WO2022166866A1 - Composé macrocyclique de dihydropyrazine et de pyrazine - Google Patents

Composé macrocyclique de dihydropyrazine et de pyrazine Download PDF

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WO2022166866A1
WO2022166866A1 PCT/CN2022/074894 CN2022074894W WO2022166866A1 WO 2022166866 A1 WO2022166866 A1 WO 2022166866A1 CN 2022074894 W CN2022074894 W CN 2022074894W WO 2022166866 A1 WO2022166866 A1 WO 2022166866A1
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compound
mmol
ethyl acetate
pharmaceutically acceptable
acceptable salt
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PCT/CN2022/074894
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Chinese (zh)
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陈新海
胡伯羽
于衍新
陈兆国
肖敏亮
李霜训
陈曙辉
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南京明德新药研发有限公司
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/4985—Pyrazines or piperazines ortho- or peri-condensed with heterocyclic ring systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/06—Peri-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D491/00—Heterocyclic compounds containing in the condensed ring system both one or more rings having oxygen atoms as the only ring hetero atoms and one or more rings having nitrogen atoms as the only ring hetero atoms, not provided for by groups C07D451/00 - C07D459/00, C07D463/00, C07D477/00 or C07D489/00
    • C07D491/02—Heterocyclic compounds containing in the condensed ring system both one or more rings having oxygen atoms as the only ring hetero atoms and one or more rings having nitrogen atoms as the only ring hetero atoms, not provided for by groups C07D451/00 - C07D459/00, C07D463/00, C07D477/00 or C07D489/00 in which the condensed system contains two hetero rings
    • C07D491/06—Peri-condensed systems

Definitions

  • the present invention relates to a class of macrocyclic compounds comprising a dihydropyrazinopyrazine structure, and specifically discloses a compound represented by formula (I) and a pharmaceutically acceptable salt thereof.
  • the present invention relates to two signaling pathways that play a key role in tumor proliferation, invasion and metastasis and anti-apoptosis, namely phosphatidylinositol 3 kinase (PI3K)-AKT-mammalian rapamycin protease mTOR signaling pathway and tumor cells Strongly dependent non-homologous DNA double-strand end joining (NHEJ) repairs DNA double-strand breaks (DSBs).
  • PI3K phosphatidylinositol 3 kinase
  • NHEJ non-homologous DNA double-strand end joining
  • mTOR is a highly conserved protein belonging to the serine/threonine protein kinase PIKK family, which also includes ATM (ataxia-telangiectasia-mutated), ATR (ataxia-telangiectasia and Rad3-related) and DNA-PKcs ( DNA-dependent protein kinase).
  • mTOR is a protein that can affect the ability of cell volume expansion. There are at least two different complexes of mTOR in the cell.
  • mTOR rapamycin-sensitive complex
  • mTORC1 rapamycin-sensitive complex
  • the main function of mTORC1 is to regulate protein synthesis and cell cycle progression, while mTORC2 plays an important role in actin cytoskeleton organization and cell survival.
  • the progress of tumor research has made us realize that different tumors have different genetic alterations and signaling pathway alterations.
  • An effective tumor treatment strategy will require genetic alterations and signaling pathway alterations tailored to individual characteristics.
  • mTOR signaling pathway In a variety of solid tumors, such as breast, prostate, lung, colon, pancreas, liver, stomach, colorectal, kidney, thyroid, meningitis and acute and chronic lymphocytic leukemia, Merkel cell tumor Wait. And it is closely related to treatment tolerance and poor prognosis, and can be used as a therapeutic target for these tumors.
  • DNA-PK is a nuclear serine/threonine protein kinase and a member of the phosphatidylinositol-3-kinase family. It consists of a complex composed of the large catalytic subunit DNA-PKcs and a Ku protein heterodimer (Ku70/80) as a regulatory factor. It plays an important role in cell signal transduction and cell cycle functions after DNA damage. In DNA damage, DNA double strand break (DNA double strand break, DSB) is the most deadly, and DSB is repaired mainly through non-homologous end joining (DNA-PK) dominated by DNA-dependent protein kinase (DNA-PK). joining, NHEJ).
  • Radiation and a number of anticancer drugs can directly or indirectly act on DNA or DNA metabolism, resulting in DNA damage and triggering a series of cellular reactions such as damaged DNA repair.
  • the result of repair is to improve cell survival, which is also a tumor.
  • DNA-PK is highly expressed in tumor tissues after radiotherapy, and the continuous repair of radiotherapy-damaged tumor cells has become one of the main reasons for radiotherapy and chemotherapy resistance.
  • DNA-PK signaling pathway By inhibiting the DNA-PK signaling pathway, it is expected to increase the effect of chemoradiotherapy. Therefore, DNA-PK is expected to become another target for the treatment of tumors.
  • DNA-PK not only contributes to the survival of tumor cells, but also activates and phosphorylates AKT protein kinase, which indirectly activates the mTOR signaling pathway, which may be involved in the clinical drug resistance of mTOR inhibitors.
  • the present invention aims to invent a class of pyrazinone macrocyclic small molecules to target mTOR and DNA-PK, and to treat various hematological tumors including non-Hodgkin's lymphoma by single drug or in combination with chemotherapeutic drugs.
  • Treatment of malignant solid tumors including lung cancer, prostate cancer, colorectal cancer, head and neck cancer, pancreatic cancer, stem cell cancer and brain cancer.
  • Such compounds have very excellent inhibitory activities against DNA-PK and mTOR, and exhibit excellent effects and effects, and have broad application prospects.
