HnRNPA2B1 agonist compound and anti-infection application thereof

Abstract

The invention relates to a nuclear glycoprotein A2B1 (hnRNPA2B1) hnRNPA2B1 agonist compound of a heterogeneous cell nucleus and an anti-infection application of the nuclear glycoprotein A2B1 (hnRNPA2B1) hnRNPA2B1 agonist compound. Specifically, the invention relates to a fused ring compound as shown in a formula (I), cis-trans isomers, enantiomers, diastereoisomers, racemes, solvates, hydrates or pharmaceutically acceptable salts or prodrugs thereof, and a composition and a product containing any one of the former, the invention also relates to the use of the above substances as hnRNPA2B1 agonist in the preparation of products for the prevention and / or treatment of infectious diseases and / or diseases and / or symptoms related to infection. The invention can be used for broad-spectrum prevention and treatment of infection (such as virus infection), and has a wide application prospect. # imgabs0 #

Description

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 are agonists of nuclear protein hnRNPA2B1 (i.e., heterogeneous nuclear ribonucleoprotein A2B1), products containing the compounds, and their effects, mechanisms of action, implementation methods, and uses in preventing or treating diseases or symptoms associated with infection and controlling damage caused by infection. Background Art

[0002] Infection, especially viral infection, is a common clinical disease with great harm. Initially, people did not fully understand the molecular mechanism of the body's resistance to viral infection. With the discovery of a type of cytokine, interferon, the body's natural immune cells and secondary acquired immune cells produced against viral infection and the molecular biological basis of their actions have gradually been understood.

[0003] Interferon (IFN) is a general term for a family of cytokines with powerful antiviral functions. In 1957, Professors Alick Isaacs and Jean Lindenmann discovered a component in the process of studying influenza virus infection of chicken embryos, which can significantly prevent the proliferation of influenza viruses. They named this component interferon (Isaacs, A. et al., Proc R Soc Lond B Biol Sci. 1957; 927: 258-267.). Since then, cytokines of the interferon family have been found to have very broad-spectrum and effective antiviral effects. At present, interferon has been widely used in clinical defense against viral infections and treatment of various diseases caused by viral infections.

[0004] Type I interferon (IFN-I) directly or indirectly activates and regulates innate and adaptive immune cells by inducing other mediators during infection with viruses, bacteria, parasites, and fungi. It is a core cytokine for host defense against pathogenic microorganisms such as viruses and plays a very important role. In infected cells and neighboring cells, type I interferon induces the expression of interferon-stimulated genes (ISGs), inhibiting the further spread of pathogens. Innate immune cells also respond to type I interferon by enhancing antigen presentation and increasing the production of cytokines and chemokines. Adaptive immunity is also affected by type I interferon: for example, type I interferon can induce B cells to produce specific antibodies and amplify the effector function of T cells.

[0005] After innate immune receptors recognize pathogen-associated molecular patterns (PAMPs) or damage-associated molecular patterns (DAMPs), they induce the production of cytokines such as type I interferon and inflammatory factors, initiating innate immune responses. Among these PAMPs and DAMPs, nucleic acids derived from pathogens and from the body itself are a class of crucial ligands. Therefore, agonists targeting these innate immune recognition receptors have become an important means of treating infectious diseases.

[0006] The natural immune recognition receptors of nucleic acids include RNA recognition receptors and DNA recognition receptors. According to the different intracellular localization, DNA recognition receptors are mainly divided into cytoplasmic and nuclear DNA recognition receptors. The main DNA recognition receptor in the cytoplasm is cyclic GMP-AMP synthase (cGAS). After recognizing DNA in the cytoplasm, it will induce the activation of the Stimulator of Interferon Genes (STING), further recruit and activate TANK-binding kinase 1 (TBK1) and interferon regulatory factor 3 (IRF3), and initiate the production of type I interferon. In addition, TLR9 recognizes single-stranded DNA containing unmethylated CpG motifs, and AIM2 (Absent in melanoma-2, AIM2) can also recognize pathogen-derived DNA in the cytoplasm, promote the formation of inflammasomes, and the maturation and release of the inflammatory factor IL-1β.

[0007] Studies have found many proteins that can recognize viral DNA and induce IFN-α / β production, such as RNA polymerase III, IFI16, DAI, LRRFIP1, LSm14A, MRE11, DNA-PK, HMGBs, DDX41, and cyclic GMP-AMP (cGAMP) synthase (cGAS) (Goubau, D. et al.; Immunity, 2013; 38, 855-869.). After these PPRs recognize the virus, they stimulate the immune response signaling pathway in the host to induce the production of a large amount of type I interferon, thereby quickly establishing the body's first line of defense against infection and resisting the infection of pathogens. However, only cGAS and DNA-PK in the cytoplasm have been confirmed as DNA recognition receptors by in vivo experiments in mice. There are also several proteins involved in the inflammatory response induced by DNA viruses, including AIM2, IFI16, Rad50, and Sox2. Therefore, in-depth research on nuclear DNA recognition is urgently needed 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 in recent years, belonging to the hnRNP family. On the one hand, hnRNP-A2B1 can sense and recognize the nucleic acid components of DNA viruses (such as HSV-1, etc.), and self-activate to form homodimers, which are demethylated by the demethylase JMJD6 and translocated from the nucleus to the cytoplasm, thereby activating the TBK1-IRF3 signaling pathway and initiating the expression of type I interferon to exert an antiviral effect. On the other hand, as an RNA-binding protein, hnRNPA2B1 can also promote m6A modification, nuclear-cytoplasmic transfer and translation on cGAS, IFI16 and STING mRNA, thereby ensuring the sufficient induction of type I interferon expression to resist viral infection.

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

[0010] 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 its prodrugs), 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.

[0011] In some aspects of the present disclosure, provided are fused ring compounds represented by formula (I), cis-trans isomers, enantiomers, diastereomers, racemates, solvates, hydrates, or pharmaceutically acceptable salts or prodrugs thereof.

[0012]

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

[0014] X is a halogen;

[0015] R 1 For-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 N 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 hydroxyl, amino, halogen, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 1-6 Alkoxy, hydroxyl C 1-6 Alkyl, C 1-6 Alkylamino, C 3-6 Cycloalkyl, C 3-6 Cycloalkyl C 1-6Alkyl, 3-6 membered heterocycloalkyl, 3-6 membered heterocycloalkylC 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.

[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 an infectious disease and / or a disease and / or symptom associated with an 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, the method comprising administering a preventive 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, there is also provided a compound or product of the present disclosure 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 in 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 above technical solutions and technical features without departing from the inventive concept and protection scope of the present disclosure. Other aspects of the present disclosure are obvious to those skilled in the art due to the disclosure of this article. 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 rather than for limiting the scope of the present disclosure.

