Perinaphthenone compounds and their use

Abstract

The present invention discloses a perinaphthenone compound and its use, which can bind to E3 ubiquitin ligase 3 motif 25 (TRIM25), promote pathogen recognition by TRIM25, and induce proteasome-dependent ubiquitination degradation of pathogen proteins, and is a promising ligand for TRIM25, and can realize a wider range of applications, such as the production of PROTAC molecules, so that the perinaphthenone compound has great potential for research and development value and future applications.

Description

[Technical Field]

[0001] The present invention relates to the pharmaceutical technology, and more specifically to perinaphthenone compounds and their use, for example, in the manufacture of drugs for preventing and / or treating diseases, as ligands for E3 ubiquitin ligase 3 motif protein 25 (tripartite motif 25, TRIM25), in the regulation of targeted ubiquitination levels, and in the manufacture of chimeras (PROTACs) that target proteolysis. [Background technology]

[0002] In recent years, cell therapy, immunotherapy, and gene editing technologies have been rapidly developing. Proteolysis-targeting chimeras (PROTACs) represent an emerging direction in drug research and development. They can directly induce the degradation of substrate proteins via the ubiquitin-proteasome protein degradation pathway in vivo, and are attracting widespread attention from researchers.

[0003] The mechanism of action of PROTAC involves linking a small molecule inhibitor with the ligand of an E3 ubiquitin ligase via a linker, forming a complex that targets and induces protein degradation. In vivo, the inhibitor portion of this bifunctional molecule recognizes the target protein, while the E3 ligand portion recognizes the E3 ubiquitin ligase. This spatially brings the target protein and the E3 ubiquitin ligase closer together, transferring ubiquitin from the E3 ubiquitin-conjugating enzyme to the target protein. After ubiquitinating the target protein, it is then degraded via the ubiquitin-proteasome pathway.

[0004] Finding small molecule ligands for E3 ubiquitin ligases is a significant advance in expanding the range of applications for PROTACs in drug discovery. In 2008, Craig et al. reported a PROTAC (SCHNEEKLOTH AR, PUCHEAULT M, TAE HS, et al. Targeted intracellular protein degradation induced by a small molecule: En route to chemical proteomics. Bioorganic & medicinal chemistry letters, 2008, 18(22): 5904-5908) in which nutlin, a ligand for E3 ubiquitin ligase (MDM2), was linked to an androgen receptor inhibitor. Crews et al. designed and modified ligands for E3 ubiquitin ligase (VHL) and discovered VHL ligands with high affinity (BUCKLEY DL, GUSTAFSON JL, VAN MOLLE I, et al. Small-molecule inhibitors of the interaction between the E3 ligase VHL and HIF1alpha. Angewandte Chemie, 2012, 51(46): 11463-11467).In 2015, the Bradner and Crews research teams reported methods for designing the dual-function molecules dBET1 and ARV-825 using E3 ubiquitin ligase (CRBN) ligands (domide-type drugs) and the BRD4 inhibitor JQ1, respectively (WINTER GE, BUCKLEY DL, PAULK J, et al. DRUG DEVELOPMENT. Phthalimide conjugation as a strategy for in vivo target protein degradation. Science, 2015, 348(6241): 1376-1381; LU J, QIAN Y, ALTIERI M, et al. Hijacking the E3 ubiquitin ligase cereblon to efficiently target brD4. Chemistry & biology, 2015, 22(6): 755-763).

[0005] While PROTAC technology is developing rapidly, it still faces many challenges in its application. For example, there are relatively few ligands for E3 ubiquitin ligases.

[0006] E3 ubiquitin ligase 3 motif 25 (TRIM25: tripartite motif 25) is a member of the 3-motif protein family in E3 ubiquitin ligases. Currently, there are few reports on TRIM25 ligands, and there are no reports of PROTACs utilizing TRIM25 ligands. [Overview of the project] [Problems that the invention aims to solve]

[0007] As a result of research, the present inventors found perinaphthenone compounds that bind to E3 ubiquitin ligase tripartite motif containing 25 (TRIM25) in vitro, promote the recognition of PA protein by TRIM25, can induce proteasome-dependent ubiquitination degradation of PA protein, are promising as ligands for E3 ubiquitin ligase TRIM25, can achieve broader applications such as the preparation of PROTAC molecules, and therefore are extremely expected to have research and development value and application prospects. [Means for Solving the Problem]

[0008] In a first aspect of the present invention, there is provided a compound having the following structure. TIFF0007915812000001.tif43170(wherein the covalent bond at position 1 or position 2 TIFF0007915812000002.tif3170 represents a single bond or a double bond, and 1 and 2 are not double bonds at the same time, R1 to R 18 are each independently selected from H, alkyl, alkenyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocyclyl, heterocyclylalkyl, halogen, -CN, -NO2, -COR A , -C(O)OR A , -OCOR A , -C(O)NR A R B , -CH=NR A , -OR A , -OC(O)R A , -S(O) t -R A , -S(O) t -NR A R B , -NR A R B , -NR A C(O)R B and optionally, any H on each group may be substituted with one or more groups selected from halogen, -CN, -CF3, -NO2, -CHO, -COOH, -C(O)NH2, -OH, -OC(O)H, -SH, -S(O)2H, -NH2, R 19 and R 20 The group is independently selected from H, alkyl, alkenyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocyclyl, and heterocyclylalkyl, where H on each group is substituted or unsubstituted heterocyclyl, halogen, -CN, -NO2, -COR A , -C(O)OR A -C(O)NR A R B -CH=NR A , -OR A ,-OC(O)R A , -S(O) t -R A , -S(O) t -NR A R B , -NR A R B , -NR A C(O)R B It may be substituted with one or more groups selected from R 19 and R 20 These simultaneously form substituted or unsubstituted cycloalkyl or heterocyclines with the carbon atoms to which they are linked, or R 17 and R 19 These simultaneously form substituted or unsubstituted cycloalkyl or heterocyclines together with the carbon atoms to which they are linked. t is selected from 0, 1, and 2. Each R A and R B Each of these is independently selected from H, alkyl, cycloalkyl, alkenyl, aryl, heterocyclyl, and halogen. Specifically, R1~R 18These are independently H, C1-10 alkyl, C1-10 haloalkyl (e.g., fluoroalkyl, e.g., trifluoromethyl), C1-10 alkenyl, C3-6 cycloalkyl, C4-10 cycloalkylalkyl, phenyl, 4-6 member heterocycloalkyl, halogen, -CN, -NO2, -CHO, -CO(C1-10 alkyl), -COOH, -C(O)O(C1-10 alkyl), -C(O)NH2, -C( Selected from O)N(C1~10alkyl)(C1~10alkyl), -OH, -O(C1~10alkyl), -OC(O)H, -OC(O)(C1~10alkyl), -SH, -S(C1~10alkyl), -S(O)2H, -S(O)2(C1~10alkyl), -NH2, -N(C1~10alkyl)(C1~10alkyl), -NHC(O)H, -N(C1~10alkyl)C(O)(C1~10alkyl).

[0009] More specifically, R1 is selected from -OH, -O(C1~10alkyl), -SH, -S(C1~10alkyl), -S(O)2H, and -S(O)2(C1~10alkyl), and in some embodiments of the present invention, R1 is -OH.

[0010] More specifically, R2 is selected from H, halogen, -CN, -CF3, -NO2, -CHO, -COOH, -C(O)NH2, and -NH2, and in some embodiments of the present invention, R2 is H.

[0011] More specifically, R3 is selected from C1-10 alkyl, C1-10 haloalkyl (e.g., fluoroalkyl, e.g., trifluoromethyl), C1-10 alkenyl, C3-6 cycloalkyl, C4-10 cycloalkylalkyl, and especially C1-10 alkyl, e.g., C1-6 alkyl, C1-3 alkyl (e.g., methyl, ethyl, n-propyl, isopropyl), and in some embodiments of the present invention, R3 is methyl.

[0012] More specifically, R4 is selected from -OH, -O(C1~10 alkyl), -SH, -S(C1~10 alkyl), -S(O)2H, and -S(O)2(C1~10 alkyl), and in some embodiments of the present invention, R4 is -OH.

[0013] More specifically, R5 is selected from H, C1-10 alkyl, C1-10 haloalkyl (e.g., fluoroalkyl, e.g., trifluoromethyl), C1-10 alkenyl, C3-6 cycloalkyl, C4-10 cycloalkylalkyl, and especially C1-10 alkyl, e.g., C1-6 alkyl, C1-3 alkyl (e.g., methyl, ethyl, n-propyl, isopropyl), and in some embodiments of the present invention, R5 is methyl, and in some other embodiments of the present invention, R5 is H.

[0014] More specifically, R6 is selected from -OH, -O(C1~10 alkyl), -SH, -S(C1~10 alkyl), -S(O)2H, and -S(O)2(C1~10 alkyl), and in some embodiments of the present invention, R6 is -OH.

[0015] More specifically, R7 is selected from -OH, -O(C1~10 alkyl), -SH, -S(C1~10 alkyl), -S(O)2H, and -S(O)2(C1~10 alkyl), and in some embodiments of the present invention, R7 is -OH.

[0016] More specifically, R8, R9, R 11 , R 12 , R 13 , R 15 , R 16 These are independently selected from H, halogen, -CN, -CF3, -NO2, -CHO, -COOH, -C(O)NH2, -NH2, and in some embodiments of the present invention, R8, R9, R 11 , R 12 , R 13 , R 15 , R 16 All of them are H.

[0017] More specifically, R 10and R 14 These are independently selected from C1-10 alkyl, C1-10 haloalkyl (e.g., fluoroalkyl, e.g., trifluoromethyl), C1-10 alkenyl, C3-6 cycloalkyl, C4-10 cycloalkylalkyl, and especially C1-6 alkyl, e.g., C1-3 alkyl (e.g., methyl, ethyl, n-propyl, isopropyl), and in some examples of the present invention, R 10 and R 14 They are all methyl.

[0018] More specifically, R 17 R is selected from H, -OH, -O(C1~10alkyl), -SH, -S(C1~10alkyl), -S(O)2H, -S(O)2(C1~10alkyl), and in some embodiments of the present invention, R 17 H is H, and in some other embodiments of the present invention, R 17 It is -OH.

[0019] More specifically, R 18These are derived from H, C1~10alkyl, C1~10haloalkyl (e.g., fluoroalkyl, e.g., trifluoromethyl), halogen, -CN, -CF3, -NO2, -CHO, -CO(C1~10alkyl), -COOH, -C(O)O(C1~10alkyl), -C(O)NH2, -C(O)N(C1~10alkyl)(C1~10alkyl), -OH, -O(C1~10alkyl), -OC(O)H, -OC(O)(C1~10alkyl), -SH, -S(C1~10alkyl), -S(O)2H, -S(O)2(C1~10alkyl), -NH2, -N(C1~10alkyl)(C1~10alkyl), -NHC(O)H, -N(C1~10alkyl)C(O)(C1~10alkyl) Selected and optionally, where one or more H on each group may be substituted with a group selected from halogens, -CN, -CF3, -NO2, -CHO, -CO(C1~10alkyl), -COOH, -C(O)O(C1~10alkyl), -C(O)NH2, -C(O)N(C1~10alkyl)(C1~10alkyl), -OH, -O(C1~10alkyl), -OC(O)H, -OC(O)(C1~10alkyl), -SH, -S(C1~10alkyl), -S(O)2H, -S(O)2(C1~10alkyl), -NH2, -N(C1~10alkyl)(C1~10alkyl), -NHC(O)H, -N(C1~10alkyl)C(O)(C1~10alkyl), more specifically, R 18 R is selected from H, C1-6 alkyl, C1-6 haloalkyl (e.g., fluoroalkyl, e.g., trifluoromethyl), C1-6 hydroxysubstituted alkyl (one or more hydroxysubstituted C1-6 alkyl, e.g., hydroxymethyl), -CHO, -CO(C1-10 alkyl), -COOH, -C(O)O(C1-10 alkyl), and in some embodiments of the present invention, R 18 The compound is selected from H, methyl, ethyl, n-propyl, isopropyl, -CH2OH, -COOH, -COOCH3, and -CHO.

