Mercaptophenol derivatives and their applications

Abstract

The present invention relates to a mercaptophenol derivative having a structure shown in formula (I) or a pharmaceutically acceptable salt thereof or a stereoisomer thereof, and use thereof in the preparation of an antibacterial drug. The mercaptophenol 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, and low drug resistance.

Description

Technical Field

[0001] The present invention belongs to the field of medical technology, and in particular relates to a mercaptophenol derivative and a preparation method and application thereof. Background Art

[0002] Currently, drug-resistant bacterial infections are increasing in clinical practice and the mortality rate is gradually increasing. The therapeutic effect of traditional antibiotics on drug-resistant bacterial infections is becoming increasingly limited and can no longer meet the needs of modern clinical treatment. The frequent emergence of drug-resistant bacteria, even multidrug-resistant bacteria, has become a pressing issue in global public health. The development of new antimicrobial drugs to meet this challenge is particularly urgent. Therefore, it is crucial to design antimicrobial drugs that have a different mechanism of action from existing antibiotics, possess new molecular entities, and are less likely to induce bacterial resistance. Summary of the Invention

[0003] Based on this, the purpose of the present invention is to provide a new antibacterial drug.

[0004] In order to achieve the above-mentioned object of the invention, the present invention includes the following technical solutions.

[0005] The present invention provides a mercaptophenol derivative having a structure represented by formula (I), or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof:

[0006]

[0007] Wherein, R1 is selected from: C1-C 20 Alkyl, C3-C 20 Unsaturated chain hydrocarbon group, halogenated C1-C 20 Alkyl, halogenated C3-C 20 Unsaturated chain hydrocarbon group;

[0008] R2 is selected from: R3 substituted C1-C 20 Alkyl, R3 substituted C3-C 20 Unsaturated chain hydrocarbon group, C1-C6 alkyl substituted by R4;

[0009] R3 is selected from: -NHR5, -NR6R7,

[0010] R4 is selected from: -C(=O)R8;

[0011] R5 is selected from: C1-C6 alkyl, R9 substituted C1-C6 alkyl;

[0012] R6 and R7 are independently selected from: C1-C6 alkyl, or R6, R7 together with the nitrogen atom to which they are connected form R 10 a substituted or unsubstituted 5-6 membered heterocyclic group;

[0013] R8 is selected from: 11 A substituted arginine group, a polypeptide group formed by 2-5 arginines, wherein the carbon terminal hydroxyl group of the polypeptide is replaced by R 11 Substitution; the arginine group or polypeptide group in R8 refers to the group after the nitrogen terminal amino group of the corresponding arginine or polypeptide loses a hydrogen;

[0014] R9 is selected from the group consisting of amino, C1-C6 alkylamino, (C1-C6 alkyl)2amino, aminoC1-C6 alkylamino, (aminoC1-C6 alkyl)2amino;

[0015] R 10 Selected from: hydrogen, C1-C6 alkyl;

[0016] R 11 Selected from: C1-C6 alkoxy, amino, C1-C6 alkylamino, (C1-C6 alkyl)2amino, benzylamino, cyano-substituted benzylamino,

[0017] In some embodiments, R1 is selected from: C4-C 12 Alkyl, C4-C 12 Unsaturated chain hydrocarbon group, halogenated C4-C 12 Alkyl, halogenated C4-C 12 Unsaturated chain hydrocarbon group.

[0018] In some embodiments, R1 is selected from: C6-C 10 Alkyl, C6-C 10 Unsaturated chain hydrocarbon group, halogenated C6-C 10 Alkyl, halogenated C6-C 10 Unsaturated chain hydrocarbon group.

[0019] In some embodiments, R1 is selected from: C7-C9 alkyl, C7-C9 unsaturated chain hydrocarbon group, halogenated C7-C9 alkyl group, halogenated C7-C9 unsaturated chain hydrocarbon group.

[0020] In some embodiments, R1 is selected from the group consisting of: 3,3,4,4,5,5,6,6,7,7,8,8,8-tridecafluorooctyl.

[0021] In some embodiments, R2 is selected from: R3 substituted C1-C 10 Alkyl, R3 substituted C3-C 10 Unsaturated chain hydrocarbon group, C1-C3 alkyl group substituted by R4.

[0022] In some embodiments, R2 is selected from: C2-C6 alkyl substituted by R3, C3-C6 unsaturated chain hydrocarbon group substituted by R3, C1-C2 alkyl substituted by R4.

[0023] In some embodiments, R2 is selected from: R3 substituted ethyl, R3 substituted propyl, R3 substituted butyl, R3 substituted pentyl, R4 substituted methyl, R4 substituted ethyl.

[0024] In some embodiments, R3 is selected from: -NHR5, -NR6R7,

[0025] R5 is selected from the group consisting of: methyl, ethyl, propyl, butyl, R9-substituted methyl, R9-substituted ethyl, and R9-substituted propyl;

[0026] R6 and R7 are independently selected from: methyl, ethyl, or R6, R7 together with the nitrogen atom to which they are connected form R 10 substituted or unsubstituted piperidinyl, piperazinyl or tetrahydropyrrolyl.

[0027] In some embodiments, R9 is selected from the group consisting of amino, methylamino, dimethylamino, and (aminoethyl)2amino.

[0028] In some embodiments, R 10 Selected from: hydrogen, methyl, ethyl.

[0029] In some embodiments, R3 is selected from the group consisting of methylamino, ethylamino, propylamino, butylamino, dimethylamino, diethylamino, 4-methylpiperazinyl, tetrahydropyrrolyl,

[0030] In some embodiments, R4 is selected from: -C(=O)R8;

[0031] R8 is selected from: -N(H)-R 12 -C(=O)R 11 ;

[0032] R 12 The residue is selected from: the residue after removing the carbon terminal carboxyl group and the nitrogen terminal amino group of arginine, and the residue after removing the carbon terminal carboxyl group and the nitrogen terminal amino group of a polypeptide formed by 2-4 arginines.

[0033] In some embodiments, R 11 Selected from: methoxy, ethoxy, propoxy, amino, methylamino, dimethylamino, benzylamino, cyano-substituted benzylamino,

[0034] In some embodiments, R8 is selected from the following structures:

[0035]

[0036] The present invention also provides the use of the mercaptophenol derivative or its pharmaceutically acceptable salt or its stereoisomer, including the following:

[0037] The present invention relates to the use of the mercaptophenol derivative or its pharmaceutically acceptable salt or its stereoisomer in the preparation of a drug for inhibiting bacterial growth.

[0038] The present invention relates to the use of the mercaptophenol derivative or its pharmaceutically acceptable salt or its stereoisomer in the preparation of a drug for preventing and / or treating bacterial infection.

[0039] In some embodiments, the bacteria are Gram-positive bacteria and / or Gram-negative bacteria, preferably Gram-positive bacteria.

[0040] In some embodiments, the bacteria are Staphylococcus, Enterococcus, Enterobacter, or Acinetobacter.

[0041] In some embodiments, the bacteria are Staphylococcus aureus, methicillin-resistant Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Acinetobacter baumannii, or Klebsiella pneumoniae.

[0042] The present invention also provides an antibacterial drug prepared from an active ingredient and pharmaceutically acceptable excipients, wherein the active ingredient comprises the mercaptophenol derivative of the present invention or a pharmaceutically acceptable salt thereof or a stereoisomer thereof.

