WO2020135715A1 - Compound for tumor immunity and application thereof - Google Patents

Abstract

Disclosed is a compound represented by Formula (I), optical isomers thereof, pharmaceutically acceptable salts thereof, and an application of the compound as a STING agonist. (I)

Description

Compounds as tumor immunity and their applications

This application claims the following priority:

CN201811635979.4, application date December 29, 2018;

CN201910350752.3, application date April 28, 2019.

Technical field

The present invention relates to the compound represented by formula (I), its optical isomer and its pharmaceutically acceptable salt, and the use of the compound as a STING agonist.

Background technique

For a long time, scientific researchers have tried to activate the patient's immune system so that their own immune system can effectively fight tumors and completely remove tumor cells. However, the probability of spontaneous remission of the tumor is extremely low, so most patients cannot benefit from it. In the 1960s and 1970s, treatment methods such as BCG vaccine injection and non-specific strengthening of immune system function appeared. In the 1980s, interferon and IL-2, which can activate T cells and NK cells, were also tried in the treatment of cancer, but these methods still have many limitations, such as the half-life of exogenous cytokines in the blood Very short, this must be compensated with frequent administration and high doses. Non-specific activation of the immune system leads to inflammation of normal tissues, cytokine storms, etc. Therefore, many treatments have very strong toxic and side effects. As an immunomodulator that triggers the production of specific therapeutic beneficial cytokines in vivo, the therapy targeting STING has brought dawn to solve this dilemma.

It is known that STING is activated in three ways: 1) by binding exogenous (3', 3') cyclic dinucleotides (c-diGMP, c-diAMP and c-GAMP) released by invading bacteria or archaea ) Activation, which shows that STING has the role of innate immune activation in anti-infection; 2) by binding (2', 3') cyclic guanosine monophosphate adenosine monophosphate (2', 3'c-GAMP) Activation, which is induced by the presence of circular GMP-AMP dinucleotide synthetase (cGAS) in the presence of foreign double-stranded DNA (eg, released by invading bacteria, viruses, or protozoa) or self-DNA in mammals Endogenous circular dinucleotide, which shows that STING has the effect of innate immunity induced by endogenous or exogenous DNA; 3) activation by binding synthetic ligands.

STING acts as a receptor for DNA in the cytoplasm, and its activation can lead to the activation of two downstream pathways, IRF3 and NF-κB, to activate the immune system. Activation of the NF-κB pathway leads to the activation of a series of proinflammatory cytokines downstream, while activation of the IRF3 pathway leads to the activation of type I interferon (IFN-α/β), dendritic cells, cytotoxic cells, NK cells, etc. Activation, thereby exerting an anti-tumor effect.

The DNA in the human body usually does not activate the STING protein, because under normal circumstances DNA can only exist in the nucleus (except mitochondrial DNA). But if DNA leaks into the cytoplasm, it will activate STING and trigger an immune response. Recently, it was found that radiotherapy and chemotherapy can also activate STING, which may also be caused by DNA leakage in dead tumor cells.

Summary of the invention

The present invention provides the compound represented by formula (I), its optical isomer and its pharmaceutically acceptable salt,

among them,

L ₁ and L ₂ are independently selected from -O-, -N(R)- and -C(RR)-;

L _{3 is} selected from

R ₁ and R _1a are independently selected from H, halogen, OH, NH ₂ , CN, N ₃ , C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _{1- 6} alkylamino and C _2-6 alkynyl, wherein the C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino or C _2-6 alkynyl Optionally substituted by 1, 2 or 3 R;

R ₂ and R _2a are independently selected from H, halogen, OH, NH ₂ , CN, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino And C _2-6 alkynyl, wherein the C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino, or C _2-6 alkynyl is optionally substituted 1, 2, or 3 R substitutions;

R ₃ and R _3a are independently selected from H, halogen, OH, NH ₂ , CN, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino And C _2-6 alkynyl, wherein the C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino, or C _2-6 alkynyl is optionally substituted 1, 2, or 3 R substitutions;

R ₄ and R _4a are independently selected from H, halogen, OH, NH ₂ , CN, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino And C _2-6 alkynyl, wherein the C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino, or C _2-6 alkynyl is optionally substituted 1, 2, or 3 R substitutions;

R ₅ and R _5a are independently selected from H, halogen, OH, NH ₂ , CN, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino And C _3-6 cycloalkyl, wherein the C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino or C _3-6 cycloalkyl Choose to be replaced by 1, 2 or 3 R;

X _{1 is} selected from BH ₃ ^- and -S(R ₆ );

R _{6 is} selected from H, CH ₂ OC(═O)R ₇ , CH ₂ OC(═O)OR ₇ , CH ₂ CH ₂ SC(═O)R ₇ and CH ₂ CH ₂ SSCH ₂ R ₇ ;

R ₇ is selected from C _6-10 aryl, 5-10 membered heteroaryl, C _1-6 heterocycloalkyl and C ₁ - ₂₀ alkyl group, wherein the C ₁ - ₂₀ alkyl optionally substituted with 1,2 , 3, 4 or 5 C _6-10 aryl, C _3-10 cycloalkyl, OH and F substitutions;

Alternatively, R ₁ and R _{4 are} connected together to form a 5-6 membered heterocycloalkyl;

Alternatively, R _1a and R _{4a are} joined together to form a 5-6 membered heterocycloalkyl;

R is independently selected from H, halogen, OH, NH ₂ , CN,

C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino, and C _1-6 alkyl-(C═O)NH-, wherein the C _{1 -6} alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino or C _1-6 alkyl-(C═O)NH- is optionally substituted by 1, 2 or 3 R'substitution;

R'is selected from F, Cl, Br, I, OH, NH ₂ and CH ₃ ;

The 5-6 membered heterocycloalkyl group, 5-10 membered heteroaryl group or C _1-6 heterocycloalkyl group contains 1, 2 or 3 independently selected from -O-, -NH-, -S-,- C(=O)-, -C(=O)O-, -S(=O)-, -S(=O) ₂ -and heteroatoms or heteroatom groups of N.

In some aspects of the present invention, the above R is independently selected from H, halogen, OH, NH ₂ , CN,

C _1-3 alkyl, C _{1- 3} alkoxy, C _1-3 alkylthio, C _1-3 alkyl - (C = O) NH- and a C _1-3 alkoxy group, wherein the C _{1 -3} alkyl, C _1-3 alkoxy, C _1-3 alkylthio, C _1-3 alkyl-(C=O)NH- or C _1-3 alkylamino is optionally substituted by 1, 2 or 3 R'instead, other variables are as defined in the present invention.

In some aspects of the present invention, the above R is independently selected from H, F, Cl, Br, I, OH, NH ₂ , CN, Me,

Where Me,

It can be optionally substituted with 1, 2 or 3 R', and other variables are as defined in the present invention.

In some aspects of the present invention, the above R is independently selected from H, F, Cl, Br, I, OH, NH ₂ , CN, Me,

Other variables are as defined in the present invention.

In some aspects of the present invention, R ₁ and R _1a are independently selected from H, F, Cl, Br, OH, NH ₂ , CN, N ₃ , C _1-3 alkyl, and C _1-3 alkoxy , C _1-3 alkylthio, C _1-3 alkylamino and C _2-3 alkynyl, wherein the C _1-3 alkyl, C _1-3 alkoxy, C _1-3 alkylthio, C _{The 1-3} alkylamino group or C _2-3 alkynyl group is optionally substituted with 1, 2, or 3 R, and other variables are as defined in the present invention.

In some aspects of the present invention, the above R ₁ and R _1a are independently selected from H, F, Cl, Br, OH, NH ₂ , CN, N ₃ , Me and

Other variables are as defined in the present invention.

In some aspects of the present invention, the above R ₅ and R _5a are independently selected from H, F, Cl, Br, OH, NH ₂ , CN, N ₃ , C _1-3 alkyl, and C _1-3 alkoxy , C _1-3 alkylthio, C _1-3 alkylamino and C _3-6 cycloalkyl, wherein the C _1-3 alkyl, C _1-3 alkoxy, C _1-3 alkylthio, The C _1-3 alkylamino group or C _3-6 cycloalkyl group is optionally substituted with 1, 2, or 3 R, and other variables are as defined in the present invention.

In some aspects of the present invention, the above R ₅ and R _5a are independently selected from H, F, Cl, Br, OH, NH ₂ , CN, N ₃ , Me,

Other variables are as defined in the present invention.

In some solutions of the present invention, the above R ₁ and R _{4 are} connected together, the structural unit

Select from

Other variables are as defined in the present invention.

In some solutions of the present invention, the above R _1a and R _{4a are} connected together, the structural unit

Select from

Other variables are as defined in the present invention.

In some aspects of the present invention, the above-mentioned L ₁ and L ₂ are independently selected from -O-, -NH- and -CH _2- , and other variables are as defined in the present invention.

In some aspects of the invention, the above L _{3 is} selected from

Other variables are as defined in the present invention.

In some solutions of the present invention, the above structural unit

Select from

Other variables are as defined in the present invention.

In some embodiments of the present invention, the above compound, its optical isomer and its pharmaceutically acceptable salt are selected from

among them,

X ₁ , R ₁ , R _1a , R ₄ , R _4a , R ₅ , R _5a , L ₁ , L ₂ , L _{3 are} as defined above.

The present invention provides compounds of the following formula, their optical isomers and their pharmaceutically acceptable salts, which are selected from

The present invention also provides a pharmaceutical composition containing the above compound or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents or excipients.

The present invention also provides the use of the above compound or a pharmaceutically acceptable salt thereof or the above pharmaceutical composition in the preparation of a medicament for preventing and/or treating STING-related diseases.

