The present application claims priority from a prior application entitled "estrogen receptor modulator compounds and uses thereof" filed on 28.8.2020, patent application No. 202010886642.1 by the national intellectual property office. The entire contents of said prior application are incorporated by reference into the present application.
Disclosure of Invention
The invention relates to a compound shown as a formula (I) or a pharmaceutically acceptable salt thereof:
wherein,
X1、X2、X3、X4independently selected from CR7Or N;
R7selected from H, F, Cl, Br, I, OH, CN, C1-C6Alkyl radical, C3-C6Cycloalkyl, 3-6 membered heterocycloalkyl, C1-C6Alkoxy radical, C3-C6Cycloalkyloxy or 3-6 membered heterocycloalkyloxy;
R1、R2Independently selected from hydrogen, OH, F, Cl, Br, I, C1-C6Alkyl or C2-C8An alkenyl group;
or, R1、R2Together with the carbon atom to which they are attached form C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl, said C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl optionally substituted with RaSubstitution;
w is selected from the group consisting of C in linear or branched form1-C3An alkylene group;
R5is selected from C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl, said C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl is optionally substituted with 1 or more groups selected from: F. cl, Br, I, CN, OH, OCH3And SO2CH3;
R6Selected from H, C1-C6Alkyl radical, C2-C8Alkenyl radical, C2-C4Alkynyl, C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl, said C1-C6Alkyl radical, C2-C8Alkenyl radical, C2-C4Alkynyl, C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl is optionally substituted with 1 or more groups selected from: F. cl, Br, I, CN, OH, OCH3And SO2CH3;
R3Selected from H, CN, COOH, C (O) ORb、OC(O)Rb、CONH2、C(O)NRbRc、SO2Rb、SO2NRbRc、C1-C6Alkyl radical, C1-C6Alkoxy radical, C3-C6Cycloalkyl radical, C3-C6Heterocycloalkyl radical, C6-C10Aryl or C5-C10Heteroaryl of said C1-C6Alkyl radical, C3-C6Cycloalkyl radical, C3-C6Heterocycloalkyl radical, C6-C10Aryl or C5-C10Heteroaryl is optionally substituted with 1 or more RdSubstitution;
R4selected from H, F, Cl, Br, I, OH, C1-C6Alkyl radical, C1-C6Alkoxy radical, C3-C6Cycloalkyl or C3-C6Heterocycloalkyl radical of said C1-C6Alkyl radical, C1-C6Alkoxy radical, C3-C6Cycloalkyl or C3-C6Heterocycloalkyl is optionally substituted with one or more groups selected from: F. cl, Br, I, CN, OH, OCH3And SO2CH3;
Ring Q is selected from C6-C10Aryl or 5-to 10-membered heteroaryl, said C6-C10Aryl or 5-10 membered heteroaryl optionally substituted with 1 or more R8Substitution;
R8selected from F, Cl, Br, I, OH, CN, COOH, C (O) ORb、OC(O)Rb、CONH2、C(O)NRbRc、NRcC(O)Rb、SO2Rb、SO2NRb、C1-C6Alkyl radical, C1-C6Alkoxy radical, C3-C6Cycloalkyl, 3-6 membered heterocycloalkyl, C6-C10Aryl or 5-to 10-membered heteroaryl, said C3-C6Cycloalkyl, 3-6 membered heterocycloalkyl, C6-C10Aryl or 5-10 membered heteroaryl is optionally substituted with 1 or more groups selected from: F. cl, Br, I, OH, CN, C1-C6Alkyl and C1-C6An alkoxy group;
Raselected from halogen, OH, CN, C1-6Alkyl radical, C1-6Alkoxy radical, C3-6Cycloalkyl or 3-6 membered heterocycloalkyl, said C1-6Alkyl radical, C1-6Alkoxy radical, C3-6Cycloalkyl or 3-6 membered heterocycloalkyl optionally substituted with halogen;
Rbis independently selected from C1-C6Alkyl radical, C2-C8Alkenyl radical, C2-C4Alkynyl, C3-C6Cycloalkyl, 3-6 membered heterocycloalkyl, C6-C10Aryl or 5-10 membered heteroaryl;
Rcindependently selected from H, C1-C6Alkyl radical, C2-C8Alkenyl radical, C2-C4Alkynyl, C3-C6Cycloalkyl, 3-6 membered heterocycloalkyl, C6-C10Aryl or 5-10 membered heteroaryl;
Rdindependently selected from F, Cl, Br, I, OH, CN, C1-C6Alkyl radical, C1-C6Alkoxy radical, C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl, said C1-C6Alkyl radical, C1-C6Alkoxy radical, C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl optionally substituted with 1 or more groups selected from F, Cl, Br, I or OH;
n is 0, 1 or 2.
In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt thereof, is selected from a compound of formula (Ia), or a pharmaceutically acceptable salt thereof:
in some embodiments, the R is1、R2Together with the carbon atom to which they are attached form C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl, said C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl optionally substituted with RaAnd (4) substitution.
In some embodiments, the R is1、R2Together with the carbon atom to which they are attached form C3-C4Cycloalkyl or 3-4 membered heterocycleAlkyl radical, said C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl optionally substituted with RaAnd (4) substitution.
In some embodiments, the R is1、R2Together with the carbon atom to which they are attached form a cyclopropane.
In some embodiments, the R is1、R2Independently selected from hydrogen, OH, F, Cl, Br, I, C1-C6Alkyl or C2-C8An alkenyl group.
In some embodiments, the R is1、R2Independently selected from hydrogen, OH, F, Cl, Br or I.
In some embodiments, the R is1、R2Selected from hydrogen.
In some embodiments, the n is selected from 0 or 1.
In some embodiments, the n is selected from 0.
In some embodiments, the R is3Is selected from C1-C6Alkyl, CN, COOH, C (O) ORb、OC(O)Rb、CONH2、C(O)NRbRcOr C6-C10Aryl radical, said C1-C6Alkyl or C6-C10Aryl is optionally substituted by one or more RdSubstituted, RdIndependently selected from F, Cl, Br, I, OH, CN or C optionally substituted with 1 or more groups selected from F, Cl, Br, I or OH1-C6An alkyl group.
In some embodiments, the R is3Is selected from C1-C6Alkyl or C6-C10Aryl radical, said C1-C6Alkyl or C6-C10Aryl is optionally substituted by one or more RdSubstituted, RdIndependently selected from F, Cl, Br, OH or C optionally substituted with 1 or more groups selected from F, Cl, Br or OH1-C6An alkyl group.
In some embodiments, the R is3Is selected from C1-C3Alkyl radical, said C1-C3Alkyl is optionally substituted by one or more RdSubstituted, RdIndependently selected from F or OH.
In some embodiments, the R is
3Is selected from-CH
2CF
3Or
In some embodiments, the R is
3Is selected from
In some embodiments, the R is4Selected from C optionally substituted by F, Cl, Br or I1-C6An alkyl group.
In some embodiments, the R is4Selected from methyl optionally substituted with F, Cl, Br or I.
In some embodiments, the R is4Is methyl.
In some embodiments, the W is selected from the linear form of C1-C3An alkylene group.
In some embodiments, the W is selected from-CH2-or-CH2CH2-。
In some embodiments, the W is selected from-CH2-。
In some embodiments, the R is5Is C3-C6Cycloalkyl radical, said C3-C6Cycloalkyl is optionally substituted with 1 or more groups selected from: F. cl, Br, I, CN, OH, OCH3And SO2CH3。
In some embodiments, the R is5Is C3-C5Cycloalkyl radical, said C3-C5Cycloalkyl is optionally substituted with 1 or more groups selected from: F. cl, Br, I, CN, OH, OCH3And SO2CH3。
In some embodiments, the R is5Is cyclopropyl or cyclobutyl, optionally substituted by 1 or more groups selected from: F. cl, Br, I,CN、OH、OCH3And SO2CH3。
In some embodiments, the R is5Is cyclopropyl or cyclobutyl, optionally substituted by 1 or more groups selected from: F. cl, Br, I and CN.