[0020] In one embodiment of the present invention, R 19 It has the following structure: TIFF0007915812000003.tif21170(here, TIFF0007915812000004.tif3170 represents a single bond or a double bond. R 21 ~R 24 These are independently H, C1-10 alkyl, C1-10 haloalkyl (e.g., fluoroalkyl, e.g., trifluoromethyl), C1-10 alkenyl, C3-6 cycloalkyl, C4-10 cycloalkylalkyl, phenyl, 4-6 member heterocycloalkyl, halogen, -CN, -NO2, -CHO, -CO(C1-10 alkyl), -COOH, -C(O)O(C1-10 alkyl), -C(O)NH2, -C(O) Selected from N(C1~10alkyl)(C1~10alkyl), -OH, -O(C1~10alkyl), -OC(O)H, -OC(O)(C1~10alkyl), -SH, -S(C1~10alkyl), -S(O)2H, -S(O)2(C1~10alkyl), -NH2, -N(C1~10alkyl)(C1~10alkyl), -NHC(O)H, -N(C1~10alkyl)C(O)(C1~10alkyl), or R 21 ~R 24 Two of these, together with the carbon atoms in the middle, form substituted or unsubstituted cycloalkyl or heterocyclyl groups. If TIFF0007915812000005.tif3170 represents a double bond, then R 24 It does not exist. R 25 and R 26 These are independently selected from C1-10 alkyl, C1-10 haloalkyl (e.g., fluoroalkyl, e.g., trifluoromethyl), C1-10 alkenyl, C3-6 cycloalkyl, C4-10 cycloalkylalkyl, phenyl, and 4-6 member heterocycloalkyl. Specifically, R 25 and R 26 Independently, C1-10 alkyl groups are selected from, for example, C1-6 alkyl groups, C1-3 alkyl groups, and in some embodiments of the present invention, R 25 and R 26 They are all methyl.

[0021] In some embodiments of the present invention, R21 H is H.

[0022] In some embodiments of the present invention, R 19 teeth, The filename is TIFF0007915812000006.tif23170.

[0023] Specifically, R 24 is selected from -OH, -O (C1~10 alkyl), or R 23 and R 24 These, together with the carbon atoms between them, form substituted or unsubstituted cycloalkyl or heterocyclines.

[0024] Specifically, R 22 The group is selected from H, -OC(O)H, and -OC(O)(C1-10 alkyl), for example, -OC(O)CH3.

[0025] Specifically, R 23 The group is selected from H, -OH, -O (C1-10 alkyl), -SH, and -S (C1-10 alkyl), for example, H or -OH.

[0026] In some other embodiments of the present invention, R 20 and R 22 These, along with the carbon atoms between them, form a heterocycline.

[0027] In some other embodiments of the present invention, R 20 and R 23 These, along with the carbon atoms between them, form a heterocycline.

[0028] Specifically, the above R 19 In the definition, heterocyclyl is a heterocycloalkyl, for example, a 4- to 6-membered heterocycloalkyl, especially an oxygen-containing 5- to 6-membered heterocycloalkyl, for example TIFF0007915812000007.tif19170, where R Cis one or more independent substituents on the ring, selected from C1-10 alkyl, C1-10 haloalkyl, C1-10 hydroxy-substituted alkyl, C1-10 alkenyl, halogen, -CN, -NO2, -CHO, -CO(C1-10 alkyl), -COOH, -C(O)O(C1-10 alkyl), -C(O)NH2, -C(O)N(C1-10 alkyl)(C1-10 alkyl), -OH, -O(C1-10 alkyl), -OC(O)H, -OC(O)(C1-10 alkyl), -SH, -S(C1-10 alkyl), -S(O)2H, -S(O)2(C1-10 alkyl), -NH2, -N(C1-10 alkyl)(C1-10 alkyl), -NHC(O)H, and -N(C1-10 alkyl)C(O)(C1-10 alkyl).

[0029] Specifically, R C is one or more independent substituents on the ring, selected from C1-6 alkyl, C1-6 haloalkyl, C1-6 hydroxy-substituted alkyl, C1-6 alkenyl, halogen, -CN, -NO2, -CHO, -COOH, -C(O)NH2, -OH, -OC(O)H, and -SH. In some embodiments of the present invention, R C is one or more independent substituents on the ring, selected from methyl, ethyl, n-propyl and isopropyl.

[0030] In some embodiments of the present invention, R 19 is selected from TIFF0007915812000008.tif21170.

[0031] In some embodiments of the present invention, R 20 is H.

[0032] In some embodiments of the present invention, R 19 and R 20 , together with the carbon atom to which they are simultaneously bonded, form the group TIFF0007915812000009.tif21170.

[0033] In some embodiments of the present invention, the valence bond at position 1 is a double bond, and the valence bond at position 2 is a single bond.

[0034] In some other embodiments of the present invention, the valence bond at position 1 is a single bond, and the valence bond at position 2 is a double bond.

[0035] In some other embodiments of the present invention, the valence bond at position 1 is a single bond, and the valence bond at position 2 is a single bond.

[0036] Specifically, the compound may have the following structure. TIFF0007915812000010.tif38170(wherein R1, R4, R5, R6, R7, R 17 , R 18 , R 19 , R 20 have the corresponding definitions described above in the present invention.)

[0037] More specifically, the compound may have the following structure. TIFF0007915812000011.tif38170(wherein R1, R4, R5, R6, R 17 , R 18 , R 19 , R 20 have the corresponding definitions described above in the present invention.)

[0038] Even more specifically, the compound may have the following structure. TIFF0007915812000012.tif31170(wherein R5, R 17 , R 18 , R 19 , R 20 have the corresponding definitions described above in the present invention.)

[0039] More specifically, the compound may be selected from the following structures. TIFF0007915812000013.tif81170

[0040] In the embodiments of the present invention, the compound has the following structure. TIFF0007915812000014.tif241170TIFF0007915812000015.tif25170

[0041] Preferably, it does not contain the following compounds. TIFF0007915812000016.tif128170

[0042] A second aspect of the present invention provides pharmaceutically acceptable salts, esters, stereoisomers, prodrugs, or solvates of the compound described in the first aspect.

[0043] Specifically, the stereoisomers of the compound may have the following structures. TIFF0007915812000017.tif36170

[0044] More specifically, the stereoisomers of the compound may have the following structures. TIFF0007915812000018.tif37170

[0045] In some embodiments of the present invention, the stereoisomers of the compound have the following structures. TIFF0007915812000019.tif236170TIFF0007915812000020.tif24170

[0046] Specifically, the compounds described in the first aspect of the present invention, and the pharmaceutically acceptable salts, esters, stereoisomers, prodrugs, and solvates described in the second aspect, can be produced by any suitable method known to those skilled in the art, such as chemical synthesis, semi-synthesis, microbial fermentation, or plant and animal extraction. For example, they can be produced by extracting and separating a fermented product of a microorganism (e.g., Aspergillus iizukae CPCC 401321, deposit number CGMCC No. 22467), or by chemically modifying (and physically processing) the compound obtained by extraction and separation (semi-synthesis), or by a series of chemical synthesis and physical processing steps from chemical raw materials with relatively simple chemical structures (total synthesis).

[0047] In some embodiments of the present invention, the method for producing the compound described in the first aspect of the present invention may include a step of extracting and separating a fermented product of a microorganism (for example, Aspergillus iizukae CPCC 401321, deposit number CGMCC No. 22467), and further, the method for producing the compound may include a step of chemically modifying the compound obtained by extraction and separation.

[0048] In some other embodiments of the present invention, the method for producing the compound described in the first aspect of the present invention may include the step of producing the compound from a chemical raw material with a relatively simple chemical structure through a series of chemical synthesis and physical processing steps (total synthesis).

[0049] A third aspect of the present invention provides a pharmaceutical composition comprising a compound described in the first aspect of the present invention or a pharmaceutically acceptable salt, ester, stereoisomer, prodrug, or solvate described in the second aspect, and one or more pharmaceutically acceptable auxiliary materials.

[0050] Specifically, the pharmaceutical composition may be in any suitable dosage form, such as tablets, sugar-coated tablets, film-coated tablets, enteric-coated tablets, capsules, hard capsules, soft capsules, oral solutions, oral preparations, granules, suppositories, pills, powders, ointments, elixirs, suspensions, powders, solutions, injections, suppositories, ointments, hard ointments, creams, sprays, drops, and patches, but is not limited to these. Oral dosage forms such as capsules, tablets, oral solutions, granules, pills, powders, elixirs, and ointments are preferred.

[0051] Specifically, pharmaceutically acceptable auxiliary materials may include, but are not limited to, adhesives, fillers, diluents, tableting agents, lubricants, disintegrants, colorants, flavoring agents, and wetting agents. Suitable fillers may include, for example, cellulose, mannitol, lactose, and other similar fillers; suitable disintegrants may include, for example, starch, polyvinylpyrrolidone, and starch derivatives such as sodium starch glycolate; suitable lubricants may include, for example, magnesium stearate; and suitable wetting agents may include, for example, sodium lauryl sulfate.

[0052] Specifically, the pharmaceutical composition may further contain one or more other components selected from inosine monophosphate dehydrogenase (IMPDH) inhibitors, interferon inducers, M2 ion channel protein inhibitors, and neuraminidase inhibitors.

[0053] Specifically, the inosine monophosphate dehydrogenase inhibitor may be, for example, ribavirin.

[0054] Specifically, the interferon inducer may be, for example, arbidol hydrochloride.

[0055] Specifically, the M2 ion channel protein inhibitor may be, for example, amantadine hydrochloride or rimantadine hydrochloride.

[0056] Specifically, the neuraminidase inhibitor may be, for example, osehamivir phosphate, oseltamivir, zanamivir, or peramivir.

[0057] A fourth aspect of the present invention provides the use of the compound described in the first aspect of the present invention or the pharmaceutically acceptable salt, ester, stereoisomer, prodrug, solvate described in the second aspect, or the pharmaceutical composition described in the third aspect, in the manufacture of a pharmaceutical composition for preventing and / or treating a disease.

[0058] In one embodiment of the present invention, the above-mentioned disease is a disease caused by a pathogen infection.

[0059] Specifically, the pathogens mentioned above include viruses, such as adenoviridae (e.g., adenoviruses), herpesviridae (e.g., HSV1 (oral herpes), HSV2 (genital herpes), VZV (varicella), EBV (Epstein-Barr virus), CMV (cytomegalovirus)), poxviridae (e.g., smallpox virus, vaccinia, etc.), papilloviridae (e.g., papillomavirus), parvoviridae (e.g., B19 virus), hepadnaviridae (e.g., hepatitis B virus), and polio. Maviridae (e.g., polyomavirus), Reoviridae (e.g., reovirus, rotavirus), Picornaviridae (e.g., enterovirus, foot-and-mouth disease virus), Caliciviridae (e.g., Norwalk virus, hepatitis E virus), Togaviridae (e.g., rubella virus), Arenaviridae (e.g., lymphocytic choriomeningitis virus), Retroviridae (HIV-1, HIV-2, HTLV-1), Flaviviridae (e.g., dengue virus, Zika virus, Japanese encephalitis virus, Chikungunya virus, yellow fever virus, hepatitis C virus, West Nile virus), Orthomyxoviridae (e.g., influenza viruses (e.g., influenza A virus, influenza B virus, influenza C virus, etc.)), Paramyxoviridae (e.g., human parainfluenza virus type 1 (HPV), type 2, type 3, type 4, Sendai virus, mumps virus, measles virus, RSV, Newcastle disease virus), Bunyaviridae (e.g., This may include, but is not limited to, California encephalitis virus, hantavirus, rhabdoviridae (e.g., rabies virus), filoviridae (e.g., Ebola virus, Marburg virus), coronavirusidae (e.g., HCoV-229E, HCoV-OC43, HCoV-NL63, HCoV-HKU1, SARS-CoV, MERS-CoV, SARS-CoV-2), stellateviridae (e.g., astrovirus), and bornaviridae (e.g., borna virus).