In some aspects of the invention, the aforementioned STING-related diseases are selected from the group consisting of lymphoma, melanoma, colorectal cancer, breast cancer, acute myeloid leukemia, colon cancer, liver cancer, prostate cancer, pancreatic cancer, renal cancer, and glioma , Bladder cancer, pleural effusion, malignant pleural effusion, head and neck cancer, fibrosarcoma, renal cell carcinoma, lung cancer, malignant ascites, gastric cancer, ovarian cancer, uterine cancer, optic neuroblastoma, bone cancer, rhabdomyosarcoma and esophageal cancer.

Definition and description

Unless otherwise stated, the following terms and phrases used herein are intended to have the following meanings. A specific term or phrase should not be considered uncertain or unclear unless specifically defined, but should be understood in its ordinary meaning. When a trade name appears in this article, it is intended to refer to its corresponding trade product or its active ingredient.

The term "pharmaceutically acceptable" as used herein refers to those compounds, materials, compositions and/or dosage forms, which are within the scope of reliable medical judgment and are suitable for use in contact with human and animal tissues Without excessive toxicity, irritation, allergic reactions or other problems or complications, commensurate with a reasonable benefit/risk ratio.

The term "pharmaceutically acceptable salt" refers to a salt of a compound of the present invention, prepared from a compound having a specific substituent and a relatively non-toxic acid or base found in the present invention. When the compounds of the present invention contain relatively acidic functional groups, base addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of base in a pure solution or a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amine or magnesium salts or similar salts. When the compounds of the present invention contain relatively basic functional groups, acid addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of acid in a pure solution or 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, Bisulfate, hydroiodic acid, phosphorous acid, etc.; and organic acid salts, such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, Fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, methanesulfonic acid and other similar acids; also includes salts of amino acids (such as arginine, etc.) , And salts of organic acids such as glucuronic acid. Certain compounds of the present invention contain basic and acidic functional groups and can be converted to any base or acid addition salt.

The pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound containing acid radicals or bases by conventional chemical methods. Generally, such salts are prepared by reacting these compounds in free acid or base form with a stoichiometric amount of the appropriate base or acid in water or an organic solvent or a mixture of both.

The compounds of the present invention may exist in specific geometric or stereoisomeric forms. The present invention contemplates all such compounds, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers Isomers, (D)-isomers, (L)-isomers, and their racemic mixtures and other mixtures, such as enantiomerically or diastereomerically enriched mixtures, all of which belong to this Within the scope of the invention. Additional asymmetric carbon atoms may be present in the substituents such as alkyl. All these isomers and mixtures thereof are included in the scope of the present invention.

Unless otherwise stated, use a solid wedge key

And wedge-shaped dotted keys

Represents the absolute configuration of a three-dimensional center, using straight solid line keys

And straight dotted keys

Represents the relative configuration of the three-dimensional center, with wavy lines

Represents a solid wedge key

Or wedge-shaped dashed key

Or with wavy lines

Represents a straight solid line key

And straight dotted keys

Dotted line

Indicates the site to be connected.

The compounds of the present invention may be present in specific. Unless otherwise stated, the term "tautomer" or "tautomeric form" means that at room temperature, the isomers of different functional groups are in dynamic equilibrium and can quickly convert to each other. If tautomers are possible (as in solution), the chemical equilibrium of tautomers can be achieved. For example, proton tautomers (also known as prototropic tautomers) include interconversion through proton migration, such as ketone-enol isomerization and imine-ene Amine isomerization. Valence tautomers (valence tautomer) include some recombination of bond-forming electrons for mutual conversion. A specific example of keto-enol tautomerization is the interconversion between two tautomers of pentane-2,4-dione and 4-hydroxypent-3-en-2-one.

Unless otherwise stated, the terms "rich in one isomer", "isomer enriched", "rich in one enantiomer" or "enantiomerically enriched" refer to one of the isomers or pairs The content of the enantiomer is less than 100%, and the content of the isomer or enantiomer is greater than or equal to 60%, or greater than or equal to 70%, or greater than or equal to 80%, or greater than or equal to 90%, or greater than or equal to 95%, or 96% or greater, or 97% or greater, or 98% or greater, or 99% or greater, or 99.5% or greater, or 99.6% or greater, or 99.7% or greater, or 99.8% or greater, or greater than or equal to 99.9%.

Unless otherwise stated, the term "isomer excess" or "enantiomeric excess" refers to the difference between the relative percentages of two isomers or two enantiomers. For example, if the content of one isomer or enantiomer is 90% and the content of the other isomer or enantiomer is 10%, the excess of isomer or enantiomer (ee value) is 80% .

The optically active (R)- and (S)-isomers and D and L isomers can be prepared by chiral synthesis or chiral reagents or other conventional techniques. If an enantiomer of a compound of the present invention is desired, it can be prepared by asymmetric synthesis or derivatization with a chiral auxiliary, where the resulting mixture of diastereomers is separated and the auxiliary groups are cleaved to provide pure The desired enantiomer. Alternatively, when the molecule contains a basic functional group (such as an amino group) or an acidic functional group (such as a carboxyl group), a diastereomer salt is formed with an appropriate optically active acid or base, and then by conventional methods known in the art The diastereomers are resolved and the pure enantiomers are recovered. In addition, the separation of enantiomers and diastereomers is usually accomplished by the use of chromatography that uses a chiral stationary phase and is optionally combined with chemical derivatization methods (eg, amino groups from amines) Formate). The compound of the present invention may contain unnatural proportions of atomic isotopes in one or more atoms constituting the compound. For example, compounds can be labeled with radioactive isotopes, such as tritium ( ³ H), iodine-125 ( ¹²⁵ I) or C-14 ( ¹⁴ C). For another example, the hydrogen can be replaced by heavy hydrogen to form a deuterated drug. The bond formed by deuterium and carbon is stronger than the bond formed by ordinary hydrogen and carbon. Compared with undeuterated drugs, deuterated drugs have lower toxicity and increase drug stability. , Strengthen efficacy, prolong the biological half-life of drugs and other advantages. The conversion of all isotopic compositions of the compounds of the present invention, whether radioactive or not, is included within the scope of the present invention. "Optional" or "optionally" means that the subsequently described event or condition may, but need not necessarily occur, and the description includes situations where the event or condition occurs and conditions where the event or condition does not occur.

The term "substituted" means that any one or more hydrogen atoms on a particular atom are replaced by a substituent, which may include heavy hydrogen and hydrogen variants, as long as the valence state of the particular atom is normal and the compound after substitution is stable of. When the substituent is oxygen (ie = O), it means that two hydrogen atoms are substituted. Oxygen substitution does not occur on aromatic groups. The term "optionally substituted" means that it may or may not be substituted. Unless otherwise specified, the type and number of substituents may be arbitrary on the basis of chemical realization.

When any variable (such as R) appears more than once in the composition or structure of a compound, its definition in each case is independent. Thus, for example, if a group is substituted with 0-2 Rs, the group can optionally be substituted with at most two Rs, and R in each case has independent options. Furthermore, combinations of substituents and/or variants thereof are only allowed if such combinations will produce stable compounds.

When the listed linking group does not indicate the connection direction, the connection direction is arbitrary, for example,

The linking group L in the middle is -MW-, then -MW- can be formed by connecting the benzene ring and cyclopentane in the same direction as the reading order from left to right

It can also be formed by connecting the benzene ring and cyclopentane in the opposite direction to the reading order from left to right

Combinations of the linking group, substituents, and/or variants thereof are only allowed if such a combination will produce a stable compound.

Unless otherwise specified, the number of atoms on a ring is usually defined as the number of members of the ring. For example, "5-6 membered ring" refers to a "ring" with 5-6 atoms arranged around it.

Unless otherwise specified, "5-6 membered ring" means cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, cycloalkynyl, heterocycloalkynyl, aryl consisting of 5 to 6 ring atoms Radical or heteroaryl. The ring includes a single ring, and also includes a bicyclic ring system such as a spiro ring, a parallel ring, and a bridge ring. Unless otherwise specified, the ring optionally contains 1, 2 or 3 heteroatoms independently selected from O, S and N. The 5-6 member ring includes 5 member, 6 member ring and the like. "5-6 membered ring" includes, for example, phenyl, pyridyl, piperidinyl, and the like; on the other hand, the term "5-6 membered heterocycloalkyl" includes piperidinyl and the like, but does not include phenyl. The term "ring" also includes ring systems containing at least one ring, where each "ring" independently conforms to the above definition.

Unless otherwise specified, the term "C _1-20 alkyl" is used to indicate a linear or branched saturated hydrocarbon group composed of 1 to 20 carbon atoms. The C _1-20 alkyl group includes C _1-10 , C _1-9 , C _1-8 , C _1-6 , C _1-5 , C _1-14 , C _1-3 , C _1-2 , C _2-16 , C _2-4 , C ₁₀ , C ₈ , C ₇ , C ₆ and C ₅ alkyl, etc.; it can be monovalent (such as methyl), divalent (such as methylene) or multivalent ( Such as methine). Examples of C _1-20 alkyl groups 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, heptyl, octyl and so on.

Unless otherwise specified, the term "C _1-6 alkyl" is used to indicate a linear or branched saturated hydrocarbon group composed of 1 to 6 carbon atoms. The C _1-6 alkyl group 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 groups; etc.; Is monovalent (such as methyl), divalent (such as methylene) or multivalent (such as methine). Examples of C _1-6 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), pentyl (including n-pentyl, isopentyl and neopentyl), hexyl and so on.

Unless otherwise specified, the term "C _1-3 alkyl" is used to indicate a linear or branched saturated hydrocarbon group composed of 1 to 3 carbon atoms. The C _1-3 alkyl group includes C _1-2 and C _2-3 alkyl groups, etc.; it may be monovalent (such as methyl), divalent (such as methylene), or polyvalent (such as methine) . Examples of C _1-3 alkyl include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), and the like.