[0025] Figure 1 : The test compounds activated the nuclear export of hnRNPA2B1 in macrophages.

[0026] Figure 2 :The test compounds activated the downstream TBK1 and IRF3 after hnRNPA2B1 was exported from the nucleus of macrophages.

[0027] Figure 3 to Figure 5 : Test compounds on the activation of type I interferon in macrophages.

[0028] Figure 6-7 : Test compounds for their effects on activation of interferon-stimulated genes in immune cells.

[0029] Figure 8: Test compounds for their inhibitory effects on the replication of herpes simplex virus HSV-1.

[0030] Fig. 9 : Test the inhibitory effect of compounds on the replication of hepatitis B virus HBV.

[0031] Fig.10 : Test compounds for their inhibitory effects on the replication of vesicular stomatitis virus (VSV). DETAILED DESCRIPTION

[0032] In the present application, through a large amount of research, development and experiments, a new type of condensed ring compound is discovered, which can specifically bind to the hnRNPA2B1 protein with high affinity and can further widely induce the production of higher levels of IFN-β, and is thus defined as a hnRNPA2B1 agonist.

[0033] In the anti-cell infection test of different types of infection sources, the hnRNPA2B1 agonist compounds disclosed in the present invention can significantly inhibit the replication of infection sources, thereby playing an anti-infection role. In addition, since the hnRNPA2B1 agonist compounds disclosed in the present invention can widely induce the production of higher levels of IFN-β, it can be expected that these compounds can play an anti-infection role through the action of type I interferon.

[0034] Thus, the present disclosure provides methods and strategies for applying the novel anti-infective molecule hnRNPA2B1 agonist compounds 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 clearly include all values ​​falling between the range endpoints and the numerical ranges therebetween. Features mentioned in the present disclosure or features mentioned in the embodiments may be combined. All features disclosed in this specification may be used in combination with any composition form, and each feature disclosed in the specification may be replaced by any alternative feature that can provide the same, equal or similar purpose. Therefore, unless otherwise specified, the disclosed features are only general examples of equal or similar features.

[0036] As used herein, “containing,” “having,” or “including” include “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 and refer to heterogeneous nuclear ribonucleoprotein A2B1. The hnRNPA2B1 protein involved in the present disclosure may be a protein encoded by the hnRNPA2B1 gene, its cDNA or CDS in an animal (such as a human or mouse, etc.), or a homologous sequence of these proteins that promotes the expression of interferon (for example, the homologous sequence of hnRNPA2B1 can be obtained through a database or comparison software known in the art), a variant or a modified form.

[0039] As used herein, the terms "hnRNPA2B1 gene", "hnRNPA2B1 encoding gene", "hnRNPA2B1 protein encoding gene" or "nucleic acid molecule encoding hnRNPA2B1" are used interchangeably and refer to a nucleotide sequence encoding the hnRNPA2B1 protein or polypeptide described in the present disclosure, which can be, for example, the human hnRNPA2B1 gene of Gene ID: 3181, the mouse hnRNPA2B1 gene of Gene ID: 53379. The term also includes molecules that hybridize with the signed nucleic acid molecule under stringent conditions, or family gene molecules that are highly homologous to the above molecules, and it is believed that the expression of the gene has a certain promoting effect on the production and effect of interferon.

[0040] The present disclosure provides a class of novel compounds that are hnRNPA2B1 "agonists" (or "promoters"). The terms "agonist" or "hnRNPA2B1 agonist compound" are used interchangeably and refer to a class of novel condensed ring compounds that can increase the level or activity of hnRNPA2B1, which can specifically bind to the hnRNPA2B1 protein with high affinity, and can further widely 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 related 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]

[0044] Wherein, L is -(CH2)n-, and n is an integer of 0-6;

[0045] X is a halogen;

[0046] R 1 For-NR 4 R 5 , where R 4 , R 5 Each independently is H or C 1-6 alkyl;

[0047] R 2 and R 3 Each independently is H or C 1-6 alkyl;

[0048] or R 2 and R 3 Together with the N atom to which they are attached, they form a 5-8 membered heterocycloalkyl group containing 1-3 heteroatoms selected from nitrogen, oxygen or sulfur;

[0049] 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 hydroxyl, amino, halogen, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 1-6 Alkoxy, hydroxyl 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 heterocycloalkylC 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.

[0050] Preferably, X is F or Cl, more preferably, X is F.

[0051] Preferably, L is -(CH2)n-, wherein n is 1, 2 or 3, more preferably, n is 2.

[0052] Preferably, R 4 , R 5 Each is independently H, methyl or ethyl.

[0053] Preferably, R 2 and R 3 Each independently is C 1-3 Alkyl, preferably methyl, ethyl or isopropyl, the 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 3Each independently is C 1-3 Alkyl, preferably methyl, ethyl or isopropyl, the C 1-3 The alkyl group is unsubstituted or substituted with a 4-6 membered heterocycloalkyl group containing 1 nitrogen atom. 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.

[0054] Preferably, R 2 and R 3 Together with the nitrogen atom to which they are attached, they form a 5-8 membered heterocycloalkyl group containing 2 nitrogen atoms, wherein the 5-8 membered heterocycloalkyl group is surrounded by a 3-8 membered heterocycloalkyl group C 1-6 Alkyl or hydroxyl 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 hydroxyl 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 hydroxyl C 1-6 Alkyl substituted; most preferably, R 2 and R 3 Together with the N atom to which they are attached, they form a piperazinyl group, which is substituted by a 5-6 membered heterocycloalkyl C containing one N atom. 1-6 Alkyl substituted or hydroxyl C 1-6 Alkyl is substituted, preferably, the piperazinyl is substituted with a pyrrolidinyl C 1-6 Alkyl substituted or hydroxyl C 1-6 Alkyl substituted, more preferably, the piperazinyl is substituted by pyrrolidinylethyl or hydroxyethyl.

[0055] Preferably, the compound has the following formula (II):

[0056]

[0057] Among them, R a 、 R b Each independently is H or C 1-6 Alkyl, preferably, R a 、 R b Each is independently H, methyl or ethyl;

[0058] M is -(CH2)n-, wherein n is 1, 2 or 3, more preferably, n is 2;

[0059] X is F or Cl, more preferably, X is F;

[0060] R c is hydroxyl, amino, 3-8 membered heterocycloalkyl containing 1-2 heteroatoms selected from nitrogen, oxygen or sulfur, more preferably, R c is hydroxyl or a 4-6 membered heterocycloalkyl group containing 1 nitrogen atom, more preferably, R c is hydroxyl or pyrrolidinyl, most preferably, R c is hydroxy or 1-pyrrolidinyl.