[0060] In some embodiments of the present invention, the virus is one or more influenza viruses, such as influenza A virus, influenza B virus, and influenza C virus, and is particularly influenza A virus.

[0061] Specifically, the influenza A virus may be an influenza A virus of the H1N1 subtype, H2N2 subtype, H3N2 subtype, H5NI subtype, H7N9 subtype, or H9N2 subtype.

[0062] Specifically, the diseases caused by the above-mentioned viral infections include, but are not limited to, influenza, SARS, COVID-19, viral hepatitis (e.g., hepatitis A, B, C, D, etc.), AIDS, rabies, dengue fever, and Ebola virus disease.

[0063] In another embodiment of the present invention, the disease is a tumor.

[0064] Specifically, the tumors listed above are malignant tumors, including, but not limited to, lung cancer (e.g., non-small cell lung cancer), prostate cancer, cervical cancer, skin cancer, pancreatic cancer, colorectal cancer, kidney cancer, liver cancer, brain cancer, lymphoma, leukemia, lung cancer (small cell lung cancer, non-small cell lung cancer), melanoma, gastric cancer, gastroesophageal adenocarcinoma, esophageal cancer, small intestine cancer, cardia cancer, bladder cancer, anal cancer, gallbladder cancer, bile duct cancer, teratomas, and cardiac tumors.

[0065] Specifically, the subjects of the drug may be mammals (e.g., humans, monkeys, primates, pigs, cattle, or sheep) or birds (domesticated birds such as chickens, ducks, or geese, or wild birds).

[0066] A fifth aspect of the present invention provides the use of the following compounds and their pharmaceutically acceptable salts, esters, stereoisomers, prodrugs, or solvates in the production of chimeras (PROTACs) that target the regulation of the target ubiquitination level and proteolysis, as ligands for the E3 ubiquitin ligase TRIM25. TIFF0007915812000021.tif44170(here, Valence bonds in one or two locations TIFF0007915812000022.tif3170 represents a single bond or a double bond, and 1 and 2 are not simultaneously double bonds. R1~R 18 These are independently H, alkyl, alkenyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocyclyl, heterocyclylalkyl, halogen, -CN, -NO2, -COR A , -C(O)OR A , -OCOR A -C(O)NR A R B -CH=NR A , -OR A ,-OC(O)R A , -S(O) t -R A , -S(O) t -NR A R B , -NR A R B , -NR A C(O)R B Selected from, optionally, where H on each group may be replaced with one or more groups selected from halogen, -CN, -CF3, -NO2, -CHO, -COOH, -C(O)NH2, -OH, -OC(O)H, -SH, -S(O)2H, -NH2.

[0067] R 19 and R 20The group is independently selected from H, alkyl, alkenyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocyclyl, and heterocyclylalkyl, where H on each group is substituted or unsubstituted heterocyclyl, halogen, -CN, -NO2, -COR A , -C(O)OR A -C(O)NR A R B -CH=NR A , -OR A ,-OC(O)R A , -S(O) t -R A , -S(O) t -NR A R B , -NR A R B , -NR A C(O)R B It may be substituted with one or more groups selected from R 19 and R 20 These simultaneously form substituted or unsubstituted cycloalkyl or heterocyclines with the carbon atoms to which they are linked, or R 17 and R 19 These simultaneously form substituted or unsubstituted cycloalkyl or heterocyclines together with the carbon atoms to which they are linked. t is selected from 0, 1, and 2. Each R A and R B Each of these is independently selected from H, alkyl, cycloalkyl, alkenyl, aryl, heterocyclyl, and halogen.

[0068] Specifically, each group has the corresponding definition described in the first aspect of the present invention.

[0069] Specifically, the target is a target protein to be degraded, and may be a protein of the body itself or an exogenous protein such as a viral protein.

[0070] In one embodiment of the present invention, the regulation of the ubiquitination level includes promoting PA protein ubiquitination, promoting the binding of PA protein to E3 ubiquitin ligase TRIM25, and using the PA protein as a ligand for E3 ubiquitin ligase TRIM25. In some embodiments of the present invention, the compound has the following structure in the above use. TIFF0007915812000023.tif227170TIFF0007915812000024.tif117170

[0071] In some embodiments of the present invention, the stereoisomers of the compound have the following structures in the above-described use. TIFF0007915812000025.tif235170TIFF0007915812000026.tif48170

[0072] Specifically, in the use described above, PROTAC has the following structure. TIFF0007915812000027.tif6170 (wherein SMI is the small molecule inhibitor moiety (which may be formed with any suitable target small molecule inhibitor known in the prior art), E3L is the ligand moiety of the E3 ubiquitin ligase (for example, the structural moiety formed with the above compound or a pharmaceutically acceptable salt, stereoisomer, ester, prodrug, or solvate thereof), and L is the binding or linking group between SMI and E3L.)

[0073] A sixth aspect of the present invention provides a PROTAC having the following structure. TIFF0007915812000028.tif10170 (Here, SMI is the small molecule inhibitor portion, E3L is the ligand portion of the E3 ubiquitin ligase and is formed from the compound described in the sixth aspect of the present invention or a pharmaceutically acceptable salt, stereoisomer, ester, prodrug, or solvate thereof. L is a bond or linking group between SMI and E3L.

[0074] Specifically, the SMI may be formed with any suitable target small molecule inhibitor known in the prior art.

[0075] A seventh aspect of the present invention provides a method for producing a PROTAC, comprising the step of using a compound described in the sixth aspect of the present invention or a pharmaceutically acceptable salt, stereoisomer, ester, prodrug, or solvate thereof.

[0076] An eighth aspect of the present invention provides an Aspergillus species deposited on July 8, 2021, with the Center for Ordinary Microorganisms of the Chinese Microbial Species Depositary Administration (address: No. 3, Building 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences), with deposit number CGMCC No. 22467 and classification name Aspergillus iizukae. A ninth aspect of the present invention provides a method for preventing and / or treating a disease, comprising the step of administering an effective amount of the compound described in the first aspect of the present invention or its pharmaceutically acceptable salt, stereoisomer, ester, prodrug, solvate, pharmaceutical composition described in the third aspect of the present invention, or PROTAC described in the sixth aspect of the present invention, to a subject in need.

[0077] Specifically, the diseases mentioned above may include diseases caused by pathogen infection, such as influenza, SARS, COVID-19, viral hepatitis (e.g., hepatitis A, B, C, D, etc.), AIDS, rabies, dengue fever, Ebola virus disease, etc., as well as tumors, such as breast cancer, prostate cancer, ovarian cancer, cervical cancer, skin cancer, pancreatic cancer, colorectal cancer, kidney cancer, liver cancer, brain cancer, lymphoma, leukemia, lung cancer (small cell lung cancer, non-small cell lung cancer), melanoma, gastric cancer, gastroesophageal adenocarcinoma, esophageal cancer, small intestine cancer, cardia cancer, bladder cancer, anal cancer, gallbladder cancer, bile duct cancer, teratomas, and cardiac tumors.

[0078] Specifically, the subjects mentioned above may be mammals (e.g., humans, monkeys, primates, pigs, cows, or sheep) or birds (domesticated birds such as chickens, ducks, or geese, or wild birds).

[0079] A tenth aspect of the present invention provides a method for adjusting a target ubiquitination level, comprising the step of administering an effective amount of the compound described in the first aspect of the present invention or a pharmaceutically acceptable salt, stereoisomer, ester, prodrug, or solvate thereof, the pharmaceutical composition described in the third aspect of the present invention, or the PROTAC described in the sixth aspect of the present invention, to a subject requiring such treatment.

[0080] The present invention provides a perinaphthenone compound that can bind to TRIM25, promote TRIM25-mediated recognition of pathogen proteins (e.g., viruses), induce proteasome-dependent ubiquitination and degradation of pathogens, is promising as a ligand for the E3 ubiquitin ligase TRIM25, and can be applied to a wider range of applications, such as the preparation of PROTAC molecules, thus offering significant research and development value and promising future application potential.

[0081] The storage information for the biomaterial of this invention is as follows:

[0082] Aspergillus iizukae CPCC 401321 (Deposited on July 8, 2021, at the Center for Ordinary Microorganisms, CGMCC (Address: No. 3, Building 1, Beichen West Road, Chaoyang District, Beijing, China; Deposit Number: CGMCC No. 22467; Taxonomic Name: Aspergillus iizukae). [Brief explanation of the drawing]