The term "heteroalkyl" by itself or in combination with another term means a stable linear or branched alkyl radical consisting of a certain number of carbon atoms and at least one heteroatom or heteroatom group or a combination thereof. In some embodiments, the heteroatom is selected from B, O, N, and S, wherein the nitrogen and sulfur atoms are optionally oxidized, and the nitrogen heteroatom is optionally quaternized. In other embodiments, the heteroatom group is selected from -C(=O)O-, -C(=O)-, -C(=S)-, -S(=O), -S(=O) ₂ -, -C(=O)N(H)-, -N(H)-, -C(=NH)-, -S(=O) ₂ N(H)- and -S(=O)N( H)-. In some embodiments, the heteroalkyl is C _1-6 heteroalkyl; in other embodiments, the heteroalkyl is C _1-3 heteroalkyl. The heteroatom or heteroatom group may be located at any internal position of the heteroalkyl group, including the attachment position of the alkyl group to the rest of the molecule, but the terms "alkoxy", "alkylamino" and "alkylthio" (or thioalkane Oxygen) is a conventional expression and refers to those alkyl groups that are connected to the rest of the molecule through an oxygen atom, an amino group, or a sulfur atom, respectively. Examples of heteroalkyl groups include, but are not limited to, -OCH ₃ , -OCH ₂ CH ₃ , -OCH ₂ CH ₂ CH ₃ , -OCH ₂ (CH ₃ ) ₂ , -CH ₂ -CH ₂ -O-CH ₃ , -NHCH ₃ , -N(CH ₃ ) ₂ , -NHCH ₂ CH ₃ , -N(CH ₃ )(CH ₂ CH ₃ ), -CH ₂ -CH ₂ -NH-CH ₃ , -CH ₂ -CH ₂ -N( CH ₃ )-CH ₃ , -SCH ₃ , -SCH ₂ CH ₃ , -SCH ₂ CH ₂ CH ₃ , -SCH ₂ (CH ₃ ) ₂ , -CH ₂ -S-CH ₂ -CH ₃ , -CH _2- CH ₂ , -S(=O)-CH ₃ , -CH ₂ -CH ₂ -S(=O) ₂ -CH ₃ , and. Up to two heteroatoms may be consecutive, for example, -CH ₂ -NH-OCH _3.

Unless otherwise specified, the term "C _1-6 alkoxy" refers to those alkyl groups containing 1 to 6 carbon atoms attached to the rest of the molecule through one oxygen atom. The C _1-6 alkoxy group includes 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. . Examples of 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, etc.

Unless otherwise specified, the term "C _1-3 alkoxy" refers to those alkyl groups containing 1 to 3 carbon atoms connected to the rest of the molecule by one oxygen atom. The C _1-3 alkoxy group includes C _1-2 , C _2-3 , C ₃ and C ₂ alkoxy groups 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.

Unless otherwise specified, the term "C _1-6 alkylamino" refers to those alkyl groups containing 1 to 6 carbon atoms that are attached to the rest of the molecule through an amino group. The C _1-6 alkylamino group 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 alkyl amino groups Wait. Examples of C _1-6 alkylamino 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 ) ( CH ₂ CH ₃ ), -NHCH ₂ CH ₂ CH ₃ , -NHCH ₂ (CH ₃ ) ₂ , -NHCH ₂ CH ₂ CH ₂ CH _3, etc.

Unless otherwise specified, the term "C _1-3 alkylamino" refers to those alkyl groups containing 1 to 3 carbon atoms attached to the rest of the molecule through an amino group. The C _1-3 alkylamino group includes 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 etc.

Unless otherwise specified, the term "C _1-6 alkylthio" refers to those alkyl groups containing 1 to 6 carbon atoms connected to the rest of the molecule through a sulfur atom. The C _1-6 alkylthio group 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 alkane Sulfur-based. Example C _{1- 6} alkylthio include, but are not limited to, _{-SCH 3, -SCH 2 CH 3,} -SCH 2 CH 2 CH 3, -SCH 2 (CH 3) 2 and the like.

Unless otherwise specified, the term "C _1-3 alkylthio" refers to those alkyl groups containing 1 to 3 carbon atoms connected to the rest of the molecule through a sulfur atom. The C _1-3 alkylthio group includes C _1-3 , C _1-2 and C ₃ alkylthio groups. Examples of C _1-3 alkylthio groups include, but are not limited to, -SCH ₃ , -SCH ₂ CH ₃ , -SCH ₂ CH ₂ CH ₃ , -SCH ₂ (CH ₃ ) ₂ and the like.

Unless otherwise specified, "C _2-6 alkynyl" is used to denote a linear or branched hydrocarbon group consisting of at least one carbon-carbon triple bond consisting of 2 to 6 carbon atoms, a carbon-carbon triple bond It can be located anywhere on the group. The C _2-6 alkynyl group includes C _2-4 , C _2-3 , C ₄ , C ₃ and C ₂ alkynyl groups. It can be monovalent, divalent or multivalent. Examples of C _2-6 alkynyl include, but are not limited to, ethynyl, propynyl, butynyl, pentynyl, and the like.

Unless otherwise specified, the term "5-6 membered heterocycloalkyl" by itself or in combination with other terms means a saturated cyclic group consisting of 5 to 6 ring atoms, with 1, 2, 3 or 4 ring atoms Are heteroatoms independently selected from O, S, and N, and the rest are carbon atoms, wherein nitrogen atoms are optionally quaternized, and nitrogen and sulfur heteroatoms may be optionally oxidized (ie, NO and S(O) _p , p Is 1 or 2). It includes single-ring and double-ring systems, where the double-ring system includes spiro ring, parallel ring and bridge ring. In addition, as far as the "5-6 membered heterocycloalkyl group" is concerned, the hetero atom may occupy the connection position of the heterocyclic alkyl group to the rest of the molecule. The 5-6 membered heterocycloalkyl includes 5-membered and 6-membered heterocycloalkyl. Examples of 5-6 membered heterocycloalkyl include, but are not limited to, pyrrolidinyl, pyrazolidinyl, imidazolidinyl, tetrahydrothienyl (including tetrahydrothien-2-yl and tetrahydrothien-3-yl, etc.) , Tetrahydrofuranyl (including tetrahydrofuran-2-yl, etc.), tetrahydropyranyl, piperidinyl (including 1-piperidinyl, 2-piperidinyl and 3-piperidinyl, etc.), piperazinyl (including 1 -Piperazinyl and 2-piperazinyl, etc.), morpholinyl (including 3-morpholinyl and 4-morpholinyl, etc.), dioxanyl, dithianyl, isoxazolidinyl, isothiazole Alkyl, 1,2-oxazinyl, 1,2-thiazinyl, hexahydropyridazinyl, homopiperazinyl or homopiperidinyl, etc.

Unless otherwise specified, the terms "C _6-10 aromatic ring" and "C _6-10 aryl group" of the present invention may be used interchangeably. The term "C _6-10 aromatic ring" or "C _6-10 aryl group" means A cyclic hydrocarbon group consisting of 6 to 10 carbon atoms with a conjugated π-electron system. It can be a monocyclic ring, fused bicyclic ring or fused tricyclic ring system, where each ring is aromatic. It may be monovalent, divalent, or polyvalent, and C _6-10 aryl groups include C _6-9 , C ₉ , C _10, and C ₆ aryl groups. Examples of C _6-10 aryl groups include, but are not limited to, phenyl, naphthyl (including 1-naphthyl, 2-naphthyl, etc.).

Unless otherwise specified, the terms "5-10 membered heteroaryl ring" and "5-10 membered heteroaryl group" of the present invention may be used interchangeably. The term "5-10 membered heteroaryl group" means from 5 to 10 rings Atom-containing cyclic groups with a conjugated π-electron system. 1, 2, 3, or 4 ring atoms are heteroatoms independently selected from O, S, and N, and the rest are carbon atoms. It can be a monocyclic ring, fused bicyclic ring or fused tricyclic ring system, where each ring is aromatic. Where nitrogen atoms are optionally quaternized, nitrogen and sulfur heteroatoms can be optionally oxidized (ie NO and S(O) _p , p is 1 or 2). The 5-10 membered heteroaryl group can be attached to the rest of the molecule through a heteroatom or carbon atom. The 5-10 membered heteroaryl group includes 5-8 membered, 5-7 membered, 5-6 membered, 5 membered, and 6 membered heteroaryl groups. Examples of the 5-10 membered heteroaryl include, but are not limited to, pyrrolyl (including N-pyrrolyl, 2-pyrrolyl, and 3-pyrrolyl, etc.), pyrazolyl (including 2-pyrazolyl and 3-pyryl Oxazolyl, 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-triazolyl, etc.), tetrazolyl, isoxazolyl (3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, etc.), thiazolyl (including 2-thiazolyl , 4-thiazolyl and 5-thiazolyl, etc.), furyl (including 2-furanyl and 3-furanyl, etc.), thienyl (including 2-thienyl and 3-thienyl, etc.), pyridyl (including 2 -Pyridyl, 3-pyridyl, 4-pyridyl, etc.), pyrazinyl, pyrimidinyl (including 2-pyrimidyl and 4-pyrimidyl, etc.), benzothiazolyl (including 5-benzothiazolyl, etc.) , Purinyl, benzimidazolyl (including 2-benzimidazolyl, etc.), benzoxazolyl, indolyl (including 5-indolyl, etc.), isoquinolinyl (including 1-isoquinolinyl And 5-isoquinolinyl, etc.), quinoxalinyl (including 2-quinoxalinyl and 5-quinoxalinyl, etc.) or quinolinyl (including 3-quinolinyl and 6-quinolinyl, etc.) .