[0061] Preferably, the compound is selected from the following group of compounds or their salts (eg, hydrochloride):

[0062]

[0063] Unless otherwise indicated, the following terms and phrases used herein are intended to have the following definitions. A particular term or phrase should not be considered to be uncertain or unclear in the absence of a special definition, but should be understood according to its ordinary meaning. When a trade name appears in this article, it is intended to refer to its corresponding commercial product or active ingredient.

[0064] The term "pharmaceutically acceptable" as used herein refers to those compounds, materials, compositions and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0065] The term "pharmaceutically acceptable salt" refers to a salt of a compound of the invention, prepared from a compound having a specific substituent discovered by the invention and a relatively nontoxic acid or base. When the compound of the invention contains a relatively acidic functional group, a base addition salt can be obtained by contacting the neutral form of such compound with a sufficient amount of base in a pure solution or a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium calcium, ammonium, organic amine or magnesium salts or similar salts. When the compound of the invention contains a relatively basic functional group, a base addition salt can be obtained by contacting the neutral form of such compound with a sufficient amount of acid in a pure solution or a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts, such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, bicarbonate, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, hydrogen sulfate, hydroiodic acid, phosphorous acid, etc., and organic acid salts, 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, and salts of amino acids (such as arginine, etc.), and salts of organic acids such as glucuronic acid. Certain specific compounds of the present invention contain basic and acidic functional groups, and can be converted into any base or acid addition salt.

[0066] Pharmaceutically acceptable salts of the present invention can be synthesized by conventional chemical methods from parent compounds containing acid radicals or bases. Generally, the preparation method of such salts is: in water or an organic solvent or a mixture of the two, these compounds in free acid or base form are reacted with a stoichiometric amount of an appropriate base or acid to prepare.

[0067] 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. All of these isomers and their mixtures are included within the scope of the present invention.

[0068] Unless otherwise indicated, the term "enantiomer" or "optical isomer" refers to stereoisomers that are mirror images of one another.

[0069] Unless otherwise indicated, the term "cis-trans isomers" or "geometric isomers" arises from the inability of a ring to rotate freely about double bonds or single bonds of ring carbon atoms.

[0070] 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.

[0071] The compounds of the present invention may exist in specific forms. Unless otherwise indicated, the term "tautomer" or "tautomeric form" means that at room temperature, different functional group isomers are in dynamic equilibrium and can quickly convert to each other. If tautomers are possible (such as in solution), it is possible to reach a chemical equilibrium of tautomers. For example, proton tautomers (proton tautomers) (also known as prototropic tautomers) include interconversions carried out by proton migration, such as keto-enol isomerization and imine-enamine isomerization. Valence isomers (valencetautomers) include interconversions carried out by the reorganization of some bonding monads. Among them, 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.

[0072] The term "prodrug" generally refers to a compound of formula (I) that has been derivatized with a functional group, and its derivative can be easily converted into a compound of formula (I) in vivo. Generally, the selection and preparation of suitable prodrugs can be described in, for example, Design of Prodrugs, ed. H. Bundgaard, Elsevier, 1985.

[0073] The compounds of the invention may contain unnatural proportions of atomic isotopes on one or more of the atoms constituting the compound, said isotopes having the same atomic number but an atomic mass or mass number different from the atomic mass or mass number that predominantly occurs 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 included within the scope of the present invention. Isotopic variants may enhance certain therapeutic advantages, such as the substitution of heavy hydrogen for hydrogen to form deuterated drugs, the bond between deuterium and carbon being stronger than the bond between ordinary hydrogen and carbon, and the deuterated drugs having advantages such as reduced toxic side effects, increased drug stability, enhanced efficacy, and prolonged drug biological half-life compared to undeuterated drugs, or providing standard compounds that can be used as characterization of biological samples. Isotopically enriched compounds within the general formula (I) may 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 routes and embodiments of the present invention, using appropriate isotopically enriched reagents and / or intermediates.

[0074] "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.

[0075] The naming convention used in the present invention is based on the IUPAC system nomenclature generated by ChemDraw software.Any open valence bonds occurring on carbon, oxygen, sulfur or nitrogen atoms in the structures given in the present invention indicate the presence of hydrogen atoms.

[0076] The term "substituted" means that any one or more hydrogen atoms on a specific atom are replaced by a substituent, which may include deuterium and hydrogen variants, as long as the valence state of the specific 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 not substituted, and unless otherwise specified, the type and number of substituents can be arbitrary on the basis of chemical realization.

[0077] 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, and each occurrence of R is an independent choice. In addition, combinations of substituents and / or variants thereof are permitted only if such combinations result in stable compounds.

[0078] When the number of a linking group is 0, such as -(CRR)0-, it means that the linking group is a single bond.

[0079] 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.

[0080] When the listed linking group does not indicate its linking direction, its linking direction is arbitrary, for example, The connecting group L is at this time You can connect the benzene ring and the cyclopentyl group 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.

[0081] Unless otherwise specified, the number of atoms in a ring is generally defined as the ring member number, for example, "3-7 membered ring" refers to a "ring" having 3-7 atoms arranged around it.

[0082] Unless otherwise specified, the term "halogen" refers to fluorine, chlorine, bromine and iodine.

[0083] Unless otherwise specified, the term “C 1-6 "Alkyl" is used to refer to 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.; it 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.

[0084] Unless otherwise specified, the term “C 1-3 "Alkyl" is used to refer to 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 may be monovalent (such as methyl), divalent (such as methylene) or polyvalent (such as methine). 1-3 Examples of the alkyl group include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), and the like.

[0085] Unless otherwise specified, the term “C 2-6"Alkenyl" is used to refer to a straight or branched hydrocarbon group consisting of 2 to 6 carbon atoms containing at least one carbon-carbon double bond. The carbon-carbon double bond can be located at any position of the group. 2-6 Alkenyl includes 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.

[0086] Unless otherwise specified, the term “C 1-6 "Alkoxy" refers to those alkyl groups containing 1 to 6 carbon atoms which are attached to the rest of the molecule through 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.

[0087] Unless otherwise specified, the term “C 1-6 "Alkylamino" refers to those alkyl groups containing 1 to 6 carbon atoms which are attached to the rest of the molecule via 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.

[0088] 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 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 include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like.