[0083] [Figure 1] The 1H-NMR spectrum of compound C1 is shown. [Figure 2] The 13C-NMR spectrum of compound C1 is shown. [Figure 3] The 1H-NMR spectrum of compound C2 is shown. [Figure 4] 13C-NMR spectrum of compound C2 [Figure 5] The 1H-NMR spectrum of compound C3 is shown. [Figure 6] The 13C-NMR spectrum of compound C3 is shown. [Figure 7] The HSQC spectrum of compound C3 is shown. [Figure 8] The HMBC spectrum of compound C3 is shown. [Figure 9] The 1H-1H COSY spectrum of compound C3 is shown. [Figure 10] The NOESY spectrum of compound C3 is shown. [Figure 11] The 1H-NMR spectrum of compound C4 is shown. [Figure 12] The 13C-NMR spectrum of compound C4 is shown. [Figure 13] The HSQC spectrum of compound C4 is shown. [Figure 14] The HMBC spectrum of compound C4 is shown. [Figure 15] The 1H-1H COSY spectrum of compound C4 is shown. [Figure 16] The NOESY spectrum of compound C4 is shown. [Figure 17] The 1H-NMR spectrum of compound C5 is shown. [Figure 18] The 13C-NMR spectrum of compound C5 is shown. [Figure 19] The HSQC spectrum of compound C5 is shown. [Figure 20] The HMBC spectrum of compound C5 is shown. [Figure 21] The 1H-1H COSY spectrum of compound C5 is shown. [Figure 22] The NOESY spectrum of compound C5 is shown. [Figure 23] The 1H-NMR spectrum of compound C6 is shown. [Figure 24] The 13C-NMR spectrum of compound C6 is shown. [Figure 25] The HSQC spectrum of compound C6 is shown. [Figure 26] The HMBC spectrum of compound C6 is shown. [Figure 27] The 1H-1H COSY spectrum of compound C6 is shown. [Figure 28]The NOESY spectrum of compound C6 is shown. [Figure 29] The 1H-NMR spectrum of compound C7 is shown. [Figure 30] The 13C-NMR spectrum of compound C7 is shown. [Figure 31] The HSQC spectrum of compound C7 is shown. [Figure 32] The HMBC spectrum of compound C7 is shown. [Figure 33] The 1H-1H COSY spectrum of compound C7 is shown. [Figure 34] The NOESY spectrum of compound C7 is shown. [Figure 35] The 1H-NMR spectrum of compound C8 is shown. [Figure 36] The 13C-NMR spectrum of compound C8 is shown. [Figure 37] The HSQC spectrum of compound C8 is shown. [Figure 38] The HMBC spectrum of compound C8 is shown. [Figure 39] The 1H-1H COSY spectrum of compound C8 is shown. [Figure 40] The NOESY spectrum of compound C8 is shown. [Figure 41] The 1H-NMR spectrum of compound C9 is shown. [Figure 42] The 13C-NMR spectrum of compound C9 is shown. [Figure 43] The HSQC spectrum of compound C9 is shown. [Figure 44] The HMBC spectrum of compound C9 is shown. [Figure 45] The 1H-1H COSY spectrum of compound C9 is shown. [Figure 46] The NOESY spectrum of compound C9 is shown. [Figure 47] The 1H-NMR spectrum of compound C10 is shown. [Figure 48] The 13C-NMR spectrum of compound C10 is shown. [Figure 49]The HSQC spectrum of compound C10 is shown. [Figure 50] The HMBC spectrum of compound C10 is shown. [Figure 51] The 1H-1H COSY spectrum of compound C10 is shown. [Figure 52] The 1H-NMR spectrum of compound C11 is shown. [Figure 53] The 13C-NMR spectrum of compound C11 is shown. [Figure 54] The HSQC spectrum of compound C11 is shown. [Figure 55] The HMBC spectrum of compound C11 is shown. [Figure 56] The 1H-1H COSY spectrum of compound C11 is shown. [Figure 57] The 1H-NMR spectrum of compound C12 is shown. [Figure 58] The 13C-NMR spectrum of compound C12 is shown. [Figure 59] The HSQC spectrum of compound C12 is shown. [Figure 60] The HMBC spectrum of compound C12 is shown. [Figure 61] The 1H-1H COSY spectrum of compound C12 is shown. [Figure 62] The NOESY spectrum of compound C12 is shown. [Figure 63] The 1H-NMR spectrum of compound C13 is shown. [Figure 64] The 13C-NMR spectrum of compound C13 is shown. [Figure 65] The HSQC spectrum of compound C13 is shown. [Figure 66] The HMBC spectrum of compound C13 is shown. [Figure 67] The 1H-1H COSY spectrum of compound C13 is shown. [Figure 68] The NOESY spectrum of compound C13 is shown. [Figure 69] The 1H-NMR spectrum of compound C14 is shown. [Figure 70] The 13C-NMR spectrum of compound C14 is shown. [Figure 71] The HSQC spectrum of compound C14 is shown. [Figure 72] The HMBC spectrum of compound C14 is shown. [Figure 73] The 1H-1H COSY spectrum of compound C14 is shown. [Figure 74] The NOESY spectrum of compound C14 is shown. [Figure 75] The experimental results regarding the effects of compounds C1-C14 on the expression of influenza virus PA protein are shown. [Figure 76] The experimental results of the PA protein degradation downregulation pathway by compound C1 are shown. [Figure 77] The experimental results of PA polyubiquitination induction by compound C1 are shown. [Figure 78] The results of an experiment in which compound C1 recognizes the E3 ligase TRIM25 and induces PA degradation are shown. [Figure 79] The results of an experiment in which compound C1 promotes the interaction between TRIM25 and PA are shown. [Figure 80] The results of experiments in which the compound binds to TRIM25 in vitro are shown. [Figure 81] This shows experimental results regarding the ability of the compound to bind to PA in vitro. [Figure 82] The results of an experiment in which compound C1 promotes the polyubiquitination level of PA protein in vitro are shown. [Modes for carrying out the invention]

[0084] Unless otherwise defined, all scientific and technical terms used in this invention have the same meanings as those generally understood by those skilled in the art.

[0085] The term "alkyl" refers to a linear or branched hydrocarbon group that does not contain unsaturated bonds, and this hydrocarbon group is linked to the rest of the molecule by single bonds. The alkyl groups used here generally contain 1 to 10 carbon atoms (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10) (i.e., C1-10 alkyl groups), preferably 1 to 6 carbon atoms (i.e., C1-6 alkyl groups). Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, isoamyl, neopentyl, tert-pentyl, n-hexyl, and isohexyl. When an alkyl group is substituted with a cycloalkyl group, it becomes a "cycloalkylalkyl" such as cyclopropylmethyl, cyclopropylethyl, cyclobutylmethyl, cyclopentylmethyl, and cyclohexylmethyl. When an alkyl group is substituted with an aryl group, it becomes an "aralkyl" such as benzyl, diphenylmethyl, or phenethyl. When an alkyl group is substituted with a heterocyclyl group, it becomes a "heterocyclylalkyl group."

[0086] The term "alkenyl" refers to a linear or branched hydrocarbon group containing at least two carbon atoms and at least one unsaturated bond, the hydrocarbon group being linked to the rest of the molecule by a single bond. As used herein, alkenyls generally contain 1 to 10 carbon atoms (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10) (i.e., C1-10 alkenyls), preferably 1 to 6 carbon atoms (i.e., C1-6 alkenyls). Examples of alkenyls include, but are not limited to, vinyl, 1-methyl-vinyl, 1-propenyl, 2-propenyl, or butenyl.

[0087] The term "cycloalkyl" refers to an alicyclic hydrocarbon, and as used herein, cycloalkyl generally comprises 1 to 4 monocyclic and / or fused rings and 3 to 18 carbon atoms, preferably 3 to 10 carbon atoms (e.g., 3, 4, 5, 6, 7, 8, 9, 10) (e.g., C3-10 cycloalkyl, C3-6 cycloalkyl), such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, or adamantyl.

[0088] The term "aryl" refers to a functional group or substituent derived from a simple aromatic ring, including monocyclic aryl groups and / or fused aryl groups, for example, comprising 1 to 3 rings, monocyclic or fused rings, and having 6 to 18 (e.g., 6, 8, 10, 12, 14, 16, 18) carbocyclic atoms. The aryls used herein are generally aryls (i.e., C6-12 aryls) comprising 1 to 2 rings, monocyclic or fused rings, and having 6 to 12 carbocyclic atoms, where the H atoms on the carbon atoms may be substituted with groups such as alkyl or halogen. Examples of such aryls include, but are not limited to, phenyl, p-methylphenyl, naphthyl, biphenyl, and indenyl.

[0089] The term "halogen" refers to bromine, chlorine, iodine, or fluorine.

[0090] The term "heterocyclyl" refers to a 3- to 18-membered non-aromatic ring group containing 2-17 carbon atoms and 1-10 heteroatoms. Heterocyclyls may be monocyclic, dicyclic, tricyclic, or tetracyclic ring systems, and may include fused rings, spirocyclic rings, or bridging ring systems. Heterocyclyls may be partially saturated (heteroaryl) or fully saturated (heterocycloalkyl). A suitable heteroaryl in the compound of the present invention comprises one, two, or three heteroatoms, the heteroatoms being selected from N, O, or S atoms, and the heteroaryl includes, for example, coumarin containing 8-coumarin, quinolinyl containing 8-quinolinyl, isoquinolinyl, pyridyl, pyrazinyl, pyrazolyl, pyrimidinyl, furyl, pyrrolyl, thienyl, thiazolyl, isothiazolyl, triazolyl, tetrazolyl, isoxazolyl, oxazolyl, imidazolyl, indolyl, isoindolyl, indazolyl, indolidinyl, phthalazinyl, pteridinyl, purinyl, oxadiazolyl, thiadiazolyl, flazanyl, pyridazinyl, triazinyl, sinnolinyl, benzimidazolyl, benzofuranyl, benzflazanyl, benzothienyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinoxalinyl, naphthilidinyl, and flopyridyl.A suitable heterocycloalkyl in the compound of the present invention comprises one, two, or three heteroatoms, wherein the heteroatoms are selected from N, O, or S atoms, and the heterocycloalkyl is, for example, pyrrolidinyl, tetrahydrofuranyl, dihydrofuranyl, tetrahydrothiophenyl, tetrahydrothiopyranyl, piperidinyl, morpholinyl, thiomorpholinyl, oxathianyl, piperazinyl, azetidinyl, oxetanyl, thietanyl, homopiperidinyl, oxyranyl, thiirane group, azepinyl, o This includes xoazepanil, diazepinil, triazepinil, 1,2,3,6-tetrahydropyridyl, 2-pyrrolinil, 3-pyrrolinil, dihydroindolyl, 2H-pyranil, 4H-pyranil, dioxanil, 1,3-dioxolanil, pyrazolinil, dithianil, dithiolanil, dihydropyranil, dihydrothienyl, pyrazolidinil, imidazolinil, imidazolidinil, 3-azabicyclo[3.1.0]hexyl, 3-azabicyclo[4.1.0]heptyl, 3H-indolyl, and quinodinil.

[0091] The pharmaceutically acceptable salts of the present invention include acid addition salts and alkali addition salts.

[0092] The aforementioned acid addition salts include, but are not limited to, salts derived from inorganic acids such as hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid, hydroiodic acid, and phosphonic acid, and salts derived from organic acids such as aliphatic monocarboxylic acids and dicarboxylic acids, phenyl-substituted alkanes, hydroxyalkanoates, alkanedioates, aromatic acids, and aliphatic and aromatic sulfonic acids. Therefore, these salts include, but are not limited to, sulfates, pyrosulfates, bisulfates, sulfites, bisulfites, nitrates, phosphates, monohydrogen phosphates, dihydrogen phosphates, metaphosphates, pyrophosphates, hydrochlorides, hydrobroms, iodates, acetates, propions, octanates, isobutyrates, ethylene glycolates, malons, succinates, octanates, sebacinates, fumarates, maleates, mandelates, benzoates, chlorobenzoates, methylbenzoates, dinitrobenzoates, phthalates, benzenesulfons, toluenesulfons, phenylacetates, citrates, lactates, maleates, tartrates, and mesylates, as well as salts of amino acids such as arginine salts, glucons, and galacturonic acid salts. Acid addition salts can be produced, as in the conventional method, by contacting a free alkali form with a desired acid in sufficient quantity to form a salt. By bringing the salt form into contact with the alkali, the free alkali form can be regenerated, and this free alkali can be separated as in conventional methods.

[0093] The alkali addition salt of the present invention refers to a salt formed with a metal or amine such as an alkali metal or alkaline earth metal hydroxide, or an organic amine. Examples of metals used as cations include, but are not limited to, sodium, potassium, magnesium, and calcium. Examples of suitable amines include, but are not limited to, N,N'-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine (ethane-1,2-diamine), N-methylglucosamine, and procaine. Alkali addition salts can be produced by contacting a free acid form with a desired alkali in a sufficient amount to form a salt, as in the conventional method. By contacting the salt form with an acid, the free acid form can be regenerated, and this free acid can be separated as in the conventional method.

[0094] The stereoisomers of the present invention exist in the form of enantiomers, diastereomers, and geometric isomers. Some compounds of the present invention have a cyclic hydrocarbon group which may be substituted with one or more carbon atoms, in which case all of its geometric forms, including cis and trans, as well as mixtures thereof, are within the scope of the present invention. The cyclic hydrocarbon group includes alicyclic hydrocarbon groups and aryl groups, the alicyclic hydrocarbon group may be a non-aromatic monocyclic, fused, crosslinked, or spirocyclic saturated or unsaturated cyclic hydrocarbon group, and the aryl group may be, for example, phenyl, naphthyl, phenantrenyl, biphenyl, etc.

[0095] The solvates of the present invention refer to physical bonds between the compound of the present invention and one or more solvent molecules. These physical bonds include covalent bonds, including ionic bonds and hydrogen bonds, to varying degrees. In some cases, solvates can be separated, for example, when one or more solvent molecules are incorporated into the lattice of a crystalline solid. Solvates include a solution phase and separable solvates. Typical solvates include ethanolates and methanolates.

