Tetraphenylethylene derivatives and their applications

Abstract

The present invention relates to a tetraphenylethylene derivative having a structure shown in formula (I) or a pharmaceutically acceptable salt thereof or a stereoisomer thereof, and an application thereof in the preparation of fluorescent antibacterial drugs. The tetraphenylethylene derivative of the present invention has many advantages such as a broad antibacterial spectrum, good biocompatibility, low cytotoxicity (high efficiency and low toxicity), a novel mechanism of action, a fast sterilization speed, low drug resistance, and has fluorescent properties, and can be used as a fluorescent probe for bacterial visualization imaging, bacterial labeling, bacterial diagnosis or bacterial detection. The tetraphenylethylene derivative provided by the present invention is used as a fluorescent antibacterial drug, which provides a new idea for reducing bacterial infection and the development of drug-resistant bacteria, and provides a new idea for the research and development of a new type of small molecule antibacterial drug for diagnosis and treatment.

Description

Technical Field

[0001] The invention belongs to the field of medical technology, and specifically relates to a tetraphenylethylene compound and a preparation method and application thereof. Background Art

[0002] At present, the number of clinical drug-resistant bacterial infections is increasing and the mortality rate is gradually rising. The therapeutic effect of traditional antimicrobial drugs on drug-resistant bacterial infections is becoming increasingly limited and can no longer meet the needs of modern clinical treatment. Therefore, it is urgent to find new antimicrobial drugs that have a different mechanism of action from existing antimicrobial drugs and are not easy to induce bacterial resistance to cope with this dilemma.

[0003] Traditional methods for pathogen detection mainly include bacterial culture and morphological observation, which have poor timeliness. Fluorescent probes can quickly and accurately detect the presence of pathogens. However, fluorescent probes with traditional structures generally have the defect of aggregation quenching effect, which greatly limits their effectiveness. Luminescent molecules with aggregation-induced emission characteristics can overcome the aggregation quenching effect and have become powerful fluorescent tools for pathogen detection and antibacterial treatment. Fluorescent antibacterial drugs have the characteristics of both antibacterial compounds and fluorescent probes. They can be used to treat bacterial infections, and can also be used to clarify the mechanism of action of antibacterial drugs, detect pathogen infections, and track the uptake of antibacterial drugs in organisms. Therefore, the development of fluorescent antibacterial drugs can provide new ideas for solving the problem of bacterial resistance. Summary of the invention

[0004] Based on this, the object of the present invention is to provide a new type of antibacterial drug.

[0005] In order to achieve the above-mentioned object of the invention, the present invention includes the following technical solutions.

[0006] The present invention provides a tetraphenylethylene derivative having a structure represented by formula (I) or a pharmaceutically acceptable salt thereof or a stereoisomer thereof:

[0007]

[0008] Wherein, R1 is selected from: C1-C 20 Alkyl, C3-C 20 Unsaturated chain hydrocarbon group;

[0009] Each R2 is independently selected from: C1-C 20 Alkyl, R3 substituted C3-C 20 Unsaturated chain hydrocarbon group, C1-C6 alkyl substituted by R4;

[0010] R3 is selected from: -NR5R6;

[0011] R4 is selected from: -C(=O)NR7R8;

[0012] R5 and R6 are independently selected from: hydrogen, C1-C6 alkyl;

[0013] R7 is selected from: hydrogen, C1-C6 alkyl;

[0014] R8 is selected from: 1 or more R9 substituted or unsubstituted C1-C 20 alkyl;

[0015] R9 is selected from: hydrogen, C1-C6 alkoxycarbonyl, -C(=O)NR5R6, amino, C1-C6 alkylamino,

[0016] In some embodiments, R1 is selected from: C1-C 12 Alkyl, C3-C 12 Unsaturated chain hydrocarbon group.

[0017] In some embodiments, R1 is selected from: C3-C 10 Alkyl, C3-C8 unsaturated chain hydrocarbon group.

[0018] In some embodiments, R1 is selected from: C3-C6 alkyl, C4-C6 unsaturated chain hydrocarbon group.

[0019] In some embodiments, R1 is selected from: 3-methyl-2-butenyl, decyl, propyl.

[0020] In some embodiments, each R2 is independently selected from: C1-C 10 Alkyl, R3 substituted C3-C 10 Unsaturated chain hydrocarbon group, C1-C3 alkyl group substituted by R4.

[0021] In some of the embodiments, each R2 is independently selected from: C2-C6 alkyl substituted by R3, C3-C6 unsaturated chain hydrocarbon group substituted by R3, C1-C2 alkyl substituted by R4.

[0022] In some embodiments, each R2 is independently selected from: R3 substituted ethyl, R3 substituted propyl, R3 substituted butyl, R3 substituted pentyl, R4 substituted methyl, R4 substituted ethyl.

[0023] In some of these embodiments, R3 is selected from: -NR5R6;

[0024] R5 and R6 are independently selected from the group consisting of hydrogen, methyl, ethyl, and propyl.

[0025] In some embodiments, R3 is selected from: methylamino, dimethylamino, diethylamino.

[0026] In some embodiments, R4 is selected from: -C(=O)NR7R8;

[0027] R7 is selected from: hydrogen, methyl, ethyl, propyl;

[0028] R8 is selected from: 1 or more R9 substituted or unsubstituted C1-C 10 alkyl;

[0029] R9 is selected from: hydrogen, C1-C3 alkoxycarbonyl, -C(=O)NR5R6, amino, C1-C3 alkylamino,

[0030] Wherein, R5 and R6 are independently selected from: hydrogen, methyl, ethyl, propyl.

[0031] In some embodiments, R8 is selected from: 1 or more R9 substituted or unsubstituted C1-C6 alkyl.

[0032] In some embodiments, R8 is selected from: 1 or more R9 substituted or unsubstituted ethyl, 1 or more R9 substituted or unsubstituted propyl, 1 or more R9 substituted or unsubstituted butyl.

[0033] In some embodiments, R9 is selected from: hydrogen, methoxycarbonyl, -C(=O)NH2,

[0034] In some embodiments, -NR7R8 is selected from the following structures:

[0035]

[0036] In some embodiments, the tetraphenylethylene derivative is selected from:

[0037]

[0038] The present invention also provides the use of the tetraphenylethylene derivative or its pharmaceutically acceptable salt or its stereoisomer, including the following:

[0039] The invention discloses an application of the tetraphenylethylene derivative or its pharmaceutically acceptable salt or its stereoisomer in preparing a drug for inhibiting bacterial growth.

[0040] The invention discloses an application of the tetraphenylethylene derivative or a pharmaceutically acceptable salt or a stereoisomer thereof in preparing a drug for preventing and / or treating bacterial infection.

[0041] The tetraphenylethylene derivatives or pharmaceutically acceptable salts or stereoisomers thereof described in the present invention are used as fluorescent probes in bacterial visualization imaging, bacterial labeling, bacterial diagnosis or bacterial detection.

[0042] In some embodiments, the bacteria are Gram-positive bacteria and / or Gram-negative bacteria, preferably Gram-positive bacteria.

[0043] In some embodiments, the bacteria are Staphylococcus, Enterococcus, Enterobacter, or Acinetobacter.

[0044] In some of the embodiments, the bacteria are Staphylococcus aureus, methicillin-resistant Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Acinetobacter baumannii, and Klebsiella pneumoniae.

[0045] The present invention also provides an antibacterial drug prepared from an active ingredient and a pharmaceutically acceptable excipient, wherein the active ingredient comprises the tetraphenylethylene derivative described in the present invention or a pharmaceutically acceptable salt thereof or a stereoisomer thereof.

