Hnrnpa2b1 agonist compound and Anti-infection use thereof

Abstract

A fused ring compound represented by formula (I), or a cis-trans isomer, an enantiomer, a diastereoisomer, a racemate, a solvate, a hydrate, a pharmaceutically acceptable salt or a prodrug thereof, a composition and a product containing any one of the foregoing, and the use of the foregoing substances as heterogeneous nuclear ribonucleoprotein A2B1 (hnRNPA2B1) agonist compounds in the preparation of a product for preventing and / or treating infectious diseases and / or diseases and / or symptoms associated with infections.

Description

hnRNPA2B1 agonist compounds and their anti-infective applications Technical Field

[0001] The present disclosure relates to the fields of medicine and biotechnology. Specifically, the present disclosure relates to a class of compounds that act as agonists of the nuclear protein hnRNPA2B1 (i.e., heterogeneous nuclear ribonucleoprotein A2B1), products containing the compounds, and their effects, mechanisms of action, methods of implementation, and uses in preventing or treating infection-related diseases or symptoms and controlling infection-induced damage. Background Art

[0002] Infections, especially viral infections, are common and extremely harmful clinical conditions. Initially, the molecular mechanisms of the body's resistance to viral infection were not fully understood. However, with the discovery of interferons, a class of cytokines, the molecular biological basis of the body's innate and acquired immune cells, as well as their effects, against viral infection, has gradually been understood.

[0003] Interferon (IFN) is a family of cytokines with potent antiviral properties. In 1957, while studying influenza virus infection in chicken embryos, Alick Isaacs and Professor Jean Lindenmann discovered a compound that significantly inhibited the proliferation of the influenza virus. They named it interferon (Isaacs, A. et al., Proc R Soc Lond B Biol Sci. 1957; 927: 258-267). Since then, members of the interferon family of cytokines have been found to possess broad-spectrum and potent antiviral effects. Currently, interferon is widely used clinically to protect against viral infections and treat various diseases caused by them.

[0004] Type I interferons (IFN-I) activate and regulate innate and adaptive immune cells directly or indirectly by inducing other mediators during infection with viruses, bacteria, parasites, and fungi. They are crucial cytokines that are core to the host's defense against pathogens such as viruses. In infected and neighboring cells, type I interferons induce the expression of interferon-stimulated genes (ISGs), inhibiting the further spread of pathogens. Innate immune cells also respond to type I interferons by enhancing antigen presentation and increasing the production of cytokines and chemokines. Adaptive immunity is also influenced by type I interferons: for example, type I interferons can induce B cells to produce specific antibodies and amplify the effector functions of T cells.

[0005] Innate immune receptors recognize pathogen-associated molecular patterns (PAMPs) or damage-associated molecular patterns (DAMPs), inducing the production of cytokines such as type I interferon and inflammatory factors, initiating an innate immune response. Among these PAMPs and DAMPs, pathogen-derived and self-derived nucleic acids are crucial ligands. Therefore, agonists targeting these innate immune recognition receptors have become an important approach for treating infectious diseases.

[0006] Innate immune receptors for nucleic acids include RNA and DNA. DNA recognition receptors are primarily categorized as cytoplasmic and nuclear, depending on their intracellular localization. The primary DNA recognition receptor in the cytoplasm is cyclic GMP-AMP synthase (cGAS). Recognizing DNA in the cytoplasm, cGAS induces activation of the stimulator of interferon genes (STING), which further recruits and activates TANK-binding kinase 1 (TBK1) and interferon-regulatory factor 3 (IRF3), initiating type I interferon production. Additionally, TLR9 recognizes single-stranded DNA containing unmethylated CpG motifs, and AIM2 (Absent in melanoma-2) can also recognize pathogen-derived DNA in the cytoplasm, promoting inflammasome formation and the maturation and release of the inflammatory cytokine IL-1β.

[0007] Studies have identified numerous proteins that can recognize viral DNA and induce IFN-α / β production, including RNA polymerase III, IFI16, DAI, LRRFIP1, LSm14A, MRE11, DNA-PK, HMGBs, DDX41, and cyclic GMP-AMP (cGAMP) synthase (cGAS) (Goubau et al.; Immunity, 2013; 38, 855-869). After recognizing the virus, these PPRs stimulate the host's immune response signaling pathways, inducing the production of large amounts of type I interferons, thereby rapidly establishing the body's first line of defense against infection and resisting pathogen infection. However, only cytoplasmic cGAS and DNA-PK have been confirmed as DNA recognition receptors in mouse experiments. Several other proteins are involved in DNA virus-induced inflammatory responses, including AIM2, IFI16, Rad50, and Sox2. Therefore, research on nuclear DNA recognition needs to be deepened in order to fully and clearly understand the innate immune response against DNA viruses, especially to find the mechanism that links the recognition of exogenous DNA in the nucleus with the activation of extranuclear innate immune signaling.

[0008] Heterogeneous nuclear ribonucleoprotein A2B1 (hnRNP-A2B1) is a newly identified DNA recognition receptor in the nucleus, belonging to the hnRNP family. hnRNP-A2B1 can sense and recognize the nucleic acid components of DNA viruses (such as HSV-1) and self-activate to form homodimers. Demethylation is mediated by the demethylase JMJD6, leading to translocation from the nucleus to the cytoplasm, where it activates the TBK1-IRF3 signaling pathway and triggers the expression of type I interferons, exerting an antiviral effect. Furthermore, as an RNA-binding protein, hnRNP-A2B1 promotes m6A modification, nucleocytoplasmic translocation, and translation of cGAS, IFI16, and STING mRNAs, thereby ensuring sufficient induction of type I interferon expression to combat viral infection.

[0009] In summary, the development of specific agonists targeting hnRNPA2B1 has great potential for treating infectious diseases. There is an urgent need for immunologically active substances that can recognize viral DNA in the cell nucleus, activate interferon production, enhance antiviral effects, effectively resist viral infection, and control the damage caused by viral infection.

[0010] Summary of the Invention

[0011] The present disclosure provides a fused ring compound having a structure shown in formula (I), its related derivatives (e.g., its cis-trans isomers, its enantiomers, its diastereomers, its racemates, its solvates, its hydrates, or its pharmaceutically acceptable salts or prodrugs thereof), and products comprising the compound or its related derivatives. The present disclosure also provides the use of the compound, derivatives, and products in anti-infection, and further provides their use in treating or preventing infectious diseases and related diseases or symptoms. The compounds, drugs, pharmaceutical compositions, or kits disclosed herein can be used to effectively resist infection and control the occurrence of infectious diseases.

[0012] In some aspects of the present disclosure, provided are fused ring compounds represented by formula (I), their cis-trans isomers, their enantiomers, their diastereomers, their racemates, their solvates, their hydrates, or their pharmaceutically acceptable salts or their prodrugs.

[0013] Wherein, L is -(CH2)n-, and n is an integer from 0 to 6;

[0014] X is a halogen;

[0015] R 1 -NR 4 R 5 , where R 4 、R 5 Each independently is H or C 1-6 alkyl;

[0016] R 2 and R 3 Each independently is H or C 1-6 alkyl;

[0017] or R 2 and R 3 Together with the nitrogen atom to which they are attached, they form a 5-8 membered heterocycloalkyl group containing 1-3 heteroatoms selected from nitrogen, oxygen or sulfur;

[0018] The C 1-6 The alkyl group and the 5-8 membered heterocycloalkyl group are unsubstituted or substituted by one or more substituents selected from the group consisting of hydroxy, amino, halogen, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 1-6 Alkoxy, hydroxy C 1-6 Alkyl, C 1-6 Alkylamino, C 3-6 Cycloalkyl, C 3-6 Cycloalkyl C 1-6 Alkyl, 3-6 membered heterocycloalkyl, 3-6 membered heterocycloalkyl C1-6 Alkyl, 6-10 membered aryl, 6-10 membered aryl C 1-6 Alkyl or 5-10 membered heteroaryl and 5-10 membered heteroaryl C 1-6 alkyl.

[0019] In some aspects of the present disclosure, there is provided use of a compound of the present invention in the preparation of a product for preventing and / or treating infectious diseases and / or diseases and / or symptoms associated with infection.

[0020] In some aspects of the present disclosure, a method for preventing and / or treating infectious diseases and / or diseases and / or symptoms associated with infection is also provided, comprising administering a preventively and / or therapeutically effective amount of a compound or product of the present disclosure to a subject in need thereof.

[0021] In some aspects of the present disclosure, compounds or products of the present disclosure are also provided for use in preventing and / or treating infectious diseases and / or diseases and / or symptoms associated with infection.

[0022] In some aspects of the present disclosure, there is provided use of a compound or product of the present disclosure for increasing the level of an interferon (eg, type I interferon, such as IFN-α and / or IFN-β).

[0023] Those skilled in the art may arbitrarily combine the aforementioned technical solutions and technical features without departing from the inventive concept and protection scope of the present disclosure. Other aspects of the present disclosure will be apparent to those skilled in the art from the disclosure herein. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] The present disclosure will be further described below in conjunction with the accompanying drawings, wherein these drawings are only for illustrating the embodiments of the present disclosure and are not intended to limit the scope of the present disclosure.

[0025] Figure 1: Activation of hnRNPA2B1 nuclear export in macrophages by test compounds.

[0026] Figure 2: Effects of test compounds on the activation of TBK1 and IRF3 downstream of hnRNPA2B1 nuclear export in macrophages.

[0027] Figures 3 to 5: Effects of test compounds on the activation of type I interferon in macrophages.

[0028] Figures 6 and 7: Activation effects of test compounds on interferon-stimulated genes in immune cells.

[0029] FIG8 : Inhibitory effect of the test compounds on the replication of herpes simplex virus HSV-1.

[0030] FIG9 : Inhibitory effect of the test compounds on the replication of hepatitis B virus HBV.

[0031] Figure 10: Inhibitory effect of test compounds on the viral replication of vesicular stomatitis virus (VSV). DETAILED DESCRIPTION

[0032] Through extensive research, development, and testing, this application discovered a new class of fused ring compounds that can specifically bind to the hnRNPA2B1 protein with high affinity and can further widely induce the production of higher levels of IFN-β, and are thus defined as hnRNPA2B1 agonists.

[0033] In anti-cell infection tests with different types of infectious agents, the hnRNPA2B1 agonist compounds disclosed herein significantly inhibited the replication of infectious agents, thereby exerting an anti-infective effect. Furthermore, since the hnRNPA2B1 agonist compounds disclosed herein can broadly induce higher levels of IFN-β production, it is anticipated that these compounds may exert anti-infective effects through the action of type I interferons.

[0034] Thus, the present disclosure provides methods and strategies for applying agonist compounds of the novel anti-infective molecule hnRNPA2B1 to inhibit infection, or for the prevention and treatment of infectious diseases, particularly for controlling viral infection, such as liver damage caused by viral infection.

[0035] All numerical ranges provided herein are intended to expressly include all values ​​falling between the endpoints of the ranges and the ranges therebetween. Features disclosed herein or in the embodiments may be combined. All features disclosed herein may be used in any combination, and each feature disclosed herein may be replaced by any alternative feature that serves the same, equivalent, or similar purpose. Therefore, unless otherwise specified, the features disclosed herein are merely general examples of equivalent or similar features.

[0036] As used herein, “containing,” “having,” or “including” encompasses “comprising,” “mainly consisting of,” “substantially consisting of,” and “consisting of”; “mainly consisting of,” “substantially consisting of,” and “consisting of” are subordinate concepts of “containing,” “having,” or “including.”