In some embodiments, the R is5Is cyclopropyl or cyclobutyl, optionally substituted by 1 or more groups selected from: F. cl, Br and I.
In some embodiments, the R is
5Is composed of
In some embodiments, said X is1、X4Selected from the group consisting of CR7Or N, X2、X3Is CR7。
In some embodiments, the R is7Selected from H, F, Cl, Br or I.
In some embodiments, said X is1、X4Selected from CH, CF or N, X2、X3Is CH.
In some embodiments, said X is1、X4Is CF, X2、X3Is CH.
In some embodiments, said X is1Is N, X2、X3、X4Is CH.
In some embodiments, the R is6Is selected from H or C1-C6An alkyl group.
In some embodiments, the R is6Is selected from H.
In some embodiments, when Het is associated with NR6When the connected atom is a chiral carbon, the configuration is (S) -configuration.
In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt thereof is selected from a compound of formula (II) or a pharmaceutically acceptable salt thereof:
wherein X1、X2、X3、X4、Het、R3、R4And R6As defined for formula (I);
R10selected from H, C1-C6Alkyl radical, C2-C8Alkenyl radical, C2-C4Alkynyl, C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl, said C1-C6Alkyl radical, C2-C8Alkenyl radical, C2-C4Alkynyl, C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl is optionally substituted with 1 or more groups selected from: F. cl, Br, I, CN, OH, OCH3And SO2CH3。
In some embodiments, R10Selected from H, C1-C6Alkyl radical, C3-C6Cycloalkyl or 3-6 membered heterocycloalkyl.
In some embodiments, R10Is H.
In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt thereof is selected from a compound of formula (IIa) or a pharmaceutically acceptable salt thereof:
wherein X1、X2、X3、X4、Het、R3、R4、R6And R10As defined in formula (II).
In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt thereof is selected from a compound of formula (III):
wherein X1、X2、X3、X4、Het、R3、R4And R6As defined in formula (I).
In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt thereof is selected from a compound of formula (IIIa) or a pharmaceutically acceptable salt thereof:
wherein X1、X2、X3、X4、Het、R3、R4And R6As defined in formula (I).
In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt thereof is selected from the following compounds or pharmaceutically acceptable salts thereof:
in one aspect, the compounds described herein exist as a racemic mixture or in enantiomerically enriched or enantiomerically pure form. In certain embodiments, the compounds described herein are prepared as their individual stereoisomers by reacting a racemic mixture of the compounds with an optically active resolving agent to form a pair of diastereomeric compounds/salts, separating the diastereomers and recovering the enantiomers in optical purity. In some embodiments, resolution of enantiomers is performed using covalent diastereomeric derivatives of the compounds described herein. In another embodiment, the diastereomers are separated by separation/resolution techniques based on solubility differences. In certain embodiments, the compounds described herein are prepared as their individual stereoisomers by enzymatic resolution. In some embodiments, resolution of individual stereoisomers is performed using a lipase or esterase. In some embodiments, resolution of individual stereoisomers is performed by lipase or esterase catalyzed asymmetric deacylation. In other embodiments, separation of stereoisomers is performed by chromatography, or by forming salts of diastereomers and separating by recrystallization or chromatography, or any combination thereof. In some embodiments, the stereoisomers are obtained by stereoselective synthesis.
The invention also provides a pharmaceutical composition which comprises the compound shown in the formula (I) or pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier and/or excipient.
In some embodiments, the pharmaceutical composition is formulated for intravenous injection, subcutaneous injection, oral administration, or topical administration.
In some embodiments, the pharmaceutical composition is a tablet, pill, capsule, liquid, suspension, gel, dispersion, solution, emulsion, ointment, or lotion.
Further, the invention relates to an application of the compound shown in the formula (I) or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof in preparing a medicament for preventing or treating estrogen receptor related diseases.
Further, the invention relates to a compound shown in a formula (I) or pharmaceutically acceptable salt thereof or a pharmaceutical composition thereof, which is used for preventing or treating estrogen receptor related diseases.
Further, the invention relates to an application of the compound shown in the formula (I) or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof in preventing or treating estrogen receptor related diseases.
Further, the present invention relates to a compound represented by formula (I) or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof for preventing or treating an estrogen receptor related disease.
The invention also relates to a method for treating estrogen receptor related diseases, which comprises administering a therapeutically effective dose of a pharmaceutical preparation containing the compound shown in the formula (I) or the pharmaceutically acceptable salt thereof to a patient.
The present invention also relates to a method of treating estrogen receptor related disorders in a mammal, comprising administering to a mammal, preferably a human, in need of such treatment a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition thereof.
The preferred embodiment of the present invention, wherein the estrogen receptor related diseases include but are not limited to cancer, autoimmune diseases and the like.
In some embodiments of the invention, the estrogen receptor related disorder is preferably a tumor.
In some embodiments of the invention, the estrogen receptor related disorder includes, but is not limited to, breast cancer.
Definition and description of terms
Unless otherwise indicated, the definitions of groups and terms described in the specification and claims of the present invention, including definitions thereof as examples, exemplary definitions, preferred definitions, definitions described in tables, definitions of specific compounds in the examples, and the like, may be arbitrarily combined and coupled with each other. The definitions of the groups and the structures of the compounds in such combinations and after the combination are within the scope of the present invention as defined in the specification. A particular term should not be considered as ambiguous or unclear without special definition, but rather construed according to ordinary meaning in the art. When a trade name appears herein, it is intended to refer to its corresponding commodity or its active ingredient.
In this context
Indicates the attachment site.
The term "stereoisomer" refers to isomers resulting from the different arrangement of atoms in a molecule, including cis-trans isomers, enantiomers, diastereomers, and conformers.
The compounds of the present invention may have an asymmetric atom such as a carbon atom, a sulfur atom, a nitrogen atom, a phosphorus atom or an asymmetric double bond, and thus the compounds of the present invention may exist in a specific geometric or stereoisomeric form. The particular geometric or stereoisomeric forms may be the cis and trans isomers, the E and Z geometric isomers, the (-) -and (+) -enantiomers, (R) -and (S) -enantiomers, diastereomers, (D) -isomers, (L) -isomers, as well as racemic or other mixtures thereof, e.g., enantiomerically or diastereomerically enriched mixtures, all of which, as well as mixtures thereof, are within the definition of the compounds of the present invention. The presence of additional asymmetric carbon, sulfur, nitrogen or phosphorus atoms in substituents such as alkyl groups is also within the definition of the compounds of the invention as regards all such isomers and mixtures thereof. The compounds of the invention containing asymmetric atoms can be isolated in optically pure form or in racemic form, the optically pure form can be resolved from a racemic mixture, or synthesized by using chiral starting materials or chiral reagents.
The enantiomers or enantiomerically pure compounds herein are illustrated by Maehr, J.chem.Ed.1985, 62: 114-120. Using wedge and dashed bonds, unless otherwise indicated (
And
) Indicating the absolute configuration of a stereocenter. When the compounds described herein contain olefinic double bonds or other centers of geometric asymmetry, they include the E, Z geometric isomer unless otherwise specified. Likewise, all tautomeric forms are included within the scope of the invention.