[0089] Unless otherwise specified, the term "3-8 membered heterocycloalkyl" by itself or in combination with other terms refers to 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 rest are 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, for the "3-8 membered heterocycloalkyl", a heteroatom may occupy the position where the heterocycloalkyl is connected to the rest of the molecule. The 3-8 membered heterocycloalkyl includes 4-6 membered, 5-6 membered, 7-8 membered, 4 membered, 5 membered and 6 membered heterocycloalkyls, etc. Examples of 3-6 membered heterocycloalkyls include, but are not limited to, azetidinyl, oxetanyl, thietanyl, pyrrolidinyl, pyrazolidinyl, imidazolidinyl, tetrahydrothienyl, tetrahydrofuranyl (including tetrahydrofuran-2-yl), piperidinyl, tetrahydropyranyl, tetrahydrothiopyranyl, piperazinyl, morpholinyl, thiomorpholinyl, etc.

[0090] Unless otherwise specified, the terms "6-10 membered aromatic ring" and "6-10 membered aryl" can be used interchangeably, and the term "6-10 aryl" means a monovalent aromatic carbocyclic ring system containing 6-10 carbon atoms and having at least one aromatic ring or multiple condensed rings in which at least one ring is an aromatic ring. Examples of aryl are, but not limited to, phenyl, naphthyl, biphenyl or indanyl, etc.

[0091] 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 with a conjugated π electron system consisting of 5 to 10 ring atoms, 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 and 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 may be attached to the rest of the molecule via a heteroatom or a carbon atom, including 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, 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 1-thiazolyl and 5-thiazolyl, etc.), furanyl (including 2-furanyl and 3-furanyl, 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.

[0092] 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 with a conjugated π electron system consisting of 5 to 6 ring atoms, 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). The 5-6 membered heteroaryl may be attached to the rest of the molecule via a heteroatom or a carbon atom, and the 5-6 membered heteroaryl includes 5-membered and 6-membered heteroaryl, etc. Examples of the 5-6 membered heteroaryl include, but are not limited to, pyrrolyl (including N-pyrrolyl, 2-pyrrolyl and 3-pyrrolyl, etc.), pyrazolyl (including 2-pyrazolyl and 3-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.), furanyl (including 2-furanyl and 3-furanyl, 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.

[0093] 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 Etc.; similarly, n-membered to n+m-membered means that the number of atoms in the ring is n to n+m, for example, 3-7-membered ring includes 3-membered, 4-membered, 5-membered, 6-membered and 7-membered rings, and also includes any range from n to n+m, for example, 3-7-membered ring includes 3-6-membered ring, 4-7-membered ring, 5-7-membered ring and 6-7-membered ring, etc.

[0094] Preparation method

[0095] The present invention also relates to a method for producing the compound of the general formula (I) defined above. In some embodiments, the synthesis method of the compound of the present application is as follows:

[0096]

[0097] When X is selected from halogens other than Br, such as F, Cl, I, etc., the method comprises the following steps:

[0098] 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) or 1,3-dibromo-5,5-dimethylhydantoin, preferably, the reaction temperature is 0-30° C.;

[0099] 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;

[0100] c) oxidizing the compound of formula Ic to the compound of formula Id using an oxidant, preferably, the oxidant includes but is not limited to K2Cr2O7, preferably, the reaction temperature is 60-150°C;

[0101] 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.;

[0102] 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.

[0103] When X is selected from Br, the method comprises:

[0104] 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;

[0105] c) oxidizing the compound of formula Ib to a compound of formula Id using an oxidant, preferably, the oxidant includes but is not limited to K2Cr2O7, preferably, the reaction temperature is 60-150°C;

[0106] 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;

[0107] 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.

[0108]

[0109] Alternatively, when X is selected from halogens other than Br, such as F, Cl, I, etc., the method comprises the following steps:

[0110] f) 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) or 1,3-dibromo-5,5-dimethylhydantoin, preferably, the reaction temperature is 0-30° C.;

[0111] 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;

[0112] h) oxidizing the compound of formula I-c' to a compound of formula I-d' using an oxidant, preferably, the oxidant includes but is not limited to K2Cr2O7, preferably, the reaction temperature is 60-150°C;

[0113] 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;

[0114] 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.

[0115] 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.

[0116] Products and Applications

[0117] The present disclosure also provides a product, which may be, for example, a medicament, a pharmaceutical composition or a kit, containing an effective amount of the 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 and refer to a hnRNPA2B1 agonist compound having the structural formula (I) or a derivative or product thereof.

[0118] As used herein, the term "pharmaceutically acceptable" ingredients are suitable for humans and / or animals without excessive adverse reactions (such as toxicity, irritation and allergic reactions), i.e., substances with a reasonable benefit / risk ratio. As used herein, the term "effective amount" refers to an amount that can produce a function or activity to humans and / or animals and can be accepted by humans and / or animals.

[0119] As used herein, the term "pharmaceutically acceptable carrier" refers to a carrier for the administration of a therapeutic agent, including various excipients and diluents. The term refers to such pharmaceutical carriers: they are not necessary active ingredients themselves and are not excessively toxic after administration. Suitable carriers are well known to those of ordinary skill in the art, and a full discussion of pharmaceutically acceptable excipients can be found in Remington's Pharmaceutical Sciences (Mack Pub. Co., NJ 1991).

[0120] The pharmaceutically acceptable carrier in the composition may contain a liquid such as water, saline, glycerol and ethanol. In addition, auxiliary substances such as fillers, disintegrants, lubricants, glidants, effervescent agents, wetting agents or emulsifiers, flavoring agents, pH buffer substances, etc. may also be present in these carriers. 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, the pH is about 6-8.

[0121] The active substance in the product disclosed herein accounts for 0.001-99.9 wt% of the total weight of the composition, preferably 1-95 wt%, more preferably 5-90 wt%, and more preferably 10-80 wt%, and the remainder is pharmaceutically acceptable carriers and other additives.

[0122] As used herein, the term "unit dosage form" means that the product of the present disclosure is prepared into a dosage form required for a single dose for the convenience of taking, including but not limited to various solid doses (such as tablets), liquid doses, capsules, and sustained-release preparations.

[0123] In another preferred embodiment of the present disclosure, the product is a unit dosage form or a multi-dose form, and the content of the active substance is 0.01-2000 mg / dose, preferably 0.1-1500 mg / dose, and more preferably 1-1000 mg / dose. In another preferred embodiment of the present disclosure, 1-6 doses of the composition of the present disclosure are administered per day, preferably 1-3 doses are administered; most preferably, the dose taken per day is 1 dose.

[0124] It should be understood that the effective dose of the active substance such as the hnRNPA2B1 agonist compound used may vary depending on the severity of the subject to be administered or treated. The specific situation is determined according to the individual situation of the subject (e.g., the subject's weight, age, physical condition, and the desired effect), which is within the scope of the judgment of a skilled physician.