[0037] hnRNPA2B1 agonist compounds

[0038] As used herein, the terms "hnRNPA2B1 protein (polypeptide)" and "hnRNPA2B1" are used interchangeably to refer to the heterogeneous nuclear riboprotein A2B1. The hnRNPA2B1 protein referred to in the present disclosure can be a protein encoded by the hnRNPA2B1 gene, cDNA, or CDS thereof, in an animal (e.g., human or mouse), or a homologous sequence of such protein that promotes interferon expression (e.g., a homologous sequence of hnRNPA2B1 can be obtained through databases or alignment software known in the art), a variant, or a modified form thereof.

[0039] As used herein, the terms "hnRNPA2B1 gene," "hnRNPA2B1-encoding gene," "hnRNPA2B1 protein-encoding gene," or "hnRNPA2B1-encoding nucleic acid molecule" are used interchangeably to refer to a nucleotide sequence encoding the hnRNPA2B1 protein or polypeptide described herein, such as the human hnRNPA2B1 gene (Gene ID: 3181) or the mouse hnRNPA2B1 gene (Gene ID: 53379). The term also encompasses molecules that hybridize under stringent conditions to a signed nucleic acid molecule or molecules of family genes that are highly homologous to such molecules. Expression of such genes is believed to promote the production and effects of interferon.

[0040] This disclosure provides a novel class of compounds that act as hnRNPA2B1 "agonists" (or "promoters"). The terms "agonist" and "hnRNPA2B1 agonist compound" are used interchangeably to refer to a class of novel fused-ring compounds that can increase the level or activity of hnRNPA2B1. These compounds specifically bind to the hnRNPA2B1 protein with high affinity and can further induce the production of higher levels of IFN-β, thereby exerting an hnRNPA2B1 agonist effect.

[0041] The hnRNPA2B1 agonist compounds disclosed herein can inhibit infection and thus can be further used to prevent or treat diseases associated with infection (especially viral infection) and / or infection-induced symptoms, as well as chronic inflammatory diseases caused by infection and / or their symptoms.

[0042] A fused ring compound represented by formula (I), its cis-trans isomers, its enantiomers, its diastereomers, its racemates, its solvates, its hydrates, or its pharmaceutically acceptable salts or prodrugs thereof,

[0043] Wherein, L is -(CH2)n-, and n is an integer from 0 to 6;

[0044] X is a halogen;

[0045] R 1-NR 4 R 5 , where R 4 、R 5 Each independently is H or C 1-6 alkyl;

[0046] R 2 and R 3 Each independently is H or C 1-6 alkyl;

[0047] or R 2 and R 3 Together with the nitrogen atom to which they are attached, they form a 5-8 membered heterocycloalkyl group containing 1-3 heteroatoms selected from nitrogen, oxygen or sulfur;

[0048] The C 1-6 The alkyl group and the 5-8 membered heterocycloalkyl group are unsubstituted or substituted by one or more substituents selected from the group consisting of hydroxy, amino, halogen, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 1- 6 alkoxy, hydroxy C 1-6 Alkyl, C 1-6 Alkylamino, C 3-6 Cycloalkyl, C 3-6 Cycloalkyl C 1-6 Alkyl, 3-6 membered heterocycloalkyl, 3-6 membered heterocycloalkyl C 1-6 Alkyl, 6-10 membered aryl, 6-10 membered aryl C 1-6 Alkyl or 5-10 membered heteroaryl and 5-10 membered heteroaryl C 1-6 alkyl.

[0049] Preferably, X is F or Cl, more preferably, X is F.

[0050] Preferably, L is -(CH2)n-, wherein n is 1, 2 or 3, more preferably, n is 2.

[0051] Preferably, R 4 、R 5 Each is independently H, methyl or ethyl.

[0052] Preferably, R 2 and R 3 Each independently is C 1-3 Alkyl, preferably methyl, ethyl or isopropyl, said C 1-3 The alkyl group is unsubstituted or substituted with a 3-8 membered heterocycloalkyl group containing 1-2 heteroatoms selected from nitrogen, oxygen or sulfur. More preferably, R 2 and R 3 Each independently is C 1-3Alkyl, preferably methyl, ethyl or isopropyl, said C 1-3 The alkyl group is unsubstituted or substituted with a 4-6 membered heterocycloalkyl group containing one nitrogen heteroatom. Most preferably, R 2 and R 3 Each independently is C 1-3 The alkyl group is preferably methyl, ethyl or isopropyl, and the C1-3 alkyl group is unsubstituted or substituted by pyrrolidinyl.

[0053] Preferably, R 2 and R 3 Together with the nitrogen atom to which they are attached, they form a 5-8 membered heterocycloalkyl containing 2 nitrogen atoms, wherein the 5-8 membered heterocycloalkyl is surrounded by a 3-8 membered heterocycloalkyl C 1-6 Alkyl or hydroxy C 1-6 Alkyl substituted; more preferably, R 2 and R 3 Together with the nitrogen atom to which they are attached, they form a 6-membered heterocycloalkyl group containing 2 nitrogen atoms, wherein the 6-membered heterocycloalkyl group is surrounded by a 4-6-membered heterocycloalkyl group C 1-6 Alkyl or hydroxy C 1-6 Alkyl substituted; further preferably, R 2 and R 3 Together with the nitrogen atom to which they are attached, they form a piperazinyl group which is substituted by a 4-6 membered heterocycloalkyl C 1-6 Alkyl or hydroxy C 1-6 Alkyl substituted; most preferably, R 2 and R 3 Together with the nitrogen atom to which they are attached, they form a piperazinyl group, which is substituted by a 5-6 membered heterocycloalkyl C containing one nitrogen atom. 1-6 Alkyl substituted or hydroxyl C 1-6 Alkyl is substituted, preferably, the piperazinyl is substituted with pyrrolidinyl C 1-6 Alkyl substituted or hydroxyl C 1-6 Alkyl substituted, more preferably, the piperazinyl is substituted by pyrrolidinylethyl or hydroxyethyl.

[0054] Preferably, the compound has the following formula (II):

[0055] Among them, R a 、R b Each independently is H or C 1-6 Alkyl, preferably, R a 、R b are each independently H, methyl or ethyl;

[0056] M is -(CH2)n-, wherein n is 1, 2 or 3, more preferably, n is 2;

[0057] X is F or Cl, more preferably, X is F;

[0058] R c is hydroxy, amino, or a 3-8 membered heterocycloalkyl group containing 1-2 heteroatoms selected from nitrogen, oxygen, or sulfur. More preferably, R c is hydroxy or a 4-6 membered heterocycloalkyl group containing one nitrogen heteroatom, more preferably, R c is hydroxy or pyrrolidinyl, most preferably, R c is hydroxy or 1-pyrrolidinyl.

[0059] Preferably, the compound is selected from the following group of compounds or their salts (eg, hydrochloride):

[0060] Unless otherwise indicated, the following terms and phrases used herein are intended to have the following definitions. The absence of a specific definition for a particular term or phrase should not be construed as indefinite or unclear, but rather as having its ordinary meaning. When a trade name appears in this document, it is intended to refer to the corresponding commercial product or active ingredient.

[0061] The term "pharmaceutically acceptable" as used herein refers to those compounds, materials, compositions and / or dosage forms which, within the scope of sound medical judgment, are suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0062] The term "pharmaceutically acceptable salt" refers to salts of the compounds of the present invention, prepared by reacting the compounds of the present invention with relatively nontoxic acids or bases, having specific substituents. 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 neat solution or in a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amine, or magnesium salts, or similar salts. When the compounds of the present invention contain relatively basic functional groups, base addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of acid in neat solution or in a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, bicarbonate, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, bisulfate, hydroiodic acid, phosphorous acid, and the like, 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, and methanesulfonic acid, as well as salts of amino acids (such as arginine), and salts of organic acids such as glucuronic acid. Certain specific compounds of the present invention contain both basic and acidic functional groups and can be converted into either base or acid addition salts.

[0063] Pharmaceutically acceptable salts of the present invention can be synthesized by conventional chemical methods from parent compounds containing acid radicals or bases. Generally, such salts are prepared by reacting these compounds in free acid or base form with a stoichiometric amount of a suitable base or acid in water or an organic solvent or a mixture of the two.

[0064] 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, (D)-isomers, (L)-isomers, and racemic mixtures and other mixtures thereof, such as enantiomerically or diastereomerically enriched mixtures, all of which are within the scope of the present invention. Additional asymmetric carbon atoms may be present in substituents such as alkyl groups. All such isomers and mixtures thereof are encompassed within the scope of the present invention.

[0065] Unless otherwise indicated, the term "enantiomer" or "optical isomer" refers to stereoisomers that are mirror images of one another.

[0066] Unless otherwise indicated, the term "cis-trans isomers" or "geometric isomers" arises from the inability to rotate freely about double bonds or single bonds forming ring carbon atoms.

[0067] Unless otherwise indicated, the term "diastereomer" refers to stereoisomers that have two or more chiral centers and that are not mirror images of each other.

[0068] The compounds of the present invention may exist in specific forms. Unless otherwise indicated, the term "tautomer" or "tautomeric form" refers to the fact that at room temperature, different functional group isomers are in dynamic equilibrium and can quickly convert into each other. If tautomerism is possible (such as in solution), it is possible to reach a chemical equilibrium of tautomers. For example, proton tautomers (also known as prototropic tautomers) include interconversions carried out by proton migration, such as keto-enol isomerization and imine-enamine isomerization. Valence tautomers include interconversions carried out by the reorganization of some bonding monomers. A specific example of keto-enol tautomerization is the interconversion between two tautomers of pentane-2,4-dione and 4-hydroxy-3-ene-2-one.

[0069] The term "prodrug" generally refers to a compound of formula (I) that has been derivatized with a functional group, and the derivative can be readily converted into the compound of formula (I) in vivo. Suitable prodrug selection and preparation can generally be found in, for example, Design of Prodrugs, ed. H. Bundgaard, Elsevier, 1985.

[0070] The compounds of the present invention may contain unnatural proportions of atomic isotopes at one or more of the atoms comprising the compound, said isotopes having the same atomic number but an atomic mass or mass number different from the atomic mass or mass number predominantly found in nature. For example, the compounds may be labeled with radioactive isotopes, such as deuterium ( 2 H), tritium ( 3 H), iodine-125 ( 125 I) or C-14( 14C). All isotopic variations of the compounds of the present invention, whether radioactive or not, are encompassed by the present invention. Isotopic variations may provide certain therapeutic advantages. For example, deuterated drugs can be formed by replacing hydrogen with heavy hydrogen. The bond formed by deuterium to carbon is stronger than the bond formed by ordinary hydrogen to carbon. Compared to non-deuterated drugs, deuterated drugs have advantages such as reduced toxic side effects, increased drug stability, enhanced therapeutic efficacy, and prolonged biological half-life. Alternatively, they may provide standard compounds useful for characterizing biological samples. Isotopically enriched compounds within Formula (I) can be prepared without undue experimentation by conventional techniques well known to those skilled in the art, or by methods similar to those described in the schemes and examples of the present invention, using appropriate isotopically enriched reagents and / or intermediates.

[0071] "Optional" or "optionally" means that the subsequently described event or circumstance may but need not occur, and that the description includes instances where said event or circumstance occurs and instances where it does not.

