CN116270564A - Application of 4,3',5' -trihydroxy resveratrol compound in preparation of coronavirus 3CL protease inhibitor - Google Patents

Abstract

The invention provides application of 4,3',5' -trihydroxy resveratrol compounds in preparing coronavirus 3CL protease inhibitors, and discovers that 4,3',5' -trihydroxy resveratrol and analogues thereof have the effect of inhibiting coronavirus 3CL protease activity, can play an anti-coronavirus role by inhibiting the activity of coronavirus 3CL protease, wherein the half inhibition concentration of piceatannol, oxidized resveratrol, danshin-leaf rhein, 2,6,3',5' -tetrahydroxy stilbene, cis-quzhazulene, 4-hydroxy resveratrol and 4,3',5' -trihydroxy resveratrol on SARS-CoV and SARS-CoV-23CL protease is lower than 10 mu M, the anti-coronavirus 3CL protease has good effect, can effectively inhibit the activity of coronavirus, can be potentially used for preventing and/or treating coronavirus infection, and further can be used for preparing medicines for preventing and/or treating coronavirus infection.

Description

Application of 4,3',5' -trihydroxy resveratrol compound in preparation of coronavirus 3CL protease inhibitor

Technical Field

The invention belongs to the field of medicines, and particularly relates to application of 4,3',5' -trihydroxy resveratrol and analogues thereof in preparation of coronavirus 3CL protease inhibitors.

Background

The covd-19 epidemic is a further highly infectious, highly pathogenic coronavirus epidemic following Severe Acute Respiratory Syndrome (SARS) and Middle Eastern Respiratory Syndrome (MERS), and has posed a serious threat to human health, social stability and economic development. By 2022, 8 months and 22 days, the global cumulative report of new coronary diagnosis cases has exceeded 5.9 billion, with cumulative deaths exceeding 644 thousands. Covd-19 is caused by the transmission of acute respiratory syndrome coronavirus type II (SARS-CoV-2). Among the numerous targets of the novel coronavirus (SARS-CoV-2), the 3CL protease (3 CLpro, 3C-like protease, also known as the major protease Mpro) is of interest to researchers due to its extremely critical role in the replication and transcription process of progeny viruses. The 3CL protease has high structural similarity and conservation in various coronaviruses and novel coronavirus variants, and is the most attractive for coronavirus targeted drug developmentOne of the targets of attraction. The 4,3',5' -trihydroxy resveratrol and the analogues thereof are antioxidants, can reduce blood viscosity, inhibit platelet coagulation and vasodilation, keep blood smooth, prevent occurrence and development of cancers, and have the prevention and treatment effects of atherosclerosis and coronary heart disease, ischemic heart disease and hyperlipidemia. Also has tumor inhibiting and estrogen-like effects, and can be used for treating chemical book breast cancer. Can delay aging and prevent cancer. In recent years, the activity of 4,3',5' -trihydroxy resveratrol and analogues thereof in antibacterial and antitumor aspects has attracted attention from the vast pharmaceutical workers. However, at present, 4,3',5' -trihydroxy resveratrol and analogues thereof have not been capable of inhibiting 3CL ^pro Is reported to be associated with coronaviruses.

Disclosure of Invention

Aiming at the problems existing in the prior art, the invention provides the application of 4,3',5' -trihydroxy resveratrol and analogues thereof in preparing coronavirus 3CL protease inhibitors and medicines for resisting new coronaviruses, which are expected to solve the current situation that no medicines for resisting coronaviruses exist at present.

The invention is realized by the following technical scheme:

the invention provides application of 4,3',5' -trihydroxy resveratrol compounds in preparing coronavirus 3CL protease inhibitors.

The invention provides application of 4,3',5' -trihydroxy resveratrol compounds in preparing medicines for preventing and/or treating diseases caused by coronaviruses.

Preferably, the structural formula of the 4,3',5' -trihydroxy resveratrol compound is shown as formula I:

wherein R is ₁ 、R ₂ 、R ₃ 、R ₄ 、R ₅ 、R ₆ 、R ₇ 、R ₈ Can be independently selected from H, OH or OCH ₃ 。

Preferably, the 4,3',5' -trihydroxy resveratrol compound is one of the following structural formulas:

preferably, the 4,3',5' -trihydroxy resveratrol compound is piceatannol, oxidized resveratrol, rheum palmatum, 2,6,3',5' -tetrahydroxy stilbene, cis-quzhazulene, 4-hydroxy resveratrol or 4,3',5' -trihydroxy resveratrol.

