ray diffraction angle (2θ ± 0.2°): 3.7°, 7.6°, 11.3°, 13.0°, 16.9°, 19.0°, 19.8°.
Example 5
Preparation of crystal of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide benzenesulfonate
To a solution of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.35 mg) in acetonitrile (2 mL) was added a solution of benzenesulfonic acid monohydrate (100 mg/mL) in acetonitrile (1.3 mL). Then, the mixture was stirred at 50 °C for 30 minutes, 1.5 mL of tert-butyl methyl ether was added, and stirred at room temperature overnight. Again, 1.5 mL of tert-butyl methyl ether was added to the mixture, it was stirred at 50 °C for an hour and at room temperature overnight. The obtained solid was filtered off, washed with tert-butyl methyl ether, and dried for 9 days to give the titled crystal (461 mg, 90%). A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 5 are shown in Figures 5, 16 and 27, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.64 (d, J=4.7 Hz, 3H), 3.24 (t, J=4.4 Hz, 2H), 3.44 (q, J=10.8 Hz, 2H), 3.76 (d, J=6.6 Hz, 2H), 4.07 (t, J=4.7 Hz, 2H), 6.15 (d, J=15.5 Hz, 1H), 6.55 (dt, J=15.5, 6.8 Hz, 1H), 6.68 (d, J=8.7 Hz, 2H), 6.85 (d, J=8.7 Hz, 2H), 7.15 (m, 1H), 7.17-7.25 (m, 5H), 7.26-7.33 (m, 3H), 7.52 (dd, J=8.7, 1.4 Hz, 1H), 7.56 (br s, 1H), 7.59 (dd, J=7.9, 1.8 Hz, 2H), 8.11 (q, J=4.6 Hz, 1H), 8.74 (br s, 2H), 12.66 (s, 1H).
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 5 are shown below.
13C NMR (100MHz, solid state): δ 27.6, 44.9, 47.1, 116.8, 132.3, 136.1, 156.7, 164.4.
Powder X-ray diffraction angle (2θ ± 0.2°): 5.5°, 9.8°, 10.9°, 17.6°, 19.2°, 19.9°, 21.9°, 24.1°, 26.5°.
Example 6
Preparation of crystal of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide p-toluenesulfonate
To a solution of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.05 mg) in acetonitrile (2 mL) was added a solution of p-toluenesulfonic acid monohydrate (100 mg/mL) in acetonitrile (1.4 mL). Then, 5 mL of acetonitrile and 2 mL of tert-butyl methyl ether were added the mixture, it was stirred at 50 °C for an hour. Then, 2 mL of tert-butyl methyl ether was added to the mixture again, it was stirred at room temperature 3 days, at 50 °C for an hour and at room temperature overnight. The obtained solid was filtered off, washed with tert-butyl methyl ether, and dried for 9 days to give the titled crystal (448 mg, 85%). A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 6 are shown in Figures 6, 17 and 28, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.27 (s, 3H), 2.65 (d, J=4.7 Hz, 3H), 3.26 (br s, 2H), 3.44 (q, J=10.8 Hz, 2H), 3.77 (br s, 2H), 4.07 (t, J=4.8 Hz, 2H), 6.16 (d, J=15.4 Hz, 1H), 6.55 (dt, J=15.5, 6.8 Hz, 1H), 6.68 (d, J=8.8 Hz, 2H), 6.85 (d, J=8.9 Hz, 2H), 7.10 (d, J=7.8 Hz, 2H), 7.15 (dddd, J=7.0, 7.0, 1.6, 1.6 Hz, 1H), 7.18-7.25 (m, 5H), 7.46 (d, J=8.1 Hz, 2H), 7.52 (dd, J=8.8, 1.5 Hz, 1H), 7.56 (br s, 1H), 8.12 (q, J=4.5 Hz, 1H), 8.83 (br s, 2H), 12.66 (s, 1H).
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 6 are shown below.
13C NMR (100MHz, solid state): δ 19.2, 27.5, 46.4, 112.3, 117.0, 125.9, 140.7, 144.2, 156.4, 166.9.
Powder X-ray diffraction angle (2θ ± 0.2°): 8.9°, 11.5°, 12.5°, 16.2°, 17.5°, 20.0°, 20.9°, 21.4°, 22.1°, 23.4°.
Example 7
Preparation of crystal of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide succinate
To a solution of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.23 mg) in tert-butyl methyl ether (2 mL) was added a solution of succinic acid (86 mg, 1eq) in acetonitrile (2 mL). Then, 4 mL of tert-butyl methyl ether was added to the mixture, it was stirred at 50 °C for 30 minutes, at room temperature overnight, and at 50 °C for 30 minutes. After cooling down to room temperature, the obtained solid was filtered off, and dried for 9 days to give the titled crystal (388 mg, 80%). A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 7 are shown in Figures 7, 18 and 29, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.38 (s, 4H), 2.61 (d, J=4.7 Hz, 3H), 2.82 (t, J=5.5 Hz, 2H), 3.33 (d, J=4.6 Hz, 2H), 3.43 (q, J=10.8 Hz, 2H), 3.89 (t, J=5.6 Hz, 2H), 5.99 (d, J=15.5 Hz, 1H), 6.56 (dt, J=15.4, 5.6 Hz, 1H), 6.63 (d, J=8.9 Hz, 2H), 6.80 (d, J=8.8 Hz, 2H), 7.14 (dddd, J=6.9, 6.9, 1.6, 1.6 Hz, 1H), 7.17-7.25 (m, 5H), 7.51 (dd, J=8.7, 1.5 Hz, 1H), 7.56 (br s, 1H), 7.87 (q, J=4.5 Hz, 1H), 12.65 (s, 1H).
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 7 are shown below.
13C NMR (100MHz, solid state): δ 26.0, 35.1, 116.0, 125.3, 139.2, 145.8, 156.1, 165.5, 175.0, 181.3.
Powder X-ray diffraction angle (2θ ± 0.2°): 5.2°, 10.4°, 12.9°, 14.8°, 17.0°, 17.5°, 18.2°, 19.1°, 21.8°, 23.6°.
Example 8
Preparation of crystal form A of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide maleate