Unless otherwise specified, C _n-n+m or C _n -C _n+m includes any specific case of n to n+m carbons, for example, C _1-6 includes C ₁ , C ₂ , C ₃ , C ₄ , C ₅ and C ₆ , including any range from n to n+m, for example, C _1-6 includes C _1-3 , C _1-6 , C _1-4 , C _3-6 , C _{3- 5} , C _{2- 5} and C _1-5, etc.; in the same way, n to n+m means that the number of atoms in the ring is n to n+m, for example, 5-6 member ring includes 5 member ring and 6 member ring .

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, the embodiments formed by the combination with other chemical synthesis methods and well known to those skilled in the art Equivalently, preferred embodiments include but are not limited to the embodiments of the present invention.

The present invention uses the following abbreviations: CDCl ₃ stands for deuterated chloroform; CD ₃ OD stands for deuterated methanol; DMSO-d ₆ stands for deuterated dimethyl sulfoxide; Bz stands for benzoyl; TBS stands for tert-butyl dimethyl Silyl; DMTr stands for 4,4'-bismethoxytrityl; CE stands for cyanoethyl; BSA stands for N,O-bistrimethylsilylacetamide; DMTrCl stands for 4,4'-bismethoxytris Benzyl chloride; DIAD stands for diisopropyl azodicarboxylate; DDTT stands for (E)-N,N-dimethyl-N'-(3-thio-3H-1,2,4-dithiazole -5-yl)formamidine; ADDP stands for azodicarbonyl dipiperidine.

Compound detection:

HPLC: Column: YMC-Pack ODS-A 150*4.6mm, 5um, mobile phase: water (0.06875% trifluoroacetic acid)-acetonitrile (0.0625% trifluoroacetic acid); flow rate: 1.0mL/min; detection wavelength: UV 220nm&215nm&254nm; column temperature: 40℃.

detailed description

The following describes the application in detail through examples, but it does not mean that there are any disadvantageous restrictions for the application. This application has described this application in detail, and its specific embodiments are also disclosed. For those skilled in the art, various changes and improvements are made to the specific implementation of this application without departing from the spirit and scope of this application Will be obvious.

Example 1: Preparation of compounds 1A and 1B

Step 1: Preparation of compound 1-2

At 0°C, compound 1-1 (3g, 8.04mmol) was dissolved in pyridine (25mL), DMTrCl (2.99g, 8.84mmol) was added, and the reaction system was heated to 25°C and stirred for 48h. After concentration under reduced pressure, a solid was obtained. The solid was dissolved in water (60 mL) and extracted with ethyl acetate (40 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (Petroleum ether/ethyl acetate (v/v) = 1/0 ~ 1/9) to give compound 1-2.

MS(ESI)m/z(M+H) ⁺ =676.2.

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 11.26 (s, 1H), 8.74 (s, 1H), 8.62 (s, 1H), 8.05 (d, J=7.2 Hz, 2H), 7.68-7.63 ( m, 1H), 7.59-7.53 (m, 2H), 7.32 (d, J = 7.2Hz, 2H), 7.26-7.18 (m, 7H), 6.86-6.79 (m, 4H), 6.43 (d, J = 19.2Hz, 1H), 5.76(d, J=6.8Hz, 1H), 5.74-5.60(m, 1H), 4.92-4.78(m, 1H), 4.15-4.13(m, 1H), 3.71(s, 6H ), 3.32-3.31 (m, 2H).

Step 2: Preparation of compound 1-3

At 5℃, compound 1-2 (2g, 2.96mmol) and imidazole (302.25mg, 4.44mmol) were dissolved in N,N-dimethylformamide (10mL), and then tert-butyldimethylchlorosilane was added (669.18mg, 4.44mmol), the reaction system was heated to 25 ℃ and stirred for 24h. Dilute with water (60 mL), extract with ethyl acetate (20 mL x 3), combine the organic phases, dry over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure, and purify the crude product by silica gel column chromatography (petroleum ether/ethyl acetate (v/ v) = 1/0 ~ 1/1) to give compound 1-3.

MS(ESI)m/z(M+H) ⁺ =790.3.

¹ H NMR (400 MHz, DMSO-d ₆ ) δ 11.27 (s, 1H), 8.79 (s, 1H), 8.69 (s, 1H), 8.03 (d, J=7.2 Hz, 2H), 7.68-7.62 ( m, 1H), 7.57-7.52 (m, 2H), 7.30-7.25 (m, 2H), 7.22-7.13 (m, 7H), 6.80 (d, J=8.4Hz, 4H), 6.48-6.41 (m, 1H), 5.88-5.72 (m, 1H), 5.23-5.12 (m, 1H), 4.08 (br d, J = 7.6Hz, 1H), 3.71 (s, 6H), 3.33-3.23 (m, 1H), 3.06 (dd, J = 4.0, 10.8 Hz, 1H), 0.81 (s, 9H), 0.11 (s, 3H), 0.02 (s, 3H).

Step 3: Preparation of compound 1-4

At 0°C, under argon protection, compound 1-3 (3.9g, 4.94mmol) was dissolved in tetrahydrofuran (45mL), and trans 1,4-butenediol (869.94mg, 9.87mmol, 813.02μL) was added in sequence. With triphenylphosphine (2.59g, 9.87mmol), then DIAD (2.00g, 9.87mmol, 1.92mL) was added dropwise. After the addition, the reaction was heated to 10°C and stirred for 16h. Dilute with water (300mL), extract with ethyl acetate (100mL x 2), combine the organic phases, dry over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure, and purify the crude product by silica gel column chromatography (petroleum ether/ethyl acetate (v/ v) = 1/0 ~ 1/1) to give compound 1-4.

MS(ESI)m/z(M+H) ⁺ =860.3.

Step 4: Preparation of compound 1-5

At 5°C, compound 1-1 (2g, 5.36mmol) and imidazole (547.04mg, 8.04mmol) were dissolved in N,N-dimethylformamide (10mL), and tert-butyldimethylchlorosilane ( 888.17mg, 5.89mmol), after addition, the reaction was warmed to 25 ℃ and stirred for 24h. Dilute with water (50mL), extract with ethyl acetate (200mL x 3), combine organic phases, dry over anhydrous sodium sulfate, filter, concentrate the filtrate under reduced pressure, and purify the crude product by silica gel column chromatography (petroleum ether/ethyl acetate (v/ v) = 1/0～1/1) to give compound 1-5.

MS(ESI)m/z(M+H) ⁺ = 488.2.

¹ H NMR (400 MHz, CDCl ₃ ) δ 8.70 (s, 1H), 8.30 (s, 1H), 7.94-7.91 (m, 2H), 7.53-7.48 (m, 1H), 7.44-7.39 (m, 2H ), 6.30 (dd, J=2.4, 15.6 Hz, 1H), 5.38-5.22 (m, 1H), 4.63-4.55 (m, 1H), 4.12-4.08 (m, 1H), 3.96 (dd, J=2.4 , 11.8Hz, 1H), 3.79 (dd, J = 2.8, 11.8Hz, 1H), 0.80 (s, 9H), 0.00 (s, 6H).

Step 5: Preparation of compound 1-6

At 0°C, under argon protection, compounds 1-4 (3g, 3.49mmol), 1-5 (1.70g, 3.49mmol) and triphenylphosphine (1.83g, 6.98mmol) were dissolved in tetrahydrofuran (40mL), DIAD (1.41g, 6.98mmol, 1.36mL) was added dropwise. After the addition was completed, the reaction was warmed to 10°C and stirred for 16h. Dilute with water (300mL), extract with ethyl acetate (100mL x 3), combine the organic phases, dry over anhydrous sodium sulfate, filter, and concentrate the filtrate under reduced pressure. The crude product is purified by silica gel column chromatography (petroleum ether/ethyl acetate (v/ v) = 1/0 ~ 5/3) to give compound 1-6.

MS (ESI) m/z (M+H) ⁺ = 1351.7.

Step 6: Preparation of compounds 1-7

At 10°C, compound 1-6 (4.8g, 3.61mmol) was dissolved in dichloromethane (50mL), water (650.35mg, 36.10mmol, 650.35μL) was added, and then dichloroacetic acid (2.47g, 10.83) was added dropwise mmol) (5% dichloroacetic acid solution), after the addition is complete, the reaction is stirred for 30 min. Triethylsilane (4.20g, 36.10mmol, 5.77mL) was added, and the reaction was continued to stir for 30min. Diluted with dichloromethane (100 mL), the organic phase was washed with saturated sodium bicarbonate solution (50 mL x 3), dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography (petroleum ether/ethyl acetate) Ester (v/v) = 1/0 ~ 5/3) to give compound 1-7.

MS (ESI) m/z (M+H) ⁺ = 1027.5.

Step 7: Preparation of compounds 1-8

At 10°C, under argon protection, compound 1-7 (386.30mg, 376.04μmol) was dissolved in tetrazole in acetonitrile solution (0.45M, 25.00mL), 4A molecular sieve (1g) was added, and then 2- A solution of cyanoethyl N,N,N',N'-tetraisopropylphosphoramidite (226.69 mg, 752.09 μmol, 238.87 μL) in acetonitrile (0.5 mL) was stirred and reacted for 2 h. Add DDTT to continue the reaction for 1h. Dilute with dichloromethane (100 mL), wash with water (50 mL x 2), dry the organic phase over anhydrous sodium sulfate, filter, and concentrate the filtrate under reduced pressure. The crude product is purified by silica gel column chromatography (dichloromethane/methanol (v/v ) = 20/1) to give compound 1-8.

MS (ESI) m/z (M+H) ⁺ = 1158.4.

Step 8: Preparation of compounds 1-9

At 10°C, compound 1-8 (450 mg, 388.48 μmol) was dissolved in tetrahydrofuran (12 mL), and acetic acid (699.87 mg, 11.65 mmol, 666.54 μL) and tetrabutylammonium fluoride (1M tetrahydrofuran solution, 1.17 mL) were added ), the reaction was stirred for 12h. Diluted with dichloromethane (100mL), the organic phase was washed with saturated sodium bicarbonate solution (30mL x 2), saturated brine (30mL x 2), dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the crude product was passed through a silica gel column Chromatographic purification (dichloromethane/methanol (v/v) = 20/1) to give compound 1-9.