  • Celgene WO2010/062571A1 discloses compound CC-115, which belongs to mTOR/DNA-PK dual kinase target inhibitor, and its structural formula is as follows:
  • the present invention provides a compound of formula (I) or a pharmaceutically acceptable salt thereof,
  • X is selected from N and CH;
  • L is selected from -(CR 6 R 7 ) 4 -, -(CR 6 R 7 ) 5 -, -(CR 6 R 7 ) 6 -, -(CR 6 R 7 ) 7 -, -(CR 6 R 7 ) 8 - and -CH 2 -CH 2 -O-CH 2 -CH 2 -;
  • L 1 and L 3 are each independently selected from -(CR 8 R 9 )-, -(CR 8 R 9 ) 2 -, -(CR 8 R 9 ) 3 -, -(CR 8 R 9 ) 4 -, - (CR 8 R 9 ) 5 -, -(CR 8 R 9 ) 6 -;
  • R 1 is selected from the group consisting of C 1-6 alkyl and 5-6 membered heteroaryl, each of which is independently optionally surrounded by 1, 2 or 3 R b replace;
  • R 2 and R 3 are each independently selected from H, F, Cl, Br, I, CN, C 1-4 alkyl and C 3-6 cycloalkyl, said C 1-4 alkyl and C 3-6
  • the cycloalkyl groups are each independently optionally substituted with 1, 2 or 3 R c ;
  • R 4 and R 5 are each independently selected from H, F, Cl, Br, I, OH, CN, NH 2 , CH 3 , OCH 3 , CH 2 CH 3 and CF 3 ;
  • R 6 and R 7 are each independently selected from H, F, Cl, Br, I and C 1-3 alkyl optionally substituted with 1 , 2 or 3 R d ;
  • R 6 and R 7 attached to the same carbon atom are each independently optionally attached to form a C 3-6 cycloalkyl optionally substituted with 1, 2 or 3 Rs;
  • R is selected from H, F, Cl, Br, I, OH, CN, NH 2 , C 1-6 alkyl, C 1-6 alkoxy and C 1-6 alkylamino, the C 1-6 alkyl , C 1-6 alkoxy and C 1-6 alkylamino are optionally substituted by 1, 2 or 3 R e ;
  • R 8 and R 9 are each independently selected from H, F, Cl, Br, I, OH, CN, NH 2 , C 1-6 alkyl, C 1-6 alkoxy and C 1-6 alkylamino, so Said C 1-6 alkyl, C 1-6 alkoxy and C 1-6 alkylamino are optionally substituted with 1, 2 or 3 R f ;
  • Ra , Rb , Rc , Rd , Re , and Rf are each independently selected from F, Cl , Br, I, OH, CN, NH2 , CH3 , OCH3 , CH2CH3 , and CF3 ;
  • the 5-6 membered heteroaryl groups respectively comprise 1, 2, 3 or 4 heteroatoms or heteroatomic groups independently selected from -O-, -NH-, -S- and N.
  • R 8 and R 9 are independently selected from H, F, Cl, Br, I, OH, CN, NH 2 , C 1-3 alkyl, C 1-3 alkoxy and C 1-3 alkylamino, the C 1-3 alkyl, C 1-3 alkoxy and C 1-3 alkylamino are optionally substituted with 1, 2 or 3 R f , R f and other variables such as as defined in the present invention.
  • R 8 and R 9 are independently selected from H, F, Cl , Br, I, OH, CN, NH 2 and CH 3 , and other The variables are as defined in the present invention.
  • the above L 1 and L 3 are each independently selected from -(CH 2 ) 2 -, -(CH 2 ) 3 -, -(CH 2 ) 4 -, -(CH 2 ) 5 - , -(CH 2 ) 6 -, -(CH 2 ) 2 -CF 2 -, -(CH 2 ) 2 -CHF-, -(CH 2 ) 3 -C(CH 3 ) 2 - and -(CH 2 ) 3 -CF 2 -, other variables are as defined in the present invention.
  • the above R 1 is selected from methyl, ethyl, n-propyl, isopropyl, pyrazolyl, 1H-1,2,4-triazolyl and 4H-1,2, 4-Triazolyl, the methyl, ethyl, n-propyl, isopropyl, pyrazolyl, 1H-1,2,4-triazolyl and 4H-1,2,4-triazolyl
  • the oxazolyl groups are each independently optionally substituted with 1, 2 or 3 R b , R b and other variables as defined herein.
  • R 1 is selected from Other variables are as defined in the present invention.
  • R 2 and formula R 3 are each independently selected from H, F, Cl, Br, I, CN, C 1-3 alkyl, and the C 1-3 alkyl is optionally 1, 2 or 3 R c substitutions, R c and other variables are as defined herein.
  • R 2 and R 3 are independently selected from H and F, respectively, and other variables are as defined herein.
  • R 1 is selected from pyrazolyl, 1H-1,2,4-triazolyl and 4H-1,2,4-triazolyl
  • the pyrazolyl, 1H -1,2,4-triazolyl and 4H-1,2,4-triazolyl are optionally substituted with 1, 2 or 3 R a , R a and other variables as defined in the present invention.
  • R 1 is selected from Other variables are as defined in the present invention.
  • R 6 and R 7 are independently selected from H, F, Cl, Br, I and CH 3 , and other variables are as defined in the present invention.
  • R are independently selected from H, F, Cl, Br, I, OH, CN, NH 2 , CH 3 , OCH 3 , CH 2 CH 3 and CF 3 , and other variables are as in invention is defined.
  • the above-mentioned R 6 and R 7 attached to the same carbon atom are each independently optionally attached to form a cyclopropyl optionally substituted with 1, 2 or 3 R, R and other variables as defined in the present invention.
  • the above-mentioned R 6 and R 7 attached to the same carbon atom are each independently optionally linked to form a cyclopropyl group, and other variables are as defined in the present invention.
  • the above L is selected from -(CH 2 ) 4 -, -(CH 2 ) 5 -, -(CH 2 ) 6 -, -(CH 2 ) 7 -, -(CH 2 ) 8 -, -CH 2 -CH 2 -O-CH 2 -CH 2 -and
  • Other variables are as defined in the present invention.
  • the compound represented by the above formula (I) is selected from
  • X, R 2 , R 3 , L 1 , L 2 and L 3 are as defined in the present invention.
  • the compound represented by the above formula (I) is selected from
  • R 2 , R 3 , R 4 , R 5 and L are as defined in the present invention.
  • the present invention also provides a compound or a pharmaceutically acceptable salt thereof, wherein the compound is selected from
  • the present invention also provides the use of the compound or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating mTOR and DNA-PK related diseases.
  • the present invention also provides the crystal form A of compound W1 hydrochloride
  • the X-ray powder diffraction pattern of Form A of the above-mentioned compound W1 hydrochloride salt has diffraction peaks at the following 2 ⁇ angles: 4.60 ⁇ 0.20°, 9.12 ⁇ 0.20°, 17.78 ⁇ 0.20°, 21.56 ⁇ 0.20°, 27.00 ⁇ 0.20°.
  • the X-ray powder diffraction pattern of Form A of the above-mentioned compound W1 hydrochloride salt has diffraction peaks at the following 2 ⁇ angles: 4.60 ⁇ 0.20°, 9.12 ⁇ 0.20°, 17.78 ⁇ 0.20°, 21.56 ⁇ 0.20°, 23.92 ⁇ 0.20°, 26.12 ⁇ 0.20°, 27.00 ⁇ 0.20°, 27.84 ⁇ 0.20°.