To (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.08 mg) was added a solution of maleic acid (84 mg, 1eq) in acetonitrile (3 mL). Then, 6 mL of acetonitrile was added the mixture, it was stirred at 50 °C for 30 minutes, at room temperature for 2 hours, at 50 °C for 30 minutes and at room temperature overnight. Again, 1 mL of acetonitrile was added to the mixture, it was stirred at 55 °C for 2 hours and room temperature overnight, at 55 °C for 3 hours and room temperature overnight, at 55 °C for 5 hours and room temperature overnight, at 55 °C for 2.5 hours and room temperature overnight, and at room temperature for 4 days. The obtained solid was filtered off, and dried overnight to give the titled crystal (353 mg, 73%). A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 8 are shown in Figures 8, 19 and 30, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.65 (d, J=4.7 Hz, 3H), 3.24 (t, J=4.7 Hz, 2H), 3.44 (q, J=10.9 Hz, 2H), 3.76 (d, J=6.6 Hz, 2H), 4.07 (t, J=4.7 Hz, 2H), 6.01 (s, 2H), 6.15 (d, J=15.5 Hz, 1H), 6.55 (dt, J=15.5, 6.7 Hz, 1H), 6.68 (d, J=8.8 Hz, 2H), 6.85 (d, J=8.9 Hz, 2H), 7.15 (dddd, J=7.0, 7.0, 1.4, 1.4 Hz, 1H), 7.18-7.25 (m, 5H), 7.52 (dd, J=8.7, 1.6 Hz, 1H), 7.56 (br s, 1H), 8.11 (q, J=4.6 Hz, 1H), 8.74 (br s, 2H), 12.66 (s, 1H).
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 8 are shown below.
13C NMR (100MHz, solid state): δ 43.4, 44.8, 47.8, 63.0, 66.0, 125.2, 133.8, 140.6, 145.0, 165.5.
Powder X-ray diffraction angle (2θ ± 0.2°): 5.4°, 11.6°, 13.3°, 14.3°, 16.8°, 18.9°, 20.2°, 21.6°.
Example 9
Preparation of crystal form B of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide maleate
To (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.31 mg) was added a solution of maleic acid (84 mg, 1eq) in acetone (2 mL). Then, 4 mL of acetone and 1 mL of heptane were added the mixture, it was stirred at room temperature overnight. The obtained solid was filtered off, and dried for 9 days to give the titled crystal (223 mg, 46%). A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 9 are shown in Figures 9, 20 and 31, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.65 (d, J=4.7 Hz, 3H), 3.24 (t, J=4.8 Hz, 2H), 3.44 (q, J=10.9 Hz, 2H), 3.76 (d, J=6.6 Hz, 2H), 4.07 (t, J=4.7 Hz, 2H), 6.01 (s, 2H), 6.15 (d, J=15.5 Hz, 1H), 6.55 (dt, J=15.5, 6.7 Hz, 1H), 6.68 (d, J=8.8 Hz, 2H), 6.85 (d, J=8.8 Hz, 2H), 7.15 (dddd, J=7.0, 7.0, 1.4, 1.4 Hz, 1H), 7.18-7.25 (m, 5H), 7.52 (dd, J=8.7, 1.5 Hz, 1H), 7.56 (br s, 1H), 8.11 (q, J=4.6 Hz, 1H), 8.75 (br s, 2H), 12.66 (s, 1H).
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 9 are shown below.
13C NMR (100MHz, solid state): δ 25.8, 48.8, 110.7, 115.7, 135.1, 136.6, 143.6, 156.3, 166.8, 172.6.
Powder X-ray diffraction angle (2θ ± 0.2°): 10.4°, 15.2°, 19.4°, 20.8°, 22.2°, 23.7°, 25.7°, 28.6°.
Example 10
Preparation of crystal of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide L-mandelate
To (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (400.17 mg) was added a solution of L-mandelic acid (110 mg, 1eq) in acetone (2 mL). Then, 6 mL of acetone and 4 mL of tert-butyl methyl ether were added to the mixture, it was stirred at 50 °C for 30 minutes, at room temperature for 2 hours, at 50 °C for 30 minutes and at room temperature overnight. Again, 2 mL of acetone was added to the mixture, it was stirred at room temperature overnight, at 50 °C for 30 minutes and room temperature overnight, and at 50 °C for 30 minutes and room temperature overnight. After adding 2 mL of acetone to the mixture, it was stirred at 40 °C for 6 days, 0.12 mL of purified water was added, stirred at 40 °C overnight, 0.36 mL of purified water was added again, stirred at 50 °C for 6 hours and 40 °C for 3 days. After cooling down to room temperature and reducing the solvent by nitrogen purging, the obtained solid was filtered off, and dried for a week to give the titled crystal (399 mg, 78%). A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 10 are shown in Figures 10, 21 and 32, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.61 (d, J=4.6 Hz, 3H), 2.85 (t, J=5.4 Hz, 2H), 3.36 (dd, J=5.6, 0.9 Hz, 2H), 3.43 (q, J=10.8 Hz, 2H), 3.90 (t, J=5.5 Hz, 2H), 4.88 (s, 1H), 6.00 (d, J=15.5 Hz, 1H), 6.56 (dt, J=15.5, 5.6 Hz, 1H), 6.63 (d, J=8.8 Hz, 2H), 6.80 (d, J=8.8 Hz, 2H), 7.14 (dddd, J=6.8, 6.8, 1.5, 1.5 Hz, 1H), 7.17-7.26 (m, 6H), 7.30 (dd, J=7.4, 7.4 Hz, 2H), 7.39 (d, J=7.2 Hz, 2H), 7.52 (dd, J=8.7, 1.5 Hz, 1H), 7.56 (br s, 1H), 7.89 (q, J=4.6 Hz, 1H), 12.66 (s, 1H).
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 10 are shown below.
13C NMR (100MHz, solid state): δ 62.9, 76.5, 116.2, 123.6, 128.1, 141.9, 146.3, 156.7, 164.6, 177.8.
Powder X-ray diffraction angle (2θ ± 0.2°): 5.1°, 8.7°, 10.3°, 13.9°, 18.2°, 20.3°.
Example 11
Preparation of crystal of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide methanesulfonate
To a solution of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (50.22 mg) in 2-butanone (1 mL) was added a solution of methanesulfonic acid (6.2 μL, 1eq) in 2-butanone (1 mL). Then, the mixture was stirred at room temperature for 2 days.