[0072] The nomenclature used in the present invention is based on the IUPAC system generated by ChemDraw software. Any open valence bonds appearing on carbon, oxygen, sulfur or nitrogen atoms in the structures given in the present invention indicate the presence of hydrogen atoms.

[0073] The term "substituted" means that any one or more hydrogen atoms on a particular atom are replaced by a substituent, which may include deuterium and hydrogen variants, as long as the valence state of the particular atom is normal and the substituted compound is stable. When the substituent is oxo (i.e., =O), it means that two hydrogen atoms are replaced, and oxygen substitution does not occur on aromatic groups. The term "optionally substituted" means that it may be substituted or unsubstituted, and unless otherwise specified, the type and number of substituents can be any on the basis of chemical achievable.

[0074] When any variable (e.g., R) occurs more than once in a compound's composition or structure, its definition at each occurrence is independent. Thus, for example, if a group is substituted with 0-2 Rs, the group may be optionally substituted with up to two Rs, with each occurrence of R being an independent choice. Furthermore, combinations of substituents and / or their variants are permissible only if such combinations result in stable compounds.

[0075] When the number of a linking group is 0, such as -(CRR)0-, it means that the linking group is a single bond.

[0076] When one of the variables is selected from a single bond, it means that the two groups it connects are directly connected. For example, when L in ALZ represents a single bond, it means that the structure is actually AZ.

[0077] When the linking group is listed without specifying its linking direction, its linking direction is arbitrary, for example, The connecting group L is at this time The benzene ring and cyclopentyl group can be connected in the same direction as reading from left to right to form It is also possible to connect phenyl and cyclopentyl groups in the opposite direction of reading from left to right to form Combinations of linkers, substituents, and / or variations thereof are permissible only if such combinations result in stable compounds.

[0078] Unless otherwise specified, the number of atoms in a ring is generally defined as the ring member number, for example, a "3-7 membered ring" refers to a "ring" having 3-7 atoms arranged around it.

[0079] Unless otherwise specified, the term "halogen" refers to fluorine, chlorine, bromine and iodine.

[0080] Unless otherwise specified, the term “C 1-6 "Alkyl" is used to represent a straight or branched chain saturated hydrocarbon group consisting of 1 to 6 carbon atoms. 1-6 Alkyl groups include C 1-5 、C 1-4 、C 1-3 、C 1-2 、C 2-6 、C 2-4 , C6 and C5 alkyl, etc.; which can be monovalent (such as methyl), divalent (such as methylene) or polyvalent (such as methine). 1-6 Examples of 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, and the like.

[0081] Unless otherwise specified, the term “C 1-3 "Alkyl" is used to represent a straight or branched chain saturated hydrocarbon group consisting of 1 to 3 carbon atoms. 1-3 Alkyl groups include C 1-2 and C 2-3 Alkyl, etc.; it can be monovalent (such as methyl), divalent (such as methylene) or polyvalent (such as methine). 1-3 Examples of alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), and the like.

[0082] Unless otherwise specified, the term “C 2-6"Alkenyl" is used to indicate a linear or branched hydrocarbon group consisting of 2 to 6 carbon atoms containing at least one carbon-carbon double bond. The carbon-carbon double bond may be located at any position of the group. 2-6 Alkenyl groups include C 2-4 、C 2-3 , C4, C3, C2 alkenyl, etc.; which may be monovalent, divalent or polyvalent. 2-6 Examples of alkenyl groups include, but are not limited to, ethenyl, propenyl, butenyl, pentenyl, hexenyl, butadienyl, piperyl, and the like.

[0083] Unless otherwise specified, the term “C 1-6 "Alkoxy" refers to an alkyl group containing 1 to 6 carbon atoms which is attached to the rest of the molecule via an oxygen atom. 1-6 Alkyl groups include C 1-4 、C 1-3 、C 1-2 、C 2-6 、C 2-4 , C6, C5, C4 and C3 alkoxy, etc.; C 1-6 Examples of alkoxy groups include, but are not limited to, methoxy, ethoxy, propoxy (including n-propoxy and isopropoxy), butoxy (including n-butoxy, isobutoxy, s-butoxy and t-butoxy), pentoxy (including n-pentoxy, isopentoxy and neopentoxy), hexyloxy, and the like.

[0084] Unless otherwise specified, the term “C 1-6 "Alkylamino" refers to those alkyl groups containing 1 to 6 carbon atoms which are linked to the rest of the molecule through an amino group. 1-6 Alkyl groups include C 1-4 、C 1-3 、C 1-2 、C 2-6 、C 2-4 , C6, C5, C4, C3 and C2 alkylamino, etc.; C 1-6 Examples of alkylamino groups include, but are not limited to, -NHCH3, -N(CH3)2, -NHCH2CH3, -N(CH3)CH2CH3, -N(CH2CH3)2, -NHCH2CH2CH3, -NHCH(CH3)2, -NHCH2CH2CH2CH3, and the like.

[0085] Unless otherwise specified, the term “C 3-6 "Cycloalkyl" means a saturated cyclic hydrocarbon group consisting of 3 to 6 carbon atoms, which is a monocyclic or bicyclic ring system. 3-6 Cycloalkyl groups include C 3-5 、C 4-5 and C 5-6 Cycloalkyl, etc.; it may be monovalent, divalent or polyvalent.3-6 Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like.

[0086] Unless otherwise specified, the term "3-8 membered heterocycloalkyl" by itself or in combination with other terms means a saturated monocyclic radical consisting of 3 to 8 ring atoms, 1, 2, 3 or 4 of which are heteroatoms independently selected from O, S and N, and the remainder being carbon atoms, wherein the nitrogen atom is optionally quaternized and the nitrogen and sulfur heteroatoms are optionally oxidized (i.e., NO and S(O)). z , z is 1 or 2). In addition, with respect to such "3-8 membered heterocycloalkyl", a heteroatom may occupy the position at which the heterocycloalkyl is attached to the rest of the molecule. Such 3-8 membered heterocycloalkyls include 4-6 membered, 5-6 membered, 7-8 membered, 4 membered, 5 membered and 6 membered heterocycloalkyls, and examples of 3-6 membered heterocycloalkyls include, but are not limited to, azetidinyl, oxetanyl, thietanyl, pyrrolidinyl, pyrazolidinyl, imidazolidinyl, tetrahydrothiophenyl, tetrahydrofuranyl (including tetrahydrofuran-2-yl), piperidinyl, tetrahydropyranyl, tetrahydrothiopyranyl, piperazinyl, morpholinyl, thiomorpholinyl, and the like.

[0087] Unless otherwise specified, the terms "6-10 membered aromatic ring" and "6-10 membered aryl" are used interchangeably. The term "6-10 aryl" refers to a monovalent aromatic carbocyclic ring system containing 6-10 carbon atoms and having at least one aromatic ring or multiple fused rings in which at least one ring is aromatic. Examples of aryl groups include, but are not limited to, phenyl, naphthyl, biphenyl, and indanyl.

[0088] Unless otherwise specified, the terms "5-10 membered heteroaromatic ring" and "5-10 membered heteroaryl" are used interchangeably. The term "5-10 membered heteroaryl" refers to a cyclic group consisting of 5 to 10 ring atoms with a conjugated π electron system, wherein 1, 2, 3 or 4 ring atoms are heteroatoms independently selected from O, S and N, and the rest are carbon atoms. It can be a monocyclic or fused bicyclic system, wherein at least one ring in the system is aromatic. The nitrogen atom is optionally quaternized, and the nitrogen and sulfur heteroatoms are optionally oxidized (i.e., NO and S(O) z, z is 1 or 2). The 5-10 membered heteroaryl group can be attached to the rest of the molecule through a heteroatom or a carbon atom, and the 5-10 membered heteroaryl group includes 5-8 membered, 5-7 membered, 5-6 membered, 5 membered and 6 membered heteroaryl groups, etc. Examples of the 5-10 membered heteroaryl group include, but are not limited to, pyrrolyl (including N-pyrrolyl, 2-pyrrolyl and 3-pyrrolyl), pyrazolyl (including 2-pyrazolyl and 3-pyrazolyl), imidazolyl (including N-imidazolyl, 2-imidazolyl, 4-imidazolyl and 5-imidazolyl), oxazolyl (including 2-oxazolyl, 4-oxazolyl and 5-oxazolyl), triazolyl (1H-1,2,3-triazolyl, 2H-1,2,3-triazolyl, 1H-1,2,4-triazolyl and 4H-1,2,4-triazolyl), tetrazolyl, isoxazolyl (including 3-isoxazolyl, 4-isoxazolyl and 5-isoxazolyl), thiazolyl (including 2-thiazolyl, 4 1-thiazolyl and 5-thiazolyl, etc.), furyl (including 2-furyl and 3-furyl, etc.), thienyl (including 2-thienyl and 3-thienyl, etc.), pyridyl (including 2-pyridyl, 3-pyridyl and 4-pyridyl, etc.), pyrazinyl, pyrimidinyl (including 2-pyrimidinyl and 4-pyrimidinyl, etc.), benzothiazolyl (including 2-benzothiazolyl, etc.), purinyl, benzimidazolyl (including 2-benzimidazolyl, etc.), benzoxazolyl, indazolyl (including 5-indazolyl, etc.), isoquinolyl (including 1-isoquinolyl and 5-isoquinolyl, etc.), quinoxalinyl (including 2-quinoxalinyl and 5-quinoxalinyl, etc.) or quinolyl (including 3-quinolyl and 6-quinolyl, etc.), etc.

[0089] Unless otherwise specified, the terms "5-6 membered heteroaromatic ring" and "5-6 membered heteroaryl" are used interchangeably in the present invention. The term "5-6 membered heteroaryl" refers to a cyclic group consisting of 5 to 6 ring atoms with a conjugated π electron system, wherein 1, 2, 3 or 4 ring atoms are heteroatoms independently selected from O, S and N, and the rest are carbon atoms. The nitrogen atom is optionally quaternized, and the nitrogen and sulfur heteroatoms are optionally oxidized (i.e., NO and S(O) z, z is 1 or 2). 5-6 yuan of heteroaryl can be connected to the rest of the molecule through a heteroatom or a carbon atom, and the 5-6 yuan of heteroaryl includes 5 yuan and 6 yuan of heteroaryl etc. The example of the 5-6 yuan of heteroaryl includes but is not limited to pyrrolyl (including N-pyrrolyl, 2-pyrrolyl and 3-pyrrolyl etc.), pyrazolyl (including 2-pyrazolyl and 3-pyrazolyl 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), tetrazolyl, isoxazolyl (including 3-isoxazolyl, 4-isoxazolyl and 5-isoxazolyl, etc.), thiazolyl (including 2-thiazolyl, 4-thiazolyl and 5-thiazolyl, etc.), furyl (including 2-furyl and 3-furyl, etc.), thienyl (including 2-thienyl and 3-thienyl, etc.), pyridyl (including 2-pyridyl, 3-pyridyl and 4-pyridyl, etc.), pyrazinyl, pyrimidinyl (including 2-pyrimidinyl and 4-pyrimidinyl, etc.), etc.

[0090] Unless otherwise specified, C n-n+m or C n -C n+m Any specific case including n to n+m carbons, such as C 1-7 Including C1, C2, C3, C4, C5, C6 and C7, and also including any range from n to n+m, such as C 1-7 Including C 1-3 、C 1-6 、C 3-6 、C 4-7 and C 5-7 Similarly, n-membered to n+m-membered means that the number of atoms in the ring is n to n+m, for example, a 3-7-membered ring includes a 3-membered, 4-membered, 5-membered, 6-membered and 7-membered ring, and also includes any range from n to n+m, for example, a 3-7-membered ring includes a 3-6-membered ring, a 4-7-membered ring, a 5-7-membered ring and a 6-7-membered ring, etc.