MS(ESI)m/z(M+H) ⁺ =930.3.

Step 9: Preparation of compounds 1-10

At 18°C, under argon protection, compound 1-9 (300mg, 322.64μmol) was dissolved in acetonitrile (3mL), followed by 4A molecular sieve (500mg), tetrazolium (0.45M acetonitrile solution, 14.03mL), then A solution of 2-cyanoethyl N,N,N',N'-tetraisopropylphosphoramidite (145.87 mg, 483.95 μmol, 153.71 μL) in acetonitrile (0.5 mL) was added dropwise, and the reaction was stirred for 1 h. Diluted with ethyl acetate (30mL), filtered, the filtrate was washed with water (30mL), saturated brine (30mL), dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, the crude product was purified by silica gel column chromatography (dichloromethane / Methanol (v/v) = 20/1) to give compound 1-10, the product was used in the next reaction without further purification. MS (ESI) m/z (M + H) ⁺ = 1029.1.

³¹ P NMR (162MHz, CD ₃ CN) δ 139.77, 136.11, 63.77, 63.36.

Step 10: Preparation of compounds 1-11

At 0°C, under argon protection, compound 1-10 (350mg, 340.18μmol) was dissolved in dichloromethane (8mL), followed by 4A molecular sieve (500mg), borane dimethyl sulfide complex (2M di Chloromethane solution, 680.36 μL), and stirred for 10 min. The reaction was quenched with water (0.5 mL), diluted with dichloromethane (20 mL), filtered, the filtrate was washed with water (30 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give crude product 1-11, without further purification of the product , Directly used in the next reaction.

Step 10: Preparation of compounds 1A and 1B

Compound 1-11 (340 mg, 326.07 μmol) was dissolved in methylamine (30.69 mg, 326.07 μmol, 10 mL, 33% ethanol solution), and the reaction was stirred at 25° C. for 48 h. After concentration under reduced pressure, the residue was dissolved in water (20mL), ethyl acetate (30mL) was back extracted, and the aqueous phase was lyophilized to obtain a crude product. The crude product was separated by high-efficiency preparation liquid separation (separation conditions: chromatographic column: Xbridge: Prep OBD C18 150*30mm 10 μm; mobile phase: [water (10 mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 0%-30%, flow rate: 25 mL/min, 11 min). Compound 1A (HPLC retention time 6.51 min) and compound 1B (HPLC retention time 7.29 min) were obtained.

1A:

MS(ESI)m/z(M+H) ⁺ =729.3.

¹ H NMR (400MHz, D ₂ O) δ 8.39-7.57 (m, 4H), 6.23 (br s, 2H), 6.04-5.23 (m, 3H), 5.20-4.74 (m, 1H), 4.56-4.03 (m,8H), 3.99-3.10(m,4H), 0.61--0.49(m,3H).

³¹ P NMR (162MHz, D ₂ O) δ 94.06-92.76, 55.53-53.99.

¹⁹ F NMR (376MHz, D ₂ O) δ-202.19--205.59.

1B:

MS (ESI) m/z (M+H) ⁺ = 729.1.

¹ H NMR (400MHz, D ₂ O) δ 8.56-7.59 (m, 4H), 6.17 (br s, 2H), 5.83-5.37 (m, 1H), 5.36-5.09 (m, 2H), 5.05-4.66 (m, 2H), 4.56-4.11 (m, 7H), 4.05-3.40 (m, 4H), 0.60--0.24 (m, 3H).

³¹ P NMR (162MHz, D ₂ O) δ 96.36-95.15, 55.61-54.04.

¹⁹ F NMR (376MHz, D ₂ O) δ-203.13--206.29.

Example 2: Preparation of compounds 2A, 2B, 2C and 2D

Step 1: Preparation of compound 2-1

At 0°C, under argon protection, compound 1-3 (7g, 8.86mmol) was dissolved in tetrahydrofuran (70mL), and 1,4-butynediol (1.53g, 17.72mmol) and ADDP (4.47g, 17.72mmol), after addition, tri-tert-butylphosphine (3.59g, 17.72mmol, 4.37mL) was added dropwise, and the temperature was raised to 30°C for 20h. The solid was filtered off, the filtrate was diluted with ethyl acetate (300 mL), the organic phase was washed with saturated aqueous sodium bicarbonate solution (150 mL x 2), saturated brine (150 mL x 2), dried over anhydrous sodium sulfate, filtered, and the filtrate was depressurized After concentration, the crude product was purified by silica gel column chromatography (petroleum ether/ethyl acetate (v/v) = 1/0 ~ 3/1) to give compound 2-1.

MS (ESI) m/z (M+H) ⁺ = 858.4.

¹ H NMR (400 MHz, CDCl ₃ ) δ 8.63 (s, 1H), 8.15 (s, 1H), 7.48-7.40 (m, 2H), 7.35-7.30 (m, 2H), 7.27-7.16 (m, 8H ), 7.11-6.98(m, 2H), 6.76(d, J=8.8Hz, 4H), 6.26-6.14(m, 1H), 5.75-5.50(m, 1H), 4.15(s, 1H), 4.02( br d, J = 6.0 Hz, 2H), 3.86-3.74 (m, 6H), 2.05 (s, 2H), 0.94-0.79 (m, 9H), 0.17-0.06 (m, 6H).

Step 2: Preparation of compound 2-2

At 10°C, under argon protection, compound 2-1 (4.8g, 5.59mmol) was dissolved in tetrahydrofuran (50mL), and compound 1-5 (3.55g, 7.27mmol) and triphenylphosphine (4.40g, 16.78mmol), after addition, DIAD (3.39g, 16.78mmol, 3.26mL) was added dropwise, and the reaction was carried out at this temperature for 16h. The reaction solution was diluted with ethyl acetate (300 mL), washed sequentially with saturated aqueous sodium bicarbonate solution (100 mL x 3), saturated brine (100 mL x 3), the organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was slurried with petroleum ether/ethyl acetate (100mL/30mL) to remove a large amount of triphenylphosphine oxide, the mother liquor was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography (petroleum ether/ethyl acetate (v/v) = 1/0 ~ 2/3) to give compound 2-2.

MS (ESI) m/z (M+H) ⁺ = 1327.6.

Step 3: Preparation of compound 2-3

At 10°C, compound 2-2 (3.5 g, 2.64 mmol) was dissolved in methylene chloride (40 mL), dichloroacetic acid (1.81 g, 7.91 mmol, 5% methylene chloride solution) was added, and the reaction solution was stirred for 1 h After that, triethylsilane (3.07g, 26.36mmol, 4.21mL) was added, and the reaction system was continued to stir for 15min. Diluted with ethyl acetate (200 mL), the organic phase was washed with saturated sodium bicarbonate solution (100 mL x 3), dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the crude product was purified by silica gel column chromatography (petroleum ether/ethyl acetate) Ester (v/v) = 1/0 to 2/3) to give compound 2-3.

MS(ESI)m/z(M+H) ⁺ = 1025.6.

¹ H NMR (400 MHz, CDCl ₃ ) δ 8.45 (s, 1H), 8.31 (s, 1H), 8.24 (s, 1H), 7.96 (s, 1H), 7.52-7.42 (m, 5H), 7.36 ( d, J = 7.2 Hz, 1H), 7.31-7.21 (m, 5H), 7.20-7.10 (m, 2H), 6.34 (dd, J = 2.4, 15.6 Hz, 1H), 6.10 (dd, J = 6.8, 11.6Hz, 1H), 5.85-5.61(m, 1H), 5.31(s, 1H), 5.08(s, 2H), 4.99-4.92(m, 2H), 4.65(br d, J=3.2Hz, 2H) , 4.27 (s, 1H), 4.19 (br d, J = 5.2 Hz, 1H), 4.05 (dd, J = 2.4, 11.6 Hz, 1H), 3.97 (br d, J = 12.8 Hz, 1H), 3.90 ( dd, J = 2.8, 11.6 Hz, 1H), 3.75 (s, 1H), 0.93 (d, J = 16.8 Hz, 18H), 0.20-0.04 (m, 12H).

Step 4: Preparation of compound 2-4

Argon protection, at 15 ℃, compound 2-3 (1.6g, 1.56mmol), 4A molecular sieve (2g) and tetrazole (0.45M acetonitrile solution, 52.02mL) were dispersed in acetonitrile (5mL), added dropwise 2-cyanoethyl N,N,N',N'-tetraisopropylphosphoramidite (940 mg, 3.12 mmol, 991 μL), the reaction system was stirred at this temperature for 2 h. DDTT (961mg, 4.68mmol) was added and the reaction was stirred for 1h. The reaction solution was diluted with dichloromethane (200 mL) and filtered. The filtrate was washed with saturated sodium bicarbonate solution (100 mL x 2) and saturated brine (100 mL x 2), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (dichloromethane/methanol (v/v) = 20/1) to give compound 2-4.

MS (ESI) m/z (M+H) ⁺ = 1156.5.

³¹ P NMR (162MHz, D ₂ O) δ 67.67, 65.17.

¹⁹ F NMR (376MHz, D ₂ O) δ-201.34--202.18.

Step 5: Preparation of compound 2-5

At 15°C, compound 2-4 (1.2g, 1.04mmol) was dissolved in tetrahydrofuran (10mL), acetic acid (1.87g, 31.13mmol, 1.78mL) was added, and then tetrabutylammonium fluoride (1M tetrahydrofuran) was added dropwise Solution, 4.15mL), after stirring the reaction solution for 24h, dichloromethane (50mL) was added to dilute, the organic phase was washed with saturated sodium bicarbonate solution (20mL x 2), dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, crude product Purified by silica gel column chromatography (dichloromethane/methanol (v/v) = 1/0 to 25/1) to obtain compound 2-5.