[0046] The present invention uses tetra-(4-hydroxyphenyl)ethylene as a starting material, introduces hydrophilic groups (such as fatty amines and arginine, etc.) and lipophilic side chains on a tetraphenylethylene skeleton, and synthesizes a series of amphiphilic cationic compounds with aggregation-induced luminescence effect. Since the bacterial cell membrane carries a negative charge, the introduced cationic groups can be gathered on the bacterial cell membrane by electrostatic attraction, so that it emits a specific fluorescent signal. At the same time, the hydrophobic side chain can be inserted into the phospholipid bilayer of the bacterial cell membrane, and the bacterial cell is caused to die due to rupture or leakage of contents by destroying the integrity of the bacterial cell membrane. The present invention uses tetraphenylethylene as a parent nucleus, so that the starting material that originally has no antibacterial activity is designed and synthesized into an antibacterial drug with a new antibacterial molecular skeleton. The tetraphenylethylene derivatives of the present invention have many advantages such as a wide antibacterial spectrum, good biocompatibility, low cytotoxicity (high efficiency and low toxicity), novel mechanism of action, fast sterilization speed, low drug resistance, and have fluorescence characteristics, and can be used as a fluorescent probe for bacterial visualization imaging, bacterial labeling, bacterial diagnosis or bacterial detection. The tetraphenylethylene derivatives provided by the present invention are used as fluorescent antibacterial drugs, which provide new ideas for reducing bacterial infections and the development of drug-resistant bacteria, and provide new ideas for the research and development of new small-molecule antibacterial drugs that integrate diagnosis and treatment.

[0047] The optimal compound TK24 of the present invention shows excellent antibacterial activity in the mouse corneal infection model caused by Staphylococcus aureus ATCC29213. Compound TK24 has the advantages of high efficiency and low toxicity, rapid bactericidal speed, good in vivo antibacterial activity, good drugability, and the ability to overcome drug resistance. It is also a new molecular entity and can avoid cross-resistance with traditional antibacterial drugs. It also has great application potential in bacterial tracking and monitoring treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1The cytotoxicity of compound TK24 on (a) mouse fibroblasts (NCTC clone 929) and (b) human embryonic kidney cells (HEK293).

[0049] Figure 2 The effect of compound TK24 on the cell membrane permeability of S. aureus ATCC29213 (a), MRSA NCTC10442 (b), E. coli ATCC25922 (c) and K. pneumoniae ATCC10031 (d).

[0050] Figure 3 The results of the drug resistance study of compound TK24 to MRSANCTC10442 (a) and E. coli ATCC25922 (b).

[0051] Figure 4 The in vivo antibacterial efficacy of compound TK24 in the mouse keratitis model induced by S. aureus ATCC29213.

[0052] Figure 5 Fluorescence images of (a) MRSANCTC10442, (b) E. coli ATCC25922, (c) A. baumannii R2889, and (d) K. pneumoniae ATCC10031 treated with compound TK24 observed using a laser confocal microscope, with an excitation wavelength of 405 nm. DETAILED DESCRIPTION

[0053] In order to facilitate the understanding of the present invention, the present invention will be described more fully below. The present invention can be implemented in many different forms and is not limited to the embodiments described herein. On the contrary, the purpose of providing these embodiments is to make the understanding of the disclosure of the present invention more thorough and comprehensive.

[0054] The experimental methods in the following examples without specifying specific conditions are usually carried out under conventional conditions or conditions recommended by the manufacturers. The various commonly used chemical reagents used in the examples are all commercially available products.

[0055] Unless otherwise defined, all technical and scientific terms used in the present invention have the same meaning as those commonly understood by those skilled in the art to which the present invention belongs. The terms used in the specification of the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention. The term "and / or" used in the present invention includes any and all combinations of one or more of the related listed items.

[0056] In addition, as used in the present invention, the term "or" is an inclusive "or" symbol and is equivalent to the term "and / or", unless the context clearly dictates otherwise. The term "based on" is not exclusive and allows for being based on other factors not described, unless the context clearly dictates otherwise. In addition, throughout the specification, the meanings of "a", "an", and "the" include plural referents. The meaning of "in..." includes "in..." and "on...".

[0057] In order to develop more new antibacterial drugs to overcome the problem that existing clinical drugs cannot meet the demand for drugs for drug-resistant bacterial infections, the present invention provides, in one embodiment of the present invention, a tetraphenylethylene derivative having a structure shown in formula (I) or a pharmaceutically acceptable salt thereof or a stereoisomer thereof:

[0058]

[0059] Wherein, R1 is selected from: C1-C 20 Alkyl, C3-C 20 Unsaturated chain hydrocarbon group;

[0060] Each R2 is independently selected from: C1-C 20 Alkyl, R3 substituted C3-C 20 Unsaturated chain hydrocarbon group, C1-C6 alkyl substituted by R4;

[0061] R3 is selected from: -NR5R6;

[0062] R4 is selected from: -C(=O)NR7R8

[0063] R5 and R6 are independently selected from: hydrogen, C1-C6 alkyl;

[0064] R7 is selected from: hydrogen, C1-C6 alkyl;

[0065] R8 is selected from: 1 or more R9 substituted or unsubstituted C1-C 20 alkyl;

[0066] R9 is selected from: hydrogen, C1-C6 alkoxycarbonyl, -C(=O)NR5R6, amino, C1-C6 alkylamino,

[0067] In the compounds of the present invention, when any variable (e.g., R2, etc.) occurs more than once in any component, its definition at each occurrence is independent of the definition at every other occurrence. Likewise, combinations of substituents and variables are permitted as long as such combinations stabilize the compound. It is to be understood that one of ordinary skill in the art can select substituents and substitution patterns for the compounds of the present invention to provide chemically stable compounds that can be readily synthesized from readily available raw materials by techniques in the art and the methods set forth below. If a substituent itself is substituted with more than one group, it is to be understood that these groups may be on the same carbon atom or on different carbon atoms as long as the structure is stable.

[0068] The term "substituted" refers to the replacement of hydrogen radicals in a particular structure with the radical of a specified substituent.

[0069] The term "alkyl" is intended to include branched and straight chain saturated aliphatic hydrocarbon groups having a specific number of carbon atoms. For example, the definition of "C1-C6" in "C1-C6 alkyl" includes groups having 1, 2, 3, 4, 5 or 6 carbon atoms in a straight or branched arrangement. For example, "C1-C6 alkyl" specifically includes methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, pentyl, hexyl.

[0070] The term "unsaturated chain hydrocarbon group" refers to branched and straight chain unsaturated aliphatic hydrocarbon groups having a specific number of carbon atoms, i.e., non-cyclic chain hydrocarbon groups, and the carbon chain contains one or more carbon-carbon double bonds, or contains carbon-carbon triple bonds, such as:

[0071] -CH2(CH=CH)(CH2)2CH3, -CH2(CH=CH)(CH3)CH3, etc.

[0072] The term "alkoxy" refers to a group in which an alkyl group is directly connected to oxygen, that is, a group having an -O-alkyl structure, such as -OCH3, -OCH2CH3, -OCH2CH2CH3, -OCH2CH(CH3)2, -OCH2CH2CH2CH3, -O-CH(CH3)2, etc.

[0073] The present invention includes free forms of compounds of formula I, as well as pharmaceutically acceptable salts and stereoisomers thereof. Some specific exemplary compounds of the present invention are amine compounds in non-salt form, i.e., "free forms". Pharmaceutically acceptable salts of the present invention can be synthesized from compounds of the present invention containing a basic or acidic moiety by conventional chemical methods. Typically, salts of basic compounds are prepared by ion exchange chromatography or by reacting a free base with a stoichiometric amount or excess of an inorganic or organic acid in the desired salt form in an appropriate solvent or a combination of multiple solvents. Similarly, salts of acidic compounds are formed by reacting with an appropriate inorganic or organic base.