[0125] The products disclosed herein may be solid (such as granules, tablets, lyophilized powders, suppositories, capsules, sublingual tablets) or liquid (such as oral liquids) or other suitable shapes. The routes of administration may include, but are not limited to: (1) traditional administration methods, such as gastrointestinal administration (such as oral administration), parenteral administration (such as 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, etc.; (2) connecting the hnRNPA2B1 agonist to the transferrin / poly-L-lysine complex to enhance its biological effect; (3) mediating the entry of drugs into cells after encapsulating them with liposomes, which is conducive to the smooth entry of compound molecules and is protected from the hydrolysis of various extracellular enzymes; (4) using liposomes to transport active substances so that they are specifically transported to target tissues and target cells.

[0126] In addition, the product of the present disclosure may also contain other active substances for improving and treating infectious diseases. The 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, chloramphenicol, macrolides, antifungal antibiotics, and anti-tuberculosis antibiotics.

[0127] In some embodiments, before, simultaneously with or after the administration of the product of the present disclosure, other active substances that regulate anti-infection are administered. 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 their symptoms.

[0128] In some embodiments, further regulating the immune system by activating hnRNPA2B1 of the present application compound or product provides the treatment of disease, including diseases caused by exogenous factors. Exemplary infections of exogenous factors that can be treated and / or prevented by the method 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 present 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).

[0129] 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.

[0130] In some embodiments, the infection that can be treated or prevented by the compounds of the present application is an infection involved and / or mediated by DNA. 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, such as a DNA virus 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, small DNA virus, adeno-associated virus, coronavirus, influenza virus, rhinovirus, parainfluenza virus, respiratory syncytial virus, coxsackievirus, echovirus, and novel enterovirus.

[0131] 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 organ is 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.

[0132] 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 at intervals before and after infection occurs.

[0133] 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 are obvious to those skilled in the art due to the disclosure of this article.

[0134] Example

[0135] The present disclosure is further described below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present disclosure and are not used to limit the scope of the present disclosure. Those skilled in the art may make appropriate modifications and changes to the present disclosure, and these modifications and changes are within the scope of the present disclosure.

[0136] The experimental methods for which the specific conditions are not specified in the following examples can be conventional methods in the art, for example, refer to "Molecular Cloning Laboratory Manual" (3rd edition, New York: Cold Spring Harbor Laboratory Press, 1989) or follow the conditions recommended by the supplier. The DNA sequencing method is a conventional method in the art and can also be tested by commercial companies.

[0137] Unless otherwise specified, percentages and parts are calculated by weight. Unless otherwise defined, all professional 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 applied to the disclosed methods. The preferred implementation methods and materials described herein are for demonstration purposes only.

[0138] I. Synthesis and Characterization Examples of Compounds

[0139] Example I.1: Synthesis of Compound A (FPC-1):

[0140]

[0141] *eq equivalent

[0142] 1-1. Preparation of Compound 2

[0143]

[0144] 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, the resulting suspension was stirred to allow it to naturally warm to room temperature and stirred overnight. The reaction solution was filtered, the filter cake was washed three times with ethanol, the dried filter cake was collected, and recrystallized with EA to obtain a pure product (10.1 g, off-white crystals).

[0145] 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).

[0146] 1-2. Preparation of Compound 3

[0147]

[0148] 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 N-fluorobisbenzenesulfonamide (NFSI) (9 g, 28.8 mmol) in 100 mL of anhydrous THF was slowly added, the mixture was stirred for another 60 minutes at -80 ° C. 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 with 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.

[0149] 1 H NMR (300 MHz, CHLOROFORM-d) δ 7.66 (d, J = 7.4 Hz, 1H), 7.21–7.09 (m, 3H), 3.35 (s, 4H).

[0150] 1-3. Preparation of Compound 4

[0151]

[0152] Compound 3 (4.5g, 17.9mmol) is added in the glacial acetic acid solution (50mL) of potassium dichromate (24g, 82.4mmol).The solution is heated to reflux 16 hours, then poured into ice water.Gained precipitate is collected by filtration and washed with water.Filter cake is air-dried, added among the DCM and refluxed 30 minutes, filtered, washed with DCM, filter cake is added among the DCM and refluxed 15 minutes, filtered and washed with DCM.Merge all filtrates and vacuum concentration to obtain compound 4 (4.1g, crude product, yellow solid).

[0153] 1-4. Preparation of INT-1

[0154]

[0155] 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 under reflux for 1 hour. The reaction mixture was concentrated, and the residue was purified by column chromatography (PE / EA=5 / 1). An off-white solid (2.8 g) was obtained.

[0156] 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).

[0157] 1-5. Preparation of FPC-1-1

[0158]

[0159] 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, and 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+NH3·H2O) to obtain 240 mg of pure yellow solid.

[0160] 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).

[0161] 1-6. Preparation of FPC-1

[0162]

[0163] Compound FPC-1-1 (240 mg, 0.44 mmol) was dissolved in 11 mL DCM / MeOH (10 / 1). 5M 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, the residue was dissolved in a small amount of methanol, precipitated with ether, centrifuged, the precipitate was washed with ether, centrifuged and dried in vacuo to obtain 240 mg of pure product (yellow solid).

[0164] 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).

[0165] MS (ESI) m / z [M+H] + =440.3.

[0166] Example I.2: Synthesis of Compound B (FPC-2)

[0167]

[0168] 2-1. Preparation of FPC-2-1

[0169]

[0170] 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.

[0171] 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).

[0172] 2-2. Preparation of FPC-2

[0173]

[0174] Compound FPC-2-1 (20 mg, mmol) was dissolved in 2 mL 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 obtain 5 mg of pure product (yellow solid).

[0175] 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).

[0176] MS (ESI) m / z [M+H] + =401.47.

[0177] Example I.3: Synthesis of Compound C (FPC-3)

[0178]

[0179] 3-1. Preparation of FPC-3-1

[0180]

[0181] 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), and bis(triphenylphosphine)palladium chloride (II) (128 mg, 0.18 mmol) was added to the mixture under argon protection, and 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+NH3·H2O) to obtain 30 mg of pure yellow solid.

[0182] 1 H NMR (400MHz, chloroform-d) δ8.63-8.52(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).

[0183] 3-2. Preparation of FPC-3

[0184]

[0185] Compound FPC-3-1 (30 mg, 0.06 mmol) was dissolved in 4.4 mL 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, centrifuged, and the precipitate was washed with ether, centrifuged and dried in vacuo to obtain 25 mg of the pure product (yellow solid).

[0186] 1 H 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).

[0187] MS (ESI) m / z [M+H] + =399.47.