MS(ESI)m/z(M+H) ⁺ =928.2.

³¹ P NMR (162MHz, D ₂ O) δ 65.63, 64.96.

¹⁹ F NMR (376MHz, D ₂ O) δ-200.71--200.85.

Step 6: Preparation of compound 2-6

Argon protection, at 15 ℃, compound 2-5 (530mg, 571.23μmol), 4A molecular sieve (1g) and tetrazole (0.45M acetonitrile solution, 25.39mL) were dispersed in acetonitrile (2mL), added dropwise 2 -Cyanoethyl N,N,N',N'-tetraisopropylphosphoramidite (344 mg, 1.14 mmol, 362.85 μL), and the reaction system was stirred and reacted for 1 h. The reaction solution was diluted with ethyl acetate (60 mL) and filtered. The filtrate was washed with saturated sodium bicarbonate solution (20 mL x 2) and saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. Preparative thin layer chromatography (dichloromethane/methanol (v/v)=20/1) was isolated and purified to obtain compound 2-6.

MS(ESI)m/z(M+H) ⁺ =1027.2.

Step 7: Preparation of compounds 2-7

At 0°C, borane dimethyl sulfide complex (2M in dichloromethane, 555 μL) was added dropwise to compound 2-6 (380 mg, 370.06 μmol) and 4A molecular sieve (0.5 g) in dichloromethane (25 mL) In the solution, the reaction system was added and stirred at 0°C for 15 min. The reaction solution was filtered to remove molecular sieves, and the filtrate was washed with saturated sodium bicarbonate solution (50 mL x 3) and saturated brine (50 mL x 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain crude product 2-7, without After further purification, it was directly used in the next reaction.

Step 8: Preparation of compounds 2A, 2B, 2C and 2D

Compound 2-7 (380 mg, 365.14 μmol) was dissolved in 30% methylamine ethanol solution (6 mL), and the reaction was stirred at 15° C. for 16 h. The reaction system was concentrated under reduced pressure. The residue was redissolved in 30% methylamine ethanol solution (6 mL) and stirred at 40°C for 10 h. The reaction system was concentrated under reduced pressure. The residue was dissolved in water (30 mL) and ethyl acetate (10 mL) x3) Back extraction, the aqueous phase is concentrated, and the resulting crude product is separated by two high-efficiency preparation liquid separation conditions (separation conditions: 1) Chromatography column: Xbridge Prep OBD C18 150*40mm 10μm; mobile phase: [water (10mM ammonium bicarbonate)- Acetonitrile]; Acetonitrile%: 0%-25%, flow rate: 25mL/min, 20min); 2) Chromatography column: Xbridge Prep OBD C18 150*40mm 10μm; Mobile phase: [water (0.05% ammonium hydroxide)-acetonitrile] ; Acetonitrile%: 0%-10%, flow rate: 25mL/min, 12min). Get:

Compound 2A (HPLC retention time 6.13min)

Compound 2B (HPLC retention time 6.51min)

Compound 2C (HPLC retention time 6.41min)

Compound 2D (HPLC retention time 6.95min)

Compound 2A:

MS (ESI) m/z (M+H) ⁺ = 726.9.

¹ H NMR (400 MHz, D ₂ O) δ 8.33 (s, 1H), 8.25 (s, 1H), 7.64 (br s, 1H), 7.54 (br s, 1H), 6.25 (dd, J=12.8, 15.2Hz, 2H), 6.12-5.80(m, 1H), 5.75-5.49(m, 1H), 4.94-4.75(m, 2H), 4.48-4.07(m, 6H), 4.01-3.68(m, 4H) , 0.70--0.46 (m, 3H).

³¹ P NMR (162MHz, D ₂ O) δ 95.01-94.02, 53.70.

¹⁹ F NMR (376MHz, D ₂ O) δ-203.04--203.76.

Compound 2B:

MS (ESI) m/z (M+H) ⁺ = 726.9.

¹ H NMR (400 MHz, D ₂ O) δ = 8.37-8.09 (m, 2H), 7.72 (br s, 1H), 7.48 (br s, 1H), 6.24 (br d, J = 14.8 Hz, 2H), 5.93-5.29(m, 2H), 4.81(br s, 2H), 4.52-4.29(m, 3H), 4.18(br d, J=12.0Hz, 3H), 4.03-3.63(m, 4H), 0.23( br s,3H).

³¹ P NMR (162MHz, D ₂ O) δ 93.40-92.15, 53.62.

¹⁹ F NMR (376MHz, D ₂ O) δ-201.91, -204.18.

Compound 2C:

MS (ESI) m/z (M+H) ⁺ = 726.9.

¹ H NMR (400 MHz, D ₂ O) δ = 8.29 (s, 1H), 8.24 (s, 1H), 7.91 (s, 1H), 7.43 (s, 1H), 6.43-6.16 (m, 2H), 6.12 -5.78(m,1H),5.59-5.28(m,1H),4.88(br s,2H),4.45-4.08(m,6H),3.85(dt,J=5.6,12.4Hz,3H),0.78- -0.28(m,3H).

³¹ P NMR (162MHz, D ₂ O) δ 95.26-91.70, 53.63.

¹⁹ F NMR (376MHz, D ₂ O) δ-202.19--202.37,-202.37--204.53;

Compound 2D:

MS (ESI) m/z (M+H) ⁺ = 727.1.

¹ H NMR (400 MHz, D ₂ O) δ = 8.46-8.05 (m, 2H), 7.94-7.32 (m, 2H), 6.17 (br d, J = 10.4 Hz, 2H), 5.69-5.17 (m, 2H) ), 4.81-4.68 (m, 2H), 4.38-4.11 (m, 6H), 3.79 (br s, 4H), 2.41 (br s, 5H), 0.17 (br s, 3H).

³¹ P NMR (162MHz, D ₂ O) δ 94.26-92.86, 57.11-53.23.

¹⁹ F NMR (376MHz, D ₂ O) δ-203.10--203.50;

Example 3: Preparation of compounds 3A and 3B

Step 1: Preparation of compound 3-1

Argon protection, at 18 ℃, compound 1-7 (1.8g, 1.75mmol), 4A molecular sieve (1g) and tetrazole (0.45M acetonitrile solution, 60mL) were dispersed in acetonitrile (5mL), added dropwise 2 -Cyanoethyl N,N,N',N'-tetraisopropylphosphoramidite (759.20 mg, 2.52 mmol, 0.8 mL), the reaction system was stirred at this temperature for 1 h. The reaction solution was diluted with ethyl acetate (80 mL) and filtered. The filtrate was washed successively with water (50 mL) and saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain crude product 3-1 without further The purification was directly used in the next reaction.

MS (ESI) m/z (M+H) ⁺ = 1126.5.

Step 2: Preparation of compound 3-2

Argon protection, at 0 ℃, borane dimethyl sulfide complex (2M in methylene chloride solution, 2.8mL) was added dropwise compound 3-1 (1.8g, 1.60mmol) and 4A molecular sieve (2g) In methyl chloride (50 mL) solution, the reaction system was added and stirred at 0°C for 10 min. The reaction solution was filtered to remove molecular sieves. The filtrate was washed with water (50 mL) and saturated brine (50 mL) in this order, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain crude product 3-2, which was directly used in the next reaction without further purification. .

MS (ESI) m/z (M+H) ⁺ = 1141.4.

Step 3: Preparation of compound 3-3

At 10°C, compound 3-2 (1.5g, 1.32mmol) was dissolved in pyridine (30mL), triethylamine trihydrofluoride (2.12g, 13.16mmol, 2.14mL) was added, and after the reaction solution was stirred for 8h, Diluted with ethyl acetate (40mL), the organic phase was washed sequentially with water (50mL) and saturated brine (50mL x 2), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was subjected to preparative thin layer chromatography (2 Chloromethane/methanol (v/v)=12/1) was isolated and purified to obtain compound 3-3.

MS (ESI) m/z (M+H) ⁺ = 913.0.

Step 4: Preparation of compound 3-4

Argon protection, at 18 ℃, compound 3-3 (350mg, 383.93μmol), 4A molecular sieve (1g) and tetrazole (0.45M acetonitrile solution, 14.00mL) were dispersed in acetonitrile (5mL) and tetrahydrofuran (1mL) To the mixed solution, add a solution of 2-cyanoethyl N,N,N',N'-tetraisopropylphosphoramidite (189.80mg, 629.71μmol, 0.2mL) in acetonitrile (0.5mL), the reaction system Stir the reaction for 1 h. The reaction solution was diluted with ethyl acetate (50 mL), filtered, and the filtrate was washed with saturated sodium bicarbonate solution (50 mL) and saturated brine (50 mL) in this order, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give the crude product 3- 4. Used for the next reaction without further purification.

Step 5: Preparation of compound 3-5

At 0°C, borane dimethyl sulfide complex (2M dichloromethane solution, 272.73 μL) was added dropwise to compound 3-4 (150 mg, 148.42 μmol) and 4A molecular sieve (0.2 g) in dichloromethane (4 mL ) In the solution, add the reaction system and stir at 0°C for 10 min. The reaction solution was diluted with dichloromethane (20 mL), filtered to remove the molecular sieve, the filtrate was washed with water (30 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product 3-5, which was directly used in the next step without further purification reaction.