[0074] Therefore, the pharmaceutically acceptable salts of the compounds of this invention include conventional non-toxic salts of the compounds of this invention formed by reacting alkaline compounds of this invention with inorganic or organic acids. For example, conventional non-toxic salts include salts derived from inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, sulfamic acid, phosphoric acid, nitric acid, etc., and also include salts prepared from organic acids such as acetic acid, propionic acid, succinic acid, glycolic acid, stearic acid, lactic acid, malic acid, tartaric acid, citric acid, ascorbic acid, pamoic acid, maleic acid, hydroxymaleic acid, phenylacetic acid, glutamic acid, benzoic acid, salicylic acid, p-aminobenzenesulfonic acid, 2-acetoxy-benzoic acid, fumaric acid, toluenesulfonic acid, methanesulfonic acid, ethanedisulfonic acid, oxalic acid, isethionic acid, trifluoroacetic acid, etc.

[0075] If the compound of the present invention is acidic, then suitable "pharmaceutically acceptable salts" refer to salts prepared from pharmaceutically acceptable non-toxic bases including inorganic bases and organic bases. Salts derived from inorganic bases include aluminum salts, ammonium salts, calcium salts, copper salts, iron salts, ferrous salts, lithium salts, magnesium salts, manganic salts, manganous salts, potassium salts, sodium salts, zinc salts, etc. Ammonium salts, calcium salts, magnesium salts, potassium salts and sodium salts are particularly preferred. Salts derived from pharmaceutically acceptable organic non-toxic bases, including salts of primary, secondary and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins such as arginine, betaine, caffeine, choline, N,N'-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, aminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucosamine, glucosamine, histidine, hydroxocobalamin, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, guazine, polyamine resins, procaine, purine, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine and the like.

[0076] Berg et al., "Pharmaceutical Salts," J. Pharm. Sci. '1977: 66: 1-19, describes in more detail the preparation of the above-mentioned pharmaceutically acceptable salts and other typical pharmaceutically acceptable salts.

[0077] The stereoisomers of the present invention are, i.e. (depending on their structure) as enantiomers, diastereomers, syn- / anti-isomers, cis- / trans-isomers, epimers and (E)- / (Z)-isomers. The compounds of formula I can be used in the context of the present invention in the form of pure stereoisomers or in the form of any mixture of stereoisomers, in the latter case the racemates are preferred.

[0078] In another embodiment of the present invention, there is provided a use of the tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof in the preparation of a drug for inhibiting bacterial growth.

[0079] In another embodiment of the present invention, there is provided a use of the tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof in the preparation of a medicament for treating and / or preventing bacterial infection.

[0080] In another embodiment of the present invention, there is provided the use of the tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof as a fluorescent probe in bacterial visualization imaging, bacterial labeling, bacterial diagnosis or bacterial detection.

[0081] The bacteria described in the present invention can be non-resistant bacteria or resistant bacteria, such as single-drug resistant bacteria or multi-drug resistant bacteria; they can be Gram-positive bacteria or Gram-negative bacteria, and especially have better antibacterial and fluorescent effects on Gram-positive bacteria.

[0082] In another embodiment of the present invention, an antibacterial drug is provided, which is prepared from an active ingredient and a pharmaceutically acceptable excipient, wherein the active ingredient comprises the tetraphenylethylene derivative described in the present invention or a pharmaceutically acceptable salt thereof or a stereoisomer thereof.

[0083] Among them, the amount of active ingredients contained is within the range of safe and effective amount. "Safe and effective amount" means: the amount of active ingredients is sufficient to significantly improve the condition without causing serious side effects.

[0084] "Pharmaceutically acceptable excipients" refer to: one or more compatible solid or liquid fillers or gel substances, which are suitable for human use and must be sufficiently pure and sufficiently low in toxicity.

[0085] "Compatibility" herein means that the components in the composition or preparation can be blended with the active ingredient of the present invention and with each other without significantly reducing the efficacy of the active ingredient.

[0086] Some examples of pharmaceutically acceptable excipients (or carriers) include cellulose and its derivatives (such as sodium carboxymethyl cellulose, sodium ethyl cellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (such as stearic acid, magnesium stearate), calcium sulfate, vegetable oils (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerol, mannitol, sorbitol, etc.), emulsifiers (such as ), wetting agents (such as sodium lauryl sulfate), colorants, flavoring agents, stabilizers, antioxidants, preservatives, pyrogen-free water, etc.

[0087] There is no particular limitation on the administration of the active ingredient, pharmaceutical composition or pharmaceutical preparation of the present invention, and representative administration methods include but are not limited to: oral, transdermal, rectal, parenteral (intravenous, intramuscular or subcutaneous), etc. That is, the dosage form of the pharmaceutical preparation includes but is not limited to: capsules, granules, tablets, pills, powders, drops, ointments, patches, liniments, sprays, powders, suppositories, sustained-release preparations, injections, etc.

[0088] Solid dosage forms for oral administration include capsules, granules, tablets, pills, powders, etc. In these solid dosage forms, the active ingredient is mixed with at least one conventional inert excipient (or adjuvant or carrier), such as sodium citrate or dicalcium phosphate, or with the following ingredients:

[0089] (a) fillers or extenders, for example, starch, lactose, sucrose, glucose, mannitol and silicic acid;

[0090] (b) binders, for example, hydroxymethylcellulose, alginate, gelatin, polyvinyl pyrrolidone, sucrose and gum arabic;

[0091] (c) humectants, for example, glycerin;

[0092] (d) disintegrating agents, for example, agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate;

[0093] (e) a buffering solvent, for example, paraffin;

[0094] (f) absorption accelerators, for example, quaternary ammonium compounds;

[0095] (g) wetting agents, for example, cetyl alcohol and glyceryl monostearate;

[0096] (h) adsorbents, for example, kaolin; and

[0097] (i) Lubricants, for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, or mixtures thereof. In capsules, tablets and pills, the dosage form may also contain a buffering agent.

[0098] The solid dosage forms can also be prepared using coatings and shell materials, such as enteric coatings and other materials known in the art. They can contain opacifying agents, and the release of the active ingredient in such compositions can be delayed in a certain part of the digestive tract. Examples of embedding components that can be used are polymeric substances and waxes.

[0099] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or tinctures. In addition to the active ingredient, the liquid dosage form may contain inert diluents conventionally used in the art, such as water or other solvents, solubilizers and emulsifiers, for example, ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butylene glycol, dimethylformamide and oils, in particular cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil or mixtures of these substances. In addition to these inert diluents, the composition may also contain adjuvants, such as wetting agents, emulsifiers and suspending agents, sweeteners, flavoring agents and spices.

[0100] In addition to the active ingredients, suspensions may contain suspending agents such as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methanol and agar, or mixtures of these substances.

[0101] Compositions for parenteral injection may include physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and non-aqueous carriers, diluents, solvents or excipients include water, ethanol, polyols and suitable mixtures thereof.

[0102] The present invention is further described in detail below with reference to specific embodiments.

[0103] The synthetic route of tetraphenylethylene derivatives is as follows:

[0104] Route 1: Synthesis of tetraphenylethylene derivatives TK07, TK09 and TK26

[0105]

[0106] Route 2: Synthesis of tetraphenylethylene derivatives TK24, TK25, TK35 and TK38

[0107]

[0108] Example 1 Preparation of Compound TK03

[0109]

[0110] Tetra-(4-hydroxyphenyl)ethylene (300 mg, 0.76 mmol), potassium carbonate (156 mg, 1.14 mmol) and 1-bromo-3-methyl-2-butene (96 μL, 0.83 mmol) were added to a 100 mL round-bottom flask, fully dissolved with 8 mL of anhydrous methanol, and stirred at room temperature for 3 h. After the reaction was completed, it was extracted twice with ethyl acetate and water, the aqueous layer was discarded and the organic layer was collected, and the crude product was obtained after rotary evaporation and concentration. The crude product was purified by silica gel chromatography to obtain a light yellow oily product (65 mg, 18%).