[0043] The present invention uses mercaptophenol as a basic skeleton to synthesize a series of mercaptophenol derivatives with antibacterial activity. The mercaptophenol derivatives of the present invention introduce hydrophilic groups such as amino, guanidine and arginine, which can be protonated and have a positive charge under physiological conditions, on the phenolic hydroxyl group, providing a positive charge for the mercaptophenol derivatives, which then interact with negatively charged lipoteichoic acid (LTA), lipopolysaccharide (LPS) and bacterial membranes through electrostatic interaction; and the (halogenated) alkylthio chain in the molecular structure constitutes a hydrophobic side chain, which can drive the compound to insert into the lipid bilayer of the bacterial membrane, resulting in the destruction of the permeability and integrity of the bacterial membrane, triggering the leakage of bacterial cell contents and leading to bacterial death. The mercaptophenol derivatives of the present invention have 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, and low drug resistance.

[0044] Compound 21, the most preferred compound of this invention, exhibits excellent antibacterial activity, high safety, rapid bactericidal ability, and salt ion stability. It can inhibit bacterial biofilm formation and disrupt established biofilms, killing bacteria through membrane-targeted action and overcoming bacterial resistance. It demonstrated excellent efficacy in a mouse corneal infection model caused by Staphylococcus aureus and has great potential for clinical application. BRIEF DESCRIPTION OF THE DRAWINGS

[0045] Figure 1 The in vitro cytotoxicity of compound 21 on human embryonic kidney HEK293T cells (a) and mouse fibroblast NCTC clone 929 cells (b).

[0046] Figure 2 This is a study on the drug resistance of compound 21 to S. aureus ATCC 29213, with norfloxacin as the control group.

[0047] Figure 3 The change in SYTOX Green fluorescence intensity caused by the action of compound 21 on MRSANCTC 10442.

[0048] Figure 4 Fluorescence images of S. aureus ATCC29213 treated with PBS (a) and compound 21 (b).

[0049] Figure 5 The in vivo antibacterial efficacy of compound 21 (0.5%) in the S. aureus ATCC 29213-induced mouse keratitis model was evaluated. Vancomycin (5%) was used as a positive control, and 5% glucose injection was used as a negative control. All data are expressed as mean ± SD (n = 5). **P < 0.05 indicates statistically significant, and ns indicates no statistically significant difference. DETAILED DESCRIPTION

[0050] To facilitate 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 present disclosure more thorough and comprehensive.

[0051] In the following examples, the experimental methods without specific conditions are generally 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.

[0052] Unless otherwise defined, all technical and scientific terms used herein have the same meanings as commonly understood by those skilled in the art to which this invention pertains. The terms used in this specification are for the purpose of describing specific embodiments only and are not intended to limit the invention. The term "and / or" as used herein includes any and all combinations of one or more of the associated listed items.

[0053] Furthermore, as used herein, the term "or" is inclusive and 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. Furthermore, throughout this specification, the meanings of "a," "an," and "the" include plural referents. The meaning of "in" includes "in" and "on."

[0054] 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, a mercaptophenol derivative having a structure represented by formula (I) or a pharmaceutically acceptable salt thereof or a stereoisomer thereof:

[0055]

[0056] Wherein, R1 is selected from: C1-C 20 Alkyl, C3-C 20 Unsaturated chain hydrocarbon group, halogenated C1-C 20 Alkyl, halogenated C3-C 20 Unsaturated chain hydrocarbon group;

[0057] R2 is selected from: R3 substituted C1-C 20 Alkyl, R3 substituted C3-C 20 Unsaturated chain hydrocarbon group, C1-C6 alkyl substituted by R4;

[0058] R3 is selected from: -NHR5, -NR6R7,

[0059] R4 is selected from: -C(=O)R8;

[0060] R5 is selected from: C1-C6 alkyl, R9 substituted C1-C6 alkyl;

[0061] R6 and R7 are independently selected from: C1-C6 alkyl, or R6, R7 together with the nitrogen atom to which they are connected form R 10 a substituted or unsubstituted 5-6 membered heterocyclic group;

[0062] R8 is selected from: 11 A substituted arginine group, a polypeptide group formed by 2-5 arginines, wherein the carbon terminal hydroxyl group of the polypeptide is replaced by R 11 replace;

[0063] R9 is selected from the group consisting of amino, C1-C6 alkylamino, (C1-C6 alkyl)2amino, aminoC1-C6 alkylamino, (aminoC1-C6 alkyl)2amino;

[0064] R 10 Selected from: hydrogen, C1-C6 alkyl;

[0065] R 11 Selected from: C1-C6 alkoxy, amino, C1-C6 alkylamino, (C1-C6 alkyl)2amino, benzylamino, cyano-substituted benzylamino,

[0066] In the compounds described herein, when any variable (e.g., R3, etc.) occurs more than once in any component, its definition at each occurrence is independent of the definition at every other occurrence. Similarly, combinations of substituents and variables are permitted so long as such combinations stabilize the compound. It will be appreciated 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 using techniques in the art and the methods set forth below. If a substituent is itself substituted with more than one group, it will be appreciated that these groups may be on the same carbon atom or on different carbon atoms, so long as the structure is stable.

[0067] The term "alkyl" is intended to include both branched and straight-chain saturated aliphatic hydrocarbon groups having a specified number of carbon atoms. For example, the definition of "C1-C6 alkyl" includes groups having 1, 2, 3, 4, 5, or 6 carbon atoms in a straight or branched chain. For example, "C1-C6 alkyl" specifically includes methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, pentyl, and hexyl.

[0068] The term "unsaturated chain hydrocarbon group" refers to a branched or straight chain unsaturated aliphatic hydrocarbon group having a specific number of carbon atoms, i.e., a non-cyclic chain hydrocarbon group, and the carbon chain contains one or more carbon-carbon double bonds, or a carbon-carbon triple bond, such as:

[0069] -CH2(CH=CH)(CH2)2CH3, -CH2(CH=CH)(CH3)CH3, etc.

[0070] The term "alkoxy" refers to a group where an alkyl group is directly connected to oxygen, i.e., a group having an -O-alkyl structure, such as -OCH3, -OCH2CH3, -OCH2CH2CH3, -OCH2CH(CH3)2, -OCH2CH2CH2CH3, -O-CH(CH3)2, etc.

[0071] The present invention includes free forms of compounds of Formula I, as well as pharmaceutically acceptable salts and stereoisomers thereof. Certain specific exemplary compounds of the present invention are amine compounds in non-salt form, i.e., "free form." 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 a suitable solvent or combination of solvents. Similarly, salts of acidic compounds are formed by reaction with a suitable inorganic or organic base.

[0072] Therefore, pharmaceutically acceptable salts of the compounds of this invention include conventional non-toxic salts of the compounds of this invention formed by reacting an alkaline compound of this invention with an inorganic or organic acid. 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, and the like, 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, and the like.

[0073] If the compound of the present invention is acidic, 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, ferric salts, ferrous salts, lithium salts, magnesium salts, manganic salts, manganous salts, potassium salts, sodium salts, zinc salts, and the like. 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, guaiac, polyamine resins, procaine, purine, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine and the like.