One mL of the mixture in 2-butanone was transferred to another test tube, the solvent was removed by nitrogen purging. Then, 0.75 mL of acetone was added and the mixture in acetone was stirred at room temperature overnight (hereafter the above obtained mixture is referred to as Mixture 1).
The rest (1 mL) of the mixture in 2-butanone was stirred additionally overnight. The solvent was removed by nitrogen purging. Then, 0.75 mL of acetone was added and the mixture in acetone was stirred at room temperature for 4 days (hereafter the above obtained mixture is referred to as Mixture 2).
To a solution of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (150.49 mg) in 2-butanone (2 mL) was added a solution of methanesulfonic acid (18.6 μL, 1eq) in 2-butanone (2 mL). Then, the mixture was stirred at room temperature overnight. Then, the solvent was removed by nitrogen purging, 4 mL of acetone and the Mixture 1 were added, and the mixture was stirred at room temperature for 3 days. Again, the solvent was replaced by 4 mL of acetone and the mixture was stirred at room temperature overnight (hereafter the above obtained mixture is referred to as Mixture 3).
To a solution of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide (200.26 mg) in 2-butanone (2 mL) was added a solution of methanesulfonic acid (24.8 μL, 1eq) in 2-butanone (2 mL). The mixture was stirred at room temperature for 3 days. Then, the solvent was removed by nitrogen purging, 5 mL of acetone was added, and the mixture was stirred at room temperature overnight. Two mL of acetone, the Mixture 2 and the Mixture 3 were added to the mixture and it was stirred at room temperature overnight. The obtained solid was filtered off, and dried for 2 days to give the titled crystal (260 mg, 55%).
A powder X-ray diffraction pattern, a solid state 13C NMR spectrum and a graph showing hygroscopicity for the crystal obtained in Example 11 are shown in Figures 11, 22 and 33, respectively.
1H NMR (600MHz, Bruker AVANCE, DMSO-d6): δ 2.29 (s, 3H), 2.65 (d, J=4.6 Hz, 3H), 3.26 (br s, 2H), 3.44 (q, J=10.8 Hz, 2H), 3.77 (br s, 2H), 4.08 (t, J=4.8 Hz, 2H), 6.16 (d, J=15.5 Hz, 1H), 6.55 (dt, J=15.4, 6.8 Hz, 1H), 6.69 (d, J=8.8 Hz, 2H), 6.85 (d, J=8.8 Hz, 2H), 7.15 (m, 1H), 7.18-7.25 (m, 5H), 7.53 (dd, J=8.7, 1.3 Hz, 1H), 7.56 (br s, 1H), 8.12 (q, J=4.6 Hz, 1H), 8.84 (br s, 2H), 12.66 (s, 1H).
Typical peaks in a solid state 13C NMR spectrum and typical diffraction peaks in a powder X-ray diffractometry for the crystal obtained in Example 11 are shown below.
13C NMR (100MHz, solid state): δ 24.8, 40.4, 47.2, 117.8, 133.9, 136.0, 140.4, 146.7, 158.3, 166.0.
Powder X-ray diffraction angle (2θ ± 0.2°): 7.5°, 12.1°, 12.7°, 17.1°, 18.9°, 19.7°, 20.9°, 21.8°, 25.6°.
The following test examples were carried out to examine the pharmacological effects of the compound (I).
(Test Example 1) Compounds that inhibit ERαWT/MUT activity in vitro
Cell culture
MCF7 BUS cells (Coser, et al., (2003) PNAS 100 (24): 13994-13999) were maintained in Dulbecco’s Modified Eagle Medium supplemented with 10% FBS, 4 mM L-glutamine and 1x non-essential amino acids. Lenti-X 293T cells (Clontech, Cat # 632180) were routinely cultured in Dulbecco’s Modified Eagle Medium supplemented with 10% FBS.
Site-direct mutagenesis and cell line engineering
The QuikChange II XL Site-Directed Mutagenesis Kit (Agilent Technologies, Cat #200523) was used to generate Y537S, Y537C, Y537N and D538G mutations within the ERα exon 8. Wild-type ESR1 cDNA (GeneCopoeia Inc., Cat# GC-A0322, accession no. NM 000125) was used as a template with the following mutagenesis primers (where the underlined nucleotides represent site mutations); 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); Y537N: F-AAG AAC GTG GTG CCC CTC AAT GAC CTG CTG CTG GAG ATG (SEQ ID NO: 3), R-CAT CTC CAG CAG CAG GTC ATT GAG GGG CAC CAC GTT CTT (SEQ ID NO: 4); Y537C: F-AAG AAC GTG GTG CCC CTC TGT GAC CTG CTG CTG GAG ATG (SEQ ID NO: 5), R-CAT CTC CAG CAG CAG GTC ACA GAG GGG CAC CAC GTT CTT (SEQ ID NO: 6); D538G: F-AAC GTG GTG CCC CTC TAT GGC CTG CTG CTG GAG ATG CTG (SEQ ID NO: 7), R-CAG CAT CTC CAG CAG CAG GCC ATA GAG GGG CAC CAC GTT (SEQ ID NO: 8). WT and mutant ESR1 cDNAs were cloned into the designation lentiviral vector pLenti6.3/V5-Dest (Invitrogen, Cat #V533-06). To make lentivirus, DNAs (WT and mutant ESR1) were co-transfected with packaging plasmids into Lenti-X 293T cells using TransIT (Mirus, Cat #MIR 2700). 48h post-transfection, virus containing media was filtered and added to MCF7 cells in the presence of 8 μg/ml polybrene overnight. Two days following infection, cells were placed under selection with 10 μg/ml blasticidin for 2 weeks for stable expression.
In vitro proliferation assays