[0188] Example I.4: Synthesis of Compound D (FPC-5)

[0189]

[0190] 4-1. Preparation of FPC-5-1

[0191]

[0192] 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), and bis(triphenylphosphine)palladium chloride (II) (32 mg, 0.0.045 mmol) was added to the mixture under argon protection. The reaction was carried out at 80°C for 16 hours, and the reaction solution was cooled to room temperature. Water was added to the reaction solution and stirred, and the organic phase was separated. The aqueous phase was extracted once with DCM, and the organic phases were combined and dried over anhydrous magnesium sulfate. The crude product was purified by TLC (DCM / MeOH=10 / 1+NH3·H2O) by vacuum concentration and column purification to obtain 10 mg of pure yellow solid.

[0193] 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).

[0194] 4-2. Preparation of FPC-5

[0195]

[0196] Compound FPC-5-1 (10 mg, 0.022 mmol) was dissolved in 1.1 mL 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, centrifuged, and the precipitate was washed with ether, centrifuged and dried in vacuo to obtain 9 mg of pure product (yellow solid).

[0197] 1 H NMR (400MHz, methanol-d4) δ8.61-8.52(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).

[0198] MS (ESI) m / z [M+H] + =344.42.

[0199] Example I.5: Synthesis of Compound E (AIR-2)

[0200]

[0201] 5-1. Preparation of Compound 5

[0202]

[0203] 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 for at least 10 minutes at the same temperature. N-fluorobisbenzenesulfonamide (NFSI) (60.09 g, 190.57 mmol) in 200 mL anhydrous THF was added dropwise, and after stirring for 1 hour at -78 ° C, 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 obtain 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) to give compound 5 (15.42 g, 63.82%) as a white solid.

[0204] 1 H NMR (300MHz, Chloroform-d) δ7.16 (d, J = 7.6Hz, 2H), 7.12–7.04 (m, 2H), 3.37 (s, 4H).

[0205] 5-2. Preparation of INT-2

[0206]

[0207] 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, and PE and ultrasonic dispersion were added. The resulting precipitate was filtered and dried to obtain crude INT-2 (10.55 g) as a brown solid, which was used without further purification.

[0208] 5-3. Preparation of AIR-2-1

[0209]

[0210] 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), 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 a brown solid AIR-2-1 (1.81 g, 27.86%). MS (ESI), m / z: 305.4 [M + H] + .

[0211] 1 H NMR (300MHz, Chloroform-d) δ8.65(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).

[0212] 5-4. Preparation of AIR-2

[0213]

[0214] 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 residue obtained was purified by column chromatography (DCM: MeOH = 15: 1 to 5: 1) to obtain a pure product dissolved in DCM. Then 5M HCl / 1,4-dioxane (1.5 mL) was added and stirring was continued, and precipitated with ether. The resulting precipitate was filtered, washed with ether, and dried to give a yellow solid AIR-2 (1.21 g). MS (ESI), m / z: 468.47 [M+H] + .

[0215] 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).

[0216] Example I.6: Synthesis of Compound F (AIR-3)

[0217]

[0218] 6-1. Preparation of Compound 2

[0219]

[0220] NBS (499.72 g, 2.81 mol) was added to DMF (1000 mL) and stirred until most of the solid was dissolved. The reaction system was replaced 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 allowed to warm to room temperature naturally 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 with EA to obtain pure compound 2 (102.57 g, 28.80%) as off-white crystals.

[0221] 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).

[0222] 6-2. Preparation of Compound 3

[0223]

[0224] 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 for at least 20 minutes at the same temperature. N-fluorobisbenzenesulfonamide (NFSI) (75.8 g, 240.38 mmol) in 250 mL anhydrous THF was added dropwise, and after stirring for 1.5 hours at -78 ° C, 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 a 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.

[0225] 1H NMR (300MHz, Chloroform-d) δ7.66 (d, J = 7.4Hz, 1H), 7.21–7.09 (m, 3H), 3.35 (s, 4H).

[0226] 6-3. Preparation of Compound 4

[0227]

[0228] 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 to 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, 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 could be used without further purification.

[0229] 6-4. Preparation of AIR-3-1

[0230]

[0231] DIPEA (358.48uL, 2.17mmol) and (2-aminoethyl)(methyl)carbamic acid tert-butyl ester hydrochloride (428.46mg, 2.03mmol) were added to a solution of compound 4 (400mg, 1.36mmol) in EtOH (20mL). The reaction system was replaced with argon 5 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 a yellow solid AIR-3-1 (244mg, 39.88%).

[0232] 6-5. Preparation of AIR-3-2

[0233]

[0234] 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) were added to a toluene solution (15 mL) of AIR-3-1 (244 mg, 540.68 mmol). The reaction system was replaced with argon 5 times, and then Pd(OAc)2 (24.28 mg, 108.14 mmol) was added. The reaction system was replaced with argon three 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 layer was washed with water and brine, dried over Na2SO4 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%).

[0235] 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).

[0236] 6-6. Preparation of AIR-3

[0237]

[0238] A DCM / MeOH=10 / 1 solution (3 mL) of AIR-3-2 (40 mg, 72.24 mmol) 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 ether, centrifuged, and the precipitate was washed with ether, centrifuged, and dried to give a crude product of 33 mg. The crude product was further purified by Prep-TLC (DCM:MeOH=20:1, NH3·H2O was added) to give a yellow solid AIR-3 (10 mg). MS (ESI), m / z: 454.58 [M+H] +.HPLC 92.439% (220nm), HPLC 96.712% (254nm).

[0239] 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).

[0240] Example II. Activity Test

[0241] Example II.1: Activation of hnRNPA2B1 and TBK1-IRF3 pathways in immune cells by test compounds

[0242] Obtaining primary peritoneal macrophages from mice: 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 (purchased from Sigma-Aldrich) solution once. Three days later, the mice were killed by cervical dislocation, and the peritoneal cavity was flushed with serum-free medium, and the cells were obtained by centrifugation after aspiration. Primary peritoneal macrophages were cultured with DMEM medium.

[0243] After stimulating primary peritoneal macrophages (cell density of 1x10^6 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.

[0244] The activation of hnRNPA2B1, TBK1, and IRF3 is shown in Figure 1 , 2 shown.

[0245] The results showed that the test compound E could significantly activate the nuclear export of hnRNPA2B1 in macrophages and the activation of downstream TBK1 and IRF3. In addition, the same experiment was carried out using other compounds prepared in Example 1, and similar results were obtained.

[0246] The above results demonstrate that the test compound has an activating effect on the hnRNPA2B1 pathway and is a hnRNPA2B1 agonist.

[0247] Example II.2: Broad induction of type I interferon production by test compounds in various cell types

[0248] 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.

[0249] After stimulating the cells with 20 μM concentration of test compound E (cell density was 2x10^5 cells per well) for 6 hours, total RNA of the cells was extracted and the transcription 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.

[0250] The test compound activates type I interferon as Figure 3 , 4 , as shown in Figure 5.