Preferably, the 4,3',5' -trihydroxy resveratrol compound is 4,3',5' -trihydroxy resveratrol or an analogue thereof or a pharmaceutically acceptable acid, base, salt, ester, solvate, stereoisomer, tautomer or prodrug of 4,3',5' -trihydroxy resveratrol or an analogue thereof.

Preferably, the coronavirus is: SARS-CoV, MERS-CoV or SARS-CoV-2.

Preferably, the disease caused by coronavirus is an infectious disease caused by SARS-CoV, MERS-CoV or SARS-CoV-2 or complications thereof.

Preferably, the coronavirus 3CL protease is SARS-CoV 3CL ^pro Or SARS-CoV-23CL ^pro 。

In another aspect, the present invention provides a medicament for treating and/or preventing diseases caused by coronaviruses, which comprises a 4,3',5' -trihydroxy resveratrol compound and pharmaceutically acceptable auxiliary materials, preferably one or more of 4,3',5' -trihydroxy resveratrol and analogues thereof per se or pharmaceutically acceptable acids, bases, salts, esters, solvates, stereoisomers, tautomers and prodrugs of 4,3',5' -trihydroxy resveratrol and analogues thereof.

Preferably, the dosage form of the medicament comprises: one or more of capsule, tablet, granule, gel, sustained release agent, oral liquid, dripping pill, emulsion, injection and nanometer preparation.

Compared with the prior art, the invention has the following beneficial effects:

the invention discovers that 4,3',5' -trihydroxy resveratrol and analogues thereof have the effect of inhibiting the activity of coronavirus 3CL protease, can play an anti-coronavirus role by inhibiting the activity of coronavirus 3CL protease, wherein the half inhibition concentration of piceatannol, oxidized resveratrol, danshin-leaf chrysin, 2,6,3',5' -tetrahydroxy stilbene, cis-quzhazulene, 4-hydroxy resveratrol and 4,3',5' -trihydroxy resveratrol on SARS-CoV and SARS-CoV-23CL protease is lower than 10 mu M, and the anti-coronavirus 3CL protease has good effect, thereby effectively inhibiting the activity of coronavirus, thus being potentially used for preventing and/or treating coronavirus infection and further being used for preparing medicaments for preventing and/or treating coronavirus infection.

Drawings

FIG. 1 shows the inhibition of SARS-CoV 3CL protease by white Pi Shachun of the present invention;

FIG. 2 is the inhibition of SARS-CoV 3CL protease by oxidized resveratrol in accordance with the present invention;

FIG. 3 shows the inhibition of SARS-CoV 3CL protease by Rheum emodi according to the present invention;

FIG. 4 shows the inhibition of SARS-CoV 3CL protease by 2,6,3',5' -tetrahydroxystilbene of the present invention;

FIG. 5 is an inhibitory effect of cis-Quzhazulene on SARS-CoV 3CL protease in the present invention;

FIG. 6 is the inhibition of SARS-CoV 3CL protease by 4-hydroxyresveratrol of the present invention;

FIG. 7 is the inhibition of SARS-CoV 3CL protease by 4,3',5' -trihydroxy resveratrol of the present invention;

FIG. 8 shows the inhibition of SARS-CoV-23CL protease by white Pi Shachun of the present invention;

FIG. 9 is the inhibition of SARS-CoV-23CL protease by oxidized resveratrol of the present invention;

FIG. 10 shows the inhibition of SARS-CoV-23CL protease by Rhein according to the present invention;

FIG. 11 shows the inhibition of SARS-CoV-23CL protease by 2,6,3',5' -tetrahydroxystilbene of the present invention;

FIG. 12 is an inhibitory effect of cis-Quzhazulene on SARS-CoV-23CL protease according to the present invention;

FIG. 13 is a graph showing the inhibition of SARS-CoV-23CL protease by 4-hydroxyresveratrol of the present invention;

FIG. 14 shows the inhibition of SARS-CoV-23CL protease by 4,3',5' -trihydroxyresveratrol according to the invention.

Detailed Description

For a further understanding of the present invention, the present invention is described below in conjunction with the following examples, which are provided to further illustrate the features and advantages of the present invention and are not intended to limit the claims of the present invention.

It is noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of exemplary embodiments according to the present invention. As used herein, the singular is also intended to include the plural unless the context clearly indicates otherwise, and furthermore, it is to be understood that the terms "comprises" and/or "comprising" when used in this specification are taken to specify the presence of stated features, steps, operations, devices, components, and/or combinations thereof.