[0096] The prodrug of this invention refers to a form of compound of formula I that is suitable for administration to patients, does not possess excessive toxicity, irritation, or allergens, and is effective for its intended purpose, and includes acetal, ester, and amphoteric forms. The prodrug is converted in the body, for example, by hydrolysis in the blood, to obtain the parent compound.

[0097] In the present invention, the terms "patient" or "subject," etc., may be used interchangeably herein and refer to any animal or its cells, whether in vitro or in situ, that are treated according to the methods described herein. Specifically, the animals include mammals, such as rats, mice, guinea pigs, rabbits, dogs, monkeys, or humans, particularly humans.

[0098] In the present invention, "treatment" refers to preventing, curing, reversing, alleviating, reducing, minimizing, suppressing, inhibiting, and / or stopping one or more clinical symptoms of a disease after its onset.

[0099] In this invention, "prevention" refers to avoiding, minimizing, or making difficult the onset or progression of a disease through treatment before the disease develops.

[0100] The various publications, patents, and disclosures of published patent specifications cited herein are incorporated herein by reference in their entirety.

[0101] The technical solutions of the present invention will be described clearly and completely below with reference to embodiments of the present invention, but it is clear that the embodiments described are only a subset of the present invention and not all embodiments. All other embodiments obtained by those skilled in the art without requiring any creative work based on embodiments of the present invention are within the scope of the present invention. Example 1: Preparation of fermented culture of Aspergillus iizukae CPCC 401321

[0102] Aspergillus iizukae CPCC 401321, a strain with high perinaphthenone production (deposit number CGMCC No. 22467), was cultured on a PDA slope at 28°C for 7 days. After culturing, hyphae were selected and inoculated into a 500 ml Erlenmeyer flask containing 100 ml of PDB seed medium. The culture was then incubated with shaking at 28-30°C for 5 days to obtain a seed solution. Next, 10 ml of the seed solution was placed in a 500 ml Erlenmeyer flask containing rice medium (100 g of rice was placed in a 500 ml Erlenmeyer flask, 100 ml of deionized water was added and immersed, and sterilized at 121°C for 20 minutes to obtain rice medium). The culture was incubated at 28°C for 20 days to obtain a solid-state fermented culture of Aspergillus iizukae CPCC 401321. Example 2: Preparation of ethyl acetate extract from fermented culture of Aspergillus iizukae CPCC 401321

[0103] 10 kg of the solid-state fermented culture of Aspergillus iizukae CPCC 401321 obtained in Example 1 was collected, stirred and pulverized with a glass rod, 20 L of ethyl acetate was added, and extraction was performed three times for 30 minutes each using room temperature ultrasound. The ethyl acetate extracts were combined and evaporated using a rotary vaporizer (temperature 40°C) to remove the ethyl acetate from the collected liquid, and the resulting product was the ethyl acetate extract of the Aspergillus iizukae CPCC 401321 fermented culture. Example 3: Separation of ethyl acetate extract

[0104] (1) The ethyl acetate extract (250 g) from Example 2 was dissolved in an ethyl acetate-methanol mixed solution and then separated by silica gel column chromatography. The silica gel used for silica gel column chromatography was silica gel H, and the size of the silica gel column used was 12 × 40 cm, with a column volume of 4522 mL. In the elution step used in silica gel column chromatography, linear gradient elution was performed as follows. The mobile phase used was a mixture of petroleum ether and acetone, and the volume ratio of petroleum ether to acetone in the mobile phase for linear gradient elution decreased linearly from 4:1 to 1:1. From the start of the elution process, 275 fractions of eluate were collected consecutively at a rate of 500 ml per fraction (200 ml / fraction), and each fraction was labeled Fr.1, Fr.2, Fr.3, Fr.4, ..., Fr.275. These fractions were then combined under guidance of TLC detection, and finally, Fr.1-5 (a mixture of fractions F.1 to Fr.5, and so on), Fr.6, Fr.7-11, Fr.12-23 , Fr.24-26, Fr.27-29, Fr.30-44, Fr.45-51, Fr.52-67, Fr.68-77, Fr.78-91, Fr.92-99, Fr.100-11 5, 19 combined components were obtained: Fr.116-127, Fr.128-137, Fr.138-196, Fr.197-227, Fr.228-258, and Fr.259-275. (2) The supernatants Fr.12-23 obtained in step (1) were combined and subjected to Sephadex LH-20 gel column chromatography. The gel column used was 3 × 120 cm in size. In the elution step, methanol was used as the mobile phase, and the eluate was collected in 25 fractions of 30 ml each (30 ml / fraction) from the start of the elution step, and labeled tube.1, tube.2, tube.3, ..., tube.25. Tubes 5-9 (Tubes 5-9 were mixed to obtain components called tubes 5-9) were separated by preparative high-performance liquid chromatography (separation was performed by RP-C18 liquid phase preparative chromatography). The packing used for this separation by preparative high-performance liquid chromatography was octadecylsilane-bonded silica gel packing with a particle size of 5 μm. The column used for this separation by preparative high-performance liquid chromatography had a diameter of 8 mm and a height of 250 mm. The mobile phase was a mixture of methanol and water (containing 0.05% formic acid), with a volume ratio of methanol to water of 90:10 and a flow rate of 4.0 ml / min. R Elution peaks were collected at retention times of 16.0 min, 18.0 min, and 19.9 min. Methanol and water were removed by rotary evaporation using a rotary vaporizer (temperature 50°C), yielding 80.5 mg of compound C3, 39.1 mg of compound C4, and 4.1 mg of compound C5 in that order. (3) MCI column chromatography was performed on Fr.92~99 obtained in step (1). The size of the MCI column used was 2 × 30 cm. In the elution step, a mixture of methanol and water in a volume ratio of 90:10 was used as the mobile phase, and the eluate was collected continuously in 20 fractions of 40 ml (40 ml / fraction) per tube from the start of the elution step, and labeled tube.1, tube.2, tube.3, ..., tube.20. Sephadex LH-20 gel column chromatography was performed on tubes 5~9 (a mixture of the fractions from tube.5~tube.9). The size of the gel column used was 2 × 80 cm. In the elution step, methanol was used as the mobile phase, and the eluate was collected continuously in 25 fractions of 20 ml (20 ml / fraction) per tube from the start of the elution step, and labeled tube.1′, tube.2′, tube.3′, ..., tube.25′. Tubes 2' to 5' (a mixture of Tubes 2' to 5') were separated by preparative high-performance liquid chromatography (RP-C18 liquid-phase separation). The packing used for this separation was octadecylsilane-bonded silica gel packing with a particle size of 5 μm. The column used for this separation had a diameter of 8 mm and a height of 250 mm. The mobile phase was a mixture of methanol and water (containing 0.05% formic acid), with a volume ratio of methanol to water of 85:15. The flow rate was 4.0 mL / min. RElution peaks were collected at retention times of 11.8 min, 12.5 min, and 17.7 min. Methanol and water were removed by rotary evaporation using a rotary vaporizer (temperature 50°C), yielding 21.6 mg of compound C9, 10.6 mg of compound C10, and 102.9 mg of compound C6 in that order. Sephadex LH-20 gel column chromatography was performed on tubes 11-15 (a mixture of fractions from tubes 11-15). The gel column size used was 2 × 80 cm. In the elution step, methanol was used as the mobile phase, and 20 ml of eluate per tube (20 ml / fraction) was collected continuously in 15 fractions from the start of the elution step, and labeled tube 1″, tube 2″, tube 3″, ..., tube 15″. The 2″~8″ tube fraction (a mixture of the 2″~8″ fraction) was separated by preparative high-performance liquid chromatography (RP-C18 liquid-phase separation). The packing used for this separation was octadecylsilane-bonded silica gel packing with a particle size of 5 μm. The column used for this separation had a diameter of 8 mm and a height of 250 mm. The mobile phase was a mixture of methanol and water (containing 0.05% formic acid), with a volume ratio of methanol to water of 87:13. The flow rate was 4.0 mL / min. R Elution peaks were collected at retention times of 17.2 min, 19.4 min, and 21.1 min. Methanol and water were removed by rotary evaporation using a rotary evaporator (temperature 50°C), yielding 40.5 mg of compound C6, 28.2 mg of compound C8, and 425.6 mg of compound C7 in that order. MCI column chromatography was performed on Fr.228-258 obtained in step (1). The size of the MCI column used was 6 × 18 cm. In the elution step, a mixture of methanol and water in a volume ratio of 80:20 was used as the mobile phase, and 39 fractions of the eluate were collected consecutively from 100 ml per tube (100 ml / fraction) from the start of the elution step, and labeled tube.1, tube.2, tube.3, ..., tube.39. ODS column chromatography was performed on tubes 16-39 (a mixture of the fractions from tube.16 to tube.39). The ODS column used was 5.5 × 28 cm in size. In the elution process, a mixture of methanol and water in a volume ratio of 75:25 was used as the mobile phase. From the start of the elution process, 100 ml of the eluate was collected from each tube in 50 fractions (100 ml / fraction), and these were labeled tube.1′, tube.2′, tube.3′, ..., tube.50′. Tubes 34 to 45 (a mixture of the Tube.34′ to Tube.55′ fractions) were rotated and evaporated in a rotary evaporator (temperature 50°C) to remove methanol and water, yielding 1.25 g of compound C1. Tubes 31 to 33 (a mixture of the Tube.31' to Tube.33' fractions) were separated by preparative high-performance liquid chromatography (RP-C18 liquid-phase separation). The packing used for this preparative high-performance liquid chromatography separation was octadecylsilane-bonded silica gel packing with a particle size of 5 μm. The column used for this preparative high-performance liquid chromatography separation had a diameter of 8 mm and a height of 250 mm. The mobile phase was a mixture of methanol and water (containing 0.05% formic acid), with a volume ratio of methanol to water in the mobile phase of 73:27. The flow rate was 4.5 mL / min. R The elution peak was collected at a retention time of 14.0 min, and methanol and water were removed by rotary evaporation using a rotary vaporizer (temperature 50°C) to obtain 14.4 mg of compound C11. MCI column chromatography was performed on Fr.30-44 obtained in step (1). The MCI column used was 3 × 23 cm in size. In the elution step, a mixture of methanol and water in a volume ratio of 90:10 was used as the mobile phase. From the start of the elution step, the eluate was collected in 20 fractions of 50 ml (50 ml / fraction) per tube and labeled tube.1, tube.2, tube.3, ..., tube.20. Sephadex LH-20 gel column chromatography was performed on tubes 7-20 (a mixture of the fractions from tube.7 to tube.20). The gel column used was 2 × 80 cm in size. In the elution step, methanol was used as the mobile phase. From the start of the elution step, the eluate was collected in 18 fractions of 20 ml (20 ml / fraction) per tube and labeled tube.1', tube.2', tube.3', ..., tube.18'. The fractions from tubes 2' to 6' (a mixture of tube 2' to tube 6' fractions) were separated by preparative high-performance liquid chromatography (RP-C18 liquid phase separation). The packing used for this separation was octadecylsilane-bonded silica gel packing with a particle size of 5 μm. The column used for this separation had a diameter of 8 mm and a height of 250 mm. The mobile phase was a mixture of methanol and water (containing 0.05% formic acid), with a volume ratio of methanol to water of 84:16. The flow rate was 4.5 mL / min. R Elution peaks were collected at retention times of 20.0 min, 23.7 min, 30.4 min, and 33.7 min. Methanol and water were removed by rotary evaporation using a rotary evaporator (temperature 50°C), yielding 3.2 mg of compound C14, 33.5 mg of compound C13, 152.3 mg of compound C12, and 206.1 mg of compound C2 in that order. Compounds C1-C14 are all brown, gel-like solids that are readily soluble in solvents such as methanol, ethanol, and DMSO, but sparingly soluble in water. Compounds C1-C14 have an Rf value of 0.4-0.8 when eluted in a chloroform-methanol-water (volume ratio 70:15:2) solvent system for silica gel thin-layer chromatography, exhibiting significant fluorescence at 254 nm and 365 nm, and turning brown-red in vanillin sulfate. Example 4: Identification of the structure