[0074] Berg et al., "Pharmaceutical Salts," J. Pharm. Sci. '1977: 66: 1-19, describes in more detail the preparation of the pharmaceutically acceptable salts described above and other typical pharmaceutically acceptable salts.

[0075] The stereoisomers according to the present invention are, depending on their structure, 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 being preferred.

[0076] In another embodiment of the present invention, there is provided use of the mercaptophenol derivative, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof in the preparation of a drug for inhibiting bacterial growth.

[0077] In another embodiment of the present invention, there is provided use of the mercaptophenol derivative or a pharmaceutically acceptable salt or a stereoisomer thereof in the preparation of a medicament for treating and / or preventing bacterial infection.

[0078] The bacteria described in the present invention can be non-resistant bacteria or drug-resistant bacteria, such as single-drug resistant bacteria or multidrug resistant bacteria; they can be Gram-positive bacteria or Gram-negative bacteria, and the antibacterial effect on Gram-positive bacteria is particularly better.

[0079] In another embodiment of the present invention, an antibacterial drug is provided, which is prepared from an active ingredient and a pharmaceutically acceptable excipient. The active ingredient comprises the mercaptophenol derivative described in the present invention, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof.

[0080] Among them, the amount of active ingredients contained is within the safe and effective amount range. "Safe and effective amount" means that the amount of active ingredients is sufficient to significantly improve the condition without causing serious side effects.

[0081] "Pharmaceutically acceptable excipients" refer to: one or more compatible solid or liquid fillers or gel substances that are suitable for human use and must be sufficiently pure and have sufficiently low toxicity.

[0082] "Compatibility" herein means that the components of the composition or preparation can be mixed with the active ingredient of the present invention and with each other without significantly reducing the efficacy of the active ingredient.

[0083] 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, flavorings, stabilizers, antioxidants, preservatives, pyrogen-free water, etc.

[0084] The administration method of the active ingredient, pharmaceutical composition, or pharmaceutical preparation of the present invention is not particularly limited. Representative administration methods include, but are not limited to, oral, transdermal, rectal, and parenteral (intravenous, intramuscular, or subcutaneous) administration. Specifically, 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 formulations, and injections.

[0085] 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:

[0086] (a) fillers or extenders, for example, starch, lactose, sucrose, glucose, mannitol and silicic acid;

[0087] (b) binders, for example, hydroxymethylcellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose and gum arabic;

[0088] (c) humectants, for example, glycerin;

[0089] (d) disintegrants, for example, agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate;

[0090] (e) a buffering solvent, for example, paraffin;

[0091] (f) absorption accelerators, for example, quaternary ammonium compounds;

[0092] (g) wetting agents, for example, cetyl alcohol and glyceryl monostearate;

[0093] (h) adsorbents, for example, kaolin; and

[0094] (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.

[0095] The solid dosage forms can also be prepared using coatings and shells, 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 portion of the digestive tract. Examples of embedding components that can be used are polymeric substances and waxes.

[0096] 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 commonly used in the art, such as water or other solvents, solubilizers and emulsifiers, for example, ethanol, isopropyl alcohol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butylene glycol, dimethylformamide, and oils, particularly cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, and sesame oil, or mixtures thereof. In addition to these inert diluents, the composition may also contain adjuvants, such as wetting agents, emulsifiers and suspending agents, sweeteners, flavoring agents, and fragrances.

[0097] In addition to the active ingredients, suspensions may contain suspending agents such as, for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these substances.

[0098] Compositions for parenteral injection may comprise 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.

[0099] The present invention is further described in detail below with reference to specific embodiments.

[0100] The synthesis method of mercaptophenol derivatives refers to the following synthesis routes 1-4.

[0101]

[0102] Synthesis Route 1

[0103]

[0104] Synthesis Route 2

[0105]

[0106] Synthesis Route 3

[0107]

[0108] Synthesis Route 4

[0109] Example 1 Preparation of Compound 01

[0110]

[0111] 4-(3,3,4,4,5,5,6,6,7,7,8,8,8-tridecafluorooctylthio)phenol (100.0 mg, 0.21 mmol) was mixed with anhydrous potassium carbonate (87.8 mg, 0.64 mmol) and thoroughly dissolved in 8 mL of anhydrous DMF. 1,4-Dibromobutane (51 μL, 0.42 mmol) was added dropwise with stirring and allowed to react at room temperature for 5 hours. The reaction mixture was then diluted with water and extracted twice with n-butanol. The crude product was concentrated under reduced pressure to obtain the crude product. The crude product was purified by silica gel chromatography using a 50:1 volume ratio of petroleum ether to ethyl acetate as the eluent and dried under reduced pressure to obtain the product 01 (107.1 mg) as a colorless oil in an 83% yield.

[0112] 1 H NMR (400MHz, CDCl3) δ7.37(d,J=8.5Hz,2H),6.86(d,J=8.5Hz,2H),3.99(t,J=5.6Hz, 2H),3.49(t,J=6.3Hz,2H),3.02-2.92(m,2H),2.43-2.22(m,2H),2.11-1.87(m,4H). 13 C NMR(100MHz, CDCl3)δ159.02,134.23,124.35,115.47,67.06,33.45,29.50,27.91,26.84,26.01.HRMS(ESI+):calculated for C 18 H 16 BrF 13 OS[M+NH4] + 624.0236,found624.9899.

[0113] Example 2 Preparation of Compound 02

[0114]

[0115] Compound 01 (50.0 mg, 0.08 mmol) was thoroughly dissolved in 8 mL of anhydrous DMF. 1 mL of ethylamine solution (68.0-72.0% in H₂O) was added dropwise with stirring and allowed to react at room temperature for 24 hours. The reaction mixture was then diluted with water and extracted twice with n-butanol. The crude product was concentrated under reduced pressure to obtain the crude product. The crude product was purified by preparative HPLC and dried under reduced pressure to yield the desired product 02 (32.9 mg) as a yellow gel in a 70% yield.

[0116] 1H NMR (400MHz, CD3OD) δ7.38(d,J=8.8Hz,2H),6.90(d,J=8.8Hz,2H),3.98(t,J=6.1Hz,2H),3.04 -2.93(m,2H),2.73-2.61(m,4H),2.42-2.26(m,2H),1.83-1.66(m,4H),1.13(t,J=7.2Hz,3H). 13 C NMR(100MHz,CD3OD)δ160.62,135.42,125.55,116.46,68.78,49.83,44.60,32.67,28.00,27.50,26.65,14.15.HRMS(ESI+):calculated forC 20 H 22 F 13 NOS[M+H] + 572.1287, found 572.1303.

[0117] Example 3 Preparation of Compound 03

[0118]

[0119] The synthesis of compound 03 was carried out according to the synthesis method of compound 02, using compound 01 (50.0 mg, 0.08 mmol) and n-butylamine (25 μL, 0.25 mmol) to prepare the white waxy target product 03 (26.7 mg) with a yield of 53%.

[0120] 1 H NMR (400MHz, CD3OD) δ7.38(d,J=8.4Hz,2H),6.90(d,J=8.4Hz,2H),3.98(t,J=6.1Hz,2H),3.05-2.94(m,2H),2.64(t,J=7.4H z,2H),2.61-2.55(m,2H),2.43-2.22(m,2H),1.84-1.64(m,4H),1.55-1.43(m,2H),1.40-1.26(m,2H),0.92(t,J=7.3Hz,3H). 13 CNMR(100MHz,CD3OD)δ160.65,135.40,125.55,116.50,101.34,68.88,50.34,5 0.30,32.70,32.43,28.09,27.53,26.89,21.52,14.31.HRMS(ESI+):calculated for C 22 H26 F 13 NOS[M+H] + 600.1600,found 600.1602.