The term "tautomer" refers to an isomer of a functional group resulting from the rapid movement of an atom in two positions in a molecule. The compounds of the invention may exhibit tautomerism. Tautomeric compounds may exist in two or more interconvertible species. Tautomers generally exist in equilibrium, and attempts to isolate a single tautomer often result in a mixture whose physicochemical properties are consistent with the mixture of compounds. The position of equilibrium depends on the chemical properties within the molecule. For example, in many aliphatic aldehydes and ketones such as acetaldehyde, the keto form predominates; whereas in phenol the enol type predominates. The present invention encompasses all tautomeric forms of the compounds.
The term "substituted" means that any one or more hydrogen atoms on a particular atom is replaced with a substituent, so long as the valence of the particular atom is normal and the substituted compound is stable. When the substituent is oxo (i.e., ═ O), meaning that two hydrogen atoms are substituted, oxo does not occur on the aryl.
The terms "optionally" or "optionally" mean that the subsequently described event or circumstance may or may not occur, and that the description includes instances where said event or circumstance occurs and instances where it does not. For example, ethyl is "optionally" substituted with halo, meaning that ethyl may be unsubstituted (CH)2CH3) Monosubstituted (e.g. CH)2CH2F) Polysubstituted (e.g. CHFCH)2F、CH2CHF2Etc.) or completely substituted (CF)2CF3). It will be appreciated by those skilled in the art that any group containing one or more substituents will not incorporate any substitution or substitution pattern which is sterically impossible and/or cannot be synthesized.
The term "halo" or "halogen" refers to fluorine, chlorine, bromine and iodine.
Herein Cm-CnRefers to having an integer number of carbon atoms in the range of m-n. E.g. "C1-C10By "is meant that the group can have 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, 6 carbon atoms, 7 carbon atoms, 8 carbon atoms, 9 carbon atoms, or 10 carbon atoms.
The term "alkyl" refers to a group of formula CnH2n+1The alkyl group may be linear or branched. The term "C1-C6Alkyl "is understood to mean a straight-chain or branched, saturated monovalent hydrocarbon radical having 1,2,3, 4,5, 6 carbon atoms. The alkyl group is as followsSuch as methyl, ethyl, propyl, butyl, pentyl, hexyl, isopropyl, isobutyl, sec-butyl, tert-butyl, isopentyl, 2-methylbutyl, 1-ethylpropyl, 1, 2-dimethylpropyl, neopentyl, 1-dimethylpropyl, 4-methylpentyl, 3-methylpentyl, 2-methylpentyl, 1-methylpentyl, 2-ethylbutyl, 1-ethylbutyl, 3-dimethylbutyl, 2-dimethylbutyl, 1-dimethylbutyl, 2, 3-dimethylbutyl, 1, 3-dimethylbutyl, or 1, 2-dimethylbutyl, and the like.
The term "alkoxy" refers to a monovalent radical derived from a straight or branched chain alcohol by loss of a hydrogen atom from a hydroxyl group, and is understood to mean an "-O-alkyl". According to the invention, suitable alkoxy is C1-6Alkoxy radicals, e.g. C1-5Alkoxy radical, C1-4Alkoxy radical, C1-3Alkoxy groups, including methoxy, ethoxy, propoxy, isopropoxy, isobutoxy, sec-butoxy, and the like.
The term "alkenyl" refers to an unsaturated aliphatic hydrocarbon group consisting of carbon atoms and hydrogen atoms, which is straight or branched, and has at least one double bond. The term "C2-C8Alkenyl "is understood to preferably mean a straight-chain or branched monovalent hydrocarbon radical which contains one or more double bonds and has 2,3,4, 5, 6,7 or 8 carbon atoms.
The term "alkynyl" refers to a straight or branched chain unsaturated aliphatic hydrocarbon group having at least one triple bond composed of carbon atoms and hydrogen atoms. The term "C2-C4Alkynyl "is understood as preferably meaning a straight-chain or branched, monovalent hydrocarbon radical which contains one or more triple bonds and has 2,3 or 4 carbon atoms.
The term "cycloalkyl" refers to a carbon ring that is fully saturated and exists as a single ring, a bicyclic ring, a bridged ring, or a spiro ring, among others. Unless otherwise indicated, the carbocycle is typically a 3 to 10 membered ring. The term "C3-C6Cycloalkyl "is understood to mean a saturated, monovalent, monocyclic or bicyclic hydrocarbon ring having 3 to 6 carbon atoms, such as cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl. The term "C3-C5Cycloalkyl is understood to mean a saturated monovalent monocyclic or bicyclic hydrocarbonA ring having 3 to 5 carbon atoms, such as cyclopropyl, cyclobutyl or cyclopentyl.
The term "heterocycloalkyl" refers to a cyclic group that is fully saturated and may exist as a single ring, a bridged ring, or a spiro ring. Unless otherwise indicated, the heterocyclic ring is typically a 3 to 7 membered ring containing 1 to 3 heteroatoms (preferably 1 or 2 heteroatoms) independently selected from sulfur, oxygen and/or nitrogen. The term "3-6 membered heterocycloalkyl" refers to heterocycloalkyl groups having 3,4, 5, 6 ring atoms, respectively. Examples of 3-membered heterocycloalkyl include, but are not limited to, oxiranyl, thietanyl, cycloazenyl, non-limiting examples of 4-membered heterocycloalkyl include, but are not limited to, azetidinyl, oxetanyl, thietanyl, examples of 5-membered heterocycloalkyl include, but are not limited to, tetrahydrofuranyl, tetrahydrothienyl, pyrrolidinyl, isoxazolidinyl, oxazolidinyl, isothiazolidinyl, thiazolidinyl, imidazolidinyl, tetrahydropyrazolyl, examples of 6-membered heterocycloalkyl include, but are not limited to, piperidinyl, tetrahydropyranyl, tetrahydrothiopyranyl, morpholinyl, piperazinyl, 1, 4-thiaxanyl, 1, 4-dioxanyl, thiomorpholinyl, 1, 3-dithianyl, 1, 4-dithianyl, and the like.
The term "aryl" refers to an all-carbon monocyclic or fused polycyclic aromatic ring group having a conjugated pi-electron system. The term "C6-C10Aryl "is understood to preferably mean a monocyclic or bicyclic hydrocarbon ring of monovalent aromaticity having from 6 to 10 carbon atoms. In particular a ring having 6 carbon atoms ("C)6Aryl "), such as phenyl; or a ring having 9 carbon atoms ("C)9Aryl group), such as indanyl or indenyl, or a ring having 10 carbon atoms ("C)10Aryl), such as tetralinyl, dihydronaphthyl, or naphthyl.
The term "heteroaryl" refers to a monocyclic or fused polycyclic aromatic ring system containing at least one ring atom selected from N, O, S, the remaining ring atoms being C. The term "5-to 10-membered heteroaryl" is understood to include such monovalent monocyclic or bicyclic aromatic ring systems: having 5 to 10 ring atoms and comprising 1 to 5 heteroatoms independently selected from N, O and S. The term "5-to 10-membered heteroaryl" is understood to include such monovalent monocyclic or bicyclic aromatic ring systems: which has 5, 6,7,8,9, 10 ring atoms, in particular 5 or 6 or 9 or 10 ring atoms, and which comprises 1 to 5, preferably 1 to 3, heteroatoms each independently selected from N, O and S and, in addition, can be benzo-fused in each case. In particular, the heteroaryl group is selected from thienyl, furyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, 1,2, 3-triazolyl, 1,2, 4-triazolyl, thiadiazolyl and the like and their benzo derivatives, such as benzofuryl, benzothienyl, benzothiazolyl, benzoxazolyl, benzisoxazolyl, benzimidazolyl, benzotriazolyl, indazolyl, indolyl, isoindolyl and the like; or pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, and the like, and benzo derivatives thereof, such as quinolyl, quinazolinyl, isoquinolyl, and the like; or azocinyl, indolizinyl, purinyl and the like and benzo derivatives thereof; or cinnolinyl, phthalazinyl, quinazolinyl, quinoxalinyl, naphthyridinyl, pteridinyl, carbazolyl, acridinyl, phenazinyl, phenothiazinyl, phenoxazinyl, and the like.