[0091] Preparation method

[0092] The present invention also relates to a method for producing the compound of formula (I) defined above. In some embodiments, the synthesis method of the compound of the present application is shown in the figure:

[0093] When X is selected from halogens other than Br, such as F, Cl, I, etc., the method comprises the following steps:

[0094] a) brominating the compound of formula Ia using a brominating agent to produce a compound of formula Ib, preferably, the brominating agent includes but is not limited to N-bromosuccinimide (NBS) or 1,3-dibromo-5,5-dimethylhydantoin, preferably, the reaction temperature is 0-30° C.;

[0095] b) one Br atom in the compound of formula Ib is replaced by another halogen atom to obtain a compound of formula Ic. Preferably, step b) uses and a metal organic salt strong base at low temperature, wherein X is a halogen other than Br, such as F, Cl, I, more preferably, wherein the metal organic salt strong base is n-butyl lithium, preferably, the reaction temperature is -78 ° C;

[0096] c) oxidizing the compound of formula Ic to a compound of formula Id using an oxidizing agent, preferably, the oxidizing agent includes but is not limited to K2Cr2O7, preferably, the reaction temperature is 60-150°C;

[0097] d) Compound of formula Id and H2N-LR 1 Reaction to obtain a compound of formula Ie, preferably, the reaction temperature is 50-150° C.;

[0098] e) reacting a compound of formula Ie with R 2 R 3 NH reaction to obtain a compound of formula I, preferably, the reaction is carried out in the presence of a catalyst bis(triphenylphosphine)palladium(II) chloride, preferably, the reaction temperature is 20-150°C.

[0099] When X is selected from Br, the method comprises:

[0100] a) brominating the compound of formula Ia using a brominating agent to generate a compound of formula Ib, preferably, the brominating agent includes but is not limited to N-bromosuccinimide (NBS), preferably, the reaction temperature is 0-30° C.;

[0101] c) oxidizing the compound of formula Ib to a compound of formula Id using an oxidizing agent, preferably, the oxidizing agent includes but is not limited to K2Cr2O7, preferably, the reaction temperature is 60-150°C;

[0102] d) reacting the compound of formula Id with H2N-L-R1 to obtain a compound of formula Ie, preferably at a reaction temperature of 50-150°C;

[0103] e) reacting a compound of formula Ie with R 2 R 3NH reaction to obtain a compound of formula I, preferably, the reaction is carried out in the presence of a catalyst bis(triphenylphosphine)palladium(II) chloride, preferably, the reaction temperature is 20-150°C.

[0104] Alternatively, when X is selected from halogens other than Br, such as F, Cl, I, etc., the method comprises the following steps:

[0105] f) brominating the compound of formula Ia using a brominating agent to produce a compound of formula Ib, preferably, the brominating agent includes but is not limited to N-bromosuccinimide (NBS) or 1,3-dibromo-5,5-dimethylhydantoin, preferably, the reaction temperature is 0-30° C.;

[0106] g) All two Br atoms in the compound of formula Ib are replaced by other halogen atoms to obtain a compound of formula Ic. Preferably, step b) uses and a metal organic salt strong base at low temperature, wherein X is a halogen other than Br, such as F, Cl or I, more preferably, wherein the metal organic salt strong base is n-butyl lithium, preferably, the reaction temperature is -78 ° C;

[0107] h) oxidizing the compound of formula I-c' to a compound of formula I-d' using an oxidizing agent, preferably, the oxidizing agent includes but is not limited to K2Cr2O7, preferably, the reaction temperature is 60-150°C;

[0108] i) Compound of formula I-d' and H2N-LR 1 Reaction to obtain a compound of formula I-e', preferably, the reaction temperature is 50-150 ° C;

[0109] j) reacting a compound of formula I-e' with R 2 R 3 NH reaction to obtain a compound of formula I, preferably, the reaction is carried out in the presence of a catalyst bis(triphenylphosphine)palladium(II) chloride, preferably, the reaction temperature is 20-150°C.

[0110] As known to those skilled in the art, certain reactive groups (eg, -NH2, -OH, etc.) require conventional protection and deprotection when necessary, which is familiar to those skilled in the art.

[0111] Products and Applications

[0112] The present disclosure also provides a product, which can be, for example, a medicament, a pharmaceutical composition, or a kit, comprising an effective amount of a hnRNPA2B1 agonist compound of the present disclosure and a pharmaceutically or immunologically acceptable carrier. As used herein, the terms "active substance" or "active substance of the present disclosure" are used interchangeably to refer to a hnRNPA2B1 agonist compound having the structural formula (I) or a derivative or product thereof.

[0113] As used herein, the term "pharmaceutically acceptable" refers to a substance that is suitable for use in humans and / or animals without excessive adverse reactions (such as toxicity, irritation, and allergic reactions), i.e., a substance with a reasonable benefit / risk ratio. As used herein, the term "effective amount" refers to an amount that can produce a function or activity in humans and / or animals and is acceptable to humans and / or animals.

[0114] As used herein, the term "pharmaceutically acceptable carrier" refers to a carrier for administering a therapeutic agent, including various excipients and diluents. The term refers to pharmaceutical carriers that are not essential active ingredients themselves and are not unduly toxic after administration. Suitable carriers are well known to those of ordinary skill in the art, and a comprehensive discussion of pharmaceutically acceptable excipients can be found in Remington's Pharmaceutical Sciences (Mack Pub. Co., NJ 1991).

[0115] The pharmaceutically acceptable carrier in the composition may contain a liquid such as water, saline, glycerol, and ethanol. In addition, these carriers may also contain auxiliary substances such as fillers, disintegrants, lubricants, glidants, effervescent agents, wetting agents or emulsifiers, flavoring agents, pH buffering substances, etc. Generally, these substances can be formulated in a non-toxic, inert, and pharmaceutically acceptable aqueous carrier medium, wherein the pH is generally about 5-8, preferably, about 6-8.

[0116] The active substance in the product of the present disclosure accounts for 0.001-99.9 wt% of the total weight of the composition, preferably 1-95 wt%, more preferably 5-90 wt%, and even more preferably 10-80 wt%, with the remainder being pharmaceutically acceptable carriers and other additives.

[0117] As used herein, the term "unit dosage form" refers to a dosage form for single administration of the product of the present disclosure for the convenience of administration, including but not limited to various solid dosage forms (such as tablets), liquid dosage forms, capsules, and sustained-release dosage forms.

[0118] In another preferred embodiment of the present disclosure, the product is in unit dosage form or multi-dose form, and the active substance content is 0.01 to 2000 mg / dose, preferably 0.1 to 1500 mg / dose, and more preferably 1 to 1000 mg / dose. In another preferred embodiment of the present disclosure, 1 to 6 doses of the composition of the present disclosure are administered daily, preferably 1 to 3 doses; most preferably, the daily dose is 1 dose.

[0119] It should be understood that the effective dosage of an active substance, such as a hnRNPA2B1 agonist compound, may vary depending on the severity of the condition being administered or treated. The specific dosage is determined based on the individual circumstances of the subject (e.g., weight, age, physical condition, and desired effect), which is within the judgment of a skilled physician.

[0120] The products disclosed herein may be in solid form (e.g., granules, tablets, lyophilized powders, suppositories, capsules, sublingual tablets) or liquid form (e.g., oral solution) or other suitable forms. The routes of administration may include, but are not limited to: (1) traditional administration methods, such as gastrointestinal administration (e.g., oral administration), parenteral administration (e.g., solution injection, intravenous infusion), such as mucosal administration, transdermal administration, respiratory tract atomization inhalation, nasal drops, spray, oral administration, intramuscular injection, and / or intravenous administration; (2) linking the hnRNPA2B1 agonist to a transferrin / poly-L-lysine complex to enhance its biological effect; (3) encapsulating the drug with liposomes to mediate its entry into cells, which is beneficial for the smooth entry of the compound molecules and protects them from the hydrolysis of various extracellular enzymes; and (4) transporting active substances with liposomes to enable their specific transport to target tissues and target cells.

[0121] In addition, the products of the present disclosure may also contain other active substances for improving and treating infectious diseases. Such other active substances include, but are not limited to, commonly used clinical antibiotics, including one or more of β-lactams (penicillins and cephalosporins), aminoglycosides, tetracyclines, chloramphenicols, macrolides, antifungal antibiotics, and anti-tuberculosis antibiotics.

[0122] In some embodiments, other active substances that regulate anti-infection are administered before, simultaneously with, or after administration of the product of the present disclosure. The other active substances have the activity of preventing or treating diseases associated with infection (especially viral infection), injuries caused by infection, chronic inflammatory diseases caused by infection, and / or symptoms thereof.

[0123] In some embodiments, hnRNPA2B1 is activated by the compounds or products of the present invention to further regulate the immune system to provide treatment of diseases, including diseases caused by exogenous factors. Exemplary infections of exogenous factors that can be treated and / or prevented by the methods of the present invention include: infections of bacteria (e.g., gram-positive or gram-negative bacteria), fungal infections, parasitic infections, and viral infections. In one embodiment of the invention, infection is bacterial infection (e.g., infection of Escherichia coli, Klebsiella pneumonia, Pseudomonas aeruginosa, Salmonella spp., Staphylococcus aureus, Streptococcus or vancomycin-resistant enterococci) or sepsis. In other embodiments, infection is fungal infection (e.g., mold, yeast or higher fungal infection). In other embodiments, the infection is a parasitic infection (e.g., an infection caused by a unicellular or multicellular parasite, including Giardia duodenalis, Cryptosporidium parvum, Cyclospora cayetanensis, and Toxoplasma gondiz). In yet other embodiments, the infection is a viral infection (e.g., a viral infection associated with AIDS, avian influenza, chickenpox, cold sores, the common cold, gastroenteritis, glandular fever, influenza, measles, mumps, pharyngitis, pneumonia, rubella, SARS, and lower or upper respiratory tract infections (e.g., respiratory syncytial virus).

[0124] In a preferred embodiment, the product can be used to prevent or treat diseases associated with infection (especially viral infection), chronic inflammatory diseases caused by infection, and / or their symptoms; for example, the pharmaceutical composition of the present disclosure can be used to prevent or treat viral infectious diseases known to be treatable or preventable in the prior art, such as tissue damage caused by viral infection; inflammatory damage to organs; and multiple organ failure.

[0125] In some embodiments, the infection that can be treated or prevented by the compounds of the present application is a DNA-involved and / or mediated infection. In some embodiments, the infection is a viral infection or an infection caused by a bacterial, fungal, or other DNA-involved infection or a combination thereof, for example, a DNA viral infection, such as an infection caused by one or more viruses selected from the group consisting of herpes simplex virus, hepatitis B virus, adenovirus, poxvirus, parvovirus, adeno-associated virus, coronavirus, influenza virus, rhinovirus, parainfluenza virus, respiratory syncytial virus, coxsackievirus, echovirus, and novel enterovirus.