[0251] The results showed that the test compound E could significantly activate multiple cells to express type I interferon. In addition, the same experiment was performed using other compounds prepared in Example I, and similar results were obtained.

[0252] The above results prove 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 type of novel compound is defined as a hnRNPA2B1 agonist in the present disclosure.

[0253] Example II.3: hnRNPA2B1 agonist significantly induces the expression of multiple interferon-induced genes

[0254] Primary peritoneal macrophages (prepared as described in Example II.1) were cultured in DMEM medium, and L929 fibroblasts were cultured in DMEM medium.

[0255] After stimulating the cells (cell density was 2x10^5 cells per well) with the hnRNPA2B1 agonist compound E at a concentration of 20 μM for 18 hours, total RNA of the cells was extracted and the expression of interferon-stimulated genes such as ISG15 and OAS1 was detected by real-time fluorescence quantitative PCR.

[0256] Transcription levels of interferon-stimulated genes Figure 6 , 7 shown.

[0257] The results showed that the hnRNPA2B1 agonist compound E could significantly activate the expression of interferon-stimulated genes in immune cells and non-immune cells. In addition, the same experiment was carried out using other compounds prepared in Example 1, and similar results were obtained.

[0258] Example II.4: hnRNPA2B1 inhibits the replication of the DNA virus HSV-1

[0259] L929 cells were cultured with DMEM medium, infected with HSV-1 (MOI, 1), and treated with hnRNPA2B1 agonist compound E (10 μM) or negative control (normal saline). Total cell RNA was collected after 24, 36, and 48 hours, and HSV-1 mRNA was detected by real-time fluorescence quantitative PCR.

[0260] The replication level of HSV-1 is Figure 8 shown.

[0261] The results showed that the hnRNPA2B1 agonist compound E could significantly inhibit the replication of DNA virus HSV-1. In addition, the same experiment was carried out using other compounds prepared in Example 1, and similar results were obtained.

[0262] Example II.5: hnRNPA2B1 agonists inhibit the replication of DNA virus HBV

[0263] HepG2.2.15 cells (cell density was 1x10^4 cells per well) were treated with hnRNPA2B1 agonist compound E (10 μM) or negative control (normal saline). Total cellular RNA was collected after 24, 36, and 48 hours, and HBV mRNA was detected by real-time fluorescence quantitative PCR.

[0264] The replication level of HBV is Fig. 9 shown.

[0265] The results showed that the hnRNPA2B1 agonist compound E could significantly inhibit the replication of DNA virus HBV. In addition, the same experiment was carried out using other compounds prepared in Example I, and similar results were obtained.

[0266] Example II.6: hnRNPA2B1 inhibits the replication of the RNA virus VSV

[0267] L929 cells were cultured in DMEM medium, infected with VSV (MOI, 1), and treated with hnRNPA2B1 agonist (10 μM) or negative control (normal saline). Total cell RNA was collected after 24, 36, and 48 hours, and VSV mRNA was detected by real-time fluorescence quantitative PCR.

[0268] The replication level of VSV is Fig.10 shown.

[0269] The results showed that the hnRNPA2B1 agonist can significantly inhibit the replication of RNA virus VSV. In addition, the same experiment was carried out using other compounds prepared in Example 1, and similar results were obtained.

[0270] Biological evaluation

[0271] The compounds disclosed herein are defined as hnRNPA2B1 agonists by: (i) binding to the hnRNPA2B1 protein, as demonstrated by the high affinity of the compounds to the hnRNPA2B1 protein as low as 20 μM as determined by Biacore; and (ii) demonstrating by cell-based assays that the compounds can broadly induce the production of higher levels of IFN-β at concentrations as low as 20 μM.

[0272] In the anti-cell infection test of different types of infection sources, the hnRNPA2B1 agonist compounds disclosed in the present invention can significantly inhibit the replication of infection sources, thereby playing an anti-infection role. In addition, since the hnRNPA2B1 agonist compounds disclosed in the present invention can widely induce the production of higher levels of IFN-β, it can be expected that these compounds can play an anti-infection role through the action of type I interferon.

[0273] (i) Biacore T200 detection of the binding between compounds and hnRNPA2B1

[0274] Using S series CM5 chip; amino coupling reagents (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide EDC and N-hydroxysuccinimide NHS);

[0275] Coupling buffer: 10 mM sodium acetate pH 4.0, pH 4.5;

[0276] Running buffer: 10× PBS-P+;

[0277] Analytical grade DMSO, deionized water (0.22 μm membrane filtration);

[0278] hnRNPA2B1 recombinant protein (concentration must be greater than 0.5 mg / mL, total protein amount must be at least 20 μg);

[0279] Compound: Stock concentration greater than 20 mM, volume greater than 30 μL, purity greater than 90%, dissolved in 100% DMSO;

[0280] Other consumables: 96-well plates and sealing films, 1.5 mL centrifuge tubes without caps, and type 2 rubber bottle caps.

[0281] Experimental steps:

[0282] 1. Place the running buffer 1× PBS and waste liquid bottle in the designated positions and place the chip.

[0283] 2. Protein coupling. Select the immobilization program in the Wizard template, check flow cell 2 or 4, select amine as the method, select specify contact time and flow rate to achieve high coupling, enter contact time 900s, flow rate 10μL / min.

[0284] 3. Prepare the required protein solution according to the instructions. Dilute the protein to the required volume with sodium acetate at pH 4.0, with a final concentration of 20 μg / mL. Place 100 μL EDC, 100 μL NHS, and 140 μL ethanolamine in the corresponding positions.

[0285] 4. The system will automatically coat the target conjugated amount of hnRNPA2B1 protein on the chip surface and automatically generate a conjugation report.

[0286] 5. Prepare the running buffer for the interaction. The running buffer for small molecule samples is 1×PBS-P+ containing 5% DMSO: 105mL 10×PBS-P+ is diluted to 1L with deionized water to make 1.05×PBS-P+.

[0287] Solvent correction stock solution containing 4.5% DMSO: 1.05×PBS-P+9.5mL+0.45mL DMSO

[0288] Solvent correction stock solution containing 5.8% DMSO: 1.05×PBS-P+9.5mL+0.58mL DMSO

[0289] Running buffer containing 5% DMSO: 1.05×PBS-P + 950mL + 50mL DMSO

[0290] Dilute 10mM small molecule stock solution with 1.05×PBS-P+ buffer without DMSO: 0.7μL small molecule stock solution + 6.3μL DMSO + 133μL 1.05×PBS-P+ buffer, total volume is 140μL, final concentration is 50μM. Add to 96-well plate and affix sealing film.

[0291] After the test, use Kinetics or Affinity mode to measure binding affinity.