[0121] Example 4 Preparation of Compound 04

[0122]

[0123] The synthesis of compound 04 was carried out according to the synthesis method of compound 02, using compound 01 (50.0 mg, 0.08 mmol) and n-octylamine (25 μL, 0.25 mmol) to prepare the yellow gel target product 04 (34.1 mg) with a yield of 63%.

[0124] 1 H NMR (400MHz, CD3OD) δ7.40(d,J=7.2Hz,2H),6.93(d,J=7.2Hz,2H),4.01(t,J=5.9Hz,2H),3.09-2.94(m,2H),2.66(dt,J =27.7,7.5Hz,4H),2.47-2.24(m,2H),1.87-1.66(m,4H),1.60-1.45(m,2H),1.36-1.26(m,10H),0.90(t,J=6.3Hz,3H). 13 C NMR(100MHz,CD3OD)δ160.63,135.40,125.58,116.50,101.34,68.84,50.52,50.16,33.00,32 .71,30.60,30.37,30.06,28.38,28.03,27.52,26.70,23.72,14.43.HRMS(ESI+):calculated for C 26 H 34 F 13 NOS[M+H] + 656.2226, found 656.2227.

[0125] Example 5 Preparation of Compound 05

[0126]

[0127] Compound 05 was synthesized according to the synthesis method of compound 02, using compound 01 (50.0 mg, 0.08 mmol) and 1 mL of dimethylamine solution (2 M in THF) to obtain the target product 05 (33.5 mg) as a light yellow gel with a yield of 71%.

[0128] 1 H NMR (400MHz, CD3OD) δ7.40 (d, J = 8.9 Hz, 2H), 6.93 (d, J = 8.9 Hz, 2H), 4.01 (t, J = 6. 1Hz,2H),3.05-2.95(m,2H),2.47-2.32(m,4H),2.29(s,6H),1.86-1.65(m,4H). 13 C NMR(100MHz,CD3OD)δ160.65,135.41,125.54,116.50,68.79,60.26,45.25,32.69,28.16,27.51,24.80.HRMS(ESI+):calculated for C 20 H 22 F 13 NOS[M+H] + 572.1287,found572.1289.

[0129] Example 6 Preparation of Compound 06

[0130]

[0131] The synthesis of compound 06 was carried out according to the synthesis method of compound 02, using compound 01 (50.0 mg, 0.08 mmol) and diethylamine (18 μL, 0.16 mmol) to prepare the target product 06 (33.5 mg) as a light yellow gel with a yield of 61%.

[0132] 1 H NMR (400MHz, CD3OD) δ7.40(d,J=8.8Hz,2H),6.93(d,J=8.8Hz,2H),4.01(t,J=6.1Hz,2H),3.06 -2.98(m,2H),2.62-2.51(m,6H),2.45-2.30(m,2H),1.81-1.61(m,4H),1.06(t,J=7.2Hz,6H). 13 C NMR(100MHz,CD3OD)δ160.67,135.43,125.52,116.50,68.85,53.34,47.65,28.37,27.52,23.70,11.14.HRMS(ESI+):calculated for C 22 H 26 F 13 NOS[M+H] + 600.1600,found 600.1601.

[0133] Example 7 Preparation of Compound 07

[0134]

[0135] The synthesis of compound 07 was carried out according to the synthesis method of compound 02, using compound 01 (50.0 mg, 0.08 mmol) and dipropylamine (23 μL, 0.16 mmol) to obtain the target product 07 (31.1 mg) as a colorless oil with a yield of 60%.

[0136] 1 H NMR (400MHz, CD3OD) δ7.40(d,J=8.7Hz,2H),6.93(d,J=8.7Hz,2H),4.01(t,J=6.1Hz,2H),3.06-2.96(m,2H),2.64 -2.53(m,2H),2.52-2.31(m,6H),1.82-1.74(m,2H),1.72-1.63(m,2H),1.56-1.47(m,4H),0.90(t,J=7.4Hz,6H). 13 C NMR (100MHz, CD3OD) δ160.66,135.44,125.46,116.48,68.82,57.04,54.63,32.67,28.29,27.50,23.79,20.41,12.21.HRMS(ESI+):calculated for C 24 H 30 F 13 NOS[M+H] + 628.1913, found 628.1914.

[0137] Example 8 Preparation of Compound 08

[0138]

[0139] The synthesis of compound 08 was carried out according to the synthesis method of compound 02, using compound 01 (50.0 mg, 0.08 mmol) and n-dibutylamine (42 μL, 0.16 mmol) to prepare the target product 08 (31.9 mg) as a colorless oil with a yield of 59%.

[0140] 1H NMR (400MHz, CD3OD) δ7.40(d,J=8.8Hz,2H),6.93(d,J=8.8Hz,2H),4.01(t,J=6.1Hz,2H),3.05-2.97(m, 2H),2.59-2.26(m,8H),1.83-1.61(m,4H),1.51-1.41(m,4H),1.37-1.26(m,4H),0.93(t,J=7.3Hz,6H). 13 C NMR (100MHz, CD3OD) δ160.66,135.44,125.48,116.48,68.77,54.73,54.49,32.68,29.40,28.25,27.50,23.67,21.82,14.39.HRMS(ESI+):calculated for C 26 H 34 F 13 NOS[M+H] + 656.2226, found 656.2245.

[0141] Example 9 Preparation of Compound 09

[0142]

[0143] The synthesis of compound 09 was carried out according to the synthesis method of compound 02, using compound 01 (50.0 mg, 0.08 mmol) and thiomorpholine (25 μL, 0.25 mmol) to prepare the target product 09 (36.9 mg) as a brown gel with a yield of 71%.

[0144] 1 H NMR (400MHz, CD3OD) δ7.40(d,J=8.0Hz,2H),6.93(d,J=8.1Hz,2H),4.01(t,J=5.8Hz,2H),3.06-2.97( m,2H),2.88-2.78(m,4H),2.74-2.66(m,4H),2.58-2.49(m,2H),2.46-2.29(m,2H),1.85-1.67(m,4H). 13 C NMR(100MHz,CD3OD)δ160.62,135.41,125.54,116.47,68.73,59.90,56.06,32.66,28.19,27.97,27.50,23.52.HRMS(ESI+):calculated for C 22 H 24 F 13NOS2[M+H] + 630.1164, found 630.1167.

[0145] Example 10 Preparation of Compound 10

[0146]

[0147] The synthesis of compound 10 was carried out according to the synthesis method of compound 02, using compound 01 (50.0 mg, 0.08 mmol) and N-methylpiperazine (27 μL, 0.25 mmol) to prepare the yellow gel target product 10 (38.9 mg) with a yield of 75%.