[0126] In some embodiments, the disease and / or symptom associated with infection is one or more selected from the following group: the disease and / or symptom associated with infection is one or more selected from the following group: pathological damage caused by infection; insufficient or excessive production of cytokines such as interferon after infection; endotoxic shock or death; inflammatory damage to organs; multiple organ failure, for example, the organs are selected from: liver, spleen, brain, kidney, heart, lung, stomach, intestine; chronic inflammatory diseases caused by viral infection (for example, autoimmune diseases such as inflammatory bowel disease, rheumatoid arthritis, systemic lupus erythematosus, chronic nephritis, tuberculosis, chronic gastrointestinal diseases). Preferably, the chronic inflammatory disease and / or its symptoms caused by viral infection include: autoimmune diseases such as inflammatory bowel disease, rheumatoid arthritis, systemic lupus erythematosus, chronic nephritis, tuberculosis, chronic gastrointestinal diseases. In some embodiments, the virus is selected from HSV, HBV and VSV.

[0127] In some embodiments, the compounds or products of the present application are administered as preventive drugs before infection (especially pathogen infection) occurs to prevent infection or reduce the severity of subsequent infection. In some embodiments, the compounds or products of the present application are administered as therapeutic drugs after infection occurs to reduce the severity of infection and disease. In some embodiments, the compounds or products of the present application are administered as both preventive drugs and therapeutic drugs, and are administered continuously or intermittently before and after infection occurs.

[0128] Those skilled in the art may arbitrarily combine the above technical solutions and technical features without departing from the inventive concept and protection scope of the present invention. Other aspects of the present invention will be obvious to those skilled in the art due to the disclosure herein.

[0129] Example

[0130] The present disclosure will be further described below in conjunction with specific examples. It should be understood that these examples are intended only to illustrate the present disclosure and are not intended to limit the scope of the present disclosure. Those skilled in the art may make appropriate modifications and variations to the present disclosure, and these modifications and variations are within the scope of the present disclosure.

[0131] For experimental procedures in the following examples where specific conditions are not specified, conventional methods in the art may be employed, for example, as described in Molecular Cloning: A Laboratory Manual (3rd ed., Cold Spring Harbor Laboratory Press, New York, 1989) or according to the conditions recommended by the supplier. DNA sequencing methods are conventional in the art and are also available from commercial companies.

[0132] Unless otherwise indicated, percentages and parts are by weight. Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those familiar to those skilled in the art. In addition, any methods and materials similar or equivalent to those described herein can be used in the disclosed methods. The preferred embodiments and materials described herein are for illustrative purposes only.

[0133] I. Synthesis and Characterization Examples of Compounds

[0134] Example I.1: Synthesis of Compound A (FPC-1):

[0135] *eq equivalent

[0136] 1-1. Preparation of Compound 2

[0137] NBS (57.5 g, 320 mmol) was added to DMF (120 mL) and cooled to 0°C. Compound 1 (20 g, 130 mmol) was added, and the resulting suspension was stirred, allowed to naturally warm to room temperature, and stirred overnight. The reaction solution was filtered, and the filter cake was washed three times with ethanol. The dried filter cake was collected and recrystallized from EA to obtain the pure product (10.1 g, off-white crystals).

[0138] 1 H NMR (300 MHz, chloroform-d) δ 7.79 (d, J = 7.4 Hz, 2H), 7.09 (d, J = 8.8 Hz, 2H), 3.30 (s, 4H).

[0139] 1-2. Preparation of Compound 3

[0140] Compound 2 (6 g, 19.2 mol) was dissolved in 300 mL of anhydrous THF and cooled to -80 ° C. Then n-butyl lithium (9.2 mL, 2.5 M) was added dropwise, and the reaction mixture was stirred at the same temperature for at least one hour. After slowly adding N-fluorobisbenzenesulfonamide (NFSI) (9 g, 28.8 mmol) in 100 mL of anhydrous THF, the mixture was stirred at -80 ° C for another 60 minutes. The reaction mixture was then heated to room temperature overnight and then poured into NH4Cl solution. The aqueous phase was then extracted with DCM (3 × 50 mL), the organic phases were combined, and dried over MgSO4. The solvent was removed under low pressure. The product was purified by column chromatography using n-hexane as eluent. Compound 3 (4.1 g, white solid) was obtained.

[0141] 1H NMR (300 MHz, chloroform-d) δ 7.66 (d, J=7.4 Hz, 1H), 7.21-7.09 (m, 3H), 3.35 (s, 4H).

[0142] 1-3. Preparation of Compound 4

[0143] Compound 3 (4.5g, 17.9mmol) is joined in the glacial acetic acid solution (50mL) of potassium dichromate (24g, 82.4mmol).Solution is heated to reflux 16 hours, then poured into frozen water.Gained precipitate is collected by filtration and washed with water.Filter cake is air-dried, joined among the DCM and refluxed 30 minutes, filtered, washed with DCM, filter cake is joined among the DCM and refluxed 15 minutes, filtered and washed with DCM.Merge all filtrates and vacuum concentration, obtain compound 4 (4.1g, crude product, yellow solid).

[0144] 1-4. Preparation of INT-1

[0145] N-Boc-ethylenediamine (3.3 g, 20.8 mmol) was added to a solution of compound 4 (4.1 g, 13.9 mmol) in EtOH (100 mL), and the mixture was heated at reflux for 1 hour. The reaction mixture was concentrated, and the resulting residue was purified by column chromatography (PE / EA = 5 / 1). An off-white solid (2.8 g) was obtained.

[0146] 1 H NMR (300MHz, chloroform-d) δ8.65 (dd, J=8.2, 4.6Hz, 1H), 8.43 (d, J=8.0Hz, 1H), 8.08 (d, J=8.0Hz, 1H), 7.50 (dd, J=12.1, 8.1Hz, 1H), 4.89 (s, 1H), 4.39-4.28 (m, 2H), 3.58-3.44 (m, 2H), 1.26 (s, 9H).

[0147] 1-5. Preparation of FPC-1-1

[0148] Cesium carbonate (5.55 g, 17 mmol) and 1-(2-pyridoneethyl)piperazine (1.05 g, 5.7 mmol) were added to a toluene solution (175 mL) of INT-1 (2.5 g, 5.7 mol), and bis(triphenylphosphine)palladium chloride (II) (400 mg, 0.57 mmol) was added to the mixture under argon protection. The mixture was reacted at 80° C. for 16 hours, the reaction solution was cooled to room temperature, water was added to the reaction solution and stirred, the organic phase was separated, the aqueous phase was extracted once with DCM, the organic phases were combined, and dried over anhydrous magnesium sulfate. The crude product was purified by Prep-TLC (DCM / MeOH=10 / 1+NH 3 ·H 2 O) to give 240 mg of a pure yellow solid.

[0149] 1 H NMR (400 MHz, CHLOROFORM-d) δ 8.56 (dd, J = 7.7, 4.3 Hz, 1H), 8.51 (d, J = 8.3 Hz, 1H), 7.34-7.27 (m, 1H), 7.17 (d, J = 8.2 Hz, 1H), 4.99 (s, 1H), 4.31 (s, 2H), 3.79-2.45 (m, 18H), 2.18 (s, 4H), 1.28 (s, 9H).

[0150] 1-6. Preparation of FPC-1

[0151] Compound FPC-1-1 (240 mg, 0.44 mmol) was dissolved in 11 mL of DCM / MeOH (10 / 1). 5 M HCl / 1,4-dioxane (5 mL) was added to the mixture and stirred at room temperature for 2 hours. The mixture was concentrated in vacuo, and the residue was dissolved in a small amount of methanol, precipitated with ether, and centrifuged. The precipitate was washed with ether, centrifuged, and dried in vacuo to give 240 mg of the pure product (yellow solid).

[0152] 1 H NMR (300 MHz, methanol-d4) δ 8.67-8.58 (m, 2H), 7.56 (dd, J = 13.2, 8.3 Hz, 1H), 7.47 (d, J = 8.3 Hz, 1H), 4.49-4.42 (m, 2H), 4.02-3.69 (m, 10H), 3.69-3.38 (m, 8H), 2.18 (s, 4H).

[0153] MS (ESI) m / z [M+H] + =440.3.

[0154] Example I.2: Synthesis of Compound B (FPC-2)

[0155] Preparation of FPC-2-1

[0156] In a single-necked flask, 10 mL of ethylene glycol monomethyl ether was added, followed by INT-1 (110 mg, 0.25 mmol) and 3-(piperazin-1-yl)propan-1-ol (44 mg, 0.3 mmol), and the reaction mixture was stirred at 100° C. After 2 hours, the solvent was removed in vacuo, and the residue was purified by TLC to give 20 mg of a yellow solid.

[0157] 1 H NMR (400 MHz, CHLOROFORM-d) δ 8.63-8.49 (m, 2H), 7.31 (dd, J = 12.8, 8.3 Hz, 1H), 7.18 (d, J = 8.3 Hz, 1H), 4.97 (s, 1H), 4.42-4.23 (m, 2H), 3.87 (t, J = 5.0 Hz, 2H), 3.60-3.28 (m, 8H), 2.92 (s, 4H), 1.89 (s, 2H), 1.29 (s, 9H).

[0158] 2-2. Preparation of FPC-2

[0159] Compound FPC-2-1 (20 mg, mmol) was dissolved in 2 mL of DCM / MeOH (1 / 1). 5 M HCl / 1,4-dioxane (2 mL) was added and the mixture was stirred at room temperature for 2 hours. The mixture was concentrated in vacuo and the residue was dissolved in a small amount of methanol, precipitated with ether, centrifuged, and the precipitate was washed with ether, centrifuged, and dried in vacuo to give 5 mg of the pure product (yellow solid).

[0160] 1 H NMR (400 MHz, methanol-d4) δ 8.72-8.46 (m, 2H), 7.50 (ddd, J = 26.1, 12.4, 8.3 Hz, 2H), 4.45 (t, J = 5.6 Hz, 2H), 3.93-3.69 (m, 6H), 3.45 (dt, J = 15.0, 9.8 Hz, 5H), 3.34 (s, 3H), 2.07 (dt, J = 12.2, 6.1 Hz, 2H).

[0161] MS (ESI) m / z [M+H] + =401.47.

[0162] Example I.3: Synthesis of Compound C (FPC-3)

[0163] Preparation of FPC-3-1

[0164] Cesium carbonate (1.78 g, 5.48 mmol) and N-ethyl-2-(pyrrolidin-1-yl)ethyl-1-amine (286 mg, 2.01 mmol) were added to a toluene solution (70 mL) of INT-1 (800 mg, 1.82 mmol). Bis(triphenylphosphine)palladium chloride (II) (128 mg, 0.18 mmol) was added to the mixture under argon protection. The mixture was reacted at 80° C. for 16 hours. The reaction solution was cooled to room temperature, water was added to the reaction solution and stirred, the organic phase was separated, the aqueous phase was extracted once with DCM, the obtained organic phases were combined, and dried over anhydrous magnesium sulfate. The crude product was purified by Prep-TLC (DCM / MeOH=10 / 1+NH 3 · H 2 O) to give 30 mg of a pure yellow solid.

[0165] 1 H NMR (400MHz, chloroform-d) δ 8.63-8.52 (m, 2H), 7.37-7.28 (m, 2H), 4.96 (s, 1H), 4.32 (t, J = 5.5Hz, 2H), 3.99 (s, 2H), 3.93-3.75 (m, 2H), 3.49 (s, 2H) ), 3.44 (d, J=6.8Hz, 2H), 3.36-3.16 (m, 2H), 2.87-2.62 (m, 2H), 2.29-2.12 (m, 2H), 2.12-1.91 (m, 2H), 1.28 (s, 9H), 1.10 (t, J=7.1Hz, 3H).