  • the X-ray powder diffraction pattern of Form A of Compound W1 hydrochloride above has diffraction peaks at the following 2 ⁇ angles: 4.60 ⁇ 0.20°, 9.12 ⁇ 0.20°, 17.78 ⁇ 0.20°, and /or 8.00 ⁇ 0.20°, and/or 10.32 ⁇ 0.20°, and/or 13.56 ⁇ 0.20°, and/or 15.28 ⁇ 0.20°, and/or 16.02 ⁇ 0.20°, and/or 18.16 ⁇ 0.20°, and/or 19.94 ⁇ 0.20°, and/or 20.72 ⁇ 0.20°, and/or 21.08 ⁇ 0.20°, and/or 21.56 ⁇ 0.20°, and/or 22.34 ⁇ 0.20°, and/or 23.58 ⁇ 0.20°, and/or 23.92 ⁇ 0.20°, and/or 25.30 ⁇ 0.20°, and/or 26.12 ⁇ 0.20°, and/or 27.00 ⁇ 0.20°, and/or 27.84 ⁇ 0.20°, and/or 28.90 ⁇
  • the X-ray powder diffraction pattern of Form A of the above-mentioned compound W1 hydrochloride salt has diffraction peaks at the following 2 ⁇ angles: 4.60 ⁇ 0.20°, and/or 8.00 ⁇ 0.20°, and/or 9.12 ⁇ 0.20°, and/or 10.32 ⁇ 0.20°, and/or 13.56 ⁇ 0.20°, and/or 15.28 ⁇ 0.20°, and/or 16.02 ⁇ 0.20°, and/or 17.78 ⁇ 0.20°, and/or 18.16 ⁇ 0.20°, and/or 19.94 ⁇ 0.20°, and/or 20.72 ⁇ 0.20°, and/or 21.08 ⁇ 0.20°, and/or 21.56 ⁇ 0.20°, and/or 22.34 ⁇ 0.20°, and/or 23.58 ⁇ 0.20° , and/or 23.92 ⁇ 0.20°, and/or 25.30 ⁇ 0.20°, and/or 26.12 ⁇ 0.20°, and/or 27.00 ⁇ 0.20°, and/or 27.84
  • the X-ray powder diffraction pattern of the crystal form A of the above-mentioned compound W1 hydrochloride has diffraction peaks at the following 2 ⁇ angles: 4.598°, 7.999°, 9.115°, 10.321°, 13.561°, 15.282 degrees 30.046°, 30.711°, 31.361°, 32.201°, 32.961°, 33.574°, 36.080° and 38.320°.
  • the X-ray powder diffraction spectrum of the crystal form A of the above-mentioned compound W1 hydrochloride is shown in FIG. 1 .
  • the above-mentioned compound W1 hydrochloride has the following structure:
  • x is selected from 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9 and 2.0.
  • the above x is selected from 1.8.
  • the present invention also provides compounds of formula (II)
  • x is selected from 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9 and 2.0.
  • the above x is selected from 1.8.
  • the present invention also provides compound W1 crystal form B
  • the X-ray powder diffraction pattern of the above compound W1 crystal form B has diffraction peaks at the following 2 ⁇ angles: 9.02 ⁇ 0.20°, 13.96 ⁇ 0.20°, 19.24 ⁇ 0.20°, 20.76 ⁇ 0.20°, 23.32 ⁇ 0.20°, 24.72 ⁇ 0.20°, 25.76 ⁇ 0.20°, 30.12 ⁇ 0.20°.
  • the X-ray powder diffraction pattern of the above-mentioned Compound W1 Form B has diffraction peaks at the following 2 ⁇ angles: 9.02 ⁇ 0.20°, 13.96 ⁇ 0.20°, 19.24 ⁇ 0.20°, and/or 4.72 ⁇ 0.20°, and/or 10.80 ⁇ 0.20°, and/or 11.80 ⁇ 0.20°, and/or 15.20 ⁇ 0.20°, and/or 17.88 ⁇ 0.20°, and/or 18.60 ⁇ 0.20°, and/or 20.12 ⁇ 0.20° , and/or 20.76 ⁇ 0.20°, and/or 21.66 ⁇ 0.20°, and/or 22.02 ⁇ 0.20°, and/or 23.32 ⁇ 0.20°, and/or 24.72 ⁇ 0.20°, and/or 25.76 ⁇ 0.20°, and /or 26.44 ⁇ 0.20°, and/or 26.96 ⁇ 0.20°, and/or 27.98 ⁇ 0.20°, and/or 29.14 ⁇ 0.20°, and/or
  • the X-ray powder diffraction pattern of the above compound W1 crystal form B has diffraction peaks at the following 2 ⁇ angles: 9.02 ⁇ 0.20°, and/or 4.72 ⁇ 0.20°, and/or 10.80 ⁇ 0.20° , and/or 11.80 ⁇ 0.20°, and/or 13.96 ⁇ 0.20°, and/or 15.20 ⁇ 0.20°, and/or 17.88 ⁇ 0.20°, and/or 18.60 ⁇ 0.20°, and/or 19.24 ⁇ 0.20°, and /or 20.12 ⁇ 0.20°, and/or 20.76 ⁇ 0.20°, and/or 21.66 ⁇ 0.20°, and/or 22.02 ⁇ 0.20°, and/or 23.32 ⁇ 0.20°, and/or 24.72 ⁇ 0.20°, and/or 25.76 ⁇ 0.20°, and/or 26.44 ⁇ 0.20°, and/or 26.96 ⁇ 0.20°, and/or 27.98 ⁇ 0.20°, and/or 29.14 ⁇ 0.20°, and/
  • the X-ray powder diffraction pattern of the above compound W1 crystal form B has diffraction peaks at the following 2 ⁇ angles: 4.719°, 9.019°, 10.801°, 11.799°, 13.960°, 15.202°, 17.880° , 18.600°, 19.241°, 20.121°, 20.761°, 21.657°, 22.022°, 23.321°, 24.719°, 25.760°, 26.440°, 26.960°, 27.980°, 29.141°, 30.118°, 31.002°, 31.624° °, 35.678° and 37.920°.
  • the X-ray powder diffraction pattern of the above-mentioned compound W1 crystal form B is shown in FIG. 2 .
  • the present invention also provides the application of the above-mentioned compound W1 hydrochloride crystal form A and/or compound W1 crystal form B in the preparation of a medicament for the treatment of mTOR and DNA-PK-related diseases.
  • the mTOR and DNA-PK-related disease described above is human glioma, esophageal cancer (esophageal cancer), or gastric cancer.
  • the compounds of the present invention have good inhibitory activities on mTOR and DNA-PK.
  • the term "pharmaceutically acceptable” refers to those compounds, materials, compositions and/or dosage forms that, within the scope of sound medical judgment, are suitable for use in contact with human and animal tissue , without excessive toxicity, irritation, allergic reactions or other problems or complications, commensurate with a reasonable benefit/risk ratio.