MCF7-WT and -Y537S cells were seeded at 1500 cells/well in black-walled 96-well plates (assay plates, Costar, Cat #3904). In parallel, cells were also seeded in a separate 96-well plate (8 wells/cell line, control plate) for which a CTG (CellTiter-Glo (registered trademark) Luminescent Viability Assay, Promega, Cat #G7572) was measured the following day (day 0 reading). The day 0 reading was used for the GI50 calculation at the termination of the experiment. The day following seeding, compounds were added to assay plates. Briefly, a 1:4 serial dilution was prepared in DMSO at 200x final concentration for a total of 10 concentrations (9 dilutions containing compound and one is DMSO only). Serially diluted compounds were pipetted into medium to prepare a compound-medium mix at 10x final concentration. 10 μl of compound-medium mix was added to MCF7-WT and -Y537S cells at 3 wells/concentration (triplicate for each concentration). On day 3, media/compound was removed and replaced with fresh media/compound as described above. On day 6, CTG was measured and compared to day 0 readings from control plate to assess GI50.
Results
Figure 34 shows that ectopic expression of ERαY537S/N/C, D538G in MCF7 cells conferred phenotypic resistance to currently marketed therapies tamoxifen (SERM), raloxifene (SERM) and fulvestrant (SERD). Similar observations were also recently published by several independent labs (Jeselsohn et al., (2014) Clin Cancer Res. Apr 1; 20 (7): 1757-67; Toy et al., (2013) Nat. Genet. 2013 Dec; 45(12):1439-45; Robinson et al., (2013) Nat. Genet. Dec; 45 (12): 1446-51; Merenbakh-Lamin et al., (2013) Cancer Res. Dec 1; 73(23): 6856-64; Yu et al., (2014) Science Jul 11; 345(6193): 216-20). Having confirmed that ERαMUT drive resistance to current endocrine therapies, identification of novel compounds that would reduce proliferation of the ERαMUT-bearing MCF7 cells more efficaciously than the corresponding clinical compound 4-hydroxytamoxifen was sought. Using the WT and mutant viability assay as a screening tool, compounds were identified that were more potent towards the Y537S-bearing MCF7 line relative to 4-hydroxytamoxifen. The results of the viability assay screen are shown in Table 1. These assays were conducted with free base and/or salt forms of the compounds identified in the table.
Figure JPOXMLDOC01-appb-T000107
Results as presented in Table 1 are presented as the average of one or more trials with standard deviations where available. The number of trials for each compound is presented in parentheses following the value. Those skilled in the art would appreciate that the examples and embodiments reported herein are for illustrative purposes only. Various modifications or changes in light thereof will be suggested to persons skilled in the art, and those modifications or changes are included within the spirit and purview of this application and scope of the appended claims. For example, those skilled in the art will appreciate that GI50 values may vary depending on the lot of fetal bovine serum (FBS), among other factors, used to supplement the culture media, due to varying concentrations of estrogen between batches.
Compounds were tested as prepared in the Examples below with regard to use of free base or salt; results for compound (I) are presented as averages of both free base and HCl salt trials.
(Test Example 2) In vivo xenograft methods
Methods and materials
Although not wishing to be bound by theory, applicant appreciates that certain in vivo xenograft studies may be useful in identifying effective compounds. Such studies may be conducted, for example, using compounds reported herein and/or their salts. In studies reported herein, the hydrochloride salt form as described herein was used. The WHIM20 xenograft study reported below has not yet been conducted with compounds reported herein, but the Y537S positive PDx xenograft study and studies in the ERα wild-type MCF7 and PDx models have been conducted with certain of the compounds as set forth below.
Y537S positive PDX xenograft study
A Patient-Derived Xenograft (PDX) tumor model representing an ESR1-Y537S mutated human ER+ breast cancer, designated as PDX-Y537S, was propagated subcutaneously in immunocompromised mice. The tumors were excised within 60 days of implantation and processed to mixed tumor fragments. Solid tumor tissues were depleted of necrotic components, cut into 70 mg fragments, mixed with matrigel and subcutaneously implanted into the right flank of 6-12 week old female athymic Nude (Crl:NU(NCr)-Foxn1nu) mice. The precise number of fragments and volume of matrigel were determined on a case by case basis. When the average tumor volume reaches approximately 125-250 mm3, animals were randomized prior to treatment. All of the primary human tumors utilized in this study had undergone approximately 5-7 passages in vivo.
Estrogen was not supplemented in the studies. All tested compounds were dosed orally every day at doses ranging from 3 to 30 mg/kg. The administration volume was calculated from the individual mouse body weights prior to dose administration. The body weights (BW) and tumor volumes (TV) were measured twice a week.