[0105] (1) Compounds C1 and C2 Table 1 shows the spectral data of compounds C1 and C2. 1 1H NMR and 13 The 13C NMR spectra are shown in Figures 1-4. The structures of compounds C1 and C2 are shown in Table 6. Table 1. Nuclear magnetic data of compounds C1 and C2 ( 1 H NMR 600 MHz, 13 ¹³C NMR 150 MHz, DMSO-d6) TIFF0007915812000029.tif249170TIFF0007915812000030.tif118170(2) Compound C3 HRESIMS (negative ion) ion peak of compound C3: m / z 573.2825 [MH] - Its molecular formula is C 35 H 42 It was shown to be O7. 1 1H NMR (Figure 5), 13 Comprehensive analysis of the 13C NMR (Figure 6) and HSQC spectra (Figure 7) revealed that the compound contains two ketone carbonyl groups, one aldehyde carbonyl group, 18 olefinic carbons, and 14 sp² groups. 3It is presumed to contain hybrid carbon. In the hydrogen spectrum, characteristic signals were observed: δ 14.33 (1H, s), 13.11 (1H, s), 9.29 (1H, s), 6.83 (1H, s), 6.50 (1H, t, J=7.3Hz), 5.05 (1H, t, J=7.2Hz), 4.94 (1H, t, J=6.8 Hz), 4.81 (1H, t, J=7.8 Hz), 2.83 (3H, s), 2.56 (2H, d, J=7.9 Hz), 2.14 (3H, s), 1.57 (3H, s), 1.49 (3H, s), 1.44 (3H, s), and 1.28 (3H, s). The characteristic signals described above are similar to those of compound C2. A detailed comparison of the nuclear magnetic signals of compounds C3 and C2 revealed that both have the same perinaphthenone three-membered ring structure, except for the diterpene branched chain. In the HMBC spectra (Figure 8), the following related signals were observed: δ1.28 (H-35) is associated with δ115.4 (C-17), 139.4 (C-18), and 39.2 (C-19); δ1.44 (H-34) is associated with 124.4 (C-21), 133.3 (C-22), and 37.4 (C-23); δ9.29 (H-33) is associated with δ155.2 (C-25), 142.1 (C-26), and 23.5 (C-27); and δ5.05 (H-29) is associated with δ23.5 ( δ6.50(H-25) is related to δ26.9(C-24), 23.5(C-27), and 194.9(C-33), δ4.94(H-21) is related to δ38.2(C-19), 37.4(C-23), and 15.3(C-34), and δ4.81(H-17) is related to δ39.2(C-19) and 15.5(C-35). 1 H- 1In the H-COSY spectrum (Figure 9), the following related signals were observed: δ4.81 (H-17) is related to δ2.56 (H-16), δ4.94 (H-21) is related to δ1.56 (H-20), δ1.56 (H-20) is related to δ1.64 (H-19), δ6.50 (H-25) is related to δ2.36 (H-24), δ2.36 (H-24) is related to δ2.02 (H-23), δ5.05 (H-29) is related to δ1.93 (H-28), and δ1.93 (H-28) is related to δ2.14 (H-27). The above HMBC and 1 H- 1 The planar structure of the diterpene branched chain was confirmed from the H COSY related signals. In the NOESY spectrum (Figure 10), the following related signals were observed: δ4.81 (H-17) was related to δ1.64 (H-19), δ4.94 (H-21) was related to δ2.02 (H-23), and δ6.50 (H-25) was related to δ9.29 (H-33). From this, it was revealed that the configurations of 17(18)-ene, 21(22)-ene, and 25(26)-ene are E-, E-, and E-, respectively. Since compound C3 has the same biosynthetic pathway as compounds C1 and C2, it was determined that the absolute configuration of the C-1 position of compound C3 is also the S configuration. Finally, compound C3 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignments are specifically shown in Table 2. (3) Compound C4 HRESIMS (negative ion) ion peak of compound C4: m / z 573.2838 [MH] - Its molecular formula is C 35 H 42 It was shown to be O7. 1 1H NMR (Figure 11), 13 Comprehensive analysis of the 13C NMR spectrum (Figure 12) and HSQC spectrum (Figure 13) revealed that the compound contains two ketone carbonyl groups, 18 olefinic carbons, and 15 sp groups. 3 It is presumed that hybrid carbon is present. The nuclear magnetic signal of compound C4 is substantially the same as that of compound C2, except for the C-25~C-28 structural fragments. 1 H- 1In the H-COSY spectrum (Figure 15), H-29 is related to H-28, and considering the HSQC-related signals, the signal δ H 4.43 and δ C 75.1 was identified as belonging to the C-28 position, and the signal chemical shift value indicated that C-28 was oxidized. In the HMBC spectrum (Figure 14), the following related signals were observed: H-25 (δ 4.26, 4.06) was associated with C-25 (δ 119.0) and C-26 (δ 138.8), and H-25 (δ 4.26, 4.06) was associated with C-28 (δ 75.1), indicating that C-28 and C-33 were linked by an ether bond. In the NOESY spectrum (Figure 16), δ 5.17 (H-25) was associated with δ 2.50 (H-27), demonstrating that the 25(26)-ene configuration is E-. Since compound C4 has the same biosynthetic pathway as compounds C1 and C2, it was determined that the C-1 configuration of compound C4 is the S configuration. Ultimately, compound C4 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignment is specifically shown in Table 2. (4) Compound C5 HRESIMS (negative ion) ion peak of compound C5: m / z 617.3372 [MH] - Its molecular formula is C 37 H 46 It was shown to be O8. 1 1H NMR (Figure 17), 13 Comprehensive analysis of the 13C NMR (Figure 18) and HSQC spectra (Figure 19) revealed that the compound contains two ketone carbonyl groups, one ester carbonyl group, 18 olefinic carbons, and 16 sp groups. 3It is presumed that hybrid carbon is present. The nuclear magnetic signal of compound C5 is similar to that of most of compound C2, but the C-25~C-30 structural fragments are different. In the HMBC spectrum (Figure 20), δ1.54 (H-33) is associated with 106.0 (C-5), 130.1 (C-26), and 44.6 (C-27); δ1.62 (H-31) is associated with 123.7 (C-29), 135.9 (C-30), and 25.2 (C-31); and δ1.65 (H-32) is associated with δ123.7 (C-29), 135.9 (C-30), and 18.0 (C-32). 1 H- 1 In the H-COSY spectrum (Figure 21), δ5.50 (H-28) was found to be associated with δ2.2, 2.06 (H-27), and 5.07 (H-29), respectively, and the C-25~C-30 skeletal structure was confirmed based on the above evidence. In the HMBC spectrum, δ5.50 (H-28) was found to be associated with δ169.3 (-COCH3), and δ1.91 (-COCH3) was found to be associated with δ169.3 (-COCH3), demonstrating that one acetyl was linked at the C-28 position. In the NOESY spectrum (Figure 22), the following associated signals were observed. δ4.82(H-17) is related to δ1.64(H-19), δ4.87(H-21) is related to δ1.83(H-23), and δ5.05(H-25) is related to δ2.20(H-27). From this, it was confirmed that the configurations of 17(18)-ene, 21(22)-ene, and 25(26)-ene are E-, E-, and E-, respectively. Since compound C5 has the same biosynthetic pathway as compounds C1 and C2, it was found that the C-1 configuration of compound C5 is also an S configuration. Ultimately, compound C5 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignments are specifically shown in Table 2. Table 2. Nuclear magnetic data of compounds C3-C5 ( 1 H NMR 600 MHz, 13 ¹³C NMR 150 MHz, DMSO-d6) TIFF0007915812000031.tif248170TIFF0007915812000032.tif166170(5) Compound C6 HRESIMS (negative ion) ion peak of compound C6: m / z 593.3115 [MH] - Its molecular formula is C 35 H 46 It was shown to be O8. 1 1H NMR (Figure 23), 13 Comprehensive analysis of the 13C NMR (Figure 24) and HSQC spectra (Figure 25) revealed that the compound contains two ketone carbonyl groups, sixteen olefinic carbons, and seventeen sp groups. 3 It is presumed that hybrid carbon is present. The nuclear magnetic signal of compound C6 was substantially identical to that of compound C1, except for the C-26 position. In the HMBC spectrum (Figure 26), the following related signals were observed: δ5.04 (H-25) was associated with δ36.5 (C-27) and 15.9 (C-33), and δ1.53 (H-33) was associated with δ123.2 (C-25), 135.0 (C-26), and 36.5 (C-27), indicating that the C-26 position has one methyl substitution. In the NOESY spectrum (Figure 28), δ5.04 (H-25) was associated with δ1.85 (H-27), demonstrating that the 25(26)-ene configuration is E-. Since compound C6 shares the same biosynthetic pathway as compounds C1 and C2, it was determined that the C-1 and C-29 positions of compound C6 are S and R configurations, respectively. Ultimately, compound C6 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignment is specifically shown in Table 3. (6) Compound C7 HRESIMS (negative ion) ion peak of compound C7: m / z 649.3383 [MH] - Its molecular formula is C 38 H 50 It was shown to be O9. 1 1H NMR (Figure 29), 13 Comprehensive analysis of the 13C NMR (Figure 30) and HSQC spectra (Figure 31) revealed that the compound contains two ketone carbonyl groups, 16 olefinic carbons, and 20 sp groups. 3It is presumed to contain hybrid carbon. The nuclear magnetic signal of compound C7 was substantially the same as that of compound C1, except for the C-28~C-32 structural fragment on the diterpene branch chain. Compound C7 has three more carbon atoms than compound C1 and contains one oxidized quaternary carbon δ 105.6 (C-1′) and two methyl carbon signals δ 28.4 (C-2′) [δ 1.29 (3H, s, H-2′)] and δ 26.7 (C-3′) [δ 1.21 (3H, s, H-3′)]. In the HMBC spectrum (Figure 46), δ1.29 (H-2′) was associated with δ105.6 (C-1′) and 26.7 (C-3′), and δ1.21 (H-3′) was associated with δ105.6 (C-1′) and 28.4 (C-2′), indicating that C-1′ was substituted with two methyl groups. Furthermore, the large chemical shift at C-1′ indicated substitution with two oxygen atoms. HMBC-related signals: δ1.15 (H-31) was associated with δ79.4 (C-30) and 82.1 (C-29), and δ0.98 (H-32) was associated with 79.4 (C-30) and 82.1 (C-29). 1 H- 1 Considering the H-COSY related signals, δ3.61 (H-29) is associated with δ2.04 (H-28), and δ2.04 (H-28) is associated with δ2.19 (H-27a) and 2.00 (H-27b), further confirming the structure of the C-28~C-32 fragment. From the large chemical shifts of C-29 and C-30 and the degree of unsaturation of compound C7, it was determined that C-1′ is linked to C-29 and C-30 respectively via an ether bond, ultimately constituting a five-membered ring structural fragment. Since it has the same biosynthetic pathway as compounds C1 and C2, it was determined that the absolute configuration of the C-1 position of compound C7 is the S configuration. Finally, compound C7 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignment is specifically shown in Table 3. (7) Compound C8 HRESIMS (negative ion) ion peak of compound C8: m / z 663.3114 [MH] - Its molecular formula is C 38 H 48 O 10 This demonstrated that... 1 1H NMR (Figure 35), 13Comprehensive analysis of the 13C NMR (Figure 36) and HSQC spectra (Figure 37) revealed that the compound contains two ketone carbonyl groups, one ester carbonyl group, sixteen olefinic carbons, and nineteen sp groups. 3 It is presumed that the compound contains hybrid carbon. The nuclear magnetic signal of compound C8 is substantially the same as that of compound C7, except for the C-26 position on the diterpene branch chain. In the HMBC spectrum (Figure 38), δ6.60 (H-25) is associated with δ23.6 (C-27), 37.8 (C-23), and 168.2 (C-33), indicating that the C-26 position is substituted with one carboxyl (-COOH) group. In the NOESY spectrum (Figure 40), δ2.34 (H-27) is associated with δ2.20 (H-24), demonstrating that the 25(26)-ene configuration is E-. Since it has the same biosynthetic pathway as compounds C1 and C2, it was determined that the absolute configuration of the C-1 position of compound C8 is also the S configuration. Finally, compound C8 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignments are specifically shown in Table 3. Table 3. Nuclear magnetic data of compounds C6-C8 ( 1 H NMR 600 MHz, 13 ¹³C NMR 150 MHz, DMSO-d6) TIFF0007915812000033.tif248170TIFF0007915812000034.tif248170TIFF0007915812000035.tif88170(8) Compound C9 HRESIMS (negative ion) ion peak of compound C9: m / z 609.3343 [MH] - Its molecular formula is C 35 H 46 It was shown to be O9. 1 1H NMR (Figure 41), 13 Comprehensive analysis of the 13C NMR (Figure 42) and HSQC spectra (Figure 43) revealed that the compound contains two ketone carbonyl groups, 16 olefinic carbons, and 17 sp groups. 3It is presumed to contain hybrid carbon. The nuclear magnetic signal of compound C9 is almost identical to that of compound C1, except for the C-24~C-29 structural fragments. In the HMBC spectrum (Figure 44), δ1.00 (H-31) is associated with δ85.9 (C-29) and 69.8 (C-30), δ0.99 (H-33) is associated with δ74.5 (C-25), 84.9 (C-26), and 34.3 (C-27), and δ3.13 (H-25) is associated with δ35.9 (C-23), 84.9 (C-26), and 34.3 (C-27). 1 H- 1 In the H-COSY spectrum (Figure 45), δ3.57 (H-29) was found to be related to δ1.71 (H-28), δ1.71 (H-28) to δ1.91 and 1.45 (H-27), δ3.13 (H-25) to δ1.56 and 1.19 (H-24), and δ1.56 and 1.19 (H-24) to δ2.07 and 1.85 (H-24). Based on this evidence, the structure of C-24 to C-29 in compound C9 was confirmed. In the NOESY spectrum (Figure 46), δ0.99 (H-33) was found to be related to δ3.57 (H-29), confirming the relative configuration of the C-26 and C-29 substituents. Ultimately, compound C9 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignments are specifically shown in Table 4. (9) Compound C10 HRESIMS (negative ion) ion peak of compound C10: m / z 609.3343 [MH] - Its molecular formula is C 35 H 46 It was shown to be O9. Compound C10 substantially matches the nuclear magnetic signal of C9, and it is presumed that both have the same planar structure. Compound C10 1 1H NMR (Figure 47), 13 ¹³C NMR (Figure 48) and HSQC spectrum (Figure 49), 1 H- 1A comprehensive analysis of the H COSY (Figure 50) and HMBC spectra (Figure 51) proved the above conclusion. A detailed comparison of the nuclear magnetic signals of compounds C10 and C9 revealed slight structural differences between C-26 and C-29, allowing for the estimation of the relative configurations of the C-26 and C-29 substituents in compound C10 (which differ from those in compound C9). Ultimately, compound C10 was identified with the structure shown in Table 6, and its nuclear magnetic signal assignments are specifically shown in Table 4. (10) Compound C11 HRESIMS (negative ion) ion peak of compound C11: m / z 595.2946 [MH] - Its molecular formula is C 34 H 44 It was shown to be O9. 1 1H NMR (Figure 52), 13 Comprehensive analysis of 13C NMR (Figure 53) and HSQC spectra (Figure 54) revealed that the compound contains two ketone carbonyl groups, sixteen olefinic carbons, and sixteen sp groups. 3It is presumed that hybrid carbon is present. Compound C11 is very similar to the nuclear magnetic signal of C1, but a detailed comparison revealed that both diterpene branched chain structures are identical, with only slight differences in the perinaphthenone three-membered ring structure. In the hydrogen spectrum of compound C11, there is one less methyl hydrogen signal at the C-14 position (at approximately δ2.13) and one more aromatic hydrogen signal (δ6.31) present as a single peak compared to the hydrogen spectrum of compound C1, indicating that there is no methyl substitution at the C-12 position in the structure of compound C11. In the HMBC spectrum (Figure 55), δ13.78 (13-OH) is associated with δ101.7 (C-3), 99.1 (C-12), and 167.0 (C-13), and δ6.32 (H-12) is associated with δ101.7 (C-3) and 112.6 (C-10), which further validates the above conclusion. Since it has the same biosynthetic pathway as compounds C1 and C2, it was estimated that the C-1 and C-29 positions in the structure of compound C11 are S and R configurations, respectively, and that the 17(18)-ene, 21(22)-ene, and 25(26)-ene configurations are E-, E-, and Z-, respectively. Finally, compound C11 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignments are specifically shown in Table 4. Table 4. Nuclear magnetic data of compounds C9-C11 ( 1 H NMR 600 MHz, 13 ¹³C NMR 150 MHz, DMSO-d6) TIFF0007915812000036.tif249170TIFF0007915812000037.tif127170(11) Compound C12 HRESIMS (negative ion) ion peak of compound C12: m / z 589.2827 [MH] - Its molecular formula is C 35 H 42 It was shown to be O8. 1 1H NMR (Figure 57), 13 Comprehensive analysis of 13C NMR (Figure 58) and HSQC spectra (Figure 59) revealed that the compound contains two ketone carbonyl groups, one ester carbonyl group, 18 olefinic carbons, and 14 sp groups. 3It is presumed that the compound contains hybrid carbon. Compound C12 is very similar to the nuclear magnetic signal of C2, but a detailed comparison revealed that the only difference between the two is the substituent at the C-26 position. In the HMBC spectrum (Figure 60), δ5.70 (H-25) is associated with δ34.3 (C-27), 38.4 (C-23), and 168.7 (C-33), confirming that the C-26 position of compound C12 is substituted with one carboxyl (-COOH) group. In the NOESY spectrum (Figure 62), δ5.70 (H-25) is associated with δ2.11 (H-27), demonstrating that the 25(26)-ene configuration is E-. In the structure of compound C12, the configurations at other positions are the same as those of compound C2. Ultimately, compound C12 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignments are specifically shown in Table 5. (12) Compound C13 HRESIMS (negative ion) ion peak of compound C13: m / z 591.2969 [MH] - Its molecular formula is C 35 H 42 It was shown to be O7. 1 1H NMR (Figure 63), 13 Comprehensive analysis of the 13C NMR (Figure 64) and HSQC spectra (Figure 65) revealed that the compound contains two ketone carbonyl groups, sixteen olefinic carbons, and seventeen sp groups. 3It is presumed that hybrid carbon is present. Compound C13 has a nuclear magnetic signal similar to that of C1, but a detailed comparison revealed that the C-25-C-29 structural fragments differ between the two. In the HMBC spectrum (Figure 66), H-33 (δ 4.26, 4.06) is associated with C-25 (δ 119.0) and C-26 (δ 138.8), H-33 (δ 4.49, 3.63) is associated with C-28 (δ 84.1), and H-33 (δ 3.06) is associated with C-28 (δ 66.1). From this, it was demonstrated that C-29 and C-30 are linked by an ether bond to form a six-membered ring structure. In the NOESY spectrum (Figure 68), δ5.12 (H-25) is associated with δ2.50 (H-27), and δ4.49 (H-33) is associated with δ1.99 (H-24), demonstrating that the 25(26)-ene configuration is Z-. The configurations at other positions in the structure are the same as those of compound C1. Ultimately, compound C13 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignments are specifically shown in Table 5. (13) Compound C14 HRESIMS (negative ion) ion peak of compound C14: m / z 591.3248 [MH] - Its molecular formula is C 35 H 44 It was shown to be O8. 1 1H NMR (Figure 69), 13 Comprehensive analysis of the 13C NMR (Figure 70) and HSQC spectra (Figure 71) revealed that the compound contains two ketone carbonyl groups, 16 olefinic carbons, and 17 sp groups. 3 It is presumed to contain hybrid carbon. The nuclear magnetic signal of compound C14 is almost identical to that of compound C1, except for the C-24-C-29 structural fragments. In the HMBC spectrum (Figure 72), δ1.03 (H-31) is associated with δ84.6 (C-29) and 70.0 (C-30), and δ1.20 (H-33) is associated with δ137.2 (C-25), 81.9 (C-26), and 37.0 (C-27). 1 H- 1In the H-COSY spectrum (Figure 73), δ3.60 (H-29) was found to be associated with δ1.78 and 1.70 (H-28), δ1.78 and 1.70 (H-28) were found to be associated with δ1.76 and 1.75 (H-27), and δ5.39 (H-24) was found to be associated with δ5.41 (H-25) and δ2.53 (H-23), respectively. Based on this evidence, the structure of C-24 to C-29 in compound C14 was confirmed. In the NOESY spectrum (Figure 74), δ5.41 (H-25) was found to be associated with δ2.53 (H-23), which demonstrated that the configuration of 24(25)-ene is E-. Finally, compound C14 was identified as having the structure shown in Table 6, and its nuclear magnetic signal assignments are specifically shown in Table 5. Table 5. Nuclear magnetic data of compounds C12-C14 ( 1 H NMR 600 MHz, 13 ¹³C NMR 150 MHz, DMSO-d6) TIFF0007915812000038.tif248170TIFF0007915812000039.tif156170Table 6. Structures of Compounds C1-C14 TIFF0007915812000040.tif246170TIFF0007915812000041.tif94170 Example 5: Effects of compounds C1-C14 on PA protein