[0148] 1 H NMR (400MHz, CD3OD) δ7.46-7.34(m,2H),6.99-6.87(m,2H),4.01(t,J=5.8Hz,2H),3.07-2.97(m,2H),2.91-2.27(m,15H),1.85-1.66(m,4H). 13 C NMR(100MHz,CD3OD)δ160.63,135.40,125.57,116.49,68.72,58.81,55.11,53.06,45.41,32.68,28.11,27.51,23.89.HRMS(ESI+):calculated for C 23 H 27 F 13 N2OS[M+H] + 627.1709, found 627.1713.

[0149] Example 11 Preparation of Compound 11

[0150]

[0151] The synthesis of compound 11 was carried out according to the synthesis method of compound 02, using compound 01 (50.0 mg, 0.08 mmol) and tetrahydropyrrole (21 μL, 0.25 mmol) to prepare the target product 11 (32.2 mg) as a brown gel with a yield of 65%.

[0152] 1H NMR (400MHz, CD3OD) δ7.41(d,J=8.9Hz,2H),6.94(d,J=8.9Hz,2H),4.03(t,J=5.5Hz,2H),3.06-2.98 (m,2H),2.95(t,J=6.6Hz,4H),2.91-2.84(m,2H),2.47-2.29(m,2H),1.94(M,4H),1.88-1.77(m,4H). 13 C NMR(100MHz,CD3OD)δ160.53,135.39,125.70,116.49,68.52,56.64,55.00,32.67,27.89,27.49,25.25,24.05.HRMS(ESI+):calculated for C 22 H 24 F 13 NOS[M+H] + 598.1444,found 598.1444.

[0153] Example 12 Preparation of Compound 13

[0154]

[0155] Compound 01 (50.0 mg, 0.08 mmol) was fully dissolved in 8 mL of anhydrous DMF, and 2 mL of ammonia-methanol solution (7 M in MeOH) was added dropwise under stirring. The reaction was allowed to proceed at 50-55°C for 24 hours. After the reaction, the residual ammonia was completely evaporated, and the mixture was concentrated under reduced pressure and dried thoroughly to obtain amine compound 12. Amine compound 12 was mixed with 1H-pyrazole-1-carboximidamide hydrochloride (40.5 mg, 0.28 mmol), fully dissolved in 8 mL of anhydrous DMF, and DIPEA (92 μL, 0.50 mmol) was slowly added dropwise under stirring. The reaction was allowed to proceed overnight at room temperature. The reaction mixture was then diluted with water and extracted twice with n-butanol. The mixture was concentrated under reduced pressure to obtain the crude product. The crude product was purified by preparative HPLC and dried under reduced pressure to obtain the target product 13 (29.7 mg) as a white solid in a yield of 55%.

[0156] 1 H NMR(400MHz,CD3OD)δ7.40(d,J=5.8Hz,2H),6.93(d,J=5.8Hz,2H),4.12-3.91(m, 2H),3.27-3.17(m,2H),3.08-2.91(m,2H),2.48-2.23(m,2H),1.93-1.68(m,4H). 13CNMR(100MHz,CD3OD)δ160.56,158.76,135.38,125.72,116.49,68.57,42.18,32.70,27.51,27.43,26.75.HRMS(ESI+):calculated for C 19 H 20 F 13 N3OS[M+H] + 586.1192,found586.1200.

[0157] Example 13 Preparation of Compound 14

[0158]

[0159] The synthesis of compound 14 was carried out according to the synthesis method of compound 02, using compound 01 (50.0 mg, 0.08 mmol) and tris(2-aminoethyl)amine (38 μL, 0.25 mmol) to prepare the yellow gel target product 14 (32.5 mg) with a yield of 58%.

[0160] 1 H NMR (400MHz, CD3OD) δ8.53(s,3H),7.41(d,J=8.8Hz,2H),6.94(d,J=8.8Hz,2H),4.04(t,J=5.6Hz,2H),3.17-3.0 5(m,8H),3.05-2.98(m,2H),2.83(t,J=5.8Hz,2H),2.78(t,J=5.7Hz,4H),2.46-2.29(m,2H),1.99-1.83(m,4H). 13 C NMR(100MHz,CD3OD)δ169.18,163.09,159.13,134.04,124.50,115.20,66.99,53.00,51.16,50.35,50.16,48.33,4 8.11,47.90,47.69,47.47,47.26,47.05,45.13,44.94,36.72,35.31,31.36,26.14,22.93.HRMS(ESI+):calculated for C 24 H 33 F 13 N4OS[M+H] + 673.2240,found673.2239.

[0161] Example 14 Preparation of Compound 15

[0162]

[0163] 4-(3,3,4,4,5,5,6,6,7,7,8,8,8-tridecafluorooctylthio)phenol (100.0 mg, 0.21 mmol) was mixed with anhydrous potassium carbonate (87.8 mg, 0.64 mmol) and dissolved in 8 mL of anhydrous DMF. Ethyl bromoacetate (47 μL, 0.42 mmol) was added dropwise with stirring and allowed to react at room temperature for 1.5 hours. The reaction mixture was then diluted with water and extracted twice with n-butanol. The crude product was concentrated under reduced pressure to obtain the crude product. The crude product was purified by silica gel chromatography using a 4:1 volume ratio of petroleum ether to ethyl acetate as the eluent and dried under reduced pressure to obtain the product 15 (104.8 mg) as a colorless oil in an 89% yield.

[0164] 1 H NMR(400MHz, CDCl3) δ7.37(d,J=8.1Hz,2H),6.88(d,J=8.0Hz,2H),4.62(s,2H ),4.33-4.19(m,2H),3.06-2.90(m,2H),2.41-2.16(m,2H),1.33-1.27(m,3H). 13 C NMR(100MHz, CDCl3)δ168.70,157.86,133.85,125.81,115.72,65.52,61.58,31.89,26.57,14.19.HRMS(ESI+):calculated for C 18 H 15 F 13 O3S[M+Na] + 581.0427,found,581.441.

[0165] Example 15 Preparation of Compound 17

[0166]

[0167] Compound 15 (50.0 mg, 0.09 mmol) was thoroughly dissolved in 4 mL of tetrahydrofuran (THF). Lithium hydroxide (17.2 mg, 0.72 mmol) dissolved in H₂O (2 mL) was added dropwise with stirring and allowed to react at room temperature for 0.5 h. The reaction mixture was then acidified with glacial acetic acid (1 mL), diluted with water, and extracted twice with n-butanol. The mixture was then concentrated under reduced pressure and dried to yield the corresponding crude hydrolysis product 16. The crude hydrolysis product 16 was mixed with H-Arg-OMe·2HCl (49.3 mg, 0.19 mmol) and thoroughly dissolved in 8 mL of anhydrous DMF. HATU (71.7 mg, 0.19 mmol) was added with stirring. After thorough dissolution, DIPEA (94 μL, 0.57 mmol) was slowly added dropwise and allowed to react at room temperature for 24 h. The reaction mixture was then diluted with water, extracted twice with n-butanol, and concentrated under reduced pressure to yield the crude product. The crude product was purified by preparative HPLC and dried under reduced pressure to give the target product 17 (44.3 mg) as a light yellow gel in a yield of 67%.