[0166] 3-2. Preparation of FPC-3

[0167] Compound FPC-3-1 (30 mg, 0.06 mmol) was dissolved in 4.4 mL of DCM / MeOH (10 / 1). 5 M HCl / 1,4-dioxane (2 mL) was added, and the mixture was stirred at room temperature for 2 hours. The mixture was concentrated in vacuo, and the residue was dissolved in a small amount of methanol, precipitated with ether, and centrifuged. The precipitate was washed with ether, centrifuged, and dried in vacuo to obtain 25 mg of the pure product (yellow solid).

[0168] 1H NMR (300MHz, methanol-d4) δ8.67-8.54(m, 2H), 7.58-7.46(m, 2H), 4.51-4.41(m, 2H), 3.87-3.77(m, 2H), 3.77-3.65(m , 2H), 3.51 (q, J=6.6Hz, 4H), 3.34 (d, J=1.8Hz, 1H), 3.20-3.05 (m, 2H), 2.20-1.92 (m, 5H), 1.10 (t, J=7.0Hz, 3H).

[0169] MS (ESI) m / z [M+H] + =399.47.

[0170] Example I.4: Synthesis of Compound D (FPC-5)

[0171] 4-1. Preparation of FPC-5-1

[0172] Cesium carbonate (460 mg, 1.41 mmol) and N, 2-dimethylpropane-1-amine (49 mg, 0.56 mmol) were added to a toluene solution (20 mL) of INT-1 (205 mg, 0.47 mmol). Under argon protection, bis(triphenylphosphine)palladium chloride (II) (32 mg, 0.0.045 mmol) was added to the mixture. The reaction was carried out at 80° C. for 16 hours. The reaction solution was cooled to room temperature, water was added to the reaction solution and stirred, the organic phase was separated, the aqueous phase was extracted once with DCM, the organic phases were combined, and dried over anhydrous magnesium sulfate. The crude product was purified by vacuum concentration and column purification through TLC (DCM / MeOH=10 / 1+NH 3 ·H 2 O) to give 10 mg of a pure yellow solid.

[0173] 1 H NMR (300MHz, chloroform-d) δ8.56 (dd, J=8.2, 4.9Hz, 1H), 8.45 (d, J=8.5Hz, 1H), 7.24-7.18 (m, 1H), 7.07 (d, J=8.5Hz, 1H), 5.05 (s, 1H), 4.39-4.2 6 (m, 2H), 3.51 (dd, J=11.9, 5.2Hz, 2H), 3.20-3.13 (m, 1H), 3.05 (d, J=4.0Hz, 3H), 2.17-2.05 (m, 2H), 1.31 (s, 9H), 0.87 (d, J=6.6Hz, 6H).

[0174] 4-2. Preparation of FPC-5

[0175] Compound FPC-5-1 (10 mg, 0.022 mmol) was dissolved in 1.1 mL of DCM / MeOH (10 / 1). 5 M HCl / 1,4-dioxane (0.5 mL) was added, and the mixture was stirred at room temperature for 2 hours. The mixture was concentrated in vacuo, and the residue was dissolved in a small amount of methanol, precipitated with ether, and centrifuged. The precipitate was washed with ether, centrifuged, and dried in vacuo to obtain 9 mg of the pure product (yellow solid).

[0176] 1 H NMR (400MHz, methanol-d4) δ 8.61-8.52 (m, 1H), 8.44 (d, J = 8.3Hz, 1H), 7.38 (dd, J = 12.5, 8.5Hz, 1H), 7.25 (d, J = 8.4Hz, 1H), 4.44 (t, J = 5.1Hz , 2H), 3.22 (dd, J=19.7, 7.1Hz, 4H), 3.10 (d, J=3.1Hz, 3H), 2.16 (dp, J=13.3, 7.2Hz, 1H), 1.31 (d, J=7.7Hz, 1H), 0.87 (d, J=6.4Hz, 6H).

[0177] MS (ESI) m / z [M+H] + =344.42.

[0178] Example I.5: Synthesis of Compound E (AIR-2)

[0179] 5-1. Preparation of Compound 5

[0180] Compound 3 (31.90 g, 127.04 mmol) was dissolved in anhydrous THF (800 mL), and the reaction system was replaced with argon three times and then cooled to -78 ° C. Then n-BuLi (60.98 mL, 2.5 M / L, 152.45 mmol) was added dropwise, and the reaction mixture was stirred at the same temperature for at least 10 minutes. N-fluorobisbenzenesulfonamide (NFSI) (60.09 g, 190.57 mmol) in 200 mL of anhydrous THF was added dropwise, and after stirring at -78 ° C for 1 hour, the reaction mixture was naturally warmed to room temperature and stirred overnight. The reaction mixture was quenched with NH4Cl solution and then extracted twice with EA. The combined organic layers were washed with water and brine, dried over Na2SO4 and filtered. The filtrate was concentrated to give a crude product, which was added to DCM and stirred. The resulting precipitate was filtered, the filter cake was washed twice with DCM, and all filtrates were combined and concentrated in vacuo to give the crude product, which was then purified by column chromatography (PE) to afford compound 5 (15.42 g, 63.82%) as a white solid.

[0181] 1 H NMR (300MHz, Chloroform-d) δ 7.16 (d, J=7.6Hz, 2H), 7.12-7.04 (m, 2H), 3.37 (s, 4H).

[0182] 5-2. Preparation of INT-2

[0183] Compound 5 (15.42 g, 81.08 mmol) was added to a glacial acetic acid solution (220 mL) of potassium dichromate (109.72 g, 372.96 mmol). The solution was stirred at 80 ° C for 12 hours. The reaction mixture was quenched with EA and water and extracted twice with EA. The combined organic layer was washed with water and brine, dried over Na2SO4 and filtered. The filtrate was concentrated to give a crude product, which was then added with PE and ultrasonic dispersion. The resulting precipitate was filtered and dried to obtain a crude INT-2 (10.55 g) as a brown solid, which was used without further purification.

[0184] 5-3. Preparation of AIR-2-1

[0185] INT-2 (5 g, 21.35 mmol) was dissolved in EtOH (120 mL), the reaction system was replaced with argon three times, and then heated to 50°C. N 1 , N 1 -dimethylethane-1,2-diamine (2.79 mL, 25.62 mmol) was added, and the mixture was heated to 80°C (reflux) for 4 hours. TLC showed that the reaction was complete. The reaction mixture was concentrated, and the obtained residue was purified by column chromatography (DCM: MeOH = 30: 1 to 10: 1) to give AIR-2-1 (1.81 g, 27.86%) as a brown solid. MS (ESI), m / z: 305.4 [M+H] + .

[0186] 1 H NMR (300MHz, Chloroform-d) δ 8.65 (t, J=2.5Hz, 1H), 8.62 (t, J=2.5Hz, 1H), 7.47-7.39 (m, 2H), 4.38 (t, J=6.7Hz, 2H), 2.89-2.81 (m, 2H), 2.49 (s, 6H).

[0187] 5-4. Preparation of AIR-2

[0188] AIR-2-1 (1.81 g, 5.95 mmol) was dissolved in 1,4-dioxane (150 mL), and the reaction system was replaced with argon three times. 1-(2-pyridone-ethyl)piperazine (Compound A) (1.20 g, 6.54 mmol) was added, and the mixture was stirred at room temperature overnight. The reaction mixture was concentrated, and the resulting residue was purified by column chromatography (DCM: MeOH = 15: 1 to 5: 1) to give the pure product dissolved in DCM. 5M HCl / 1,4-dioxane (1.5 mL) was then added and stirring was continued, and the mixture was precipitated with diethyl ether. The resulting precipitate was filtered, washed with diethyl ether, and dried to give AIR-2 (1.21 g) as a yellow solid. MS (ESI), m / z: 468.47 [M+H] + .

[0189] 1 H NMR (300MHz, D2O) δ8.36 (dd, J=8.3, 4.7Hz, 1H), 8.29 (d, J=8.3Hz, 1H), 7.40 (dd, J=13.1, 8.3Hz, 1H), 7.27 (d, J=8.4Hz, 1 H), 4.42 (t, J=6.0Hz, 2H), 3.90-3.54 (m, 12H), 3.47 (t, J=6.0Hz, 2H), 3.38-3.12 (m, 4H), 2.99 (s, 6H), 2.21-2.02 (m, 4H).

[0190] Example I.6: Synthesis of Compound F (AIR-3)

[0191] 6-1. Preparation of Compound 2

[0192] NBS (499.72 g, 2.81 mol) was added to DMF (1000 mL) and stirred until most of the solid dissolved. The reaction system was purged with argon twice and then cooled to 9°C. Compound 1 (176 g, 1.14 mol) was added and stirred at 9°C for 2 hours. The reaction mixture was then allowed to warm to room temperature and stirred over the weekend. The reaction solution was filtered and the filter cake was washed twice with PE / EA = 1 / 1. The dried filter cake was collected and recrystallized from EA to obtain pure compound 2 (102.57 g, 28.80%) as off-white crystals.

[0193] 1 H NMR (300 MHz, Chloroform-d) δ 7.79 (d, J = 7.4 Hz, 2H), 7.09 (d, J = 8.8 Hz, 2H), 3.30 (s, 4H).

[0194] 6-2. Preparation of Compound 3

[0195] Compound 2 (50 g, 160.26 mmol) was dissolved in anhydrous THF (950 mL), the reaction system was replaced with argon three times, and then cooled to -78 ° C. Then n-BuLi (76.92 mL, 2.5 M / L, 192.31 mmol) was added dropwise, and the reaction mixture was stirred at the same temperature for at least 20 minutes. N-fluorobisbenzenesulfonamide (NFSI) (75.8 g, 240.38 mmol) in 250 mL of anhydrous THF was added dropwise, and after stirring at -78 ° C for 1.5 hours, the reaction mixture was naturally warmed to room temperature and stirred overnight. The reaction mixture was quenched with NH4Cl solution and then extracted twice with EA. The combined organic layer was washed with water and brine, dried and filtered with Na2SO4. The filtrate was concentrated to give a crude product, added to DCM and stirred. The resulting precipitate was filtered, the filter cake was washed twice with DCM, and all filtrates were combined and concentrated in vacuo to give the crude product, which was then purified by column chromatography (PE:EA=50:1) to give compound 3 (31.90 g, 79.27%) as a white solid.

[0196] 1 H NMR (300MHz, Chloroform-d) δ 7.66 (d, J=7.4Hz, 1H), 7.21-7.09 (m, 3H), 3.35 (s, 4H).

[0197] 6-3. Preparation of Compound 4

[0198] Compound 3 (9.29 g, 37.00 mmol) was added to a glacial acetic acid solution (100 ml) of potassium dichromate (50.07 g, 170.19 mmol). The solution was heated at reflux for 16 hours and then poured into ice water. The resulting precipitate was collected by filtration and washed with water. The filter cake was air-dried, added to DCM and refluxed for 30 minutes, filtered, washed with DCM, and the filter cake was added to DCM and refluxed for 15 minutes, filtered and washed with DCM. All filtrates were concentrated to give crude compound 3 (6.72 g), which can be used without further purification.

[0199] 6-4. Preparation of AIR-3-1

[0200] To a solution of compound 4 (400 mg, 1.36 mmol) in EtOH (20 mL) was added DIPEA (358.48 uL, 2.17 mmol) and tert-butyl (2-aminoethyl)(methyl)carbamate hydrochloride (428.46 mg, 2.03 mmol). The reaction system was replaced with argon five times and then stirred (refluxed) at 80° C. for 2 hours. TLC showed that the reaction was complete. The reaction mixture was concentrated and the obtained residue was purified by column chromatography (PE: EA = 10: 1 to 4: 1) to give AIR-3-1 (244 mg, 39.88%) as a yellow solid.