  • salts refers to salts of the compounds of the present invention, prepared from compounds with specific substituents discovered by the present invention and relatively non-toxic acids or bases.
  • base addition salts can be obtained by contacting such compounds with a sufficient amount of base in neat solution or in a suitable inert solvent.
  • Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amine or magnesium salts or similar salts.
  • acid addition salts can be obtained by contacting such compounds with a sufficient amount of acid in neat solution or in a suitable inert solvent.
  • Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts including, for example, hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, bicarbonate, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, Hydrogen sulfate, hydroiodic acid, phosphorous acid, etc.; and organic acid salts including, for example, acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, Similar acids such as fumaric, lactic, mandelic, phthalic, benzenesulfonic, p-toluenesulfonic, citric, tartaric, and methanesulfonic acids; also include salts of amino acids such as arginine, etc. , and salts of organic acids such as glucuronic acid. Certain specific compounds of the present invention contain both basic and acidic functional groups and thus can be converted into either base
  • the pharmaceutically acceptable salts of the present invention can be synthesized from the acid or base containing parent compound by conventional chemical methods. Generally, such salts are prepared by reacting the free acid or base form of these compounds with a stoichiometric amount of the appropriate base or acid in water or an organic solvent or a mixture of the two.
  • the compounds of the present invention may contain unnatural proportions of atomic isotopes at one or more of the atoms that constitute the compound.
  • compounds can be labeled with radioisotopes, such as tritium ( 3 H), iodine-125 ( 125 I) or C-14 ( 14 C).
  • deuterated drugs can be formed by replacing hydrogen with deuterium, and the bonds formed by deuterium and carbon are stronger than those formed by ordinary hydrogen and carbon. Compared with non-deuterated drugs, deuterated drugs can reduce toxic side effects and increase drug stability. , enhance the efficacy, prolong the biological half-life of drugs and other advantages. All transformations of the isotopic composition of the compounds of the present invention, whether radioactive or not, are included within the scope of the present invention.
  • substituted means that any one or more hydrogen atoms on a specified atom are replaced by a substituent, which may include deuterium and hydrogen variants, as long as the valence of the specified atom is normal and the substituted compound is stable.
  • oxygen it means that two hydrogen atoms are substituted. Oxygen substitution does not occur on aromatic groups.
  • optionally substituted means that it may or may not be substituted, and unless otherwise specified, the type and number of substituents may be arbitrary on a chemically achievable basis.
  • any variable eg, R
  • its definition in each case is independent.
  • the group may optionally be substituted with up to two Rs, with independent options for R in each case.
  • combinations of substituents and/or variants thereof are permissible only if such combinations result in stable compounds.
  • linking group When the number of a linking group is 0, such as -(CRR) 0 -, it means that the linking group is a single bond.
  • substituents When a substituent is vacant, it means that the substituent does not exist. For example, when X in A-X is vacant, it means that the structure is actually A. When the listed substituents do not indicate through which atom it is attached to the substituted group, such substituents may be bonded through any of its atoms, for example, pyridyl as a substituent may be through any one of the pyridine ring The carbon atom is attached to the substituted group.
  • the direction of attachment is arbitrary, for example,
  • the linking group L in the middle is -MW-, at this time -MW- can connect ring A and ring B in the same direction as the reading order from left to right. It is also possible to connect ring A and ring B in the opposite direction to the reading order from left to right.
  • Combinations of the linking groups, substituents and/or variants thereof are permissible only if such combinations result in stable compounds.
  • any one or more sites in the group can be linked to other groups by chemical bonds.
  • connection method of the chemical bond is not located, and there is an H atom at the linkable site, when the chemical bond is connected, the number of H atoms at the site will be correspondingly reduced with the number of chemical bonds connected to the corresponding valence. the group.
  • the chemical bond connecting the site to other groups can be represented by straight solid line bonds straight dotted key or wavy lines express.
  • a straight solid bond in -OCH 3 indicates that it is connected to other groups through the oxygen atom in this group;
  • the straight dashed bond in the group indicates that it is connected to other groups through the two ends of the nitrogen atom in the group;
  • the wavy line in the phenyl group indicates that it is connected to other groups through the 1 and 2 carbon atoms in the phenyl group;
  • the number of atoms in a ring is generally defined as the number of ring members, eg, "5-7 membered ring” refers to a “ring” of 5-7 atoms arranged around it.
  • Cn-n+m or Cn - Cn+m includes any particular instance of n to n+ m carbons, eg C1-12 includes C1 , C2 , C3, C 4 , C 5 , C 6 , C 7 , C 8 , C 9 , C 10 , C 11 , and C 12 , also including any one range from n to n+m, eg C 1-12 includes C 1-3 , C 1-6 , C 1-9 , C 3-6 , C 3-9 , C 3-12 , C 6-9 , C 6-12 , and C 9-12 , etc.; in the same way, n yuan to n +m-membered means that the number of atoms in the ring is from n to n+m, for example, 3-12-membered ring includes 3-membered ring, 4-membered ring, 5-membered ring, 6-membered ring, 7-membered ring, 8-membered
  • C 1-6 alkyl is used to denote a straight or branched chain saturated hydrocarbon group consisting of 1 to 6 carbon atoms.
  • the C 1-6 alkyl includes C 1-5 , C 1-4 , C 1-3 , C 1-2 , C 2-6 , C 2-4 , C 6 and C 5 alkyl and the like; it can be Is monovalent (eg methyl), divalent (eg methylene) or polyvalent (eg methine).
  • C 1-6 alkyl examples include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), butyl (including n-butyl, isobutyl , s-butyl and t-butyl), pentyl (including n-pentyl, isopentyl and neopentyl), hexyl, etc.
  • C 1-4 alkyl is used to denote a straight or branched chain saturated hydrocarbon group consisting of 1 to 4 carbon atoms.
  • the C 1-4 alkyl includes C 1-2 , C 1-3 and C 2-3 alkyl, etc.; it can be monovalent (such as methyl), divalent (such as methylene) or polyvalent (such as methine).
  • Examples of C 1-4 alkyl include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), butyl (including n-butyl, isobutyl , s-butyl and t-butyl) and so on.
  • C 1-3 alkyl is used to denote a straight or branched chain saturated hydrocarbon group consisting of 1 to 3 carbon atoms.
  • the C 1-3 alkyl group includes C 1-2 and C 2-3 alkyl groups, etc.; it can be monovalent (such as methyl ), divalent (such as methylene) or polyvalent (such as methine) .
  • Examples of C1-3 alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), and the like.