Tumor volumes (TV) were calculated based on the formula:
TV = length ´ width2 ´ 0.5
length: largest diameter of tumor (mm)
width: diameter perpendicular to length (mm)
The Tumor Growth Inhibition % (TGI) was calculated according to the following formula:
Figure JPOXMLDOC01-appb-M000108
 Where Day X is the endpoint measurement.
Results for Y537S Positive PDx Xenograft Studies
Figure 35 shows the anti-tumor and body weight effects of compound (I), prepared as an HCl salt, in the PDX-Y537S model bearing a heterozygous ERαY537/WT xenograft. Compound (I) dosed daily inhibited xenograft growth in a dose dependent manner with 3 mg/kg QD, 10 mg/kg QD, and 30 mg/kg QD treatments significantly inhibiting growth on day 44 relative to vehicle control (TGI of 43%, 74%, and 77%, and p<0.05, respectively). All doses and regimens were well tolerated with no significant body weight loss.
Compound (I) was given orally once daily for the duration of the study. Data represent the mean ± SEM (Tumor Volume) or mean ± SEM (Body Weight) (N=6 for all groups). *p<0.05 versus vehicle control on Day 44 (repeated measures t-test, Holm-Sidak method with α=0.05 without assuming a consistent SD).
MCF7 Xenograft Studies
The ESR1 wild-type human ER+ breast cancer cell line MCF7 (ATCC) was cultured in DMEM media supplemented with 10% FBS at 37°C in a 5% CO2 atmosphere and kept in the exponential growth phase. The cells were collected in trypsin and re-suspended in a 1:1 mixture of matrigel and HBSS at a final concentration of 5 x107 cells/ mL. A 0.2mL aliquot of cells was injected subcutaneously into the 3rd mammary fat pad of 6-8 week old female Balb/c nude mice, giving 1x 107 cells/ mouse. When the average tumor volume reached approximately about 200 mm3, animals were randomized prior to treatment.
Estrogen was supplemented for the duration of the study. Compound (I) was dosed orally every day at doses ranging from 1 to 10 mg/kg. Each treatment was started on Day 0 and the administration schedule was continued for 28 days. The administration volume was calculated from the individual mouse body weights prior to dose administration. The body weights (BW) were measured daily while the tumor volumes were measured twice a week. Tumor volumes (TV) were calculated based on the above formula.
Results for MCF7 Xenograft Studies
Figure 36 shows the anti-tumor and body weight effects of compound (I), prepared as an HCl salt, in the MCF7 tumor model bearing ERαWT/WT xenograft. Compound (I) dosed daily inhibited xenograft growth in a dose dependent manner with 1 mg/kg QD, 3 mg/kg QD, and 10 mg/kg QD treatments significantly inhibiting growth on day 28 relative to vehicle control (TGI of 9.2%, 52.4%, and 69.3%, and p<0.05 for 10 and 30 mg/kg groups, respectively). All doses and regimens were well tolerated with no significant body weight loss.
Compound (I) was given orally once daily for the duration of the study. Data represent the mean ± SEM (Tumor Volume) or mean ± SEM (Body Weight) (N=8 for all groups). *p<0.05 versus vehicle control on Day 28 (repeated measures t-test, Holm-Sidak method with α=0.05 without assuming a consistent SD).
WHIM20 Xenograft Studies
The Patient-Derived Xenograft (PDX) tumor model, WHIM20, representing an ESR1-Y537S mutated human ER+ breast cancer is propagated in mice. The tumors are excised and processed to mixed tumor fragments and the fragments are re-implanted subcutaneously into new recipient mice. Solid tumor tissues are depleted of necrotic components, cut into fragments, mixed with matrigel and subcutaneously implanted into the right flank of 6-8 week old female SCID-bg mice. The precise number of fragments and volume of matrigel are determined on a case by case basis. When the average tumor volume reaches approximately 200 mm3, animals are randomized prior to treatment. All of the primary human tumors utilized in this study undergo approximately 4 passages in vivo.
Estrogen is not supplemented in WHIM20 studies. Compounds are dosed orally every day at the indicated doses. Each treatment is started on Day 0 and the administration schedule is continued for the indicated days. The administration volume is calculated from the individual mouse body weights prior to dose administration. The body weights are measured daily while the tumor volumes are measured twice a week. Tumor volumes are calculated based on the previously described formula.
ERα WT PDX Xenograft Studies