[0111] 1 H NMR (400MHz, CD3OD) δ6.90-6.85(m,2H),6.82-6.76(m,6H),6.62(d,J=8.8Hz,2H),6. 54-6.47(m,6H),5.48-5.37(m,1H),4.43(d,J=6.6Hz,2H),1.77(s,3H),1.71(s,3H). 13 C NMR (100MHz, CD3OD) δ158.42,156.64,156.61,156.60,139.97,139.38,138.49,138.44,137.33,137.32,137 .27,133.70,133.68,133.59,121.25,115.36,115.33,114.77,65.69,25.89,18.20.HRMS(ESI+):calculated for C 31 H 28 O4[M+H] + 465.2060,found465.2028.

[0112] Example 2 Preparation of Compound TK06

[0113]

[0114] Compound TK03 (50 mg, 0.11 mmol) was fully dissolved in 8 mL of acetone in a 100 mL round-bottom flask, potassium carbonate (74.37 mg, 0.54 mmol) and 1,4-dibromobutane (52 μL, 0.43 mmol) were added, and refluxed for 8 h. After the reaction, the mixture was extracted twice with ethyl acetate and water, the aqueous layer was discarded, and the organic layer was collected and concentrated by rotary evaporation to obtain a crude product, which was purified by silica gel chromatography to obtain a yellow-brown oily product (73 mg, 78%).

[0115] 1H NMR (400MHz, CDCl3) δ6.94-6.89(m,8H),6.67-6.59(m,8H),5.51-5.42(m,1H),4.43(d,J=6.8Hz,2H),3.92(t,J =6.0Hz,6H),3.48(t,J=6.6Hz,6H),2.09-2.00(m,6H),1.95-1.86(m,6H),1.80-1.77(m,3H),1.73-1.70(m,3H). 13 C NMR (100MHz, CDCl3) δ157.22,157.14,138.50,138.35,138.07,137.07,137.05,136.85,132.65,132.5 8,119.80,113.81,113.61,66.68,64.69,33.62,29.59,28.01,25.93,18.28.HRMS(ESI+):calculated forC 43 H 49 Br3O4[M+H] + 867.1088, found 867.1097.

[0116] Example 3 Preparation of Compound TK07

[0117]

[0118] Compound TK06 (50 mg, 0.06 mmol) was fully dissolved in 8 mL DMF in a 100 mL round-bottom flask, and 100 μL of methylamine solution (33 wt.% in MeOH) was added and stirred at room temperature overnight. After the reaction, it was extracted twice with n-butanol and water, the aqueous layer was discarded and the organic layer was collected and concentrated by rotary evaporation to obtain a crude product, which was purified by preparative HPLC to obtain a pale yellow oil product (25 mg, 43%).

[0119] 1 H NMR (400MHz, CD3OD) δ6.85(d,J=7.8Hz,8H),6.62(d,J=8.1Hz,8H),5.43-5.34(m,1H),4.41(d,J=5.6Hz,2H),4.0 0-3.83(m,6H),3.40-3.30(m,2H),3.07-2.93(m,5H),2.86-2.78(m,2H),2.69-2.63(m,6H),1.92-1.62(m,18H). 13C NMR (100MHz, CD3OD) δ165.07,158.61,140.01,139.79,138.52,138.50,138.32,138.05,133.67,133.65,133.64,133.60,133.55,121.20,11 4.83,114.64,114.58,67.92,65.67,50.09,50.07,34.98,33.47,29.66,27.28,27.26,25.89,25.71,24.26,18.20.HRMS(ESI+):calculated for C 46 H 61 N3O4[M+H] + 720.4735, found 720.4716.

[0120] Example 4 Preparation of Compound TK08

[0121]

[0122] Compound TK03 (50 mg, 0.11 mmol) was fully dissolved in 8 mL of acetone in a 100 mL round-bottom flask, potassium carbonate (74.37 mg, 0.54 mmol) and ethyl bromoacetate (48 μL, 0.43 mmol) were added, and refluxed for 8 h. After the reaction, the mixture was extracted twice with ethyl acetate and water, the aqueous layer was discarded, and the organic layer was collected and concentrated by rotary evaporation to obtain a crude product, which was purified by silica gel chromatography to obtain a yellow-brown oily product (62 mg, 80%).

[0123] 1 H NMR(400MHz,)δ6.94-6.85(m,8H),6.67-6.60(m,8H),5.51-5.42(m,1H),4.58-4.51(m,6H) ,4.42(d,J=6.8Hz,2H),4.25(q,J=7.1Hz,6H),1.78(s,3H),1.71(s,3H),1.30-1.26(m,9H). 13C NMR(100MHz,)δ169.04,169.01,157.37,156.21,156.19,138.94,138.21,138.11,137.79,137.75,137.74,136.36,132.67,132. 65,132.55,119.75,113.91,113.90,113.88,113.82,65.47,65.45,64.67,61.40,25.91,18.27,14.23.HRMS(ESI+):calculated for C 43 H 46 O 10 [M+H] + 723.3164, found 723.3144.

[0124] Example 5 Preparation of Compound TK09

[0125]

[0126] Compound TK06 (50 mg, 0.06 mmol) was fully dissolved in 8 mL DMF in a 100 mL round-bottom flask, and 100 μL dimethylamine solution (2 M in THF) was added and stirred at room temperature overnight. After the reaction was completed, it was extracted twice with n-butanol and water, the aqueous layer was discarded and the organic layer was collected and concentrated by rotary evaporation to obtain a crude product, which was purified by preparative HPLC to obtain a pale yellow oil product (28 mg, 63%).

[0127] 1 H NMR (400MHz, CD3OD) δ6.93-6.82(m,8H),6.70-6.60(m,8H),5.42(t,J=6.7Hz,1H),4.44(d,J=6.6Hz,2H),3 .96(t,J=5.6Hz,6H),3.20-3.11(m,6H),2.90-2.82(m,18H),1.95-1.79(m,12H),1.77(s,3H),1.72(s,3H). 13C NMR (100MHz, CD3OD) δ158.67,158.61,158.60,140.01,139.80,138.49,138.33,138.30,138.05,133.70,133.65,133.63,133. 55,121.20,114.84,114.60,101.32,67.88,65.67,58.59,43.24,27.29,27.26,25.89,22.76,18.21.HRMS(ESI+):calculated for C 49 H 67 N3O4[M+H] + 762.5204, found 762.5192.

[0128] Example 6 Preparation of Compound TK24

[0129]

[0130] Compound TK08 (50 mg, 0.08 mmol) was fully dissolved in 4 mL of tetrahydrofuran and added to a 100 mL round-bottom flask. LiOH (24 mg, 1.00 mmol) dissolved in water (2 mL) was added. The reaction was stirred at room temperature until the hydrolysis was complete. 2 mL of glacial acetic acid was added to the reaction flask and stirred thoroughly. After the reaction was completed, it was extracted twice with n-butanol and water, the water layer was discarded, and the organic layer was collected. After rotary evaporation and concentration, it was placed in a vacuum drying oven and fully dried to obtain a crude intermediate product after hydrolysis.

[0131] After adding 8 mL of anhydrous DMF to the reaction bottle containing the crude intermediate product, L-arginine methyl ester dihydrochloride (102 mg, 0.39 mmol), HATU (89 mg, 0.23 mmol), and DIPEA (77 μL, 0.47 mmol) were added and stirred overnight at room temperature. After the reaction was completed, the organic phase was extracted twice with n-butanol and water and collected, and the crude product was obtained after rotary evaporation and concentration. The crude product was purified by preparative HPLC to obtain a yellow-green oily product (43 mg, 48%).