  • the terms “5-6 membered heteroaryl ring” and “5-6 membered heteroaryl” are used interchangeably in the present invention, and the term “5-6 membered heteroaryl” means from 5 to 6 ring atoms It is composed of a monocyclic group with a conjugated ⁇ electron system, wherein 1, 2, 3 or 4 ring atoms are heteroatoms independently selected from O, S and N, and the rest are carbon atoms. Where the nitrogen atom is optionally quaternized, the nitrogen and sulfur heteroatoms may be optionally oxidized (ie, NO and S(O) p , p is 1 or 2).
  • a 5-6 membered heteroaryl group can be attached to the remainder of the molecule through a heteroatom or a carbon atom.
  • the 5-6 membered heteroaryl groups include 5- and 6-membered heteroaryl groups.
  • Examples of the 5-6 membered heteroaryl include, but are not limited to, pyrrolyl (including N-pyrrolyl, 2-pyrrolyl and 3-pyrrolyl, etc.), pyrazolyl (including 2-pyrazolyl and 3-pyrrolyl, etc.) azolyl, etc.), imidazolyl (including N-imidazolyl, 2-imidazolyl, 4-imidazolyl and 5-imidazolyl, etc.), oxazolyl (including 2-oxazolyl, 4-oxazolyl and 5- oxazolyl, etc.), triazolyl (1H-1,2,3-triazolyl, 2H-1,2,3-triazolyl, 1H-1,2,4-triazolyl and 4H-1, 2,4
  • C1-6alkoxy refers to those alkyl groups containing 1 to 6 carbon atoms attached to the remainder of the molecule through an oxygen atom.
  • the C 1-6 alkoxy groups include C 1-4 , C 1-3 , C 1-2 , C 2-6 , C 2-4 , C 6 , C 5 , C 4 and C 3 alkoxy groups, etc. .
  • C 1-6 alkoxy groups include, but are not limited to, methoxy, ethoxy, propoxy (including n - propoxy and isopropoxy), butoxy (including n-butoxy, isobutoxy) oxy, s-butoxy and t-butoxy), pentyloxy (including n-pentyloxy, isopentyloxy and neopentyloxy), hexyloxy and the like.
  • C1-3alkoxy refers to those alkyl groups containing 1 to 3 carbon atoms attached to the remainder of the molecule through an oxygen atom.
  • the C 1-3 alkoxy group includes C 1-2 , C 2-3 , C 3 and C 2 alkoxy and the like.
  • Examples of C 1-3 alkoxy groups include, but are not limited to, methoxy, ethoxy, propoxy (including n-propoxy and isopropoxy), and the like.
  • C 1-6 alkylamino refers to those alkyl groups containing 1 to 6 carbon atoms attached to the remainder of the molecule through an amino group.
  • the C 1-6 alkylamino includes C 1-4 , C 1-3 , C 1-2 , C 2-6 , C 2-4 , C 6 , C 5 , C 4 , C 3 and C 2 alkylamino Wait.
  • C 1-6 alkylamino examples include, but are not limited to, -NHCH 3 , -N(CH 3 ) 2 , -NHCH 2 CH 3 , -N(CH 3 )CH 2 CH 3 , -N(CH 2 CH 3 )( CH2CH3 ) , -NHCH2CH2CH3 , -NHCH2 ( CH3 ) 2 , -NHCH2CH2CH2CH3 , etc.
  • C 1-3 alkylamino refers to those alkyl groups containing 1 to 3 carbon atoms attached to the remainder of the molecule through an amino group.
  • the C 1-3 alkylamino groups include C 1-2 , C 3 and C 2 alkylamino groups and the like.
  • Examples of C 1-3 alkylamino include, but are not limited to, -NHCH 3 , -N(CH 3 ) 2 , -NHCH 2 CH 3 , -N(CH 3 )CH 2 CH 3 , -NHCH 2 CH 2 CH 3 , - NHCH 2 (CH 3 ) 2 and the like.
  • C 3-6 cycloalkyl means a saturated cyclic hydrocarbon group consisting of 3 to 6 carbon atoms, which are monocyclic and bicyclic ring systems, said C 3-6 cycloalkyl including C 3-5 , C 4-5 and C 5-6 cycloalkyl and the like; it may be monovalent, divalent or polyvalent.
  • Examples of C3-6 cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like.
  • the compounds of the present invention can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed in combination with other chemical synthesis methods, and those well known to those skilled in the art Equivalent to alternatives, preferred embodiments include, but are not limited to, the embodiments of the present invention.
  • the structure of the compound of the present invention can be confirmed by conventional methods well known to those skilled in the art. If the present invention relates to the absolute configuration of the compound, the absolute configuration can be confirmed by conventional technical means in the art. For example, single crystal X-ray diffraction method (SXRD), the cultured single crystal is collected by Bruker D8 venture diffractometer, the light source is CuK ⁇ radiation, and the scanning mode is: After scanning and collecting relevant data, the crystal structure was further analyzed by the direct method (Shelxs97), and the absolute configuration could be confirmed.
  • SXRD single crystal X-ray diffraction method
  • the cultured single crystal is collected by Bruker D8 venture diffractometer
  • the light source is CuK ⁇ radiation
  • the scanning mode is: After scanning and collecting relevant data, the crystal structure was further analyzed by the direct method (Shelxs97), and the absolute configuration could be confirmed.
  • the solvent used in the present invention is commercially available.
  • the solvent ratios used in the silica gel column and thin layer chromatography of the present invention are all volume ratios.
  • BOC stands for tert-butoxycarbonyl, which is an amine protecting group
  • Boc 2 O stands for di-tert-butyl dicarbonate
  • DMAP stands for 4-N,N-lutidine
  • HATU stands for 2-(7-benzotriazole oxide)-N,N,N',N'-tetramethylurea hexafluorophosphate
  • DMA stands for N,N-dimethylacetamide
  • DMF stands for N,N - dimethylformamide
  • 4M represents 4 mol/liter
  • Pd(PPh 3 ) 4 represents catalyst tetrakistriphenylphosphine palladium
  • m-CPBA represents 3-chloroperoxybenzoic acid
  • DHP represents 2,3-dihydropyridine pyran
  • THP stands for tetrahydropyran
  • DMF-DMA stands for N,N-dimethyldimethylacetal
  • DCE stands for dichloroethane
  • Test Method Approximately 20 mg of sample was used for XRPD detection.
  • Fig. 1 is the XRPD pattern of the crystal form A of compound W1 hydrochloride.
  • Figure 2 is the XRPD pattern of compound W1 crystal form B.
  • the present invention will be described in detail by the following examples, but it does not mean any unfavorable limitation of the present invention.