[0106] 2.5 × 10 5HEK 293T cell suspension at a concentration of cells / mL was inoculated into 6-well plates at 2 ml per well. When the cells had grown to 80%, the HEK 293T cell population was transfected with 500 ng pHW2000-PA plasmid per well. Four hours after transfection, the medium was replaced with DMEM medium containing 10% fetal bovine serum (FBS). One group was given 2 μL of 5.00 mM of the target compound per well, while the other group was given DMSO (dimethyl sulfoxide) as a negative control. These groups were then cultured for 24 hours. The medium was discarded, 80 μL of RIPA lysis solution was added to each well, the lysis solution was transferred to 1.5 ml EP tubes and dissolved on ice for 20 minutes, and 20 μL of 5X protein loading buffer was added to each tube. The tubes were boiled in a 100°C metal bath for 30 minutes. PA protein expression levels were detected using Western Blot. The detection results are shown in Figure 75. Furthermore, HEK293T cells were transfected with the pHW2000-PA-Luc plasmid, treated with different concentrations of the compound, and the expression level of the Luc protein was detected after 24 hours. The EC (Emission Control) of the compound was used to determine whether the compound degraded the PA protein. 50 The values ​​were calculated (Table 7). Table 7. Compounds that degrade influenza virus PA protein EC 50 result TIFF0007915812000042.tif56170 Example 6: Effective inhibition of PA protein degradation by compound C1 by protease inhibitor MG132