Step 6: Preparation of compounds 3A and 3B

Compound 3-5 (150 mg, 146.42 μmol) was dissolved in 30% methylamine ethanol solution (5 mL), and the reaction was stirred at 25°C for 40 h. The temperature was raised to 40°C and stirring was continued for 12h. The reaction system was concentrated under reduced pressure. The residue was dissolved in water (10mL) and back-extracted with ethyl acetate (30mL). The aqueous phase was lyophilized. Conditions: 1) Chromatography column: Xbridge Prep OBD C18 150*40 mm 10μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 0%-40%, flow rate: 25mL/min, 25min); 2) Chromatography column: Xbridge, Prep, OBD, C18, 150*40mm, 10μm; mobile phase: [water (10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 0%-50%, flow rate: 25mL/min, 30min)}. Get:

Compound 3A (HPLC retention time 6.02min)

Compound 3B (HPLC retention time 7.47min)

Compound 3A:

MS (ESI) m/z (MH) ⁺ = 708.7.

¹ H NMR (400MHz, D ₂ O) δ 8.51-7.85 (m, 4H), 6.29 (br s, 2H), 6.12-5.21 (m, 4H), 4.67-4.17 (m, 8H), 3.91 (br s, 2H), 3.83-3.50 (m, 2H), 0.11 (br s, 6H).

³¹ P NMR (162MHz, D ₂ O) δ 94.64-92.92.

¹⁹ F NMR (376MHz, D ₂ O) δ-202.49--209.11.

Compound 3B:

MS (ESI) m/z (MH) ⁺ = 708.7.

¹ H NMR (400MHz, D ₂ O) δ 8.52-7.83 (m, 4H), 6.28 (br s, 2H), 5.83-5.29 (m, 3H), 5.00-4.71 (m, 3H), 4.35 (br s, 6H), 4.01-3.54 (m, 4H), 0.25 (br s, 6H).

³¹ P NMR (162MHz, D ₂ O) δ 97.52–96.14.

¹⁹ F NMR (376MHz, D ₂ O) δ-203.02--206.23.

Comparative Example 4: Preparation of compounds 4A, 4B and 4C

Step 1: Preparation of compound 4-1

At 10°C, under argon protection, compound 1-9 (400mg, 430.18umol) was added to the mixed solution of 4A molecular sieve (500mg) and tetrazolium (0.45M acetonitrile solution, 9.56mL), and then added dropwise 2- A solution of cyanoethyl N,N,N',N'-tetraisopropylphosphoramidite (259.32 mg, 860.36 umol, 273.26 uL) in acetonitrile (2.0 mL) was stirred and reacted for 2 h. DDTT (264.98mg, 1.29mmol) was added to continue stirring the reaction for 1h. The reaction solution was diluted with methylene chloride (100 mL), then washed with water (50 mL x 2), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (dichloromethane/methanol (v/ v) = 20/1) to give compound 4-1.

MS(ESI)m/z(M+H) ⁺ =1061.3

Step 2: Preparation of compounds 4A, 4B and 4C

Compound 4-1 (300 mg, 282.77 umol) was dissolved in methanol (6 mL), ammonia water (5.46 g, 40.51 mmol, 6 mL, 26% aqueous solution) was added, and the reaction was stirred at 50° C. for 8 h. The reaction system was concentrated under reduced pressure, the residue was dissolved in water (10mL), back-extracted with ethyl acetate (5mL x 3), the aqueous phase was lyophilized, and the resulting crude product was subjected to two efficient preparations for liquid phase separation {separation conditions: 1) chromatography column : Xbridge Prep OBD C18 150*40mm 10μm; mobile phase: [water (0.04% ammonia + 10mM ammonium bicarbonate)-acetonitrile]; acetonitrile%: 0%-30%, flow rate: 25mL/min, 15min); 2) chromatography Column: YMC-Actus Triart C18 150*30mm*5um; mobile phase: [water (0.05% ammonia)-acetonitrile]; acetonitrile%: 5%-25%, flow rate: 25mL/min, 15min)}. Get:

Compound 4A (HPLC retention time 6.03min)

Compound 4B (HPLC retention time 6.28min)

Compound 4C (HPLC retention time 6.76min)

Compound 4A:

MS(ESI)m/z(M+H) ⁺ =747.2.

¹ H NMR (400 MHz, D ₂ O) δ 8.53-8.11 (m, 2H), 8.02 (br, s, 2H), 6.28 (br, s, 2H), 6.15-5.77 (m, 1H), 5.45 ( br s, 2H), 4.37 (br s, 7H), 4.04-3.50 (m, 2H), 3.79-3.35 (br s, 2H).

³¹ P NMR (162MHz, D ₂ O) δ 53.78, 53.63.

¹⁹ F NMR (376MHz, D ₂ O) δ-203.49--204.35, -204.53--207.74.

Compound 4B:

MS(ESI)m/z(M+H) ⁺ =747.2.

¹ H NMR (400MHz, D ₂ O) δ 8.49-7.52 (m, 4H), 6.22 (br d, J = 11.2 Hz, 2H), 5.74-5.17 (m, 3H), 4.38 (br s, 8H) ,4.13-3.44(m,1H).

³¹ P NMR (162MHz, MeOD) δ 55.57, 55.14.

¹⁹ F NMR (376MHz, MeOD) δ-203.86--204.04, -206.05--206.18.

Compound 4C:

MS (ESI) m/z (M+H) ⁺ = 747.1.

¹ H NMR (400MHz, D ₂ O) δ 8.31-7.65 (m, 4H), 6.21 (br s, 2H), 5.77-5.23 (m, 3H), 5.01-4.68 (m, 2H), 4.53-4.18 (m,7H),3.94-3.49(m,4H).

³¹ P NMR (162MHz, D ₂ O) δ 55.60, 55.36.

¹⁹ F NMR (376MHz, D ₂ O) δ-201.63--201.75, -206.08--206.19.

Biological activity test experiment

Experimental example 1: STING in vitro binding test experiment

Fluorescence polarization test (fluorescence polarization assay, FP assay) was used to detect the affinity of compounds for human STING protein. There is a certain amount of fluorescein-labeled c-di-GMP and different concentrations of test compounds in the reaction system. When the recombinant human STING C-terminal protein is added, the two small molecules competitively bind to the protein. The bound fluorescein-labeled c-di-GMP rotates slowly in the liquid phase, and the degree of fluorescence polarization detected is also high at this time. The degree of fluorescence polarization is inversely proportional to the concentration and affinity of the test compound. By detecting the magnitude of polarized light in the reaction system, we can accurately know the affinity of the test compound for human STING.

The soluble human STING protein sequence used in the experiment was intercepted from the C-terminal part of human wild-type endoplasmic reticulum binding protein STING, from 140 amino acids to 379 amino acids. Human STING protein has a variety of alleles with different sequence differences. Different alleles have different affinity for CDN (Yi, et.al., "Single Nucleotide Polymorphisms of Human STING can affect immunological responses" to cyclic dinucleotides"PLOSONE.2013, 8(10), e77846). Wild-type STING sequences (G230, R232, R293) account for approximately 57.9% of the total. The N-terminus of the recombinant STING protein is a 6His-SUMO sequence, which facilitates the correct folding and purification of the protein. After protease excision, the C-terminus STING is used for FP testing.

For the FP test, a 384-well plate was used, and a 10 μl reaction system was added with a final concentration of 30 nM fluorescein-labeled c-di-GMP, 10 μM human STING protein, and different concentrations of reference compound or test compound. Centrifuge at 1000g for 1 minute, incubate at room temperature in the dark for 30 minutes, and read the plate with Envision.

The results of the STING in vitro binding assay experiment described above are shown in Table 1.

Table 1

Compound number	FP affinity test IC50 (μM)
2’,3’-cGAMP	6.7
1A	3.1
1B	1.7
2A	5.7
2B	1.9
2C	3.7
2D	1.9
3A	3.3
3B	1.8

Conclusion: In the FP affinity test, the compounds of the present invention showed high affinity for human wild-type STING protein.

Experimental Example 2: THP1-dual reporter gene activity test experiment

The THP1-Dual ^TM cells (InvivoGen catalog code: thpd-nfis) used in the test were constructed by stably integrating two inducible reporter genes in the human monocyte cell line THP1. The promoter sequence of the secreted embryonic alkaline phosphatase (SEAP) reporter gene consists of an IFN-β basic promoter and 5 copies of the NF-κB consensus transcriptional response element upstream (NF-κB consensus transcriptional response element) And 3 copies of the c-Rel binding site. The secreted luciferase (Lucia) reporter gene is driven by five interferon (IFN)-stimulated response elements and an ISG54 basic promoter. This makes it possible to study the two main downstream signaling pathways of STING at the same time: to study the NFκB pathway by detecting the activity of SEAP: and to study the IRF pathway by evaluating the activity of Lucia luciferase.

The compound was diluted with PB buffer (50 mM HEPES, 100 mM KCl, 3 mM MgCl2, 0.1 mM DTT, 85 mM Sucrose, 1 mM ATP, 0.1 mM GTP, 0.2% BSA). Add 20 μL of the reference or test compound to each well of the 96-well plate, and then add 180 μL of THP1-Dual cells (approximately 100,000 cells/well) suspended in PB buffer. After incubating the plate at 37°C under 5% CO ₂ for 30 minutes, centrifuge at 1000 rpm for 10 minutes, discard the supernatant, wash twice with 200 μL/well RPMI-1640, and add 200 μL per well RPMI-1640 for 18 hours. The supernatant was collected and the activation of IRF3 pathway was quantified using QUANTI-Luc ^™ according to the manufacturer's instructions.

The THP1-dual in vitro binding assay results as described above are shown in Table 2.

Table 2

Compound number	EC 50 (μM)
2’,3’-cGAMP	20.19
ADU-S100	23.39
1A	6.2
1B	7.0
2A	41.9
2B	6.5
2C	10.5
2D	11.8
3A	7.4
3B	5.1
4A	30.4
4B	35.3
4C	30.2

Conclusion: In the human monocyte cell line THP1, the compounds of the present invention can activate STING and promote the production of interferon beta.