[0132] 1H NMR (400MHz, CD3OD) δ6.94(d,J=6.7Hz,6H),6.88(d,J=8.6Hz,2H),6.74(d,J=8.6Hz,6H),6.64(d,J=8.7Hz,2H),5.42(t,J=7.0Hz,1 H),4.58-4.50(m,9H),4.44(d,J=6.6Hz,2H),3.72(s,9H),3.19(t,J=6.0Hz,6H),2.04-1.74(m,9H),1.72(s,3H),1.69-1.56(m,6H). 13 C NMR (100MHz, CD3OD) δ173.36,173.35,173.33,171.39,171.36,168.30,158. 83,158.61,157.57,140.53,139.70,139.14,138.60,137.59,133.72,133.67 ,133.52,121.12,115.15,115.11,114.93,67.95,65.70,53.00,52.99,52.9 8,52.87,52.82,41.75,29.41,26.21,25.88,18.21.HRMS(ESI+):calculated for C 58 H 76 N 12 O 13 [M+H] + 1149.5728, found 1149.5733.

[0133] Example 7 Preparation of Compound TK25

[0134]

[0135] Compound TK08 (50 mg, 0.08 mmol) was fully dissolved in 4 mL of tetrahydrofuran and added to a 100 mL round-bottom flask. LiOH (24 mg, 1.00 mmol) dissolved in water (2 mL) was added. The reaction was stirred at room temperature until the hydrolysis was complete. 2 mL of glacial acetic acid was added to the reaction flask and stirred thoroughly. After the reaction was completed, it was extracted twice with n-butanol and water, the water layer was discarded, and the organic layer was collected. After rotary evaporation and concentration, it was placed in a vacuum drying oven and fully dried to obtain a crude intermediate product after hydrolysis.

[0136] After adding 8 mL of anhydrous DMF to the reaction bottle containing the crude intermediate product, add L-arginine amide dihydrochloride (135 mg, 0.55 mmol), HATU (89 mg, 0.23 mmol), and DIPEA (77 μL, 0.47 mmol). Stir overnight at room temperature. After the reaction is completed, extract twice with n-butanol and water and collect the organic phase. After rotary evaporation and concentration, the crude product is obtained. The crude product is purified by preparative HPLC to obtain a yellow-green oily product (38 mg, 44%).

[0137] 1 H NMR (400MHz, CD3OD) δ6.96-6.90(m,6H),6.87(d,J=8.8Hz,2H),6.79-6.72(m,6H),6.64(d,J=8.8Hz,2H),5.45 -5.39(m,1H),4.57-4.42(m,11H),3.23-3.14(m,6H),1.98-1.87(m,3H),1.80-1.69(m,9H),1.68-1.56(m,6H). 13 C NMR (100MHz, CD3OD) δ176.08,176.06,171.11,171.08,170.44,158.86,158.64,158.63,157.50,157.48,139.69,139.17,139.12,138.61,1 37.53,133.78,133.53,121.13,115.10,114.93,101.33,67.97,53.40,52.29,41.83,30.52,26.16,25.90,18.22.HRMS(ESI+):calculated for C 55 H 73 N 15 O 10 [M+H] + 1104.5738, found 1104.5744.

[0138] Example 8 Preparation of Compound TK26

[0139]

[0140] Compound TK06 (50 mg, 0.06 mmol) was fully dissolved in 8 mL DMF in a 100 mL round-bottom flask, and diethylamine (100 μL, 0.97 mmol) was added and stirred overnight at room temperature. After the reaction, it was extracted twice with n-butanol and water, the aqueous layer was discarded and the organic layer was collected and concentrated by rotary evaporation to obtain a crude product, which was purified by preparative HPLC to obtain a light yellow oily product (22 mg, 45%).

[0141] 1 H NMR (400MHz, CD3OD) δ6.92-6.84(m,8H),6.70-6.60(m,8H),5.45-5.38(m,1H),4.44(d,J=6.6Hz,2H),4.0 5-3.89(m,6H),3.16-3.05(m,18H),1.91-1.80(m,12H),1.77(s,3H),1.72(s,3H),1.27(t,J=7.2Hz,18H). 13 C NMR (100MHz, CD3OD) δ158.68,158.65,158.64,140.03,138.48,138.32,138.31,138.30,138.08,138.05,133.67,133.56,121.21,114.86,11 4.84,114.60,114.59,114.56,101.33,68.01,65.68,52.68,48.08,27.55,27.50,25.89,22.29,22.25,18.21,9.44.HRMS(ESI+):calculated for C 55 H 79 N3O4[M+H] + 846.6143, found 846.6123.

[0142] Example 9 Preparation of Compound TK33

[0143]

[0144] Tetrakis-(4-hydroxyphenyl)ethylene (200 mg, 0.50 mmol), potassium carbonate (104 mg, 0.76 mmol) and 1-iododecane (118 μL, 0.55 mmol) were added to a 100 mL round-bottom flask, fully dissolved with 8 mL of anhydrous methanol, and stirred at room temperature for 3 h. After the reaction was completed, it was extracted twice with ethyl acetate and water, the aqueous layer was discarded and the organic layer was collected, and the crude product was obtained after rotary evaporation and concentration. The crude product was purified by silica gel chromatography to obtain a light yellow oil product (48 mg, 16%).

[0145] 1 H NMR(400MHz,CD3OD)δ6.87(d,J=8.5Hz,2H),6.82-6.76(m,6H),6.64-6.59(m,2H),6.53-6.47(m,6H),3 .87(t,J=6.4Hz,2H),1.76-1.67(m,2H),1.48-1.39(m,2H),1.37-1.25(m,14H),0.90(t,J=6.8Hz,3H). 13 C NMR (100MHz, CD3OD) δ158.74,156.63,156.61,156.60,139.94,139.43,138.36,137.36,137.31,133.70,133.68,133.60, 115.37,115.34,114.54,68.84,33.07,30.73,30.70,30.55,30.47,30.44,27.17,23.75,14.48.HRMS(ESI+):calculated for C 36 H 40 O4[M+H] + 537.2999, found 537.5356.

[0146] Example 10 Preparation of Compound TK34

[0147]

[0148] Compound TK33 (35 mg, 0.07 mmol) was fully dissolved in 8 mL of acetone in a 100 mL round-bottom flask, potassium carbonate (45 mg, 0.33 mmol) and ethyl bromoacetate (29 μL, 0.26 mmol) were added, and refluxed for 8 h. After the reaction, the mixture was extracted twice with ethyl acetate and water, the aqueous layer was discarded, and the organic layer was collected and concentrated by rotary evaporation to obtain a crude product, which was purified by silica gel chromatography to obtain a yellow-brown oil product (45 mg, 87%).

[0149] 1 H NMR (400MHz, CDCl3) δ6.94-6.84(m,8H),6.66-6.57(m,8H),4.54(d,J=1.0Hz,6H),4.27-4.23(m,4H),3 .86(t,J=6.6Hz,2H),1.78-1.69(m,2H),1.46-1.37(m,2H),1.33-1.24(m,21H),0.87(t,J=6.9Hz,3H). 13C NMR (100MHz, CDCl3) δ169.07,157.60,156.15,156.13,156.11,138.94,138.12,137.80,137.77,137.75,136.21,132.70,132.59,113.8 6,113.84,113.61,67.87,65.42,65.40,61.46,32.01,29.69,29.67,29.54,29.44,29.43,26.18,22.80,14.27.HRMS(ESI+):calculated for C 48 H 58 O 10 [M+H] + 795.4103, found 795.4122.