The term "therapeutically effective amount" means an amount of a compound of the invention that (i) treats or prevents a particular disease, condition, or disorder, (ii) alleviates, ameliorates, or eliminates one or more symptoms of a particular disease, condition, or disorder, or (iii) prevents or delays the onset of one or more symptoms of a particular disease, condition, or disorder described herein. The amount of a compound of the present invention that constitutes a "therapeutically effective amount" varies depending on the compound, the disease state and its severity, the mode of administration, and the age of the mammal to be treated, but can be routinely determined by those skilled in the art with their own knowledge and this disclosure.
The term "pharmaceutically acceptable" is intended to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.
The term "pharmaceutically acceptable salts" refers to pharmaceutically acceptable salts of non-toxic acids or bases, including salts of inorganic acids and bases, organic acids and bases.
The term "adjuvant" refers to a pharmaceutically acceptable inert ingredient. Examples of classes of the term "excipient" include, without limitation, binders, disintegrants, lubricants, glidants, stabilizers, fillers, diluents, and the like. Excipients enhance the handling characteristics of the pharmaceutical formulation, i.e., make the formulation more amenable to direct compression by increasing flowability and/or cohesiveness. Examples of typical "pharmaceutically acceptable carriers" suitable for use in the above formulations are: saccharides, starches, cellulose and its derivatives and the like are commonly used as excipients in pharmaceutical preparations.
The term "pharmaceutical composition" denotes a mixture of one or more compounds described herein or a physiologically/pharmaceutically acceptable salt or prodrug thereof with other chemical components, such as physiologically/pharmaceutically acceptable carriers and excipients. The purpose of the pharmaceutical composition is to facilitate the administration of the compound to an organism.
The words "comprise", "comprises", "comprising" or "including" and variations thereof such as "comprises" or "comprising" are to be interpreted in an open, non-exclusive sense, i.e., "including but not limited to".
The pharmaceutical composition of the present invention can be prepared by combining the compound of the present invention with suitable pharmaceutically acceptable excipients, and can be formulated into solid, semi-solid, liquid or gaseous preparations, such as tablets, pills, capsules, powders, granules, pastes, emulsions, suspensions, suppositories, injections, inhalants, gels, microspheres, aerosols, and the like.
Typical routes of administration of the compounds of the present invention or pharmaceutically acceptable salts thereof or pharmaceutical compositions thereof include, but are not limited to, oral, rectal, topical, inhalation, parenteral, sublingual, intravaginal, intranasal, intraocular, intraperitoneal, intramuscular, subcutaneous, intravenous administration.
The pharmaceutical compositions of the present invention may be manufactured by methods well known in the art, such as conventional mixing, dissolving, granulating, emulsifying, lyophilizing, and the like.
In some embodiments, the pharmaceutical composition is in an oral form. For oral administration, the pharmaceutical compositions may be formulated by mixing the active compounds with pharmaceutically acceptable excipients well known in the art. These adjuvants enable the compounds of the present invention to be formulated as tablets, pills, dragees, capsules, liquids, gels, slurries, suspensions and the like, for oral administration to a patient.
Solid oral compositions may be prepared by conventional mixing, filling or tableting methods. For example, it can be obtained by the following method: the active compounds are mixed with solid adjuvants, optionally the mixture obtained is milled, if desired with further suitable adjuvants, and the mixture is then processed to granules, to give tablets or dragee cores. Suitable excipients include, but are not limited to: binders, diluents, disintegrants, lubricants, glidants or flavoring agents, and the like.
The pharmaceutical compositions may also be adapted for parenteral administration, as sterile solutions, suspensions or lyophilized products in suitable unit dosage forms.
In all methods of administration of the compounds of formula (I) described herein, the daily dose is from 0.01mg/kg to 200mg/kg body weight, preferably from 0.05mg/kg to 50mg/kg body weight, more preferably from 0.1mg/kg to 30mg/kg body weight, in single or divided doses.
The invention also includes isotopically-labeled compounds of the present invention, which are identical to those recited herein, but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of isotopes that can be incorporated into compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, iodine and chlorine, such as respectively2H、3H、11C、13C、14C、13N、15N、15O、17O、18O、31P、32P、35S、18F、123I、125I and36cl, and the like.
Certain isotopically labelledCompounds of the invention (e.g. with3H and14c-label) can be used in compound and/or substrate tissue distribution assays. Tritiated (i.e. by tritiation)3H) And carbon-14 (i.e.14C) Isotopes are particularly preferred for their ease of preparation and detectability. Positron emitting isotopes, such as15O、13N、11C and18f can be used in Positron Emission Tomography (PET) studies to determine substrate occupancy. Isotopically labeled compounds of the present invention can generally be prepared by following procedures analogous to those disclosed in the schemes and/or in the examples below, by substituting an isotopically labeled reagent for a non-isotopically labeled reagent.
The term "IC50"is the half maximal inhibitory concentration, and refers to the concentration of a particular compound required to obtain 50% inhibition of a biological process in vitro. IC (integrated circuit)50The values can be logarithmically converted into pIC50Value (-log IC)50) Wherein a higher value indicates a greater efficacy in an exponential manner.
The compounds of the present invention may be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combinations thereof with other chemical synthetic methods, and equivalents thereof known to those skilled in the art, with preferred embodiments including, but not limited to, examples of the present invention.
The chemical reactions of the embodiments of the present invention are carried out in a suitable solvent that is compatible with the chemical changes of the present invention and the reagents and materials required therefor. In order to obtain the compounds of the present invention, it is sometimes necessary for a person skilled in the art to modify or select the synthesis steps or reaction schemes based on the existing embodiments.
For clarity, the present application is further illustrated by examples, which do not limit the scope of the present application. All reagents used herein were commercially available and used without further purification.
Detailed Description
The following examples illustrate the technical solutions of the present invention in detail, but the scope of the present invention includes but is not limited thereto. Having described the invention in detail and having disclosed specific embodiments thereof, it will be apparent to one skilled in the art that various changes and modifications can be made therein without departing from the spirit and scope thereof.
The structure of the compounds is determined by Nuclear Magnetic Resonance (NMR) and/or Mass Spectrometry (MS). NMR shift in units of 10-6(ppm). Solvents for NMR measurement were deuterated dimethyl sulfoxide, deuterated chloroform, deuterated methanol, etc., and an internal standard was Tetramethylsilane (TMS).
Example 1: 3- (((1R, 3R) -1- (2, 6-difluoro-4- ((1- ((((1R, 2R) -2-fluorocyclopropyl) methyl) azetidin-3-yl) amino) phenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyrido [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol
Step 1: synthesis of 5-bromo-2- (diethoxymethyl) -1, 3-difluorobenzene
4-bromo-2, 6-difluorobenzaldehyde (2.19g, 10.0mmol) was added to 50mL of ethanol, triethyl orthoformate (1.48g, 10.0mmol) was added dropwise, 1 drop of concentrated sulfuric acid was added dropwise and the mixture was heated to 60 ℃ with stirring, after 4 hours the reaction was complete, and the reaction was spin-dried to obtain the title compound.