Experimental Example 3: Raw-Dual reporter gene activity test experiment

The RAW-Dual ^TM cells (InvivoGen catalog code: rawd-ismip) used in the test were constructed by stably integrating two inducible reporter genes in the mouse macrophage cell line RAW264.7: the NF-KB pathway was studied by detecting SEAP activity, And study the IRF3 pathway by evaluating Lucia luciferase activity. The suspension of cells (50,000 cells per well) was added to a 96-well plate (Corning 3599 flat bottom plate) at 200 μL per well, and cultured in a 37°C incubator for 18 to 24 hours. The next day, the culture medium was discarded, and 200 μL of the compound solution prepared with the medium was added to each well, incubated at room temperature for 30 minutes, the treatment liquid was aspirated, washed twice with serum-free culture medium, and then 200 μL of culture medium was added to each well, 37 Cultivate in an incubator for 18 to 24 hours. On the third day, 20 μL of supernatant was taken from each well, and the activation of IRF3 pathway was quantified using QUANTI-LucTM according to the manufacturer's instructions.

The results of the RAW cell activity measurement as described above are shown in Table 3.

table 3

Compound number	Raw, EC50 (μM)
ADU-S100	47.08
1A	28.3
1B	25.0
2A	>100
2B	39.0
2C	41.7
2D	70.0
3A	28.4
3B	27.0

Conclusion: It was found in the test that the compound of the present invention can activate STING in the test experiment of RAW reporter gene test of mouse macrophage cell line.

Claims (18)

1. The compound represented by formula (I), its optical isomer and its pharmaceutically acceptable salt,

   

   among them,

   L ₁ and L ₂ are independently selected from -O-, -N(R)- and -C(RR)-;

   L _{3 is} selected from

   

   R ₁ and R _1a are independently selected from H, halogen, OH, NH ₂ , CN, N ₃ , C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _{1- 6} alkylamino and C _2-6 alkynyl, wherein the C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino or C _2-6 alkynyl Optionally substituted by 1, 2 or 3 R;

   R ₂ and R _2a are independently selected from H, halogen, OH, NH ₂ , CN, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino And C _2-6 alkynyl, wherein the C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino, or C _2-6 alkynyl is optionally substituted 1, 2, or 3 R substitutions;

   R ₃ and R _3a are independently selected from H, halogen, OH, NH ₂ , CN, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino And C _2-6 alkynyl, wherein the C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino, or C _2-6 alkynyl is optionally substituted 1, 2, or 3 R substitutions;

   R ₄ and R _4a are independently selected from H, halogen, OH, NH ₂ , CN, C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino And C _2-6 alkynyl, wherein the C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino, or C _2-6 alkynyl is optionally substituted 1, 2, or 3 R substitutions;

   R ₅ and R _5a are independently selected from H, halogen, OH, NH ₂ , CN, N ₃ , C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _{1- 6} alkylamino and C _3-6 cycloalkyl, wherein the C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino or C _3-6 ring The alkyl group is optionally substituted with 1, 2, or 3 R;

   X _{1 is} selected from BH ₃ ^- and -S(R ₆ );

   R _{6 is} selected from H, CH ₂ OC(═O)R ₇ , CH ₂ OC(═O)OR ₇ , CH ₂ CH ₂ SC(═O)R ₇ and CH ₂ CH ₂ SSCH ₂ R ₇ ;

   R ₇ is selected from C _6-10 aryl, 5-10 membered heteroaryl, C _1-6 heterocycloalkyl and C ₁ - ₂₀ alkyl group, wherein the C ₁ - ₂₀ alkyl optionally substituted with 1,2 , 3, 4 or 5 C _6-10 aryl, C _3-10 cycloalkyl, OH and F substitutions;

   Alternatively, R ₁ and R _{4 are} connected together to form a 5-6 membered heterocycloalkyl;

   Alternatively, R _1a and R _{4a are} joined together to form a 5-6 membered heterocycloalkyl;

   R is independently selected from H, halogen, OH, NH ₂ , CN,

   

   C _1-6 alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino, and C _1-6 alkyl-(C═O)NH-, wherein the C _{1 -6} alkyl, C _1-6 alkoxy, C _1-6 alkylthio, C _1-6 alkylamino or C _1-6 alkyl-(C═O)NH- is optionally substituted by 1, 2 or 3 R'substitution;

   R'is selected from F, Cl, Br, I, OH, NH ₂ and CH ₃ ;

   The 5-6 membered heterocycloalkyl group, 5-10 membered heteroaryl group or C _1-6 heterocycloalkyl group contains 1, 2 or 3 independently selected from -O-, -NH-, -S-,- C(=O)-, -C(=O)O-, -S(=O)-, -S(=O) ₂ -and heteroatoms or heteroatom groups of N.
2. The compound according to claim 1, an optical isomer thereof and a pharmaceutically acceptable salt thereof, wherein R is independently selected from H, halogen, OH, NH ₂ , CN,

   

   C _1-3 alkyl, C _1-3 alkoxy, C _1-3 alkylthio, C _1-3 alkyl-(C=O)NH- and C _1-3 alkylamino, wherein the C _{1 -3} alkyl, C _1-3 alkoxy, C _1-3 alkylthio, C _1-3 alkyl-(C=O)NH- or C _1-3 alkylamino is optionally substituted by 1, 2 or 3 R'replaced.
3. The compound according to claim 2, an optical isomer thereof, and a pharmaceutically acceptable salt thereof, wherein R is independently selected from H, F, Cl, Br, I, OH, NH ₂ , CN, Me,

   

   

   Where Me,

   

   

   It is optionally substituted with 1, 2 or 3 R'.
4. The compound according to claim 3, its optical isomer and its pharmaceutically acceptable salt, wherein R is independently selected from H, F, Cl, Br, I, OH, NH ₂ , CN, Me,

   

   
5. The compound according to any one of claims 1 to 4, an optical isomer thereof, and a pharmaceutically acceptable salt thereof, wherein R ₁ and R _1a are independently selected from H, F, Cl, Br, OH, NH ₂ , CN, N ₃ , C _1-3 alkyl, C _1-3 alkoxy, C _1-3 alkylthio, C _1-3 alkylamino and C _2-3 alkynyl, wherein the C _{1 -3} alkyl, C _1-3 alkoxy, C _1-3 alkylthio, C _1-3 alkylamino or C _2-3 alkynyl is optionally substituted with 1, 2 or 3 R.
6. The compound according to claim 5, an optical isomer thereof, and a pharmaceutically acceptable salt thereof, wherein R ₁ and R _1a are independently selected from H, F, Cl, Br, OH, NH ₂ , CN, N ₃ , Me and

   
7. The compound according to any one of claims 1 to 4, its optical isomer and a pharmaceutically acceptable salt thereof, wherein R ₅ and R _5a are independently selected from H, F, Cl, Br, OH, NH ₂ , CN, N ₃ , C _1-3 alkyl, C _1-3 alkoxy, C _1-3 alkylthio, C _1-3 alkylamino, and C _3-6 cycloalkyl, wherein the C _{The 1-3} alkyl group, C _1-3 alkoxy group, C _1-3 alkylthio group, C _1-3 alkylamino group, or C _3-6 cycloalkyl group is optionally substituted with 1, 2, or 3 R.
8. The compound according to claim 7, an optical isomer thereof, and a pharmaceutically acceptable salt thereof, wherein R ₅ and R _5a are independently selected from H, F, Cl, Br, OH, NH ₂ , CN, N ₃ , Me,

   
9. The compound according to claim 1, its optical isomer and its pharmacologically acceptable salt, wherein R ₁ and R _{4 are} linked together, the structural unit

   
10. The compound according to claim 1, an optical isomer thereof and a pharmaceutically acceptable salt thereof, wherein R _1a and R _{4a are} linked together, and the structural unit

    

    Select from

    
11. The compound according to claim 1, an optical isomer thereof, and a pharmaceutically acceptable salt thereof, wherein L ₁ and L ₂ are independently selected from -O-, -NH-, and -CH _2- .
12. The compound according to claim 1, an optical isomer thereof and a pharmaceutically acceptable salt thereof, wherein L _{3 is} selected from

    
13. The compound according to claim 11 or 12, an optical isomer thereof and a pharmaceutically acceptable salt thereof, wherein the structural unit

    

    Select from

    

    
14. The compound according to any one of claims 1 to 3, its optical isomer and its pharmaceutically effective salt, which is selected from

    

    among them,

    X _{1 is} as defined in claim 1;

    R _{1 is} as defined in claims 1, 5, 6 or 9;

    R _{1a is} as defined in claims 1, 5, 6 or 10;

    R _{4 is} as defined in claim 1 or 9;

    R _{4a is} as defined in claim 1 or 10;

    R ₅ and R _{5a are} as defined in claims 1, 7 or 8;

    L ₁ and L _{2 are} as defined in claims 1, 11 or 13;

    L _{3 is} as defined in claims 1, 12 or 13.
15. The compound of the formula, its optical isomer and its pharmaceutically acceptable salt are selected from:

    

    

    

    

    

    

    

    

    

    

    

    
16. A pharmaceutical composition containing the compound according to any one of claims 1 to 15 or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents or excipient.
17. Use of the compound according to any one of claims 1 to 15 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition according to claim 16 in the preparation of a medicament for preventing and/or treating STING-related diseases.
18. The use according to claim 17, wherein the STING-related diseases are selected from lymphoma, melanoma, colorectal cancer, breast cancer, acute myeloid leukemia, colon cancer, liver cancer, prostate cancer, pancreatic cancer, renal cancer and Glioma, bladder cancer, pleural effusion, malignant pleural effusion, head and neck cancer, fibrosarcoma, renal cell carcinoma, lung cancer, malignant ascites, gastric cancer, ovarian cancer, uterine cancer, optic neuroblastoma, bone cancer, rhabdomyosarcoma, and esophageal cancer.