[0292] (ii) Total cellular RNA can be extracted using TRIzol or RNA rapid extraction kit.

[0293] 24-well plate cells (about 2-3×10 5 10 cells / well) RNA rapid extraction: discard the culture medium, add 500μL RA2 lysis buffer to each well, pipet and lyse several times, transfer to the inner sleeve of the special adsorption tube, and centrifuge at 12000rpm for 1min at room temperature. Discard the filtrate, add 500μL washing buffer, and centrifuge at 12000rpm for 1min at room temperature. Repeat the washing operation once. Discard the filtrate, put the inner sleeve back, do not add Wash Buffer, and centrifuge at 12000rpm for 2min at room temperature. Transfer the inner sleeve to a new 1.5mL centrifuge tube, add 25μL Elution Buffer to the center of the membrane, let it stand at room temperature for 5min, and centrifuge at 12000rpm for 1min at room temperature. The filtrate obtained is RNA, and the RNA concentration is measured using Nanodrop One.

[0294] qRT-PCR: After the concentration of each sample was determined, 1 μg of total RNA was taken and ReverTra qRT-PCR RTMaster Mix reverse transcribes RNA into cDNA. The reverse transcription system and reaction conditions are as follows:

[0295] Reverse transcription reaction system (20 μL system)

[0296]

[0297] Reverse transcription reaction conditions (20 μL system)

[0298]

[0299] Add 60 μL of deionized water to the cDNA sample obtained by reverse transcription for dilution, mix well and centrifuge instantly. The primer sequence information used for qRT-PCR is shown in the table. SYBR Green Realtime PCR Master Mix is ​​used to perform relative quantitative detection of the mRNA level of the gene.

[0300] qRT-PCR reaction system (20 μL system)

[0301]

[0302] qRT-PCR reaction conditions

[0303]

[0304] qRT-PCR primers

[0305]

[0306] All documents mentioned in this disclosure are cited as references in this application, just as each document is cited as reference individually. In addition, it should 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 these equivalent forms also fall within the scope defined by the claims attached to this application.

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 prodrugs thereof, in, L is -(CH2)n-, n is an integer from 0 to 6; X is a halogen; R 1 For-NR 4 R 5 , where R 4 , R 5 Each independently is 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, they form a 5-8 membered heterocycloalkyl group containing 1-3 heteroatoms selected from nitrogen, oxygen or sulfur; 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 hydroxyl, amino, halogen, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, C 1-6 Alkoxy, hydroxyl 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 heterocycloalkylC 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.

2. The compound according to claim 1, its cis-trans isomers, its enantiomers, its diastereomers, its racemates, its solvates, its hydrates, or its pharmaceutically acceptable salts or prodrugs thereof, in, X is F or Cl; L is -(CH2)n-, wherein n is 1, 2 or 3; R 4 , R 5 Each is independently H, methyl or ethyl; R 2 and R 3 Each independently is C 1-3 Alkyl, the 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; 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 2 nitrogen atoms, wherein the 5-8 membered heterocycloalkyl group is surrounded by a 3-8 membered heterocycloalkyl group C 1-6 Alkyl or hydroxyl C 1-6 Alkyl substitution.

3. The compound according to claim 1, its cis-trans isomers, its enantiomers, its diastereomers, its racemates, its solvates, its hydrates, or its pharmaceutically acceptable salts or prodrugs thereof, in, X is F; L is -(CH2)n-, wherein n is 2; R 4 , R 5 Each is independently H, methyl or ethyl; R 2 and R 3 Each independently is C 1-3 Alkyl, the C 1-3 The alkyl group is unsubstituted or substituted with a 4-6 membered heterocycloalkyl group containing 1 nitrogen heteroatom; or 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 hydroxyl C 1-6 Alkyl substitution.

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

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

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

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

8. The product of claim 7, wherein: The product is selected from: a pharmaceutical composition, a medicine, a kit or a medicine box; and / or Wherein, the product is a product for inducing the production of type I interferon in a subject and / or for preventing and / or treating an infectious disease and / or a disease and / or symptom associated with infection in a subject; and / or Among them, the product also contains: one or more other active substances for preventing or treating infectious diseases and related diseases and / or symptoms, for example, the other active substances are one or more selected from the following groups: commonly used clinical antibiotics, including β-lactams (penicillins and cephalosporins), aminoglycosides, tetracyclines, chloramphenicol, macrolides, antifungal antibiotics, anti-tuberculosis antibiotics; commonly used clinical antiviral drugs (tricyclic amines, pyrophosphates, protease inhibitors, nucleoside drugs and interferons, antisense oligonucleotides, etc.); commonly used clinical immunosuppressants (including glucocorticoids, cyclophosphamide, chloroquine, cyclosporine A, traditional Chinese medicine preparations (such as tripterygium wilfordii), anti-TNF monoclonal antibodies).

9. Use of a 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 a product for preventing and / or treating infectious diseases and / or diseases and / or symptoms associated with infection.

10. The use according to claim 9, wherein: The subject is a human or 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 deer, etc.); and / or wherein the production of type I interferon is mediated by hnRNPA2B1; and / or wherein the infectious disease in the subject can benefit from the production of type I interferon in vivo or an increase in its level; and / or Wherein, the infectious disease is caused by chemical, physical, or biological infection, such as caused by viruses, bacteria, parasites, and / or fungi, such as the infection is a DNA virus 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, small DNA virus, adeno-associated virus; and / or Wherein, the diseases and / or symptoms associated with infection are one or more selected from the following groups: 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 infection (for example, autoimmune diseases such as inflammatory bowel disease, rheumatoid arthritis, systemic lupus erythematosus, chronic nephritis, tuberculosis, chronic gastrointestinal diseases); and / or Wherein, the product is selected from: a pharmaceutical composition, a medicine, a test kit or a medicine box; and / or Wherein, the product is a product for inducing the production of type I interferon in a subject and / or for preventing and / or treating an infectious disease and / or a disease and / or symptom associated with infection in a subject; and / or Among them, the product also contains: one or more other active substances for preventing or treating infectious diseases and related diseases and / or symptoms, for example, the other active substances are one or more selected from the following groups: commonly used clinical antibiotics, including β-lactams (penicillins and cephalosporins), aminoglycosides, tetracyclines, chloramphenicol, macrolides, antifungal antibiotics, anti-tuberculosis antibiotics; commonly used clinical antiviral drugs (tricyclic amines, pyrophosphates, protease inhibitors, nucleoside drugs and interferons, antisense oligonucleotides, etc.); commonly used clinical immunosuppressants (including glucocorticoids, cyclophosphamide, chloroquine, cyclosporine A, traditional Chinese medicine preparations (such as tripterygium wilfordii), anti-TNF monoclonal antibodies).

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