Step 2: synthesis of tert-butyl 3- (((4- (diethoxymethyl) -3, 5-difluorophenyl) amino) azetidine-1-carboxylate
5-bromo-2- (diethoxymethyl) -1, 3-difluorobenzene (2.94g, 10.0mmol), 1-tert-butoxycarbonyl-3-aminocyclobutylamine (2.06g, 12.0mmol), tris (dibenzylideneacetone) dipalladium (458mg, 0.5mmol), 4, 5-bis (diphenylphosphino) -9, 9-dimethylxanthene (579mg, 1.00mmol), cesium carbonate (9.75g,30.0mmol) were added to 40mL1, 4-dioxane, heated to 80 ℃ under argon protection, stirred for 6 hours, and after completion of the reaction, the reaction solution was concentrated, 100mL ethyl acetate was added, suction filtered, and the filtrate was concentrated to give a crude product.
And step 3: synthesis of tert-butyl 3- ((3, 5-difluoro-4-formylphenyl) amino) azetidine-1-carboxylate
And (3) adding the crude product obtained in the last step into 50mL of tetrahydrofuran, adding 20mL of 2N diluted hydrochloric acid solution, stirring at room temperature, reacting after 2 hours, adjusting the pH of the reaction solution to 8-9 by using saturated sodium bicarbonate aqueous solution, adding 50mL of water, extracting by using ethyl acetate, combining organic layers, concentrating, and performing column chromatography to obtain the title compound.
And 4, step 4: synthesis of tert-butyl 3- ((4- ((1R, 3R) -2- (2, 2-difluoro-3-hydroxypropyl) -3-methyl-2, 3,4, 9-tetrahydro-1H-pyridinyl [3,4-b ] indol-1-yl) -3, 5-difluorophenyl) amino) azetidine-1-carboxylate
Adding tert-butyl 3- ((3, 5-difluoro-4-formylphenyl) amino) azetidine-1-carboxylate (2.50g, 8.00mmol), (R) -3- (((1- (1H-indol-3-yl) propan-2-yl) amino) -2, 2-difluoropropan-1-ol (2.18g, 8.00mmol) to 40mL of toluene, adding 4mL of acetic acid, heating to 80 ℃, stirring for 20 hours, then after the reaction is completed, concentrating the reaction solution, adding 100mL of water, adjusting the pH to 8-9 with a saturated sodium bicarbonate solution, extracting with ethyl acetate, combining organic layers, concentrating, and performing column chromatography to obtain the title compound.
And 5: synthesis of 3- ((1R, 3R) -1- (4- (azetidin-3-ylamino) -2, 6-difluorophenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyridinyl [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol
3- ((1R, 3R) -1- (4- (azetidin-3-ylamino) -2, 6-difluorophenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyridinyl [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol (2.88g, 5.12mmol) was dissolved in 30mL of tetrahydrofuran, cooling to 0 ℃, slowly dropwise adding concentrated sulfuric acid (2.78mL, 52.0mmol), transferring to room temperature after dropwise addition, stirring for 2 hours, then finishing reaction, cooling the reaction liquid to 0 ℃, slowly dropwise adding a saturated sodium bicarbonate aqueous solution, adjusting the pH to 8-9, adding 50mL of water, extracting with ethyl acetate, combining organic layers, drying with anhydrous sodium sulfate, and concentrating after suction filtration to obtain the title compound.
Step 6: synthesis of (3- ((4- ((1R, 3R) -2- (2, 2-difluoro-3-hydroxypropyl) -3-methyl-2, 3,4, 9-tetrahydro-1H-pyridinyl [3,4-b ] indol-1-yl) -3, 5-difluorophenyl) amino) azetidin-1-yl) ((1R, 2R) -2-fluorocyclopropyl) methanone
3- ((1R, 3R) -1- (4- (azetidin-3-ylamino) -2, 6-difluorophenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyridinyl [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol (2.34g, 5.06mmol), 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethyluronium hexafluorophosphate (2.30g, 6.07mmol), (1R, 2R) -2-fluorocyclopropane-1-carboxylic acid (526mg, 5.06mmol) were dissolved in 30mL of N, N-dimethylformamide, N, N-diisopropylethylamine (1.96g, 15.2mmol) was stirred at room temperature for 1 hour, the reaction was completed, the reaction solution was added to 100mL of water, extracted with ethyl acetate, the organic layers were combined, dried over anhydrous sodium sulfate, concentrated and subjected to column chromatography to obtain the title compound.
And 7: synthesis of 3- (((1R, 3R) -1- (2, 6-difluoro-4- ((1- ((((1R, 2R) -2-fluorocyclopropyl) methyl) azetidin-3-yl) amino) phenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyrido [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol
(3- ((4- ((1R, 3R) -2- (2, 2-difluoro-3-hydroxypropyl) -3-methyl-2, 3,4, 9-tetrahydro-1H-pyridinyl [3,4-b ] indol-1-yl) -3, 5-difluorophenyl) amino) azetidin-1-yl) ((1R, 2R) -2-fluorocyclopropyl) methanone (0.60g, 1.09mmol) was added to 10mL anhydrous tetrahydrofuran, cooling to 0 ℃, slowly dropwise adding a tetrahydrofuran solution (2.00mL,1.00mol/L) of lithium aluminum hydride, moving to room temperature after dropwise adding, stirring for 2 hours, then finishing the reaction, cooling the reaction liquid to 0 ℃, slowly dropwise adding 10mL of an ethyl acetate solution, and slowly dropwise adding methanol after dropwise adding: 2mL of a 10:1 solution of water, then the reaction solution was added dropwise to ice water, extracted with ethyl acetate, the organic layers were combined, dried over anhydrous sodium sulfate, concentrated and subjected to column chromatography to give the title compound.
DMSO-d6δH 10.52(s,1H),7.38-7.37(m,1H),7.19-7.17(m,1H),7.00-6.91(m,2H),6.72-6.71(m,1H),6.12-6.09(m,2H),5.26-5.23(m,1H),5.05(s,1H),4.83-4.79(m,0.5H),4.66-4.63(m,0.5H),3.96-3.93(m,1H),3.69-3.60(m,3H),3.47-3.37(m,2H),3.14-3.03(m,1H),2.85-2.80(m,3H),2.64-2.50(m,3H),2.47-2.42(m,1H),1.07-1.06(m,3H),0.89-0.85(m,1H),0.79-0.73(m,1H),0.60-0.50(m,1H).
LC/MS(m/z,MH+):535.2
Example 2: 3- (((1R, 3R) -1- (2, 6-difluoro-4- ((1- ((((1S, 2S) -2-fluorocyclopropyl) methyl) azetidin-3-yl) amino) phenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyrido [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol
The title compound was obtained in a similar manner to example 1 except that (1R, 2R) -2-fluorocyclopropane-1-carboxylic acid in step 6 in example 1 was replaced with (1S, 2S) -2-fluorocyclopropane-1-carboxylic acid and the last two reactions of example 1 were carried out.
DMSO-d6δH 10.50(s,1H),7.38-7.36(m,1H),7.19-7.17(m,1H),7.00-6.91(m,2H),6.70-6.68(m,1H),6.11-6.08(m,2H),5.25-5.20(m,1H),5.05(s,1H),4.82-4.78(m,0.5H),4.65-4.62(m,0.5H),3.95-3.92(m,1H),3.69-3.60(m,3H),3.47-3.37(m,2H),3.14-3.03(m,1H),2.85-2.80(m,3H),2.64-2.50(m,3H),2.47-2.42(m,1H),1.07-1.05(m,3H),0.91-0.85(m,1H),0.79-0.71(m,1H),0.59-0.49(m,1H).
LC/MS(m/z,MH+):535.2
Example 3: 3- (((1R, 3R) -1- (2, 6-difluoro-4- ((1- ((((1S, 2R) -2-fluorocyclopropyl) methyl) azetidin-3-yl) amino) phenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyrido [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol
The title compound was obtained in a similar manner to example 1 except that (1R, 2R) -2-fluorocyclopropane-1-carboxylic acid in step 6 in example 1 was replaced with (1S, 2R) -2-fluorocyclopropane-1-carboxylic acid and the last two reactions of example 1 were carried out.