[0168] 1 H NMR (400MHz, METHANOL-D4) δ7.44(d,J=8.6Hz,2H),7.02(d,J=8.7Hz,2H),4.61(s,2H),4.58-4.53(m,1H),3.73(s,3H ),3.23-3.14(m,2H),3.09-3.02(m,2H),2.48-2.31(m,2H),2.05-1.94(m,1H),1.86-1.75(m,1H),1.70-1.54(m,2H). 13 C NMR(100MHz,)δ173.34,171.13,159.13,158.68,134.96,127.42,116.94,68.12,52.93,41.78,32.64,32.43,29.47,27.23,26.25.HRMS(ESI+): calculated for C 23 H 25 F 13 N4O4S[M+H] + 701.1402, found 701.1470.

[0169] Example 16 Preparation of Compound 18

[0170]

[0171] The synthesis of compound 18 was carried out according to the synthesis method of compound 17, using intermediate 15 (50.0 mg, 0.09 mmol), H-His-OMe·2HCl (45.7 mg, 0.19 mmol) and HATU (71.7 mg, 0.19 mmol), and DIPEA (94 μL, 0.57 mmol) was added dropwise to obtain the target product 18 (27.7 mg) as a light yellow gel with a yield of 43%.

[0172] 1 H NMR (400MHz, CD3OD) δ8.13(s,1H),7.43(d,J=8.9Hz,2H),7.05(s,1H),6.97(d,J=8.9Hz,2H ),4.86-4.80(m,1H),4.57-4.51(m,2H),3.73(s,3H),3.28-3.01(m,4H),2.47-2.30(m,2H). 13 C NMR(100MHz,CD3OD)δ172.42,170.92,159.05,135.89,134.95,133.10,127.36,11 8.09,116.88,68.04,53.05,53.03,32.59,28.80,27.17.HRMS(ESI+):calculated for C 23 H 20 F 13 N3O4S[M+H] + 682.1041, found 682.1039.

[0173] Example 17 Preparation of Compound 19

[0174]

[0175] The synthesis of compound 19 was carried out according to the synthesis method of compound 17, using compound 15 (50.0 mg, 0.09 mmol), H-Arg-NH2·2HCl (46.4 mg, 0.19 mmol) and HATU (71.7 mg, 0.19 mmol), and DIPEA (94 μL, 0.57 mmol) was added dropwise to obtain the target product 19 (38.4 mg) as a yellow gel with a yield of 59%.

[0176] 1H NMR (400MHz, CD3OD) δ7.44(d,J=8.5Hz,2H),7.03(d,J=8.5Hz,2H),4.66-4.56(m,2H),4.54-4.47(m, 1H),3.23-3.16(m,2H),3.09-3.03(m,2H),2.46-2.34(m,2H),1.98-1.72(m,2H),1.68-1.58(m,2H). 13 C NMR(100MHz,CD3OD)δ176.04,170.84,159.05,158.66,134.98,127.45,116.89,1 01.32,68.13,53.41,41.86,32.61,30.47,27.20,26.19.HRMS(ESI+):calculated for C 22 H 24 F 13 N5O3S[M+H] + 686.1465, found 686.1461.

[0177] Example 18 Preparation of Compound 20

[0178]

[0179] The synthesis of compound 20 was carried out according to the synthesis method of compound 17, using compound 15 (50.0 mg, 0.09 mmol), H-Arg-Arg-OMe·3HCl (65.0 mg, 0.19 mmol) and HATU (107.6 mg, 0.28 mmol), and DIPEA (94 μL, 0.57 mmol) was added dropwise to obtain the target product 20 (49.1 mg) as a light yellow gel with a yield of 60%.

[0180] 1 H NMR (400MHz, CD3OD) δ7.44(d,J=7.6Hz,2H),7.01(d,J=7.7Hz,2H),4.60(s,2H),4.51-4.42(m, 2H),3.73(s,3H),3.26-3.12(m,4H),3.10-3.02(m,2H),2.48-2.31(m,2H),1.99-1.55(m,8H). 13C NMR(100MHz,CD3OD)δ173.88,173.53,170.90,169.95,159.08,158.68,158.50,134.96,127.43,116.86 ,67.98,54.01,53.19,52.90,41.90,41.76,32.58,30.24,29.46,26.19,26.11.HRMS(ESI+):calculated for C 29 H 37 F 13 N8O5S[M+H] + 857.2473,found857.2471.

[0181] Example 19 Preparation of Compound 21

[0182]

[0183] The synthesis of compound 21 was carried out according to the synthetic method of 17, using intermediate 15 (50.0 mg, 0.09 mmol), H-Arg-Arg-NH2·3HCl (62.1 mg, 0.19 mmol) and HATU (107.6 mg, 0.28 mmol), and DIPEA (94 μL, 0.57 mmol) was added dropwise to obtain the target product 21 (48.6 mg) as a light yellow gel with a yield of 61%. 1 H NMR(400MHz,CD3OD)δ7.45(d,J=8.4Hz,2H),7.02(d,J=8.4Hz,2H),4.61(s,2H),4.52-4 .33(m,2H),3.25-3.14(m,4H),3.10-3.02(m,2H),2.49-2.31(m,2H),1.97-1.59(m,8H). 13 C NMR(100MHz,CD3OD)δ176.28,173.69,170.64,168.60,162.12,159.07,158.68,134.96,127.51,117.15, 116.87,68.02,53.88,51.11,41.87,30.37,30.02,26.26,26.24,26.11,26.06.HRMS(ESI+):calculated for C 28 H 36 F 13 N9O4S[M+H] + 842.2476, found 842.2494.

[0184] Example 20 Preparation of Compound 22

[0185]

[0186] The synthesis of compound 22 was carried out according to the synthesis method of compound 17, using compound 15 (50.0 mg, 0.09 mmol), H-Arg-Arg-Arg-OMe·4HCl (141.6 mg, 0.28 mmol) and HATU (107.6 mg, 0.28 mmol), and DIPEA (94 μL, 0.57 mmol) was added dropwise to obtain the target product 21 (63.7 mg) as a light yellow gel with a yield of 66%.

[0187] 1 H NMR (400MHz, CD3OD) δ7.70(d,J=8.7Hz,2H),7.25(d,J=8.5Hz,2H),4.81-4.67(m,2H),4. 50-4.35(m,3H),3.72(s,3H),3.30-3.03(m,8H),2.76-2.26(m,2H),1.96-1.55(m,12H). 13 C NMR (100MHz, CD3OD) δ174.04,173.82,173.60,171.10,169.85,159.05,158.65,134.96,127.42,116.84,67.97,54.24 ,54.17,53.32,52.88,41.90,41.85,41.80,30.10,30.07,29.42,27.18,27.14,26.23,26.15.HRMS(ESI+):calculated forC 35 H 49 F 13 N 12 O6S[M+H] + 1013.3484, found 1013.3475.

[0188] Example 21 Preparation of Compound 24

[0189]

[0190] The synthesis of compound 24 was similar to that of compound 17. Compound 17 (50 mg, 0.07 mmol) was first hydrolyzed with lithium hydroxide (13.7 mg, 0.57 mmol) at room temperature for 1.5 hours to obtain the crude hydrolysis product 23. The crude hydrolysis product 23 was then used with H-Arg-Arg-NH2·3HCl (71.9 mg, 0.22 mmol), HATU (83.1 mg, 0.22 mmol), and DIPEA (73 μl, 0.44 mmol) to obtain the desired product 24 (30.9 mg) as a pale yellow gel in a 42% yield.