Term interpretation:

the "pharmaceutically acceptable": within the scope of sound medical judgment, compounds, materials, compositions and/or dosage forms suitable for contact with the tissues of humans and animals without causing additional toxicity, irritation, allergic response, or other problem commensurate with a reasonable benefit/risk ratio.

The term "pharmaceutically acceptable salt": the biological effectiveness and properties of the compounds of the present invention are preserved and are not biologically or otherwise undesirable salts. In some cases, the compounds of the present invention are capable of forming salts in the presence of phenols and/or carboxyl groups. Pharmaceutically acceptable base addition salts can be prepared with inorganic and organic bases. Salts derived from inorganic bases include, for example: sodium, potassium, lithium, ammonium, calcium and magnesium salts. Salts derived from organic bases include, for example: primary, secondary and tertiary amines.

The term "prodrug": inactive forms of the compounds, which must be metabolized in vivo, e.g. by biological fluids or enzymes, and/or by the individual after administration, into the active form of the compound in order to produce the desired pharmacological effect. Prodrugs may be metabolized prior to absorption, during absorption, after absorption, or at specific sites. Prodrug forms of the compounds may be utilized to enhance bioavailability; improving individual acceptability, such as masking or reducing unpleasant characteristics: such as bitter taste, malodor or gastrointestinal irritation; changing the solubility; providing prolonged or slow release or delivery; making the formulation easy; and/or provide localized delivery of the compound. Reference to a compound in the present invention includes a prodrug form of the compound.

It should be noted that the raw materials used in the present invention are all common commercial products, and the sources thereof are not particularly limited.

The following raw material sources are exemplary illustrations:

Assay Reagent(Assay Buffer：SARS-CoV M ^pro /3CL ^pro ＝90：1)、Substrate

(100. Mu.M Dabcyl-KTSAVLQSGFRKME-Edans) were all obtained from Shanghai Biyun Biotechnology Co., ltd

Example 1 in vitro enzyme Activity inhibition assay of 4,3',5' -Trihydroxyresveratrol and its analogues for SARS-CoV 3CL protease

The 3CLpro inhibitor screening kit using the principle of fluorescence resonance energy transfer (PRET) simply, rapidly and sensitively completes the test of the inhibitory activity of the tested medicament. And preparing a proper amount of Assay Reagent and a positive control medicament according to the number of samples, and sequentially adding a detection Reagent and a sample solvent into a 96-well plate according to the blank control, the 100% enzyme activity control, the positive inhibitor control and the group setting of the test medicament. And (3) rapidly adding 4 mu L of subbate into each hole at a low temperature by using a gang gun, uniformly mixing, incubating for 5 minutes at 37 ℃ in a dark place, performing fluorescence measurement by using a multifunctional enzyme label (lambda excitation=340 nm, lambda emission=490 nm), calculating Relative Fluorescence Units (RFU) in each hole and a blank control hole, and recording as RFU blank control, RFU100% enzyme activity control, RFU positive control and RFU tested medicine respectively, thereby calculating the percent inhibition (%) and IC50 value of the target compound, and evaluating the inhibition capability of the inhibitor on 3CLpro activity.

As shown in Table 1, the example 22 compounds have inhibitory activity against SARS-CoV 3CL protease, wherein piceatannol, resveratrol oxide, rhein, 2,6,3',5' -tetrahydroxy stilbene, cis-quzhazhiside, 4-hydroxy resveratrol, 4,3',5' -trihydroxy resveratrol has strong inhibitory effect against SARS-CoV 3CL protease, and IC ₅₀ IC of 4,3',5' -trihydroxy resveratrol with value below 10 μm ₅₀ The value reached 1.69. Mu.M (FIGS. 1-7 show the inhibition of SARS-CoV 3CL protease by piceatannol, resveratrol oxide, rhein, 2,6,3',5' -tetrahydroxy stilbene, cis-aspergilloside, 4-hydroxy resveratrol, 4,3',5' -trihydroxy resveratrol, respectively).