  • the compounds of the present invention can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed in combination with other chemical synthesis methods, and those well known to those skilled in the art Equivalent to alternatives, preferred embodiments include, but are not limited to, the embodiments of the present invention. It will be apparent to those skilled in the art that various changes and modifications can be made to the specific embodiments of the present invention without departing from the spirit and scope of the invention.
  • the reaction solution was cooled to 25°C, the mixture was washed three times with 1500 mL of water each time, the aqueous phase was extracted once with dichloromethane (1000 mL), the organic phases were combined, washed once with saturated brine (2000 mL), and dried over anhydrous sodium sulfate. , filtered and concentrated.
  • Phthalimide potassium salt (11.51 g, 62.12 mmol), WB-8 (10 g, 56.47 mmol) was added to 100 mL of DMF and stirred at 70°C for 18 hours. Filter, pour the filtrate into 100 mL of water, and extract three times with ethyl acetate (30 mL), combine the organic phases, wash once with 50 mL of saturated brine, dry over anhydrous sodium sulfate, filter, and concentrate to obtain crude product WB- 9. (M+H: 244.2)
  • WB-9 (10 g, 41.10 mmol) was dissolved in 150 mL of absolute ethanol, hydrazine monohydrate (6.30 g, 123.30 mmol, 6.12 mL, purity 98%) was added, and then the mixture was heated to 60° C. Stir at 60°C for 18 hours.
  • the hydrochloride salt WB-10 (5.30 g, 35.40 mmol) was dissolved in 100 mL of N-methylpyrrolidone, N,N-diisopropylethylamine (15.25 g, 118.80 mmol) was added, and the solution was heated at 25°C. After stirring for 10 minutes, B-7 (10 g, 29.50 mmol) was added, heated to 130°C and stirring continued for 18 hours.
  • WB-12 (7.0 g, 21.52 mmol), hexamethyldistanane (10.58 g, 32.29 mmol, 6.7 mL), Pd( PPh3 ) 4 (2.49 g, 2.15 mmol) were dissolved in 100 mL of In dioxane and replaced with nitrogen three times, the mixture was heated to 120 °C and stirred for 4 hours.
  • 1 H NMR 400 MHz, CDCl 3 -d
  • WB-23 (10.00 g, 73.46 mmol) was dissolved in glacial acetic acid (160 mL) and N-iodosuccinimide (16.53 g, 73.46 mmol) was added portionwise at 0°C. The reaction mixture was stirred at 25°C for 16 hours. The reaction solution was concentrated under reduced pressure to half the volume, filtered, and the filter cake was washed twice with petroleum ether (60 mL), and then the solid was removed in vacuo to remove the solvent to obtain WB-24, which was directly used in the next step.
  • the reaction solution was cooled to 25°C, water (60 mL) was added, extracted three times with ethyl acetate (100 mL), the organic phases were combined, washed once with saturated brine (50 mL), dried over anhydrous sodium sulfate, and pumped with a Buchner funnel. After filtration, the filtrate was concentrated in vacuo.
  • cuprous bromide (6.21 g, 43.29 mmol) and potassium bromide (206.06 g, 2.66 mol) were dissolved in water (60 ml), and the above reaction solution was slowly added to cuprous bromide and bromine The potassium chloride suspension was added dropwise for about 30 minutes. After the addition, the reaction solution was continued to react at 0° C. for 1.5 hours.
  • reaction solution was poured into saturated aqueous ammonium chloride solution (200 mL), extracted twice with 100 mL of ethyl acetate each time, the organic phases were combined, dried over anhydrous sodium sulfate, filtered and concentrated.
  • BB-3 (2.75 g, 9.25 mmol), hexamethyldistanane (4.42 g, 13.50 mmol) and Pd( PPh3 ) 4 (470 mg, 406.73 mmol) were added to 1,4-dichlorohexanol In a ring (30 mL), the reaction mixture was sparged with nitrogen and heated to 120°C with stirring for 2 hours.
  • reaction solution was slowly poured into 50 mL of saturated ammonium chloride solution, extracted three times with 20 mL of ethyl acetate, and concentrated.
  • reaction solution was concentrated, diluted with 10 milliliters of ethyl acetate, filtered, and the filtrate was concentrated, and the filtrate and the filter cake were respectively subjected to preparative high performance liquid chromatography (separating column model: Phenomenex Synergi C18 150*30mm*4 ⁇ m; mobile phase: [water ( 0.05% hydrochloric acid)-acetonitrile]; acetonitrile%: 13%-33%, 12 minutes) after purification, combined and concentrated to obtain the hydrochloride salt of compound 7.
  • the crude product was purified by prep-HPLC (column: Waters Xbridge 150*25mm*5 ⁇ m; mobile phase: [water (0.05% ammonia water, volume ratio)-acetonitrile]; acetonitrile%: 32%-58%, 10 minutes) to obtain compound 9.
  • Step ten
  • Step ten
  • 1 H NMR 400 MHz, CDCl 3 -d
  • BB-2 (700 mg, 1.91 mmol) and 14-6 (703.42 mg, 1.95 mmol), tris(o-methylphenyl)phosphine (116.10 mg, 381.44 ⁇ mol), Pd 2 (dba) 3 ( 174.64 mg, 190.72 ⁇ mol), triethylamine (578.96 mg, 5.72 mmol) was suspended in 1,4-dioxane (10 mL), replaced with nitrogen three times, and the reaction mixture was stirred at 90-100 °C At 4 hours, LCMS showed that the reaction was essentially complete, with only a small amount of BB-2 not being consumed.
  • Step ten
  • the number of chloride ions molecular weight of free alkali/ ⁇ (1-chloride ion content) ⁇ chloride ion content ⁇ chloride ion molecular weight, the calculated chloride ion number is 1.8.
  • reaction solution was cooled to 25°C, water (5 mL) and ethyl acetate (10 mL) were added, the organic phase was washed with saturated brine (5 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent.
  • Step ten
  • reaction solution was stirred under nitrogen protection at 0-25°C for 13 hours.
  • Step ten
  • reaction solution was cooled to 0°C, water (3.5 mL), 15% sodium hydroxide solution (3.5 mL), water (10.5 mL) were slowly added dropwise in sequence, filtered, and the filtrate was extracted twice with ethyl acetate (50 mL each) , the organic phases were combined, the pH was adjusted to 2-3 with ethyl acetate hydrochloride (4M), and concentrated to obtain compound W11-3.
  • Step ten
  • ligand beads were blocked with excess biotin and washed with buffer (seablock, 1% bovine serum albumin, 0.05% Tween 20, 1 mM DTT (dithiothreitol)) to wash away unbound ligand. ligands and non-specifically bound ligands;
  • test compound is dissolved in pure dimethyl sulfoxide
  • Affinity beads were resuspended in buffer (IX PBS, 0.05% Tween 20, 0.5 ⁇ m of non-biotin affinity ligand) and incubated for 30 minutes at room temperature.