[0150] Example 11 Preparation of Compound TK35

[0151]

[0152] Compound TK34 (50 mg, 0.07 mmol) was fully dissolved in 4 mL of tetrahydrofuran and added to a 100 mL round-bottom flask. LiOH (24 mg, 1.00 mmol) dissolved in water (2 mL) was added. The reaction was stirred at room temperature until the hydrolysis was complete. 2 mL of glacial acetic acid was added to the reaction flask and stirred thoroughly. After the reaction was completed, it was extracted twice with n-butanol and water, the water layer was discarded, and the organic layer was collected. After rotary evaporation and concentration, it was placed in a vacuum drying oven and fully dried to obtain a crude intermediate product after hydrolysis.

[0153] After adding 8 mL of anhydrous DMF to the reaction bottle containing the crude intermediate product, add L-arginine methyl ester dihydrochloride (128 mg, 0.49 mmol), HATU (80 mg, 0.21 mmol), and DIPEA (70 μL, 0.42 mmol). Stir overnight at room temperature. After the reaction is completed, extract twice with n-butanol and water, discard the water layer and collect the organic layer, and concentrate by rotary evaporation to obtain a crude product. The crude product is purified by preparative HPLC to obtain a yellow-green oil product (39 mg, 45%).

[0154] 1H NMR (400MHz, CD3OD) δ6.93(d,J=8.2Hz,6H),6.87(d,J=8.6Hz,2H),6.74(d,J=8.5Hz,6H),6.63(d,J=8.6Hz,2H),4.58-4.46(m,9H),3.87(t,J=6.3 Hz,2H),3.75-3.67(m,9H),3.25-3.12(m,6H),2.04-1.77(m,6H),1.75-1 .56(m,8H),1.49-1.40(m,2H),1.37-1.27(m,12H),0.90(t,J=6.8Hz,3H). 13 C NMR (100MHz, CD3OD) δ173.36,173.35,173.33,171.37,171.35,171.34,159.12,158.67,157.56,140 .53,139.63,139.15,139.13,137.46,133.78,133.74,133.73,133.67,133.55,133.53,115.15,115 .14,115.10,115.06,114.69,114.59,101.31,68.84,67.95,52.98,52.89,52.85,49.85,41.74,33. 08,30.75,30.71,30.56,30.48,30.44,29.41,27.20,26.24,23.76,14.50.HRMS(ESI+):calculated forC 63 H 88 N 12 O 13 [M+H] + 1221.6667,found1221.6647

[0155] Example 12 Preparation of Compound TK36

[0156]

[0157] Tetrakis-(4-hydroxyphenyl)ethylene (200 mg, 0.50 mmol), potassium carbonate (104 mg, 0.76 mmol) and 1-iodopropane (53 μL, 0.55 mmol) were added to a 100 mL round-bottom flask, fully dissolved with 8 mL of anhydrous methanol, and stirred at room temperature for 3 h. After the reaction was completed, it was extracted twice with ethyl acetate and water, the aqueous layer was discarded and the organic layer was collected, and the crude product was obtained after rotary evaporation and concentration. The crude product was purified by silica gel chromatography to obtain a light yellow oil product (40 mg, 18%).

[0158] 1 H NMR(400MHz,CD3OD)δ6.89-6.85(m,2H),6.82-6.76(m,6H),6.64-6.60(m,2H),6. 53-6.48(m,6H),3.84(t,J=6.5Hz,2H),1.79-1.68(m,2H),1.01(t,J=7.4Hz,3H). 13 CNMR(100MHz,CD3OD)δ158.75,156.64,156.62,156.61,139.95,139.43,138.38,137.36,137.34,137 .31,133.69,133.66,133.59,115.36,115.34,114.52,70.37,23.67,10.86.HRMS(ESI+):calculated for C 29 H 26 O4[M+H] + 439.1904, found 439.1866.

[0159] Example 13 Preparation of Compound TK37

[0160]

[0161] Compound TK36 (35 mg, 0.08 mmol) was fully dissolved in 8 mL of acetone in a 100 mL round-bottom flask, potassium carbonate (55 mg, 0.40 mmol) and ethyl bromoacetate (35 μL, 0.32 mmol) were added, and refluxed for 8 h. After the reaction, the mixture was extracted twice with ethyl acetate and water, the aqueous layer was discarded, and the organic layer was collected and concentrated by rotary evaporation to obtain a crude product, which was purified by silica gel chromatography to obtain a yellow-brown oil product (43 mg, 77%).

[0162] 1 H NMR(400MHz, CDCl3)δ6.96-6.84(m,8H),6.68-6.56(m,8H),4.53(s,6H),4.29-4.20(m, 6H),3.83(t,J=6.6Hz,2H),1.79-1.71(m,2H),1.30-1.25(m,9H),1.00(t,J=7.4Hz,3H). 13C NMR (100MHz, CDCl3) δ169.04,157.58,156.17,156.15,156.13,138.94,138.14,137.78,137.76,137.73,136.24,132.70,1 32.68,132.59,113.87,113.84,113.63,69.32,68.26,65.42,65.40,61.44,22.71,14.27,10.68.HRMS(ESI+):calculated forC 41 H 44 O 10 [M+H] + 697.3007, found 697.3005.

[0163] Example 14 Preparation of Compound TK38

[0164]

[0165] Compound TK37 (50 mg, 0.07 mmol) was fully dissolved in 4 mL of tetrahydrofuran and added to a 100 mL round-bottom flask. LiOH (24 mg, 1.00 mmol) dissolved in water (2 mL) was added. The reaction was stirred at room temperature until the hydrolysis was complete. 2 mL of glacial acetic acid was added to the reaction flask and stirred thoroughly. After the reaction was completed, it was extracted twice with n-butanol and water, the water layer was discarded, and the organic layer was collected. After rotary evaporation and concentration, it was placed in a vacuum drying oven and fully dried to obtain a crude intermediate product after hydrolysis.

[0166] After adding 8 mL of anhydrous DMF to the reaction bottle containing the crude intermediate product, add L-arginine methyl ester dihydrochloride (149 mg, 0.57 mmol), HATU (93 mg, 0.24 mmol), and DIPEA (81 μL, 0.49 mmol). Stir overnight at room temperature. After waiting for the reaction to be completed, add n-butanol to the reaction solution and extract twice with water. Concentrate the organic phase to obtain a crude product. The crude product is purified by preparative HPLC to obtain a yellow-green oil product (32 mg, 35%).

[0167] 1H NMR (400MHz, CD3OD) δ6.93(d,J=8.6Hz,6H),6.87(d,J=8.7Hz,2H),6.78-6.71(m,6H),6.64(d,J=8.6Hz,2H),4.5 7-4.49(m,9H),3.84(t,J=6.4Hz,2H),3.71(s,9H),3.25-3.12(m,6H),2.06-1.54(m,14H),1.01(t,J=7.4Hz,3H). 13 C NMR(100MHz,CD3OD)δ173.36,173.35,173.33,171.40,171.38,171.36,171.32,159.15,158.67 ,157.60,140.56,139.66,139.17,139.15,137.48,133.77,133.73,133.71,133.68,133.64,133 .55,133.53,115.19,115.15,115.14,115.11,114.69,101.33,70.37,67.97,57.89,52.99,52. 98,52.96,52.95,52.88,52.86,41.75,29.43,26.24,23.68,10.90.HRMS(ESI+):calculatedfor C 56 H 74 N 12 O 13 [M+H] + 1123.5571, found1123.5540.

[0168] Example 15 In vitro antibacterial activity test

[0169] In this example, the antibacterial activity of the tetraphenylethylene derivatives of the present invention against Gram-positive bacteria and Gram-negative bacteria was evaluated by determining the minimum inhibitory concentration (MIC value) by the broth dilution method according to the Clinical and Laboratory Standards Institute (CLSI) guidelines.