DMSO-d6δH 10.51(s,1H),7.38-7.35(m,1H),7.19-7.17(m,1H),7.00-6.91(m,2H),6.70-6.68(m,1H),6.11-6.08(m,2H),5.25-5.20(m,1H),5.05(s,1H),4.82-4.78(m,0.5H),4.65-4.62(m,0.5H),3.95-3.92(m,1H),3.69-3.60(m,3H),3.47-3.37(m,2H),3.14-3.03(m,1H),2.85-2.80(m,3H),2.64-2.49(m,3H),2.47-2.42(m,1H),1.07-1.05(m,3H),0.91-0.85(m,1H),0.79-0.71(m,1H),0.59-0.49(m,1H).
LC/MS(m/z,MH+):535.2
Example 4: 3- ((1R, 3R) -1- (4- ((1- ((2, 2-difluorocyclopropyl) methyl) azetidin-3-yl) amino) -2, 6-difluorophenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyrido [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol
The title compound was obtained in a similar manner to example 1 except that (1R, 2R) -2-fluorocyclopropane-1-carboxylic acid in step 6 of example 1 was replaced with 2, 2-difluorocyclopropane-1-carboxylic acid and the last two reactions of example 1 were carried out in the same manner.
DMSO-d6δH 10.51(s,1H),7.38-7.36(m,1H),7.19-7.17(m,1H),7.00-6.91(m,2H),6.71-6.70(m,1H),6.12-6.09(m,2H),5.25-5.22(m,1H),5.05(s,1H),3.98-3.90(m,1H),3.71-3.60(m,3H),3.47-3.38(m,2H),3.14-3.03(m,1H),2.83-2.80(m,3H),2.68-2.55(m,3H),2.43-2.38(m,1H),1.75-1.62(m,1H),1.57-1.48(m,1H),1.20-1.12(m,1H),1.07-1.06(m,3H).
LC/MS(m/z,MH+):553.2
Example 5: 3- ((1R, 3R) -1- (4- ((1- ((3, 3-difluorocyclobutyl) methyl) azetidin-3-yl) amino) -2, 6-difluorophenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyridinyl [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol
The title compound was obtained in a similar manner to example 1 except that (1R, 2R) -2-fluorocyclopropane-1-carboxylic acid in step 6 of example 1 was replaced with 3, 3-difluorocyclobutane-1-carboxylic acid and the last two reactions of example 1 were carried out in the same manner.
DMSO-d6δH 10.51(s,1H),7.38-7.36(m,1H),7.19-7.17(m,1H),7.08-6.91(m,2H),6.69-6.68(m,1H),6.12-6.08(m,2H),5.24-5.21(m,1H),5.05(s,1H),3.97-3.90(m,1H),3.72-3.62(m,3H),3.49-3.36(m,3H),3.13-3.03(m,1H),2.84-2.77(m,3H),2.68-2.55(m,4H),2.27-2.15(m,3H),2.08-2.06(m,1H),1.07-1.06(m,3H).
LC/MS(m/z,MH+):567.2
Example 6: 3- ((1R, 3R) -1- (2, 6-difluoro-4- ((1- ((3-fluorocyclobutyl) methyl) azetidin-3-yl) amino) phenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyrido [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol
The title compound was obtained in a similar manner to example 1 except that (1R, 2R) -2-fluorocyclopropane-1-carboxylic acid in step 6 of example 1 was replaced with 3-fluorocyclobutanecarboxylic acid and the last two reactions of example 1 were carried out in the same manner.
DMSO-d6δH 10.51(s,1H),7.38-7.36(m,1H),7.19-7.17(m,1H),7.00-6.91(m,2H),6.69-6.67(m,1H),6.11-6.08(m,2H),5.24-5.21(m,1H),5.05(s,1H),3.95-3.90(m,1H),3.72-3.62(m,4H),3.43-3.36(m,3H),3.13-3.03(m,1H),2.84-2.77(m,3H),2.68-2.55(m,4H),2.27-2.15(m,3H),2.08-2.06(m,1H),1.07-1.05(m,3H).
LC/MS(m/z,MH+):549.2
Example 7: 3- (((1R, 3R) -1- (2, 6-difluoro-4- (((S) -1- ((((1S, 2S) -2-fluorocyclopropyl) methyl) pyrrolidin-3-yl) amino) phenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyridine [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol
The title compound was obtained in analogy to example 1, except that the last six steps of example 1 were carried out in analogy to example 1, substituting 1-tert-butoxycarbonyl-3-aminocyclobutylamine for (S) -3-aminopyrrolidine-1-carboxylic acid tert-butyl ester in step 2 of example 1.
DMSO-d6δH 10.51(s,1H),7.38-7.36(m,1H),7.19-7.17(m,1H),7.00-6.91(m,2H),6.22-6.19(m,3H),5.25-5.22(m,1H),5.03(s,1H),4.85-4.81(m,0.5H),4.68-4.64(m,0.5H),3.76-3.62(m,2H),3.46-3.36(m,2H),3.12-3.02(m,2H),2.83-2.78(m,1H),2.70-2.60(m,3H),2.46-2.38(m,4H),1.07-1.05(m,3H),0.82-0.57(m,5H).
LC/MS(m/z,MH+):549.2
Example 8: 3- ((1R, 3R) -1- (2, 6-difluoro-4- (((S) -5- ((((1R, 2R) -2-fluorocyclopropyl) methyl) -5-azaspiro ] [2.4] heptan-7-yl) amino) phenyl) -3-methyl-1, 3,4, 9-tetrahydro-2H-pyrido [3,4-b ] indol-2-yl) -2, 2-difluoropropan-1-ol
The title compound was obtained in a similar manner to example 1 except that 1-tert-butoxycarbonyl-3-aminocyclobutylamine in step 2 of example 1 was replaced with tert-butyl (S) -7-amino-5-azaspiro [2.4] heptane-5-carboxylate by the latter six-step reaction of example 1.
DMSO-d6δH 10.51(s,1H),7.38-7.36(m,1H),7.19-7.17(m,1H),7.00-6.91(m,2H),6.22-6.19(m,3H),5.25-5.22(m,1H),5.03(s,1H),4.85-4.81(m,0.5H),4.68-4.64(m,0.5H),3.76-3.62(m,2H),3.46-3.36(m,2H),3.12-3.02(m,2H),2.83-2.78(m,1H),2.70-2.54(m,4H),2.46-2.38(m,3H),1.07-1.05(m,3H),1.00-0.94(m,1H),0.81-0.71(m,2H),0.65-0.41(m,4H).
LC/MS(m/z,MH+):575.2
Example 9: n- (1- ((((1R, 2R) -2-fluorocyclopropyl) methyl) azetidin-3-yl) -6- ((6S, 8R) -8-methyl-7- (2,2, 2-trifluoroethyl) -6,7,8, 9-tetrahydro-3H-pyrazolo [4,3-f ] isoquinolin-6-yl) pyridin-3-amine
Step 1: synthesis of tert-butyl 3- ((6- ((6S, 8R) -8-methyl-3- (tetrahydro-2H-pyran-2-yl) -7- (2,2, 2-trifluoroethyl) -6,7,8, 9-tetrahydro-3H-pyrazolo [4,3-f ] isoquinolin-6-yl) pyridin-3-yl) amino) azetidine-1-carboxylate
The intermediate (6S, 8R) -6- (5-bromopyridin-2-yl) -8-methyl-3- (tetrahydro-2H-pyran-2-yl) -7- (2,2, 2-trifluoroethyl) -6,7,8, 9-tetrahydro-3H-pyrazolo [4,3-f ] isoquinoline was prepared according to the procedure of WO2018077630a1, example 17.