[0191] 1 H NMR(400MHz,CD3OD)δ7.45(d,J=8.6Hz,2H),7.02(d,J=8.7Hz,2H),4.62(s,2H),4.48-4.41(m,1H) ,4.41-4.31(m,2H),3.24-3.12(m,6H),3.11-3.03(m,2H),2.49-2.32(m,2H),1.95-1.60(m,12H). 13 C NMR (100MHz, CD3OD) δ176.33,174.02,173.84,171.15,170.05,159.06,158.67,158.65,134.97,127.43,116.84,101.33,67.9 9,54.43,54.28,53.92,41.90,41.85,41.81,30.46,30.04,29.92,29.89,27.18,26.25,26.18,26.14.HRMS(ESI+):calculated for C 34 H 48 F 13 N 13 O5S[M+H] + 998.3487, found 998.3478.

[0192] Example 22 Preparation of Compound 25

[0193]

[0194] The synthesis of compound 25 was carried out according to the synthesis method of compound 24. Compound 17 (50 mg, 0.07 mmol) was first used to prepare the crude hydrolysis product 23. The crude hydrolysis product 23 was then used, H-Arg-Arg-Arg-OMe·4HCl (109.4 mg, 0.22 mmol), and HATU (83.1 mg, 0.22 mmol), and DIPEA (73 μL, 0.44 mmol) was added dropwise to obtain the desired product 25 (33.4 mg) as a pale yellow gel in a 39% yield.

[0195] 1 H NMR (400MHz, CD3OD) δ7.44(d,J=8.8Hz,2H),7.02(d,J=8.8Hz,2H),4.63(s,2H),4.49-4.33(m, 4H),3.72(s,3H),3.26-3.14(m,8H),3.11-3.02(m,2H),2.49-2.31(m,2H),1.99-1.57(m,16H). 13 C NMR (100MHz, CD3OD) δ174.10,173.99,173.88,173.62,171.20,170.43,159.05,158.68,158.67,158.64,134.94,127.44,116.86,68.02,54.3 7,54.32,54.25,53.38,52.90,41.93,41.88,41.84,41.81,30.21,30.20,29.97,29.95,29.42,26.29,26.19,26.13.HRMS(ESI+):calculated for C 41 H 61 F 13 N 16 O7S[M+H] + 1169.4495,found1169.4495.

[0196] Example 23 Preparation of Compound 26

[0197]

[0198] The synthesis of compound 26 was carried out according to the synthesis method of compound 24. Compound 17 (50 mg, 0.07 mmol) was first used to prepare the crude hydrolysis product 23. The crude hydrolysis product 23 was then used, along with 1-[3-(1H-tetrazol-5-yl)phenyl]methanamine (38.3 mg, 0.22 mmol) and DMTMM (64.4 mg, 0.22 mmol), and DIPEA (73 μL, 0.44 mmol) was added dropwise to obtain the desired product 26 (35.8 mg) as a pale yellow gel in a 58% yield.

[0199] 1 H NMR (400MHz, CD3OD) δ7.97-7.88(m,2H),7.47-7.38(m,3H),7.33(d,J=7.7Hz,1H),7.01(d,J=8.8Hz,2H),4.67 -4.55(m,4H),4.43-4.34(m,1H),3.25-3.16(m,2H),3.10-3.01(m,2H),2.49-2.29(m,2H),2.04-1.63(m,4H). 13 C NMR (100MHz, DMSO-D6) δ171.30,167.63,161.11,158.01,157.33,140.03,133.37,131.89,129.07,127.05,125.56,125.11,123.70,116 .25,67.16,52.57,42.24,41.61,40.64,40.43,40.22,40.01,39.80,39.59,39.39,31.02,30.20,25.95,25.54.HRMS(ESI+):calculated for C 30 H 30 F 13 N9O3S[M+H] + 844.2057, found 844.2039.

[0200] Example 24 Preparation of Compound 27

[0201]

[0202] The synthesis of compound 27 was carried out according to the synthesis method of compound 24. Compound 17 (50 mg, 0.07 mmol) was first used to prepare the crude hydrolysis product 23. The crude hydrolysis product 23, isoniazid (20.0 mg, 0.15 mmol), and HATU (83.1 mg, 0.22 mmol) were then added dropwise with DIPEA (73 μL, 0.44 mmol) to obtain the desired product 27 (23.3 mg) as a yellow gel in a 39% yield.

[0203] 1 H NMR (500MHz, CD3OD) δ8.72(d,J=5.7Hz,2H),7.82(d,J=5.7Hz,2H),7.45(d,J=8.6Hz,2H),7.03(d,J=8.6Hz,2H),4 .70-4.59(m,3H),3.26-3.14(m,2H),3.08-3.03(m,2H),2.46-2.32(m,2H),2.07-1.83(m,2H),1.77-1.59(m,2H). 13 C NMR(100MHz,CD3OD)δ172.75,170.90,166.81,159.09,158.69,151.14,141.73,134.97,127.44,1 23.09,116.90,101.33,68.11,52.48,41.92,32.62,30.37,27.20,25.99.HRMS(ESI+):calculated forC 28 H 28 F 13 N7O4S[M+NH4] + 823.2054, found 822.1738.

[0204] Example 25 Preparation of Compound 28

[0205]

[0206] The synthesis of compound 28 was carried out according to the synthesis method of compound 24. Compound 17 (50 mg, 0.07 mmol) was first used to prepare the crude hydrolysis product 23. The crude hydrolysis product 23, 4-cyanobenzylamine (19.3 mg, 0.15 mmol), and HATU (83.1 mg, 0.22 mmol) were then added dropwise with DIPEA (73 μL, 0.44 mmol) to obtain the desired product 28 (36.9 mg) as a yellow gel in a 63% yield.

[0207] 1H NMR (400MHz, CD3OD) δ7.66(d,J=8.1Hz,2H),7.50-7.37(m,4H),7.02(d,J=8.8Hz,2H),4.68-4.58(m, 2H),4.53-4.40(m,3H),3.24-3.14(m,2H),3.10-3.01(m,2H),2.51-2.29(m,2H),2.01-1.53(m,4H). 13 C NMR (100MHz, CD3OD) δ173.83,171.11,159.06,158.65,145.72,134.91,133.43,129.23,127.45,119. 65,116.87,111.95,68.11,54.08,43.69,41.83,32.61,30.16,27.18,26.33.HRMS(ESI+):calculated for C 30 H 29 F 13 N6O3S[M+H] + 801.1887, found801.1883.

[0208] Example 26 In vitro antibacterial activity assay

[0209] According to the standards and guidelines of the American Clinical and Laboratory Standards Institute, the minimum inhibitory concentration (MIC) values ​​of mercaptophenol derivatives were determined by the broth microdilution method.

[0210] The strains used in this example: Staphylococcus aureus ATCC29213 and Escherichia coli ATCC25922 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.