[0170] The strains used in this example: Staphylococcus aureus S. aureus ATCC29213, Escherichia coli E. coli ATCC25922, Acinetobacter baumannii A. baumannii ATCC 17978, Klebsiella pneumoniae K. pneumoniae ATCC10031 were purchased from Guangdong Huankai Microbiology Technology Co., Ltd. Methicillin-resistant Staphylococcus aureus MRSAN315 and methicillin-resistant Staphylococcus aureus MRSANCTC 10442 were provided by Qingyuan Hospital Affiliated to Guangzhou Medical University.

[0171] The specific operations are as follows:

[0172] The strain to be tested was inoculated into MHA medium and incubated in a 37°C incubator for 24 h. An appropriate amount of bacteria was picked up with a sterile inoculation loop and dispersed in PBS buffer and its OD 600 was adjusted to 0.1. Finally, the suspension was diluted with MHB to a density of about 1×10 6 CFU / mL. The compound to be tested was dissolved in DMSO / H2O to a 1 mg / mL stock solution, diluted with MHB and added to the bacterial suspension to make the working concentration of the compound on the 96-well plate 0.19-25 μg / mL. The 96-well plate was placed in a 37°C constant temperature incubator and incubated for 24 hours. The MIC value of the compound was determined by measuring OD600 with an enzyme reader. The experiment was repeated at least twice.

[0173] The antibacterial activity results are shown in Table 1. Among them, compounds TK07, TK09, TK24, and TK38 showed excellent in vitro antibacterial activity against Gram-positive bacteria and Gram-negative bacteria (including drug-resistant bacteria) (MIC=0.39-6.25μg / mL); compounds TK25, TK26, and TK35 showed excellent in vitro antibacterial activity against Gram-positive bacteria (including drug-resistant bacteria), and the antibacterial activity against Gram-negative bacteria was weaker than that against Gram-positive bacteria; among them, compound TK24 showed the best broad-spectrum antibacterial activity.

[0174] Table 1. In vitro antibacterial activity of tetraphenylethylene derivatives (μg / mL)

[0175]

[0176] Note: TK00 in the table refers to the raw material tetra-(4-hydroxystyrene)ethylene.

[0177] Example 16 In vitro cytotoxicity test

[0178] In this example, the CCK-8 method was used to determine the cytotoxicity of compound TK24 on mouse fibroblasts (NTCT clone 929) and human embryonic kidney cells (HEK293). 50 The specific method is as follows:

[0179] The cells in good condition were inoculated into a 96-well plate containing 100 μL of DMEM medium (containing 10% fetal bovine serum), placed in a 37°C (5% CO2) incubator for 24 hours, and then the medium was carefully removed. The compound TK24 solution prepared with DMSO / DMEM was slowly added to the experimental group wells to make the working concentration on the 96-well plate 3.125-100 μg / mL. The control group was not treated with drugs, and the blank group only contained culture medium. After continuing to culture for 24 hours under the same conditions as above, 10 μL of CCK-8 reagent was added to each well, and after continuing to incubate for 1 hour, the absorbance value at 450nm was recorded on the Biotek multi-function microplate reader. The final cell viability calculation formula: % cell viability = [(Abs 样品 -Abs 阴性对照 ) / (Abs 阳性对照 -Abs 阴性对照 )] × 100. This experiment was performed with at least two biological replicates.

[0180] The results are as follows Figure 1 As shown in Figure 2, after the cells were treated with 50 μg / mL of compound TK24, the survival rates of NCTC clone 929 cells and HEK293 cells remained at 65.67±4.14% and 78.36±3.99%, respectively. 50 The values ​​were all greater than 50 μg / mL, indicating that it has low cytotoxicity to mammalian cells and high biosafety.

[0181] Example 17 SYTOX Green Test

[0182] This example studies the cell membrane permeability of the compound to Gram-positive bacteria (S. aureus ATCC29213, MRSANCTC10442) and Gram-negative bacteria (E. coli ATCC25922, K. pneumoniae ATCC10031) by testing the fluorescence intensity change of the cell nucleic acid dye SYTOX Green. The specific operation is as follows:

[0183] Take a single colony and culture it in 25mL of MHB for 6-8h (37℃, 220rpm), then wash the bacteria 3 times with PBS buffer (5000rpm, 5min), prepare a bacterial suspension (OD600=0.2), add 0.3μM SYTOX Green dye under light-proof conditions, evenly disperse it in the suspension and add it to a 24-well plate. Monitor the fluorescence of the mixture using a Biotek multi-function microplate reader (excitation wavelength 504nm, emission wavelength 523nm). After the fluorescence signal stabilizes, different concentrations of compounds (dissolved in DMSO) are added to the experimental group to make their working concentrations 4 and 8 times the MIC, respectively, and the positive control group is treated with 0.5% Triton solution. Continue to monitor and record the changes in fluorescence intensity for about 1h. The experiment was repeated at least twice.

[0184] In this example, SYTOX Green staining was used to study the membrane-permeabilizing effect of compound TK24. Figure 2 As shown in the figure, after adding 4× and 8× MIC concentrations of compound TK24 to the bacterial solution of S. aureus ATCC29213, MRSANCTC10442, E. coli ATCC25922 and K. pneumoniae ATCC10031 containing SYTOX Green dye, the fluorescence intensity in the mixed solution increased significantly with the increase of drug concentration, while the fluorescence intensity of the control group (not treated with drug) remained basically unchanged. The results show that compound TK24 can damage the integrity of bacterial cell membranes, thereby achieving a bactericidal effect.

[0185] Example 18 Drug resistance development trend assessment test

[0186] This experiment was used to evaluate the tendency of MRSANCTC10442 and E. coliATCC25922 to develop resistance to compound TK24. The initial MIC values ​​of the compounds against the two bacteria were tested on a 96-well plate, and the control group drugs were norfloxacin and amoxicillin. The next day, the bacteria in the sub-inhibitory concentration treatment wells were selected to prepare a suspension (1×10 6 CFU / mL) for the next MIC value determination, and so on, repeat the test 21 times. A change in MIC value greater than 4 times is defined as bacterial resistance to the compound.

[0187] Test results such as Figure 3 As shown in the figure, on the 12th day, the MIC value of ciprofloxacin to MRSANCTC10442 increased to 32 times, indicating that MRSANCTC10442 quickly developed resistance to ciprofloxacin. The MIC value of compound TK24 remained within 4 times within 21 days, indicating that MRSANCTC10442 is not likely to develop bacterial resistance to compound TK24 ( Figure 3a); On the 9th day, the MIC value of amoxicillin against E. coli ATCC25922 increased to 8 times, while the MIC value of compound TK24 remained within 4 times within 21 days, indicating that E. coli ATCC25922 quickly developed resistance to amoxicillin, but was not easy to develop resistance to compound TK24 ( Figure 3 b). The above results indicate that compound TK24 is not likely to induce drug resistance in bacteria and has great potential in clinical applications.