(6S, 8R) -6- (5-bromopyridin-2-yl) -8-methyl-3- (tetrahydro-2H-pyran-2-yl) -7- (2,2, 2-trifluoroethyl) -6,7,8, 9-tetrahydro-3H-pyrazolo [4,3-f ] isoquinoline (5.08g, 10.0mmol), 1-tert-butoxycarbonyl-3-aminocyclobutylamine (2.06g, 12.0mmol), tris (dibenzylideneacetone) dipalladium (458mg, 0.5mmol), 4, 5-bis-diphenylphosphine-9, 9-dimethylxanthene (579mg, 1.00mmol), cesium carbonate (9.75g,30.0mmol) were added to 40mL1, 4-dioxane, and the reaction was completed after heating to 80 ℃ under argon protection and stirring for 6 hours, and concentrating the reaction solution, adding 100mL of ethyl acetate, performing suction filtration, and concentrating the filtrate to obtain a crude product.
Step 2: synthesis of N- (azetidin-3-yl) -6- (((6S, 8R) -8-methyl-7- (2,2, 2-trifluoroethyl) -6,7,8, 9-tetrahydro-3H-pyrazolo [4], 3-f ] isoquinolin-6-yl) pyridin-3-amine
Dissolving tert-butyl 3- ((6- ((6S, 8R) -8-methyl-3- (tetrahydro-2H-pyran-2-yl) -7- (2,2, 2-trifluoroethyl) -6,7,8, 9-tetrahydro-3H-pyrazolo [4,3-f ] isoquinolin-6-yl) pyridin-3-yl) amino) azetidine-1-carboxylate (600mg, 1.00mmol) in 30mL of methanol, cooling to 0 ℃, slowly adding a 4M/L hydrogen chloride dioxane solution (2.50mL, 10.0mmol), after the completion of the addition, moving to room temperature, stirring for 2 hours, cooling the reaction solution to 0 ℃, slowly adding a saturated sodium bicarbonate aqueous solution, adjusting the pH to 8-9, after addition of 150mL of water, extraction was performed with ethyl acetate, and the organic layers were combined, dried over anhydrous sodium sulfate, filtered with suction and concentrated to give the title compound.
And step 3: synthesis of ((1R, 2R) -2-fluorocyclopropyl) (3- ((6- ((6S, 8R) -8-methyl-7- (2,2, 2-trifluoroethyl) -6,7,8, 9-tetrahydro-3H-pyrazolo [4,3-f ] isoquinolin-6-yl) pyridin-3-yl) amino) azetidin-1-yl) methanone
N- (azetidin-3-yl) -6- (((6S, 8R) -8-methyl-7- (2,2, 2-trifluoroethyl) -6,7,8, 9-tetrahydro-3H-pyrazolo [4], 3-f ] isoquinolin-6-yl) pyridin-3-amine (416mg, 1.00mmol), 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate (380mg, 1.00mmol), (1R, 2R) -2-fluorocyclopropane-1-carboxylic acid (104mg, 1.00mmol) was dissolved in 30mL of N, N-dimethylformamide, N, N-diisopropylethylamine (258mg, 2.00mmol) was added thereto, and the reaction was allowed to proceed to completion after stirring at room temperature for 1 hour, the reaction solution was added to 100mL of water, extracted with ethyl acetate, the organic layers were combined, dried over anhydrous sodium sulfate, concentrated, and subjected to column chromatography to give the title compound.
And 4, step 4: synthesis of N- (1- ((((1R, 2R) -2-fluorocyclopropyl) methyl) azetidin-3-yl) -6- ((6S, 8R) -8-methyl-7- (2,2, 2-trifluoroethyl) -6,7,8, 9-tetrahydro-3H-pyrazolo [4,3-f ] isoquinolin-6-yl) pyridin-3-amine
(3- ((4- ((1R, 3R) -2- (2, 2-difluoro-3-hydroxypropyl) -3-methyl-2, 3,4, 9-tetrahydro-1H-pyridinyl [3,4-b ] indol-1-yl) -3, 5-difluorophenyl) amino) azetidin-1-yl) ((1R, 2R) -2-fluorocyclopropyl) methanone (250mg, 0.50mmol) was added to 10mL anhydrous tetrahydrofuran, cooling to 0 ℃, slowly dropwise adding a tetrahydrofuran solution (1.00mL,1.00M/L) of borane, moving to 70 ℃ after dropwise adding, heating and stirring for 2 hours, then finishing reaction, cooling the reaction liquid to 0 ℃, adding 20mL of methanol, concentrating, and carrying out column chromatography to obtain the title compound.
DMSO-d6δH 13.02(s,1H),8.06(s,1H),7.79-7.78(m,1H),7.24-7.22(m,1H),7.06-7.04(m,1H),6.98-6.96(m,1H),6.80-6.78(m,1H),6.58-6.56(m,1H),4.96(s,1H),4.52-4.51(m,0.5H),4.35-4.34(m,0.5H),3.95(m,1H),3.64-3.60(m,2H),3.55-3.45(m,2H),3.09-3.04(m,1H),3.00-2.94(m,1H),2.87-2.81(m,1H),2.74-2.71(m,2H),2.47-2.41(m,2H),1.07-1.06(m,3H),0.98-0.82(m,3H).
LC/MS(m/z,MH+):489.2。
Examples of biological Activity and related Properties
Experimental example 1: detection of degradation effect of compound of the invention on MCF7 intracellular estrogen receptor
The test principle is as follows: determination of the degradative Activity of the Compounds of the invention on endogenously expressed Estrogen receptors in MCF7 cells, according to IC50And evaluation of maximum degradation efficiencyThe activity of the test compound.
The test method comprises the following steps:
MCF7 cells (purchased from ATCC, HTB-22) were cultured in DMEM (purchased from Gibco, 11995-. The first day of experiment, MCF7 cells were seeded at 3000 cells/well in 384-well plates at 37 ℃ with 5% CO using complete medium2And (5) culturing in a cell culture box. Test compounds were dissolved in DMSO at a stock concentration of 10mM, diluted with Echo550 (available from Labcyte Inc.) and added to the cell culture plate, each compound treatment was initiated at 100nM, diluted in a 3-fold gradient, 9 concentration points, a blank containing 0.5% DMSO was set up, and a duplicate well control was set up for each concentration point. 37 ℃ and 5% CO2The cell culture box was cultured for 24 hours. Adding paraformaldehyde into each cell culture well, fixing cells at a final concentration of about 3.7%, reacting for 30min, removing supernatant, and washing with 50 μ L PBS once per well; the cells were treated for 30 minutes with PBS (containing 0.5% v/v Tween-20) and washed once with PBS; adding blocking solution (self-made, 5% BSA and 0.05% v/v Tween-20 in PBS) and incubating for 1 hour at room temperature; removing the blocking solution, adding primary antibody mixture (anti-ER monoclonal antibody, Estrogen Receptor alpha (D8H8) Rabbit mAb purchased from GST, #8644S, diluted at 1: 1000; anti-GAPDH monoclonal antibody, GAPDH (D4C6R) Mouse mAb purchased from GST, #97166S, diluted at 1: 2000) and incubating at room temperature for 3 hours; washing 3 times with PBST (homemade, PBS containing 0.05% v/v Tween-20); adding a detection secondary antibody (800 CW-goat anti-rabbit IgG, purchased from LI-COR, P/N:926-32211, diluted 1: 1000; 680 RD-goat anti-mouse IgG, purchased from LI-COR, #925-68070, diluted 1: 1000), and incubating for 45 minutes at room temperature in a dark place; PBST was washed 3 times and the fluorescence signal from each well was read using the Odyssey CLx. XLFit was used for data processing, and the inhibitory activity IC of each compound was calculated from the concentration of the compound and the fluorescence signal value50And maximum degradation efficiency.