A PDX tumor model representing an ESR1-WT human ER+ breast cancer, designated as PDX-WT, was propagated subcutaneously in immunocompromised mice. The tumors were excised within 60 days of implantation and processed to mixed tumor fragments. Solid tumor tissues were depleted of necrotic components, cut into 70 mg fragments, mixed with matrigel and subcutaneously implanted into the right flank of 6-12 week old female athymic Nude (Crl:NU(NCr)-Foxn1nu) mice. The precise number of fragments and volume of matrigel were determined on a case by case basis. When the average tumor volume reached approximately 125-250 mm3, animals were randomized prior to treatment.
Estrogen was supplemented for the duration of the study. Compound (I) was dosed orally every day at doses ranging from 1 to 30 mg/kg. The administration volume was calculated from the individual mouse body weights prior to dose administration. The body weights (BW) and tumor volumes (TV) were measured twice a week.
Results for ERα WT PDX Xenograft Studies
Figure 37 shows the anti-tumor and body weight effects of compound (I), prepared as an HCl salt, in a PDX model bearing ERαWT/WT xenograft. Compound (I) dosed daily inhibited xenograft growth in a dose dependent manner with 1 mg/kg QD, 3 mg/kg QD, 10 mg/kg QD, and 30 mg/kg QD treatments significantly inhibiting growth on day 39 relative to vehicle control (TGI of 78.5%, 92.3%, 93.1%, and 90.7%, and p<0.05, respectively). All doses and regimens were well tolerated with no significant body weight loss.
Compound (I) was given orally once daily for the duration of the study. Data represent the mean ± SEM (Tumor Volume) or mean ± SEM (Body Weight) (N=6 for all groups). *p<0.05 versus vehicle control on Day 39 (repeated measures t-test, Holm-Sidak method with α=0.05 without assuming a consistent SD).
(Test Example 3) Time-Dependent Inhibition Assays
To demonstrate that a compound is (or is not) a CYP inactivator, abbreviated experimental designs are commonly used for screening in drug development. One of these approaches uses multiple test compound concentrations at a single incubation time, e.g. the “IC50 shift” approach. In an IC50 shift experiment, the IC50 is determined for a CYP marker activity before and after the test compound has been incubated with enzyme and the co-factor nicotinamide adenine dinucleotide phosphate (NADPH) for a set preincubation time (Grimm et al, 2009).
The IC50 shift approach was used to determine whether compounds are time-dependent inhibitors of human CYP3A4 using liver microsomes (0.1 mg/mL). A 30-minute pre-incubation time point was selected, where compounds (9 concentrations, 0 to 30 μmol/L) are incubated at 37°C in presence and absence of 1 mmol/L NADPH. Following the pre-incubation period, 5 μmol/L midazolam (the probe substrate) was added and formation of hydroxymidazolam was measured by high-performance liquid chromatography-mass spectrometry (LC-MS/MS) analysis following a 5 minute incubation period. Any decrease in the formation of hydroxymidazolam, in peak area ratios to vehicle control, was used to calculate three IC50 values (0 min pre-incubation, 30 min pre-incubation with NADPH, and 30 min pre-incubation without NADPH). Whereas an IC50 shift less than 3-fold is accepted as demonstration that a test compound does not possess a TDI risk, an IC50 shift 3-fold or more after pre-incubation is indicative of a CYP3A4 TDI risk. Assays were performed in duplicate, and mifepristone was used as positive control. Results are shown in Table 2. In Table 2, “Yes” indicates a TDI shift greater than or equal to 3. “No” indicates a TDI shift greater than or equal to 1 and less than 3.
Figure JPOXMLDOC01-appb-T000109
By way of comparison with the results above, a TDI assay was also conducted for the following compound, which is reported as Compound 69 of PCT International Application Publication No. WO/2016/196346:
Figure JPOXMLDOC01-appb-C000110
 The IC50 shift approach was also used to determine whether Compound 69 of PCT International Application Publication No. WO/2016/196346 is a time-dependent inhibitor of human CYP3A4 using liver microsomes. Similar to the experimental design described above, a 30-minute pre-incubation time point was selected. Minor variations to the method described above were that Compound 69 of PCT International Application Publication No. WO/2016/196346 was tested at 8 concentrations (0 to 10 μmol/L). Following the pre-incubation period, 3 μmol/L midazolam was added and formation of hydroxymidazolam was measured following a 2 minute incubation period. Since under these experimental conditions, the formation of hydroxymidazolam followed first order kinetics (considered as a linear process), these changes in assay design are not expected to impact the IC50 determinations. In parallel, the IC50 shift assay was also performed with 15.6 μmol/L testosterone (0.05 mg/mL liver microsomes, 10 minute incubation) as second probe substrate. Assays for both probe substrates were performed in triplicate, and mifepristone was used as positive control.
The CYP3A4 TDI shift result for Compound 69 of PCT International Application Publication No. WO/2016/196346 was greater than 3, indicating it to be a TDI risk.