  • the compounds of the present invention have significant mTOR kinase inhibitory activity.
  • HTRF homogeneous time-resolved fluorescence
  • the compounds of the present invention have significant DNA-PK kinase inhibitory activity.
  • Promega's Luminescent Cell Viability Assay is a quantitative method that reflects the number of viable cells by detecting ATP content.
  • the number of cells is proportional to the fluorescence value. The higher the fluorescence reading, the higher the intracellular ATP content, and the more the number of living cells.
  • the assay plate was analyzed using PerkinElmer's Envision, the assay mode was fluorescence detection, and the data was represented by the reading of the chemiluminescent signal at 400-700 nm. Therefore, we can monitor the degree of inhibition of cell proliferation by the compound by the fluorescence value.
  • Test method Digest the cells in the isolated culture flask, resuspend with medium, count, and adjust the cell concentration to 1.25 ⁇ 10 4 cells with medium (500 mL RPMI1640, 55 mL fetal bovine serum, 5.5 mL penicillin/streptomycin) /mL, add 100 ⁇ L of phosphate buffer to the peripheral wells of the 384-well plate, add 40 ⁇ L of cell suspension to other wells, let stand for 10 min at room temperature, and then incubate in a 37°C, 5% CO 2 incubator for 24 h; compounds with 10-point 3-fold serial dilution of DMSO on ECHO, and transfer 100nL compound to 384-well plate respectively, transfer 100nL DMSO for positive control and negative control, centrifuge at 1000rpm for 10s, and incubate in 37°C 5% CO2 incubator 5 days; then add 20 ⁇ L of Cell Titer-Glo to each well of a 384-well plate, centrifuge at 1000
  • Inhibition rate (%) 100 ⁇ (compound signal value-negative control average signal value)/(positive control average signal value-negative control average signal value)
  • the calculated inhibition rate will be analyzed by curve fitting using GraphPad Prism software to obtain a dose-response curve, and the IC50 of the compound's inhibition of cell proliferation will be calculated.
  • the compound of the present invention can effectively inhibit the proliferation of U87MG cells.
  • OBJECTIVE In this experiment, the EC9706 cell subcutaneous xenograft tumor BALB/c nude mouse model was used to evaluate the antitumor effect of the compounds.
  • mice Female BALB/c nude mice, 6-8 weeks old, weighing 18-22 grams: Supplier: Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd.
  • Human esophageal cancer EC9706 cells (BNCC, 39892) were cultured in vitro in monolayer, cultured in EMEM medium and 10% fetal bovine serum, 100U/mL penicillin and 100 ⁇ g/mL streptomycin, 37°C, 5% CO2 incubator . Conventional digestion with trypsin-EDTA was performed twice a week for passage. When the cell saturation is 80%-90% and the number reaches the requirement, the cells are collected, counted, and seeded.
  • EC9706 cells 0.1 mL (5 ⁇ 10 6 cells) of EC9706 cells were subcutaneously inoculated into the right back of each mouse, and the pharmacodynamics experiment started when the average tumor volume reached 156 mm 3 .
  • the vehicle was 10% DMSO plus 10% Solutol plus 80% water, and the test compound was formulated into a clear solution.
  • Solutol is polyethylene glycol-15 hydroxystearate
  • Tumor diameters were measured with vernier calipers twice a week.
  • TGI percent or relative tumor proliferation rate T/C (%).
  • Relative tumor proliferation rate T/C (%) T RTV /C RTV ⁇ 100% (T RTV : the average RTV of the treatment group; C RTV : the average RTV of the negative control group).
  • TGI (%) reflecting tumor growth inhibition rate.
  • TGI(%) [1-(average tumor volume at the end of administration of a certain treatment group-average tumor volume at the beginning of administration of this treatment group)/(average tumor volume at the end of treatment in the solvent control group-average at the beginning of treatment in the solvent control group tumor volume)] ⁇ 100%.
  • the compounds of the present invention have significant antitumor activity.
  • ligand beads were blocked with excess biotin and washed with buffer (seablock, 1% bovine serum albumin, 0.05% Tween 20, 1 mM DTT (dithiothreitol)) to wash away unbound ligand. ligands and non-specifically bound ligands;
  • test compound is dissolved in pure dimethyl sulfoxide
  • Affinity beads were resuspended in buffer (IX PBS, 0.05% Tween 20, 0.5 ⁇ m of non-biotin affinity ligand) and incubated for 30 minutes at room temperature.
  • the compounds of the present invention have significant mTOR kinase inhibitory activity.
  • DNA-PK (h) was incubated in assay buffer containing 50 nM GST-cMyc-p53 and Mg/ATP (concentration as required). The reaction was initiated by the addition of a Mg/ATP mixture. After 30 minutes of incubation at room temperature, the reaction was stopped by the addition of a stop solution containing EDTA. Finally, detection buffer (containing labeled anti-GST monoclonal antibody and europium-labeled anti-phospho-Ser15 antibody against phosphorylated p53) was added.
  • HTRF homogeneous time-resolved fluorescence
  • the compounds of the present invention have significant DNA-PK kinase inhibitory activity.
  • test compound was mixed with 10% DMSO/10% Solutol/80% water, stirred and vortexed to prepare a homogeneous suspension at 1 mg/ml. Take 400 ⁇ L of 1mg/ml test compound solution and add 600 ⁇ L of 10% DMSO/10% polyethylene glycol stearyl acetic acid/80% water to obtain a 0.4mg/ml clear solution for IV group administration, microporous membrane Use after filtering. Test compounds were mixed with 5% DMSO/95% (10% HP-[beta]-CD), stirred and vortexed to prepare a clear solution at 1 mg/ml for PO administration. (Solutol is polyethylene glycol-15 hydroxystearate)
  • mice Four male CD-1 mice were divided into two groups. Group 1 animals were given a single intravenous dose of 2 mg/kg in a vehicle of 10% DMSO/10% Solutol/80% water in a volume of 5 ml/kg. The animals in the second group were given a single oral gavage of 10 mg/kg of the test compound, the oral vehicle was 5% DMSO/95% (10% HP- ⁇ -CD), and the oral volume was 10 mL/kg.
  • Whole blood was collected at 0.083 (iv injection only), 0.25, 0.5, 1, 2, 4, 8 and 24 hours post-dose. Whole blood was centrifuged at 3200 g for 10 min at 4°C to obtain plasma. The concentrations of the test compounds in the plasma were determined by LC/MS/MS, and the pharmacokinetic parameters were calculated by Phoenix WinNonlin software.