And (3) test results:
under the present experimental conditions, the test compounds have good inhibitory activity against ER level at a cell-based level. The results of the ER level activity assay for the test compounds are shown in Table 1.
TABLE 1 results of in vitro cell-level-based ER level Activity assay for test Compounds
Test compounds
|
ER level DC50(nM)
|
Maximum ER Down-Regulation (%)
|
Example 1
|
0.07
|
103.36
|
Example 2
|
0.14
|
95.76
|
Example 3
|
0.45
|
79.50
|
Example 4
|
0.12
|
95.84
|
Example 5
|
0.36
|
90.21
|
Example 6
|
0.23
|
84.93
|
Example 7
|
0.22
|
90.95
|
Example 8
|
0.31
|
95.37
|
Example 9
|
0.22
|
107.81 |
Experimental example 2: examination of inhibitory Effect of the Compound of the present invention on proliferation of T47D cells
The test principle is as follows: determining the inhibitory Effect of the Compounds of the invention on the in vitro proliferation of T47D cells, according to IC50The activity of the compounds was evaluated.
The test method comprises the following steps:
T47D (T-47D) cells (purchased from ATCC, HTB-133) were cultured in RPMI-1640 (purchased from Gibco, A10491-01) complete medium containing 10% fetal bovine serum. On the first day of experiment, T47D cells were seeded at a density of 500 cells/well in 384-well plates at 37 ℃ with 5% CO using complete medium
2The cell culture box was cultured overnight. The next day, test compound was added for drug treatment, and a10 mM stock solution of each compound was diluted with Echo550 (available from labcell Inc.) and transferred to each cell culture well at an initial intracellular treatment concentration of 100nM, 3-fold gradient dilution, 10 concentration points, a blank containing 0.3% DMSO was set, and a duplicate well control was set for each concentration point. 37 ℃ and 5% CO
2The cell culture box is used for 7 days, and on the eighth day, the cell culture plate is taken out. Adding into
Luminescennt Cell visual Assay (available from Promega, G7573) was allowed to stand at room temperature for 10 minutes, and then the luminescence signal value was read using a multi-label microplate reader EnVision (available from PerkinElmer), and the inhibitory activity IC of each compound was calculated from the concentration of the compound and the luminescence signal value using XLfit
50。
And (3) test results:
under the conditions of this experiment, the test compound showed good inhibitory activity against T47D breast cancer cells. The results of the inhibitory activity of the test compounds on T47D cell proliferation are detailed in table 2.
TABLE 2 inhibitory Activity of Compounds on T47D cell proliferation
Test compounds
|
T47D IC50(nM)
|
Example 1
|
0.17 |
Experimental example 3: examination of inhibitory Effect of the Compound of the present invention on MCF7 cell proliferation
The test principle is as follows: determining the inhibitory Effect of the Compounds of the invention on the in vitro proliferation of MCF7 cells, according to IC50And maximum inhibition efficiency to evaluate the activity of the compound.
The test method comprises the following steps:
MCF7 cells (ATCC, HTB-22) were cultured in DMEM (Gibco, 11995-065) complete medium containing 10% fetal bovine serum. The first day of experiment, MCF7 cells were seeded at a density of 500 cells/well in 384-well plates at 37 ℃ with 5% CO using complete medium
2The cell culture box was cultured overnight. The next day, test compound was added for drug treatment, and a compound solution with a stock concentration of 10mM was diluted with Echo550 (labcell Inc.) and transferred to each cell culture well, with the start concentration of each compound in the cells being 100nM, 3-fold gradient dilution, 9 concentration points, a blank containing 0.3% DMSO was set, and a duplicate well control was set for each concentration point. 37 ℃ and 5% CO
2The cell culture box is used for 7 days, and on the eighth day, the cell culture plate is taken out. Adding into
Luminescent Cell visual Assay (Promega, G7573), left at room temperature for 10 minutes, the luminescence signal value was read using a multi-label microplate reader envision (perkinelmer), and the inhibitory activity IC of each compound was calculated from the concentration of the compound and the luminescence signal value using XLfit
50。
And (3) test results:
under the experimental condition, the test compound has better inhibitory activity on MCF-7 cell proliferation. The results are shown in Table 3.
TABLE 3 inhibitory Activity of Compounds on MCF-7 cell proliferation
Compound numbering
|
MCF-7 cell proliferation inhibition IC50(nM)
|
Example 1
|
0.07 |
Experimental example 4: pharmacokinetic Property testing of Compounds of the invention
The test principle is as follows: the drug concentration in the blood plasma of the mice at different times after the mice are subjected to gastric lavage and intravenous administration of the compound is measured by an LC/MS/MS method by taking the mice as test animals. The pharmacokinetic behavior of the compounds of the invention in mice was studied and their pharmacokinetic profile was evaluated.
The test method comprises the following steps:
4.1 test animals
24 healthy adult BALB/c mice, female, were divided into 4 groups on average, 6 mice per group, 3 gavage, 3 veins, mice purchased from Beijing vitamin Tonglihua laboratory animal technology, Inc., animal production license number: SCXK (Zhe) 2019-.
4.2 pharmaceutical preparation
Weighing a certain amount of medicine, dissolving in DMSO 5% + PG 20% + absolute ethyl alcohol 5% + solutol 10% + water 60%, preparing into 10mg/ml, and using for intragastric administration. Weighing a certain amount of medicine, dissolving in DMSO 1% + PG 4% + absolute ethyl alcohol 1% + solutol 2% + water 92%, and preparing into 1mg/ml for intravenous injection.
4.3 administration of drugs
And (3) gastric lavage group: BALB/c mice were fasted overnight and then gavaged at a dose of 10mg/kg and a volume of 1 mL/kg.
Vein group: BALB/c mice were administered intravenously after overnight fasting at a dose of 1mg/kg and a volume of 1 mL/kg.
4.4 operation
After the mice are administrated by gastric lavage or intravenous injection, 40 mu L of blood collected from orbit 5min, 15min, 30min, 1h, 2h, 4h and 24h after the administration, 5 mu L of EDTA-K2 is anticoagulated, blood plasma is centrifugally separated at 12000rpm and 4 ℃ for 5 minutes, and the blood plasma is preserved at-20 ℃.
Determining the content of the compound to be tested in the plasma of the mouse after the drugs with different concentrations are administered by gastric lavage or vein: melting the sample at room temperature, and vortexing for 1 min; quantitatively transferring 15. mu.L to a 2ml 96-well plate, adding 150. mu.L of internal standard precipitant, and shaking (1200 rpm. times.3 min); centrifugation (4000rpm 15min) and transfer of 100. mu.L of supernatant into 1ml 96-well plates; blow-drying with nitrogen, adding 100. mu.L of the double solution (acetonitrile 1: 9), shaking (900 rpm. times.3 min), and analyzing 20. mu.L of the sample. LC/MS/MS conditions: mobile phase A, 0.1% aqueous formic acid, mobile phase B: 0.1% formic acid acetonitrile, column: ACE C185 μm (3.0mm x 50mm), column temperature: flow rate 0.5ml/min at 35 ℃.
And (3) test results:
under the conditions of the experiment, the tested compounds show better pharmacokinetic properties, and the results are shown in table 4.
TABLE 4 pharmacokinetic parameters after single gavage administration of Compounds in mice
The embodiments of the present invention have been described above. However, the present invention is not limited to the above embodiment. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.