Claims (68)

  1. A salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, and an acid selected from the group consisting of hydrobromic acid, phosphoric acid, sulfuric acid, benzenesulfonic acid, p-toluenesulfonic acid, succinic acid, maleic acid, mandelic acid and methanesulfonic acid.
    Figure JPOXMLDOC01-appb-C000001
  2. A crystal of hydrochloride salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 12.2°, 16.8°, 18.2°, 19.7° and 23.3° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000002
  3. The crystal according to claim 2, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 19.7° in a powder X-ray diffractometry.
  4. The crystal according to claim 2, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 16.8°, 18.2° and 19.7° in a powder X-ray diffractometry.
  5. A crystal of hydrochloride salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having peaks at chemical shifts (± 0.5 ppm) of 144.5 ppm, 156.8 ppm and 165.8 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000003
  6. A crystal of hydrochloride salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 1.
    Figure JPOXMLDOC01-appb-C000004
  7. A crystal of hydrochloride salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 12.
    Figure JPOXMLDOC01-appb-C000005
  8. A crystal of hydrobromide salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 18.4°, 19.7°, 21.6°, 22.9° and 25.6° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000006
  9. The crystal according to claim 8, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.4° in a powder X-ray diffractometry.
  10. The crystal according to claim 8, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 18.4°, 19.7° and 25.6° in a powder X-ray diffractometry.
  11. A crystal of hydrobromide salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 144.2 ppm, 156.7 ppm and 165.2 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000007
  12. A crystal of hydrobromide salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 2.
    Figure JPOXMLDOC01-appb-C000008
  13. A crystal of hydrobromide salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 13.
    Figure JPOXMLDOC01-appb-C000009
  14. A crystal of phosphate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 6.5°, 13.5°, 16.7°, 19.9° and 20.4° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000010
  15. The crystal according to claim 14, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 6.5° in a powder X-ray diffractometry.
  16. The crystal according to claim 14, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 6.5°, 16.7° and 19.9° in a powder X-ray diffractometry.
  17. A crystal of phosphate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 135.3 ppm, 140.9 ppm and 169.8 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000011
  18. A crystal of phosphate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 3.
    Figure JPOXMLDOC01-appb-C000012
  19. A crystal of phosphate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 14.
    Figure JPOXMLDOC01-appb-C000013
  20. A crystal of sulfate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 7.6°, 11.3°, 13.0°, 19.0° and 19.8° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000014
  21. The crystal according to claim 20, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 19.0° in a powder X-ray diffractometry.
  22. The crystal according to claim 20, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 7.6°, 19.0° and 19.8° in a powder X-ray diffractometry.
  23. A crystal of sulfate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 117.8 ppm, 157.4 ppm and 165.0 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000015
  24. A crystal of sulfate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 4.
    Figure JPOXMLDOC01-appb-C000016
  25. A crystal of sulfate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 15.
    Figure JPOXMLDOC01-appb-C000017
  26. A crystal of benzenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 10.9°, 17.6°, 19.9°, 21.9° and 24.1° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000018
  27. The crystal according to claim 26, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 10.9° in a powder X-ray diffractometry.
  28. The crystal according to claim 26, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 10.9°, 17.6° and 24.1° in a powder X-ray diffractometry.
  29. A crystal of benzenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 136.1 ppm, 156.7 ppm and 164.4 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000019
  30. A crystal of benzenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 5.
    Figure JPOXMLDOC01-appb-C000020
  31. A crystal of benzenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 16.
    Figure JPOXMLDOC01-appb-C000021
  32. A crystal of p-toluenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 8.9°, 12.5°, 21.4°, 22.1° and 23.4° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000022
  33. The crystal according to claim 32, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 8.9° in a powder X-ray diffractometry.
  34. The crystal according to claim 32, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 8.9°, 21.4° and 22.1° in a powder X-ray diffractometry.
  35. A crystal of p-toluenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 140.7 ppm, 156.4 ppm and 166.9 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000023
  36. A crystal of p-toluenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 6.
    Figure JPOXMLDOC01-appb-C000024
  37. A crystal of p-toluenesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 17.
    Figure JPOXMLDOC01-appb-C000025
  38. A crystal of succinate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 14.8°, 17.0°, 17.5°, 18.2° and 19.1° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000026
  39. The crystal according to claim 38, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.2° in a powder X-ray diffractometry.
  40. The crystal according to claim 38, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 17.0°, 18.2° and 19.1° in a powder X-ray diffractometry.
  41. A crystal of succinate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 156.1 ppm, 175.0 ppm and 181.3 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000027
  42. A crystal of succinate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 7.
    Figure JPOXMLDOC01-appb-C000028
  43. A crystal of succinate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 18.
    Figure JPOXMLDOC01-appb-C000029
  44. A crystal form A of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 5.4°, 11.6°, 16.8°, 18.9° and 20.2° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000030
  45. The crystal according to claim 44, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.9° in a powder X-ray diffractometry.
  46. The crystal according to claim 44, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 16.8°, 18.9° and 20.2° in a powder X-ray diffractometry.
  47. A crystal form A of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 43.4 ppm, 44.8 ppm and 145.0 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000031
  48. A crystal form A of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 8.
    Figure JPOXMLDOC01-appb-C000032
  49. A crystal form A of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 19.
    Figure JPOXMLDOC01-appb-C000033
  50. A crystal form B of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 10.4°, 15.2°, 19.4°, 20.8° and 22.2° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000034
  51. The crystal according to claim 50, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 19.4° in a powder X-ray diffractometry.
  52. The crystal according to claim 50, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 15.2°, 19.4° and 20.8° in a powder X-ray diffractometry.
  53. A crystal form B of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 25.8 ppm, 143.6 ppm and 166.8 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000035
  54. A crystal form B of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 9.
    Figure JPOXMLDOC01-appb-C000036
  55. A crystal form B of maleate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 20.
    Figure JPOXMLDOC01-appb-C000037
  56. A crystal of L-mandelate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 5.1°, 8.7°, 10.3°, 18.2° and 20.3° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000038
  57. The crystal according to claim 56, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 18.2° in a powder X-ray diffractometry.
  58. The crystal according to claim 56, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 5.1°, 10.3° and 18.2° in a powder X-ray diffractometry.
  59. A crystal of L-mandelate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 128.1 ppm, 156.7 ppm and 177.8 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000039
  60. A crystal of L-mandelate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 10.
    Figure JPOXMLDOC01-appb-C000040
  61. A crystal of L-mandelate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 21.
    Figure JPOXMLDOC01-appb-C000041
  62. A crystal of methanesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having one or more diffraction peaks at diffraction angles (2θ ± 0.2°) selected from the group consisting of 12.7°, 18.9°, 19.7°, 21.8° and 25.6° in a powder X-ray diffractometry.
    Figure JPOXMLDOC01-appb-C000042
  63. The crystal according to claim 62, characterized by having a diffraction peak at a diffraction angle (2θ ± 0.2°) of 12.7° in a powder X-ray diffractometry.
  64. The crystal according to claim 62, characterized by having diffraction peaks at diffraction angles (2θ ± 0.2°) of 12.7°, 18.9° and 25.6° in a powder X-ray diffractometry.
  65. A crystal of methanesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, which is characterized by having peaks at chemical shifts (± 0.5 ppm) of 40.4 ppm, 158.3 ppm and 166.0 ppm in a solid state 13C NMR spectrum.
    Figure JPOXMLDOC01-appb-C000043
  66. A crystal of methanesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a powder X-ray diffraction pattern substantially the same as the powder X-ray diffraction pattern shown in Figure 11.
    Figure JPOXMLDOC01-appb-C000044
  67. A crystal of methanesulfonate salt of (E)-N-methyl-4-((2-(4-((E)-4,4,4-trifluoro-1-(3-fluoro-1H-indazol-5-yl)-2-phenylbut-1-en-1-yl)phenoxy)ethyl)amino)but-2-enamide represented by the formula I, characterized by having a solid state 13C NMR spectrum substantially the same as the solid state 13C NMR spectrum shown in Figure 22.
    Figure JPOXMLDOC01-appb-C000045
  68. A pharmaceutical composition comprising the salt or the crystal thereof according to any one of claims 1 to 67.
PCT/JP2019/019947 2018-05-22 2019-05-20 Salts of indazole derivative and crystals thereof Ceased WO2019225552A1 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2018097653 2018-05-22
JP2018-097653 2018-05-22