TABLE 1 inhibitory Activity of 22 Compounds against SARS-CoV 3CL protease

Example 2 in vitro enzyme Activity inhibition assay of 4,3',5' -Trihydroxyresveratrol and its analogues for SARS-CoV-23CL protease

The 3CLpro inhibitor screening kit using the principle of fluorescence resonance energy transfer (PRET) simply, rapidly and sensitively completes the test of the inhibitory activity of the tested medicament. According to the number of samples, the configuration is adapted according to the descriptionAnd sequentially adding a detection Reagent and a sample solvent into the 96-well plate according to the blank control, the 100% enzyme activity control, the positive inhibitor control and the group setting of the test drugs. And (3) rapidly adding 4 mu L of subbate into each hole at a low temperature by using a gang gun, uniformly mixing, incubating for 5 minutes at 37 ℃ in a dark place, performing fluorescence measurement by using a multifunctional enzyme label (lambda excitation=340 nm, lambda emission=490 nm), calculating Relative Fluorescence Units (RFU) in each hole and a blank control hole, and recording as RFU blank control, RFU100% enzyme activity control, RFU positive control and RFU tested medicine respectively, thereby calculating the percent inhibition (%) and IC50 value of the target compound, and evaluating the inhibition capability of the inhibitor on 3CLpro activity.

As shown in Table 2, the compounds of example 22 have inhibitory activity against SARS-CoV-23CL protease, wherein piceatannol, oxyresveratrol, rhein, 2,6,3',5' -tetrahydroxy stilbene, cis-Quzhazhigan, 4-hydroxyresveratrol, 4,3',5' -trihydroxy resveratrol has strong inhibitory effect against SARS-CoV-23CL protease, and IC ₅₀ IC of 4,3',5' -trihydroxy resveratrol with value below 10 μm ₅₀ The value reached 1.65. Mu.M (FIGS. 8-14 show the inhibition of SARS-CoV-23CL protease by piceatannol, resveratrol oxide, rhein, 2,6,3',5' -tetrahydroxy stilbene, cis-aspergilloside, 4-hydroxy resveratrol, 4,3',5' -trihydroxy resveratrol, respectively).

TABLE 2 inhibitory Activity of 22 Compounds against SARS-CoV-23CL protease

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In conclusion, piceatannol, oxidized resveratrol, danshinolic acid, 2,6,3',5' -tetrahydroxy stilbene, cis-quzhazhiside, 4-hydroxy resveratrol and 4,3',5' -trihydroxy resveratrol show excellent inhibitory activity on 3CL proteases of two coronaviruses of SARS-CoV and SARS-CoV-2, and can be used as anti-coronavirus medicaments for development and research.

Claims (11)

1. Application of 1.4,3',5' -trihydroxy resveratrol compound in preparing coronavirus 3CL protease inhibitor.
2. Application of 2.4,3',5' -trihydroxy resveratrol compound in preparing medicament for preventing and/or treating diseases caused by coronavirus.
3. 3. The use according to claim 1 or 2, wherein the structural formula of the 4,3',5' -trihydroxy resveratrol compound is as shown in formula i:

   

   wherein R is ₁ 、R ₂ 、R ₃ 、R ₄ 、R ₅ 、R ₆ 、R ₇ 、R ₈ Can be independently selected from H, OH or OCH ₃ 。
4. 4. The use according to claim 1 or 2, wherein the 4,3',5' -trihydroxy resveratrol compound is of one of the following structural formulae:

   
5. 5. the use according to claim 1 or 2, wherein the 4,3',5' -trihydroxy resveratrol compound is piceatannol, oxidized resveratrol, rhein, 2,6,3',5' -tetrahydroxy stilbene, cis-aspergilloside, 4-hydroxy resveratrol or 4,3',5' -trihydroxy resveratrol.
6. 6. The use according to claim 1 or 2, wherein the 4,3',5' -trihydroxy resveratrol compound is 4,3',5' -trihydroxy resveratrol or an analogue thereof or a pharmaceutically acceptable acid, base, salt, ester, solvate, stereoisomer, tautomer or prodrug of 4,3',5' -trihydroxy resveratrol or an analogue thereof.
7. 7. The use according to claim 1, wherein the coronavirus 3CL protease is SARS-CoV 3CL ^pro Or SARS-CoV-2-3 CL ^pro 。
8. 8. The use according to claim 2, wherein the coronavirus is: SARS-CoV, MERS-CoV or SARS-CoV-2.
9. 9. The use according to claim 2, wherein the disease caused by coronavirus is an infectious disease caused by SARS-CoV, MERS-CoV or SARS-CoV-2 or a complication thereof.
10. 10. The medicine is characterized by comprising a 4,3',5' -trihydroxy resveratrol compound and pharmaceutically acceptable auxiliary materials.
11. 11. The medicament according to claim 10, wherein the dosage form is one of capsules, tablets, granules, gels, sustained release agents, oral liquids, drop pills, emulsions, injections and nano-preparations.

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