[0201] 6-5. Preparation of AIR-3-2

[0202] To a toluene solution (15 mL) of AIR-3-1 (244 mg, 540.68 mmol) was added NaOt-Bu (103.92 mg, 1.08 mol), 1-(2-pyridone-ethyl)piperazine (118.92 mg, 648.81 mmol) and BINAP (101.00 mg, 162.20 mmol). The reaction system was purged with argon 5 times, and then Pd(OAc) 2 (24.28 mg, 108.14 mmol) was added. The reaction system was purged with argon 3 times, and the reaction mixture was stirred at 90° C. for 12 hours under argon protection. The reaction mixture was quenched with water and extracted twice with EA. The combined organic layers were washed with water and brine, dried over Na 2 SO 4 and filtered. The filtrate was concentrated to give a crude product, which was purified by column chromatography (DCM:MeOH=100:1 to 10:1, with addition of NH3·H2O) and Prep-TLC (DCM:MeOH=10:1, with addition of NH3·H2O) to give AIR-3-2 (40 mg, 13.36%).

[0203] 1 H NMR (300MHz, Chloroform-d) δ8.56 (dd, J=8.2, 4.7Hz, 1H), 8.51 (d, J=8.3Hz, 1H), 7.29 (dd, J=7.7, 5.1Hz, 1H), 7.15 (d, J=8.3Hz, 1H), 4.36-4.29(m, 2H), 3.62-3.55(m, 2H), 3.45-3.11(m, 6H), 2.95(s, 3H), 2.82-2.62(m, 10H), 1.82-1.89(m, 4H), 1.14(s, 9H).

[0204] 6-6. Preparation of AIR-3

[0205] A solution of AIR-3-2 (40 mg, 72.24 mmol) in DCM / MeOH = 10 / 1 (3 mL) was replaced with argon three times, and then 5M HCl / 1,4-dioxane (1.5 mL) was added. The resulting mixture was stirred at room temperature for 1.5 hours. TLC showed that the reaction was complete. The reaction mixture was precipitated with diethyl ether, centrifuged, and the precipitate was washed with diethyl ether, centrifuged, and dried to give 33 mg of crude product. The crude product was further purified by Prep-TLC (DCM: MeOH = 20: 1, NH3·H2O was added) to give AIR-3 (10 mg) as a yellow solid. MS (ESI), m / z: 454.58 [M+H] + .HPLC 92.439% (220nm), HPLC 96.712% (254nm).

[0206] 1 H NMR (300MHz, D2O) δ8.35 (dd, J=8.4, 4.7Hz, 1H), 8.28 (d, J=8.3Hz, 1H), 7.39 (dd, J=13.2, 8.5Hz, 1H), 7.26 (d, J=8.3Hz, 1H), 4. 39-4.31 (m, 2H), 3.86-3.62 (m, 14H), 3.37-3.31 (m, 2H), 3.25-3.15 (m, 2H), 2.70 (s, 3H), 2.24-2.13 (m, 2H), 2.08-1.98 (m, 2H).

[0207] Example II. Activity Test

[0208] Example II.1: Activation of hnRNPA2B1 and TBK1-IRF3 pathways in immune cells by test compounds

[0209] To obtain primary mouse peritoneal macrophages: C57BL / 6 mice (6-8 weeks old, female, purchased from Shanghai Bikai Laboratory Animal Co., Ltd.) were intraperitoneally injected with 2 ml of 3% thioglycolate solution (purchased from Sigma-Aldrich). Three days later, the mice were sacrificed by cervical dislocation. The peritoneal cavity was flushed with serum-free medium, and the cells were aspirated and centrifuged to obtain the cells. Primary peritoneal macrophages were cultured in DMEM medium.

[0210] After stimulating primary peritoneal macrophages (cell density of 1 x 10^6 cells per well) with 20 μM concentration of test compound E (prepared as in Example 1.5, dissolved in physiological saline) for 30 minutes, the subcellular localization of hnRNPA2B1 was detected by immunofluorescence, and the activation of TBK1 and IRF3 was detected by SDS-PAGE electrophoresis and immunoblotting.

[0211] The activation of hnRNPA2B1, TBK1, and IRF3 are shown in Figures 1 and 2 .

[0212] The results showed that the test compound E could significantly activate the nuclear export of hnRNPA2B1 in macrophages and the downstream activation of TBK1 and IRF3. In addition, similar results were obtained by performing the same experiment using other compounds prepared in Example 1.

[0213] The above results demonstrate that the test compound has an activating effect on the hnRNPA2B1 pathway and is a hnRNPA2B1 agonist.

[0214] Example II.2: Broad induction of type I interferon production by test compounds in various cell types

[0215] Mouse primary peritoneal macrophages (prepared as described in Example II.1) were cultured with DMEM medium, human THP1 monocytes (purchased from ATCC) were cultured with 1640 medium, and L929 fibroblasts were cultured with DMEM medium.

[0216] After stimulating the cells with 20 μM of test compound E (cell density of 2 x 10^5 cells per well) for 6 hours, total RNA was extracted and the transcript level of type I interferon (IFN-β) was detected by real-time fluorescence quantitative PCR. After 18 hours of stimulation, the cell culture supernatant was collected and the IFN-β protein level was detected by ELISA.

[0217] The activation of type I interferon by the test compounds is shown in Figures 3, 4, and 5.

[0218] The results showed that the test compound E could significantly activate multiple cells to express type I interferon. In addition, similar results were obtained by performing the same experiment using other compounds prepared in Example 1.

[0219] The above results demonstrate that the test compound can induce the production of type I interferon by activating the hnRNPA2B1 pathway and is a biologically active hnRNPA2B1 agonist. Therefore, this disclosure defines such novel compounds as hnRNPA2B1 agonists.

[0220] Example II.3: hnRNPA2B1 agonist significantly induces the expression of multiple interferon-induced genes

[0221] Primary peritoneal macrophages (prepared as described in Example II.1) were cultured in DMEM medium, and L929 fibroblasts were cultured in DMEM medium.

[0222] After 18 hours of stimulation with the hnRNPA2B1 agonist compound E at a concentration of 20 μM (cell density of 2×10^5 cells per well), total RNA was extracted from the cells, and the expression of interferon-stimulated genes such as ISG15 and OAS1 was detected by real-time fluorescence quantitative PCR.

[0223] The transcription levels of interferon-stimulated genes are shown in Figures 6 and 7 .

[0224] The results showed that the hnRNPA2B1 agonist compound E could significantly activate the expression of interferon-stimulated genes in both immune cells and non-immune cells. In addition, similar results were obtained by performing the same experiment using other compounds prepared in Example 1.

[0225] Example II.4: hnRNPA2B1 inhibits the replication of the DNA virus HSV-1

[0226] L929 cells were cultured in DMEM medium, infected with HSV-1 (MOI, 1), and treated with the hnRNPA2B1 agonist compound E (10 μM) or negative control (normal saline). Total cellular RNA was collected after 24, 36, and 48 hours, and HSV-1 mRNA was detected by real-time fluorescence quantitative PCR.

[0227] The replication levels of HSV-1 are shown in FIG8 .

[0228] The results showed that the hnRNPA2B1 agonist compound E could significantly inhibit the replication of the DNA virus HSV-1. In addition, similar results were obtained by performing the same experiment using other compounds prepared in Example 1.

[0229] Example II.5: hnRNPA2B1 agonists inhibit the replication of the DNA virus HBV

[0230] HepG2.2.15 cells (cell density, 1x10^4 cells per well) were treated with the hnRNPA2B1 agonist compound E (10 μM) or negative control (normal saline). Total cellular RNA was collected 24, 36, and 48 hours later, and HBV mRNA was detected by real-time fluorescence quantitative PCR.

[0231] The HBV replication level is shown in FIG9 .

[0232] The results showed that the hnRNPA2B1 agonist compound E could significantly inhibit the replication of the DNA virus HBV. In addition, similar results were obtained by performing the same experiment using other compounds prepared in Example 1.

[0233] Example II.6: hnRNPA2B1 inhibits the replication of the RNA virus VSV

[0234] L929 cells were cultured in DMEM medium and infected with VSV (MOI, 1). They were also treated with hnRNPA2B1 agonist (10 μM) or negative control (normal saline). Total RNA was collected after 24, 36, and 48 hours, and VSV mRNA was detected by real-time fluorescence quantitative PCR.

[0235] The replication levels of VSV are shown in FIG10 .

[0236] The results showed that the hnRNPA2B1 agonist can significantly inhibit the replication of the RNA virus VSV. In addition, the same experiment was carried out using other compounds prepared in Example 1, and similar results were obtained.

[0237] Biological evaluation

[0238] The compounds of the present disclosure are defined as hnRNPA2B1 agonists by: (i) binding to the hnRNPA2B1 protein, as demonstrated by high affinity of the compounds to the hnRNPA2B1 protein at concentrations as low as 20 μM as determined by Biacore; and (ii) demonstrating by cell-based assays that the compounds can broadly induce higher levels of IFN-β production at concentrations as low as 20 μM.

[0239] In anti-cell infection tests with different types of infectious agents, the hnRNPA2B1 agonist compounds disclosed herein significantly inhibited the replication of infectious agents, thereby exerting an anti-infective effect. Furthermore, since the hnRNPA2B1 agonist compounds disclosed herein can broadly induce higher levels of IFN-β production, it is anticipated that these compounds may exert anti-infective effects through the action of type I interferons.

[0240] (i) Biacore T200 detection of the binding between compounds and hnRNPA2B1

[0241] Using S series CM5 chip; amino coupling reagents (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide EDC and N-hydroxysuccinimide NHS);

[0242] Coupling buffer: 10 mM sodium acetate pH 4.0, pH 4.5;

[0243] Running buffer: 10× PBS-P+;

[0244] Analytical grade DMSO, deionized water (0.22 μm membrane filtration);

[0245] hnRNPA2B1 recombinant protein (concentration must be greater than 0.5 mg / mL, total protein volume must be at least 20 μg);

[0246] Compound: Stock concentration greater than 20 mM, volume greater than 30 μL, purity greater than 90%, dissolved in 100% DMSO;

[0247] Other consumables: 96-well plates and sealing film, 1.5 mL centrifuge tubes without caps, and type 2 rubber bottle caps.

[0248] Experimental steps:

[0249] 1. Place the running buffer 1× PBS and waste liquid bottle in the designated positions and place the chip.

[0250] 2. Protein coupling. Select the immobilization program in the Wizard template, check flow cell 2 or 4, select amine as the method, and specify contact time and flow rate to achieve high coupling. Enter a contact time of 900 s and a flow rate of 10 μL / min.

[0251] 3. Prepare the required protein solution according to the instructions. Dilute the protein to the required volume with sodium acetate (pH 4.0) to a final concentration of 20 μg / mL. Place 100 μL of EDC, 100 μL of NHS, and 140 μL of ethanolamine in the corresponding positions.

[0252] 4. The system will automatically coat the chip surface with the target conjugated amount of hnRNPA2B1 protein and generate a conjugation report.

[0253] 5. Prepare the running buffer for the interaction. For small molecule samples, use 1× PBS-P+ containing 5% DMSO as the running buffer: dilute 105 mL of 10× PBS-P+ to 1 L with deionized water to make 1.05× PBS-P+.