Publications (1)

Publication Number Publication Date
WO2019225552A1 true WO2019225552A1 (en) 2019-11-28

Family

ID=68616803

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP2019/019947 Ceased WO2019225552A1 (en) 2018-05-22 2019-05-20 Salts of indazole derivative and crystals thereof

Country Status (1)

Country Link
WO (1) WO2019225552A1 (en)

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012037411A2 (en) * 2010-09-16 2012-03-22 Aragon Pharmaceuticals, Inc. Estrogen receptor modulators and uses thereof
WO2013142266A1 (en) * 2012-03-20 2013-09-26 Seragon Pharmaceuticals, Inc. Estrogen receptor modulators and uses thereof
WO2016196346A1 (en) * 2015-05-29 2016-12-08 Eisai R&D Management Co., Ltd. Tetrasubstituted alkene compounds and their use
US20180141913A1 (en) * 2016-11-24 2018-05-24 Eisai R & D Management Co., Ltd. Tetrasubstituted alkene compounds and their use
WO2018097273A1 (en) * 2016-11-28 2018-05-31 Eisai R&D Management Co., Ltd. Salts of indazole derivative and crystals thereof

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012037411A2 (en) * 2010-09-16 2012-03-22 Aragon Pharmaceuticals, Inc. Estrogen receptor modulators and uses thereof
WO2013142266A1 (en) * 2012-03-20 2013-09-26 Seragon Pharmaceuticals, Inc. Estrogen receptor modulators and uses thereof
WO2016196346A1 (en) * 2015-05-29 2016-12-08 Eisai R&D Management Co., Ltd. Tetrasubstituted alkene compounds and their use
US20180141913A1 (en) * 2016-11-24 2018-05-24 Eisai R & D Management Co., Ltd. Tetrasubstituted alkene compounds and their use
WO2018097273A1 (en) * 2016-11-28 2018-05-31 Eisai R&D Management Co., Ltd. Salts of indazole derivative and crystals thereof

Similar Documents

Publication Publication Date Title
EP3544967B1 (en) Salts of indazole derivative and crystals thereof
JP6744489B2 (en) 6,7,8,9-Tetrahydro-3H-pyrazolo[4,3-f]isoquinoline derivatives useful in the treatment of cancer
EP3814341B1 (en) Erbb receptor inhibitors
AU2014208964B2 (en) Chemical compounds
AU2017230437B2 (en) Urea compound, preparation method therefor and medical use thereof
US20240327384A1 (en) Salts and solid state forms of a kif18a inhibitor compound
BR112016022060B1 (en) COMPOUND, PROCESS FOR PREPARING A COMPOUND, PHARMACEUTICAL COMPOSITION, AND USE OF A COMPOUND
PL208401B1 (en) Piperazinyl-, piperidinyl- and morpholinyl-derivatives as novel inhibitors of histone deacetylase
TWI863667B (en) Allosteric chromenone inhibitors of phosphoinositide 3-kinase (pi3k) for the treatment of disease
US20200246365A1 (en) Compounds, pharmaceutical compositions and use thereof as inhibitors of ran gtpase
CN110467637B (en) Bisaminyl chloropyrimidine compound containing phosphine oxide substituted aniline, preparation method and application thereof
US20240034742A1 (en) Dosing regimens for cyclin-dependent kinase 7 (cdk7) inhibitors
BR112019008489B1 (en) SALTS OF INDAZOLE DERIVATIVE AND CRYSTALS THEREOF
CN120192251A (en) A sulfonamide compound for treating prostate cancer and its application
HK40104230A (en) Salt and crystal forms of an epidermal growth factor receptor inhibitor
HK1214592B (en) Chemical compounds

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: 19807604

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

NENP Non-entry into the national phase

Ref country code: JP

122 Ep: pct application non-entry in european phase

Ref document number: 19807604

Country of ref document: EP

Kind code of ref document: A1