  • the tested compounds exhibit excellent pharmacokinetic data, and the compounds of the present invention have high oral bioavailability and are well developed molecules for oral administration.
  • test compound was mixed with 5% DMSO + 95% (10% HP-[beta]-CD in water), stirred and vortexed to prepare a 0.4 mg/ml homogeneous suspension for PO administration.
  • 5% DMSO + 95% 10% HP-[beta]-CD in water
  • stirred and vortexed a 0.4 mg/ml homogeneous suspension for PO administration.
  • the microporous membrane filter is used for later use.
  • the tested compounds exhibit excellent pharmacokinetic data, and the compounds of the present invention have high oral bioavailability and are well developed molecules for oral administration.
  • OBJECTIVE This experiment used EC9706 cell subcutaneous xenograft tumor BALB/c nude mouse model to evaluate the antitumor effect of WuXi series compounds.
  • mice Female BALB/c nude mice, 6-8 weeks old, weighing 18-22 grams: Supplier: Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd.
  • Human esophageal cancer EC9706 cells (BNCC, 39892) were cultured in vitro in monolayer, cultured in EMEM medium and 10% fetal bovine serum, 100U/mL penicillin and 100 ⁇ g/mL streptomycin, 37°C, 5% CO2 incubator . Conventional digestion with trypsin-EDTA was performed twice a week for passage. When the cell saturation is 80%-90% and the number reaches the requirement, the cells are collected, counted, and seeded.
  • EC9706 cells 0.1 mL (5 ⁇ 10 6 cells) of EC9706 cells were subcutaneously inoculated into the right back of each mouse, and the pharmacodynamics experiment started when the average tumor volume reached 156 mm 3 .
  • the vehicle was 10% DMSO plus 10% Solutol plus 80% water, and the test compound was formulated into a clear solution.
  • Tumor diameters were measured with vernier calipers twice a week.
  • TGI percent or relative tumor proliferation rate T/C (%).
  • Relative tumor proliferation rate T/C (%) T RTV /C RTV ⁇ 100% (T RTV : the average RTV of the treatment group; C RTV : the average RTV of the negative control group).
  • TGI (%) reflecting tumor growth inhibition rate.
  • TGI(%) [1-(average tumor volume at the end of administration of a certain treatment group-average tumor volume at the beginning of administration of this treatment group)/(average tumor volume at the end of treatment in the solvent control group-average at the beginning of treatment in the solvent control group tumor volume)] ⁇ 100%.
  • the compounds of the present invention have good tumor-inhibiting effect.
  • RPMI-1640 medium fetal bovine serum, penicillin/streptomycin antibiotics were purchased from Vicente.
  • the NCI-N87 cell line was purchased from Nanjing Kebai Biotechnology Co., Ltd. Nivo Multi-Label Analyzer (PerkinElmer).
  • NCI-N87 cells were seeded in a white 96-well plate, 80 ⁇ L of cell suspension per well, which contained 8000 NCI-N87 cells. Cell plates were incubated overnight in a carbon dioxide incubator.
  • the compounds to be tested were diluted 3-fold to the 8th concentration with a row gun, that is, from 200 ⁇ M to 92 nM, and a double-well experiment was set up. Add 78 ⁇ L of medium to the middle plate, and then transfer 2 ⁇ L of each well of the compound to the middle plate according to the corresponding position. After mixing, transfer 20 ⁇ L of each well to the cell plate.
  • the cell plates were placed in a carbon dioxide incubator for 3 days.
  • the IC 50 value can be obtained by curve fitting with four parameters ("log(inhibitor) vs. response--Variable slope" mode).
  • the following table provides the inhibitory activity of the compounds of the present invention on NCI-N87 cell proliferation.
  • the compound of the present invention has very excellent anti-proliferation activity of gastric cancer cell N87.
  • FBS fetal bovine serum
  • DMSO dimethyl sulfoxide
  • EC9706 cells were seeded in Greiner CELLSTAR 96-well plate (flat black plate with lid and transparent bottom), 90 ⁇ L of cell suspension per well, which contained 6000 EC9706 cells. Cell plates were incubated overnight in a carbon dioxide incubator.
  • the compound to be tested was diluted 3.5 times to the ninth concentration, that is, from 4000 ⁇ M to 0.17764 nM, and a double-well experiment was set up.
  • the culture plate was placed at room temperature for 10 minutes to stabilize the luminescence signal.
  • IR(%) (1 ⁇ (RLU compound ⁇ RLU blank control)/(RLU vehicle control ⁇ RLU blank control)*100%.
  • the inhibition rates of different concentrations of compounds were calculated in Excel, and then the GraphPad Prism software was used to plot the inhibition curves and calculate the relevant parameters.
  • the following table provides the IC 50 of the compounds in the examples.
  • the example compounds have significant anti-proliferative activity against esophageal cancer cells EC9706, which is significantly better than CC-115.

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Abstract

L'invention concerne une classe de composés macrocycliques ayant une structure de dihydropyrazine et de pyrazine, et, en particulier, un composé représenté par la formule (I) et un sel pharmaceutiquement acceptable de celui-ci.
PCT/CN2022/074894 2021-02-08 2022-01-29 Composé macrocyclique de dihydropyrazine et de pyrazine WO2022166866A1 (fr)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2010062571A1 (fr) * 2008-10-27 2010-06-03 Signal Pharmaceuticals, Llc INHIBITEURS DE mTOR KINASE ET LEUR EMPLOI DANS LES CAS D’INDICATIONS ONCOLOGIQUES ET POUR DES MALADIES EN RAPPORT AVEC LA VOIE mTOR/P13K/AKT
CN102686225A (zh) * 2009-10-26 2012-09-19 西格诺药品有限公司 杂芳基化合物的合成和纯化方法

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2010062571A1 (fr) * 2008-10-27 2010-06-03 Signal Pharmaceuticals, Llc INHIBITEURS DE mTOR KINASE ET LEUR EMPLOI DANS LES CAS D’INDICATIONS ONCOLOGIQUES ET POUR DES MALADIES EN RAPPORT AVEC LA VOIE mTOR/P13K/AKT
CN102686225A (zh) * 2009-10-26 2012-09-19 西格诺药品有限公司 杂芳基化合物的合成和纯化方法

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
MORTENSEN D.S. ET AL.,: "Optimization of a Series of Triazole Containing Mammalian Target of Rapamycin (mTOR) Kinase Inhibitors and the Discovery of CC-115,", J. MED. CHEM., vol. 58, no. 14, 23 June 2015 (2015-06-23), pages 5599 - 5608, XP055233504, DOI: 10.1021/acs.jmedchem.5b00627 *

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