[0254] Solvent correction stock solution containing 4.5% DMSO: 1.05×PBS-P + 9.5mL + 0.45mL DMSO

[0255] Solvent correction stock solution containing 5.8% DMSO: 1.05×PBS-P + 9.5mL + 0.58mL DMSO

[0256] Running buffer containing 5% DMSO: 1.05×PBS-P + 950 mL + 50 mL DMSO

[0257] Dilute a 10 mM small molecule stock solution in 1.05× PBS-P+ buffer without DMSO: 0.7 μL small molecule stock solution + 6.3 μL DMSO + 133 μL 1.05× PBS-P+ buffer, for a total volume of 140 μL, for a final concentration of 50 μM. Add to a 96-well plate and seal with film.

[0258] After the test, use Kinetics or Affinity mode to measure binding affinity.

[0259] (ii) Total cellular RNA can be extracted using TRIzol or a rapid RNA extraction kit.

[0260] 24-well plate cells (about 2-3×10 5 Rapid RNA extraction (100 cells / well): Discard the culture medium, add 500 μL RA2 lysis buffer to each well, pipette and lyse the cells several times, transfer the cells to the inner tube of the dedicated adsorption tube, and centrifuge at 12,000 rpm for 1 minute at room temperature. Discard the filtrate, add 500 μL wash buffer, and centrifuge at 12,000 rpm for 1 minute at room temperature. Repeat the wash operation once. Discard the filtrate, replace the inner tube, centrifuge at 12,000 rpm for 2 minutes at room temperature without adding wash buffer. Transfer the inner tube to a new 1.5 mL centrifuge tube, add 25 μL Elution Buffer to the center of the membrane, let it stand at room temperature for 5 minutes, and centrifuge at 12,000 rpm for 1 minute at room temperature. The resulting filtrate is RNA, and the RNA concentration is measured using Nanodrop One.

[0261] qRT-PCR: After the concentration of each sample was determined, 1 μg of total RNA was taken and ReverTra qRT-PCR RT Master Mix reverse transcribes RNA into cDNA. The reverse transcription system and reaction conditions are as follows:

[0262] Reverse transcription reaction system (20 μL system)

[0263] Reverse transcription reaction conditions (20 μL system)

[0264] Dilute the reverse-transcribed cDNA sample with 60 μL of deionized water, mix thoroughly, and centrifuge briefly. See the table for primer sequences used for qRT-PCR. Use SYBR Green Realtime PCR Master Mix for relative quantification of gene mRNA levels.

[0265] qRT-PCR reaction system (20 μL system)

[0266] qRT-PCR reaction conditions

[0267] qRT-PCR primers

[0268] All documents mentioned in this disclosure are incorporated herein by reference, just as if each document were incorporated herein by reference individually. It should also be understood that after reading the above teachings of this disclosure, those skilled in the art may make various changes or modifications to this disclosure, and that such equivalents also fall within the scope of the claims appended hereto.

Claims

1. A fused-ring compound represented by formula (I), its cis-trans isomers, its enantiomers, its diastereomers, its racemates, its solvates, its hydrates, or its pharmaceutically acceptable salts or its prodrugs, Among them, L is -(CH2)n-, where n is an integer from 0 to 6; X is a halogen; R 1 is -NR 4 R 5 , where R 4 and R 5 are each independently H or C 1-6 alkyl; R 2 and R 3 each independently is H or C 1-6 alkyl; or R 2 and R 3 together with the N atom to which they are attached form a 5- to 8-membered heterocycloalkyl group containing 1 to 3 heteroatoms selected from nitrogen, oxygen or sulfur; The C 1-6 alkyl group and the 5- to 8-membered heteroalkyl group are unsubstituted or substituted by one or more substituent groups selected from the group consisting of: hydroxyl, amino, halogen, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, C 1-6 alkoxy, hydroxy C 1-6 alkyl, C 1-6 alkylamino, C 3-6 cycloalkyl, C 3-6 cycloalkyl C 1-6 alkyl, 3- to 6-membered heteroalkyl, 3- to 6-membered heteroalkyl C 1-6 alkyl, 6- to 10-membered aryl, 6- to 10-membered aryl C 1-6 alkyl or 5- to 10-membered heteroaryl and 5- to 10-membered heteroaryl C 1-6 alkyl.

2. The compound according to claim 1, its cis-trans isomers, its enantiomers, its diastereoisomers, its racemates, its solvates, its hydrates or its pharmaceutically acceptable salts or its prodrugs, Among them, X is F or Cl; L is -(CH2)n-, where n is 1, 2 or 3; R 4 、R 5 are each independently H, methyl or ethyl; R 2 and R 3 each independently is C 1-3 alkyl, and the C 1-3 alkyl is unsubstituted or substituted by a 3- to 8-membered heterocyclic alkyl group containing 1 to 2 heteroatoms selected from nitrogen, oxygen, or sulfur; or R 2 and R 3 together with the N atom to which they are attached form a 5- to 8-membered heterocycloalkyl group containing 2 nitrogen atoms, said 5- to 8-membered heterocycloalkyl group being substituted by a 3- to 8-membered heterocycloalkyl C 1-6 alkyl or hydroxy C 1-6 alkyl substituted.

3. The compound according to claim 1, its cis-trans isomers, its enantiomers, its diastereoisomers, its racemates, its solvates, its hydrates or its pharmaceutically acceptable salts or its prodrugs, Among them, X is F; L is -(CH2)n-, where n is 2; R 4 、R 5 are each independently H, methyl or ethyl; R 2 and R 3 each independently is C 1-3 alkyl, and the C 1-3 alkyl is unsubstituted or substituted by 4- to 6-membered heterocycloalkyl containing 1 nitrogen heteroatom; or R 2 and R 3 together with the N atom to which they are attached form a piperazinyl group, said piperazinyl group being substituted by a 4- to 6-membered heterocycloalkyl C 1-6 alkyl or hydroxy C 1-6 alkyl substituted.

4. The compound according to claim 1, its cis-trans isomers, its enantiomers, its diastereoisomers, its racemates, its solvates, its hydrates or its pharmaceutically acceptable salts or its prodrugs, Among them, X is F; L is -(CH2)n-, where n is 2; R 4 、R 5 Each independently is H, methyl or ethyl; R 2 and R 3 each independently is methyl, ethyl or isopropyl, and the C 1-3 alkyl group is unsubstituted or unsubstituted or substituted by pyrrolidinyl; or R 2 and R 3 together with the N atom to which they are attached form a piperazinyl group, which is substituted by pyrrolidinoethyl or by hydroxyethyl.

5. The compound according to claim 1, its cis-trans isomers, its enantiomers, its diastereoisomers, its racemates, its solvates, its hydrates or its pharmaceutically acceptable salts or its prodrugs, said compound having the following formula (II): Among them, R a 、R b Each independently is H or C 1-6 alkyl; M is -(CH2)n-, where n is 1, 2 or 3; X is F or Cl; R c is a hydroxyl group, an amino group, a 3- to 8-membered heterocycloalkyl group containing 1 to 2 heteroatoms selected from nitrogen, oxygen or sulfur, more preferably R c is a hydroxyl group or a 4- to 6-membered heterocycloalkyl group containing 1 nitrogen heteroatom, more preferably, R c is a hydroxyl group or pyrrolidinyl, most preferably, R c is a hydroxyl group or 1-pyrrolidinyl.

6. The compound according to claim 1, its cis-trans isomers, its enantiomers, its diastereoisomers, its racemates, its solvates, its hydrates or its pharmaceutically acceptable salts or its prodrugs, wherein the compound is selected from the group consisting of: or its hydrochloride.

7. A product comprising the compound according to any one of claims 1 to 6, and a pharmaceutically or immunologically acceptable carrier or excipient.

8. The product according to claim 7, wherein: The product is selected from: pharmaceutical compositions, medicaments, kits or cartridges; and / or wherein the product is for inducing the production of type I interferon in a subject and / or for preventing and / or treating infectious diseases and / or diseases and / or symptoms associated with infection in a subject; and / or wherein the product further comprises: one or more other active substances for preventing or treating infectious diseases and their related conditions and / or symptoms, for example, the other active substances are one or more selected from the group consisting of: clinically commonly used antibiotics, including β-lactams (penicillins and cephalosporins), aminoglycosides, tetracyclines, chloramphenicols, macrolides, antifungal antibiotics, anti-tuberculosis antibiotics; clinically commonly used antiviral drugs (tricyclic amines, pyrophosphates, protease inhibitors, nucleoside drugs and interferons, antisense oligonucleotides, etc.); clinically commonly used immunosuppressants (including glucocorticoids, cyclophosphamide, chloroquine, cyclosporin A, traditional Chinese medicine preparations (such as Tripterygium wilfordii), anti-TNF monoclonal antibodies).

9. Use of the compound according to any one of claims 1 to 6 in the preparation of a product for inducing the production of type I interferon in a subject or for preventing and / or treating infectious diseases and / or diseases and / or symptoms associated with infection.

10. The application according to claim 9, wherein, The subject is a human or a non-human mammal, such as a non-human primate (gorilla, chimpanzee), a pet (such as a pet cat, dog, guinea pig, rabbit), a livestock animal (such as a cow, horse, donkey, mule, camel, pig, sheep, chicken, duck, goose, rabbit, deer, mink, otter, musk, etc.); and / or wherein the production of the type I interferon is mediated by hnRNPA2B1; and / or wherein the infectious disease in the subject can benefit from the production or increase in the level of type I interferon in vivo; and / or Among them, the infectious disease is caused by chemical, physical, or biological infection, such as caused by virus, bacterium, parasite, and / or fungus. If the infection is a DNA virus infection, it is an infection caused by one or more viruses selected from the following group: herpes simplex virus, hepatitis B virus, adenovirus, poxvirus, parvovirus, adeno-associated virus; and / or Among them, the disease and / or symptom related to infection is one or more selected from the following group: pathological damage caused by infection; insufficient or excessive production of cytokines (such as interferon) after infection; endotoxin shock or death; inflammatory damage of organs; multiple organ failure, for example, the organs are selected from: liver, spleen, brain, kidney, heart, lung, stomach, intestine; chronic inflammatory diseases caused by infection (such as autoimmune diseases like inflammatory bowel disease, rheumatoid arthritis, systemic lupus erythematosus, chronic nephritis, tuberculosis, chronic gastrointestinal diseases); and / or Among them, the product is selected from: pharmaceutical composition, medicine, kit, or medicine box; and / or Among them, the product is a product for inducing the production of type I interferon in a subject and / or for preventing and / or treating the infectious disease and / or the disease and / or symptom related to infection of the subject; and / or Among them, the product further comprises: one or more other active substances for preventing or treating infectious diseases and their related diseases and / or symptoms. For example, the other active substances are one or more selected from the following group: clinically commonly used antibiotics, including β-lactams (penicillins and cephalosporins), aminoglycosides, tetracyclines, chloramphenicols, macrolides, antifungal antibiotics, anti-tuberculosis antibiotics; clinically commonly used antiviral drugs (tricyclic amines, pyrophosphates, protease inhibitors, nucleoside drugs, interferons, antisense oligonucleotides, etc.); clinically commonly used immunosuppressants (including glucocorticoids, cyclophosphamide, chloroquine, cyclosporin A, traditional Chinese medicine preparations (such as Tripterygium wilfordii), anti-TNF monoclonal antibodies).

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