[0107] HEK239T cells were transfected with the pHW183-PA plasmid and treated with different concentrations of compound C1 (2 μM and 10 μM). Simultaneously, the lysosomal inhibitor ConA (Concanavalin A) or the proteasome inhibitor MG-132 was added, and PA protein expression was observed. As a result, treatment with the proteasome inhibitor MG-132 almost completely restored the PA protein expression level (Figure 76A), while treatment with the lysosomal inhibitor ConA had little effect on PA protein recovery (Figure 76B), indicating that the degradation of PA protein by compound C1 is mainly carried out via the proteasome pathway. Example 7: Enhancement of PA protein ubiquitination level by compound C1

[0108] HEK293T cells were transfected with PA and myc-CW7 ubiquitin plasmid. 3.5 hours after transfection, the cells were treated with different concentrations of compound C1, and incubated with MG-132 8 hours before cell reception. Samples were received 24 hours after cell transfection, captured using PA protein antibodies, and the expression level of ubiquitinated PA protein was detected by Western blotting. The results showed that compound C1 promoted the polyubiquitination level of PA protein (Figure 77). Example 8: Discovery of E3 ubiquitin ligase TRIM25 using surface plasmon resonance (SPR) technology

[0109] SPR (Small Molecule-Protein Resorption) is a classic method for detecting the binding of small molecules to proteins. Its advantages include the absence of molecular labeling of the sample (i.e., preserving the properties of the small molecules) and high sensitivity. The basic principle involves immobilizing small molecules on a chip surface, continuously flowing a cell degradation solution over the chip surface, and recording the changes in molecular concentration on the chip surface during the binding and dissociation processes of the small molecules to the protein using LC-MS (Liquid Chromatography-Mass Spectrometry), thereby monitoring the interaction between small molecules and proteins in real time. Using SPR, proteins involved in the polyubiquitination of PA proteins were detected. The results of this experiment yielded seven host proteins involved in protein polyubiquitination, including KEAP1, HERC5, RBP2, UBA7, TRIM25, ISG15, and UB2E2. Only TRIM25 significantly affected the anti-influenza activity of compound C1 after knockdown (Figure 78A). By overexpressing TRIM25 in TRIM25 knockout cell lines, we were able to restore the anti-influenza activity of compound C1 to normal cell levels (Figure 78B). Example 9: Enhancement of the interaction between TRIM25 and PA by compound C1

[0110] HEK293T cells were transfected with PA and TRIM25 plasmids. 3.5 hours after transfection, the cells were treated with different concentrations of compound C1, and incubated with MG-132 for 8 hours before cell reception. Samples were received 24 hours after cell transfection, captured using a TRIM25 protein antibody, and the expression level of ubiquitinated PA protein was detected by Western blotting. The results showed that compound C1 promotes the interaction between TRIM25 and PA proteins (Figure 79). Example 10: Direct binding of compounds C1-C14 to TRIM25 and PA proteins

[0111] To further investigate the mechanism by which compound C1 degrades PA protein, the inventors used biofilm layer surface interference (BLI) to detect the compounds' binding ability to TRIM25 protein and PA protein. The results showed that compounds C1-C14 all bound to TRIM25 protein, with KD values ​​ranging from 12 to 43 μM (Figure 80). This suggests that the compounds' ability to bind to TRIM25 is directly related to their function in inducing PA degradation. Simultaneously, the compounds also possessed the ability to bind to PA in vitro; for example, compounds C1 and C2 could bind to PA protein, with KD values ​​of 11 μM and 58 μM, respectively (Figure 81). These results suggest that such compounds can recruit TRIM25 to PA by binding to it, thereby inducing its ubiquitination and degradation. Example 11: In vitro enhancement of PA protein ubiquitination level by compound C1

[0112] In vitro ubiquitination tests showed that compound C1 can promote the polyubiquitination level of PA protein in vitro (Figure 82). Example 12: Anti-influenza virus activity

[0113] (1)Cell culture Human embryonic kidney epithelial cells 293T and the 293T-derived cell line 293T-Gluc were cultured in DMEM medium containing 10% fetal bovine serum (FBS). (2) Preparation of recombinant influenza A virus 1.8 × 10 in a 10 cm cell culture dish 6 293T cells and 0.6 × 10¹ 6MDCK cells were inoculated in a 3:1 ratio. After 24 hours of incubation, eight plasmids of influenza A virus (IAV) A / WSN / 33 (H1N1) (pHW181-PB2, pHW182-PB1, pHW183-PA, pHW184-HA, pHW185-NP, pHW186-NA, pHW187-M, pHW188-NS) were transfected using the transfection reagent Lipofectamine 2000 at a transfection dose of 1.2 μg, with 40 μl used per dish according to the instructions. After 6 hours of transfection, the medium was replaced with fresh DMEM medium. After 24 hours of transfection, TPCK-trypsin at a final concentration of 1 μg / mL was added. After 48 hours, the supernatant was collected, centrifuged at 1000 rpm for 5 minutes to remove cell debris, filtered through a 0.45 μM filtration membrane, and divided into smaller portions to obtain recombinant A / WSN / 33(H1N1) influenza virus, which was stored in a refrigerator at -80°C. Among these, the reverse genetic system for eight influenza A virus (IAV) plasmids was donated to Dr. Robert G. Webster, and they are pHW181-PB2, pHW182-PB1, pHW183-PA, pHW184-HA, pHW185-NP, pHW186-NA, pHW187-M, and pHW188-NS (Hoffmann, E., G. Neumann, et al. A DNA transfection system for generation of influenza A virus from eight plasmids[J]. Proc Natl AcadSci USA, 2000, 97:6108-6113). (3) EC by 293T Gluc cells 50 measurement 293T-Gluc cells (Gao Q, Wang Z, Liu Z, et al. A cell-based high-throughput approach to identify inhibitors of influenza A virus[J]. Acta Pharmaceutica Sinica B, 2014, 4(4): 301-306) were plated into 96-well plates, with 2.5 × 10⁶ cells per well. 4 Cells were inoculated and cultured in 100 μl of DMEM culture medium containing 10% FBS. 24 hours after plating the cells, 1 μl of the target compound (dissolved in DMSO (dimethyl sulfoxide) and diluted with DMSO) was added per well. 1 hour after adding the target compound, viral infection was performed according to an MOI of 0.25. After 24 hours, 10 μl of the supernatant was collected and the Gluc protein content was detected. 50 The concentration needed to inhibit the virus by 50% was calculated. The experiment was repeated three times. Detection of Gasussia cyferase activity 250 μg of lyophilized Coelenterazine-h substrate powder was dissolved in 600 μL of anhydrous ethanol to prepare a 1.022 mM substrate mother liquor, which was stored at -20°C. Before measurement, the mother liquor was diluted with PBS in a 1:60 ratio to prepare the substrate working solution. The working solution was allowed to stand at room temperature for 30 minutes to stabilize. Since the substrate becomes unstable when exposed to light, it is necessary to avoid light throughout the entire process. 10 μL of cell culture supernatant (cell supernatant after 24 hours of culture following transfection in the Western Blot experiment described above) was collected in a white opaque 96-well plate and measured using a Centro XS microplate reader. 3 Using an LB 960 autosampler, a substrate working solution incubated under light protection was added to each well at a rate of 60 μL per well, and the signal was collected continuously for 0.5 seconds. The measurement results are expressed in Relative Light Units (RLU). Three sets of experiments were set up in parallel. Experimental data The data was represented as TIFF0007915812000043.tif4170. Plotting and statistical analysis were performed using GraphPad Prism 5.0. (4) Cell activity measurement The CCK-8 (Cell Counting Kit-8) kit is a rapid and highly sensitive test kit based on WST-8 (water-soluble tetrazolium salt, chemical name: 2-(2-methyloxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfobenzene)-2H-tetrazolium monosodium salt), widely applied to cell proliferation and cytotoxicity. WST-8 is a compound similar to MTT and can be reduced by mitochondrial dehydrogenases in the presence of an electron-bonding reagent to produce orange formazan. The more cells there are, the faster the proliferation and the darker the color; the greater the cytotoxicity, the lighter the color. In the same cell, there is a linear relationship between the intensity of the color and the number of cells. By measuring its light absorption value at a wavelength of 450 nm using an enzyme-linked immunosorbent assay, the number of viable cells can be indirectly reflected. 293T-Gluc cells per well: 2.5 × 10⁶ 4 Cells were inoculated into a 96-well plate and cultured in 100 μl of DMEM culture medium containing 10% FBS. 24 hours after plating the cells, 1 μl of the target compound (dissolved in DMSO and diluted with DMSO) was added to each well. A blank control (100 μl of DMEM medium only), a positive control (1 μl of ribavirin added), and a negative control (1 μl of DMSO added) were simultaneously set up, and incubated at 37°C for 48 hours. The 96-well plate was removed, 10 μl of CCK-8 was added to each well, and incubation continued at 37°C for 1-2 hours. The light absorption value at a wavelength of 450 nm was then measured for each well using an Enspire 2300 multifunction microplate reader to determine the half-cytotoxic concentration (CC). 50 The concentration of the drug that kills 50% of cells was calculated. The experiment was repeated three times. Table 8. Results of anti-IAV activity of compounds TIFF0007915812000044.tif101170

[0114] These results indicate that although compounds C1 to C14 differ in activity degree, all of them have good anti-influenza A virus activity, and the EC of their anti-influenza virus 50 values ranged from 0.45 to 2.22 μM. In addition, for these compounds, the CC against 293T-Gluc cells 50 are all greater than 100 μM. Therefore, the perinaphthenone compounds are characterized by strong antiviral activity and low cytotoxicity.

[0115] The above are only preferred embodiments of the present invention, and are not intended to limit the present invention. Any modifications and equivalent substitutions made within the scope of the spirit and principles of the present invention shall be included within the protection scope of the present invention.

[0116] The foregoing embodiments and methods described in the present invention may vary based on the ability, experience and preference of those skilled in the art.

[0117] In the present invention, the steps of the method are enumerated in a certain order, but this does not impose any limitation on the order of the steps of the method.

Claims

1. A compound selected from the following structures, or a pharmaceutically acceptable salt, stereoisomer, or solvate thereof.

2. A compound selected from the following structures, or a pharmaceutically acceptable salt or solvate thereof.

3. A pharmaceutical composition comprising a compound according to any one of claims 1 to 2, or a pharmaceutically acceptable salt, stereoisomer, or solvate thereof, and one or more pharmaceutically acceptable auxiliary materials.

4. The use of a compound, or a pharmaceutically acceptable salt, stereoisomer, or solvate thereof, in the manufacture of a pharmaceutical composition for the prevention and / or treatment of a disease, wherein the disease is a disease caused by a viral infection, and the compound is selected from the following structures.

5. The use according to claim 4, wherein the stereoisomer is selected from the following structures.

6. The use according to claim 4, wherein the disease caused by the aforementioned viral infection is selected from influenza, SARS, COVID-19, viral hepatitis, AIDS, rabies, dengue fever, and Ebola virus disease.

7. The use according to claim 4, wherein the disease is influenza.

8. The use according to claim 4, wherein the disease is AIDS.

9. The use of a compound, or a pharmaceutically acceptable salt, stereoisomer, or solvate thereof, as a ligand for the E3 ubiquitin ligase TRIM25, or in the production of a chimeric PROTAC targeting proteolysis, wherein the compound is selected from the following structures.

10. The use according to claim 9, wherein the stereoisomer is selected from the following structures.

11. Aspergillus iizukae, deposit number CGMCC No. 22467.

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