[0188] Example 19 Evaluation of in vivo antibacterial activity

[0189] The in vivo efficacy evaluation experiment has been approved by the Experimental Animal Center of South China Agricultural University and was conducted in accordance with the policy of the Ministry of Health. Female C57BL / 6 mice of appropriate age (6-8 weeks) and uniform weight (18-20g) were selected, and an immunosuppressive mouse model was established by intraperitoneal injection of cyclophosphamide (100 mg / kg), with one day between each injection, for a total of 3 injections. S. aureus ATCC29213 inoculated on the MHA plate was evenly dispersed in 25 mL of MHB and cultured overnight (37°C, 220 rpm), and the bacteria were washed 3 times with PBS buffer (5000 rpm, 5 min) to prepare a bacterial suspension (OD600 = 1.0). After the mice were anesthetized with 2.5% avertin anesthetic (500 mg / kg), the right cornea of ​​the mouse was scratched with a sterile needle under a microscope, and the bacterial suspension (20 μL) was dripped into the injured corneal surface to establish a corneal infection model in mice. The next day, the infected mice were randomly divided into 4 groups (5 mice in each group) and treated with the corresponding solution (5% vancomycin, 0.3% compound TK24, 0.5% compound TK24 or 5% glucose) 4 times a day for 3 consecutive days. Finally, the mice were euthanized, the right cornea of ​​the mice was collected, and the number of live bacteria on the cornea was counted using the MHA plate counting method. This experiment was performed using SPSS25.0 software for independent sample t-tests, and p<0.05 was considered statistically significant.

[0190] The experimental results are as follows Figure 4As shown, compared with the negative control group, both the experimental group and the positive control group can significantly reduce the number of S. aureus ATCC29213 in the infected cornea. After treatment with 5% vancomycin, it was reduced by 5.89 log (p < 0.05); after treatment with 0.3% and 0.5% compound TK24, it was reduced by 3.74 log (p < 0.05) and 5.64 log (p < 0.05), respectively. These results show that under the condition that the therapeutic concentration is 10 times lower than that of vancomycin, 0.5% compound TK24 showed a therapeutic effect comparable to vancomycin in the treatment of the S. aureus ATCC29213 corneal infection model in mice, and after reducing the dose, 0.3% compound TK24 still showed an excellent bactericidal effect, with a bactericidal rate of 99.9%. It can be seen that compound TK24 has excellent in vivo antibacterial activity.

[0191] Example 20 Laser confocal imaging experiment

[0192] The "lighting up" effect of compound TK24 on MRSANCTC10442, E.coliATCC25922, A.baumannii R2889 and K.pneumoniaeATCC10031 was observed by laser confocal microscopy. Take appropriate amount of bacteria and place them in MHB for shaking culture for 6-8h (37°C, 220rpm). Wash twice with PBS buffer and adjust the OD value of the bacterial solution to 1.0, add compound TK24 prepared with DMSO to a final concentration of 10μM in the bacterial suspension. Incubate in a 37°C incubator for 30min. After incubation, wash twice with PBS buffer and resuspend. Pipette 2μL of bacterial solution onto a glass slide and observe its fluorescence image under a laser confocal microscope with an excitation wavelength of 405nm.

[0193] Compound TK24 can generate fluorescent signals after interacting with bacteria, and has the ability to "light up" bacteria. Figure 5 As shown, compound TK24 was incubated with MRSANCTC10442, E. coli ATCC25922, A. baumannii R2889 and K. pneumoniae ATCC10031 for 30 minutes and then observed under a laser confocal microscope. The results showed that compound TK24 could interact with all four bacteria to produce visible fluorescence. Among them, all Gram-positive bacteria could be "lit up" and the fluorescence signal was the strongest. In contrast, the fluorescence signal of Gram-negative bacteria was unevenly distributed, and some bacterial cells failed to be stained. Under the same conditions, the staining degree of Gram-positive bacteria was better than that of Gram-negative bacteria. Compound TK24 can quickly "light up" bacteria, causing them to emit visible fluorescence signals visible to the naked eye in a short time, providing a powerful tool for pathological studies of bacterial infections and applied to bacterial diagnosis and treatment.

[0194] The above-mentioned embodiments only express several implementation methods of the present invention, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the scope of the invention patent. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention shall be subject to the attached claims.

Claims

1. A tetraphenylethylene derivative having a structure represented by formula (I) or a pharmaceutically acceptable salt thereof or a stereoisomer thereof: in, R1 is selected from: C1-C 20 Alkyl, C3-C 20 Unsaturated chain hydrocarbon group; Each R2 is independently selected from: C1-C 20 Alkyl, R3 substituted C3-C 20 Unsaturated chain hydrocarbon group, C1-C6 alkyl substituted by R4; R3 is selected from: -NR5R6; R4 is selected from: -C(=O)NR7R8; R5 and R6 are independently selected from: hydrogen, C1-C6 alkyl; R7 is selected from: hydrogen, C1-C6 alkyl; R8 is selected from: 1 or more R9 substituted or unsubstituted C1-C 20 alkyl; R9 is selected from: hydrogen, C1-C6 alkoxycarbonyl, -C(=O)NR5R6, amino, C1-C6 alkylamino, 2. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 1, characterized in that: R1 is selected from: C1-C 12 Alkyl, C3-C 12 Unsaturated chain hydrocarbon group.

3. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 2, characterized in that: R1 is selected from: C3-C 10 Alkyl, C3-C8 unsaturated chain hydrocarbon group.

4. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 3, characterized in that: R1 is selected from: C3-C6 alkyl, C4-C6 unsaturated chain hydrocarbon group.

5. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 4, characterized in that: R1 is selected from: 3-methyl-2-butenyl, decyl, propyl.

6. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 1, characterized in that: Each R2 is independently selected from: C1-C 10 Alkyl, R3 substituted C3-C 10 Unsaturated chain hydrocarbon group, C1-C3 alkyl group substituted by R4.

7. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 6, characterized in that: Each R2 is independently selected from: C2-C6 alkyl substituted by R3, C3-C6 unsaturated chain hydrocarbon group substituted by R3, C1-C2 alkyl substituted by R4.

8. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 7, characterized in that: Each R2 is independently selected from the group consisting of: R3 substituted ethyl, R3 substituted propyl, R3 substituted butyl, R3 substituted pentyl, R4 substituted methyl, and R4 substituted ethyl.

9. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to any one of claims 1 to 8, characterized in that: R3 is selected from: -NR5R6; R4 is selected from: -C(=O)NR7R8; R5 and R6 are independently selected from: hydrogen, methyl, ethyl, propyl; R7 is selected from: hydrogen, methyl, ethyl, propyl; R8 is selected from: 1 or more R9 substituted or unsubstituted C1-C 10 alkyl; R9 is selected from: hydrogen, C1-C3 alkoxycarbonyl, -C(=O)NR5R6, amino, C1-C3 alkylamino, 10. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 9, characterized in that: R3 is selected from the group consisting of methylamino, dimethylamino, and diethylamino.

11. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 9, characterized in that: R8 is selected from: 1 or more R9 substituted or unsubstituted C1-C6 alkyl; R9 is selected from: hydrogen, methoxycarbonyl, -C(=O)NH2, 12. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 11, characterized in that: R8 is selected from: one or more R9 substituted or unsubstituted ethyl groups, one or more R9 substituted or unsubstituted propyl groups, and one or more R9 substituted or unsubstituted butyl groups.

13. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 9, characterized in that: -NR7R8 is selected from the following structures:

14. The tetraphenylethylene derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 1, characterized in that: The tetraphenylethylene derivative is selected from:

15. Use of the tetraphenylethylene derivative according to any one of claims 1 to 14, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof in the preparation of a drug for inhibiting bacterial growth, preventing bacterial infection, or treating bacterial infection.

16. The use according to claim 15, characterized in that The bacteria are Gram-positive bacteria and / or Gram-negative bacteria.

17. The use according to claim 16, characterized in that The bacteria are Staphylococcus, Enterococcus, Enterobacter and Acinetobacter.

18. The use according to claim 17, characterized in that The bacteria are Staphylococcus aureus, methicillin-resistant Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Acinetobacter baumannii, and Klebsiella pneumoniae.

19. An antibacterial drug, characterized in that: The invention is prepared from active ingredients and pharmaceutically acceptable excipients, wherein the active ingredients include the tetraphenylethylene derivative according to any one of claims 1 to 14 or a pharmaceutically acceptable salt thereof or a stereoisomer thereof.

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