[0211] The specific operations are as follows:

[0212] The Gram-positive strains to be tested were inoculated on Mueller-Hinton agar (MHA) plates, while the Gram-negative strains were inoculated on Luria-Bertani agar (LA) plates. The test compound was fully dissolved in DMSO / H2O to prepare a stock solution with a concentration of 1 mg / mL, and diluted with 4 times the volume of Mueller-Hinton Broth (MHB) to obtain a test compound solution. An equal volume of the test compound solution was added to the first row of a 96-well plate containing 100 μL of MHB in each well and a 2-fold gradient dilution was performed downward to obtain a test solution containing 100 μL of a concentration of 0.78 to 100 μg / mL in each well. Afterwards, a disposable sterile inoculation loop was used to pick the above-mentioned test strain into PBS buffer to adjust its OD 600 The value was 0.1, and diluted with MHB to obtain 1 × 10 6 100 μL of the test solution was added to each well of the test sample group and the positive control group, and incubated at 37°C for 18 to 24 hours. The OD value was measured using a microplate reader at 0 hours and 18 to 24 hours after the addition of the test solution. 600 The minimum inhibitory concentration (MIC) value was defined as the lowest concentration of compound required to inhibit 90% of bacterial growth.

[0213] The results are shown in Table 1: Most compounds have good antibacterial activity against Gram-positive bacteria, among which compounds 14, 20-22 show excellent antibacterial activity against both Gram-positive and Gram-negative bacteria (MIC ≤ 6.25 μg / mL), and compound 21 shows the best broad-spectrum antibacterial activity.

[0214] Table 1 In vitro antibacterial activity of mercaptophenol and its derivatives (μg / mL)

[0215]

[0216]

[0217] Example 27 In vitro cytotoxicity test

[0218] The CCK-8 method was used to determine the half-maximal inhibitory concentration (CC) of compound 21 on mammalian cells. 50). NCTCclone929 mouse fibroblasts and HEK293 human embryonic kidney cells were selected as the tested cell lines. The cells were washed twice with 3 mL of PBS buffer, and then 1.5-2 mL of 0.25% trypsin was added to digest until the cells fell off the wall of the culture flask. 3-4 mL of DMEM (or MEM) culture medium containing 10% fetal bovine serum (FBS) was added to neutralize the trypsin. The digested cells were transferred to a 15 mL centrifuge tube and centrifuged. After that, the upper layer of culture medium was discarded, and DMEM (or MEM) culture medium containing 10% FBS was added again to resuspend and dilute to obtain 2×10 cells per milliliter. 5 100 μL of the test cell suspension was added to each well of a 96-well plate for seeding, and the plate was placed in a 37°C constant temperature incubator containing 5% CO2 until the cells covered 70% to 80% of the bottom of each well. Compound 21 was dissolved in DMEM (or MEM) culture medium containing 10% FBS to prepare a 1 mg / mL stock solution, and then graded dilution was performed to obtain 6 concentration gradients (3.125 to 100 μg / mL) of the test solution, with the final concentration of DMSO not exceeding 0.1% and the final concentration of Tween 80 not exceeding 0.5%. The DMEM (or MEM) culture medium in the 96-well plate with cells was slowly aspirated, and then the test solution of the corresponding concentration was added. DMEM (or MEM) culture medium containing 10% FBS was added to each well of the control group and the blank group. The control group contained cells, while the blank group did not contain cells. Then, the 96-well plate containing cells and the test solution was placed in a 37°C constant temperature incubator with 5% CO2 for 24 hours, and then taken out and CCK-8 reagent was added to each well at a volume of 10 μL. The plate was incubated for 1 hour in the dark, and the OD was measured using a microplate reader. 450 Mouse fibroblast NCTC clone 929 cells were cultured in DMEM medium containing 10% FBS, and human embryonic kidney cells HEK293 cells were cultured in MEM medium containing 10% FBS. The cell survival rate was calculated as follows: % cell survival rate = [(Abs 受测样品 -Abs 空白 ) / (Abs 对照 -Abs 空白 )] × 100. All experiments were performed with at least 2 biological replicates.

[0219] The test results are as follows Figure 1 As shown, the half-maximal inhibitory concentration (CC 50 ) value was 59.75±2.74μg / mL, and the CC of NCTC clone 929 cells was 50The value was 37.26±2.79 μg / mL, which was much higher than the MIC value of compound 21, indicating that compound 21 had a low toxic effect on mammalian cells and was very safe within the range of its antibacterial activity.

[0220] Example 28 Drug resistance test

[0221] The initial MIC values ​​of Compound 21 and norfloxacin against the test strain S. aureus ATCC 29213 were determined using the in vitro antibacterial activity assay described in Example 26. Bacterial cells treated with Compound 21 and norfloxacin at 0.5× the MIC were then aspirated from a 96-well plate as the test solution. The MIC values ​​of the compounds and the reference drug were re-determined according to the MIC assay. This experimental procedure was repeated for 20 consecutive days, and daily changes in the MIC values ​​were recorded. The tendency of the test compounds to induce the development of drug resistance was evaluated by serial bacterial passage. All experiments were performed in at least two biological replicates.

[0222] The results are as follows Figure 2 As shown, after 20 days of continuous subculture with S. aureus ATCC 29213, the MIC value of compound 21 remained less than 4 times the initial MIC value, while the control drug norfloxacin increased to 64 times the initial value. These experimental results indicate that compound 21 has very low resistance-inducing ability under laboratory conditions simulating drug resistance.

[0223] Example 29 Study on Antibacterial Mechanism

[0224] (1) SYTOX Green assay was used to study the effect of compound 21 on the membrane permeability of MRSANCTC10442 cells.

[0225] To the bacterial suspension (OD 600 =0.2) was added with SYTOX Green dye at a final concentration of 10 μM and incubated for 5 minutes in the dark. At the same time, the excitation wavelength of the fluorescence microplate reader was set to 504 nm and the emission wavelength was set to 523 nm. After the bacterial suspension was incubated, 796 μL was drawn into a 24-well plate according to the experimental concentration group, and the change in fluorescence intensity was measured. After it stabilized, 4 μL of the test solution of compound 21 prepared with DMSO was immediately added to each well to a final concentration of 1×, 2×, 4× and 8×MIC. The change in fluorescence intensity was then continuously monitored using a fluorescence microplate reader for about 60 minutes. The control group was set to a bacterial culture well that was not treated with the compound, and the blank group was set to a well containing only MHB culture medium. Finally, the fluorescence intensity value was recorded and a time-fluorescence intensity curve was drawn. All experiments were repeated at least twice.

[0226] SYTOX Green is a nucleic acid-binding dye that can be used to detect bacterial membrane integrity. When a significant increase in fluorescence intensity is observed, it indicates that the integrity of the bacterial membrane has been destroyed. The dye enters the membrane-damaged bacterial cell and binds to its DNA. Figure 3 As shown, compound 21 increased the fluorescence intensity of the mixed solution after acting on the test strain MRSANCTC 10442, with the change being particularly pronounced at concentrations of 4× and 8× the MIC. The fluorescence intensity was positively correlated with the compound concentration. This indicates that compound 21 can disrupt the integrity of the bacterial cell membrane and increase its permeability.

[0227] (2) Using LIVE / DEAD BacLight TM Bacterial viability assay kit, used to study the effect of compound 21 on the cell membrane integrity of the tested S. aureus ATCC29213.

[0228] Use a disposable sterile inoculation loop to pick up an appropriate amount of the test strain from the MHA plate, resuspend it in MHB medium, and place it in a shaking incubator at 37°C until it reaches the logarithmic growth phase. Then, centrifuge and discard the upper layer of culture medium to separate the bacterial cells. Wash the bacterial cells with sterile PBS buffer and centrifuge and discard the upper layer of PBS buffer, repeating this three times. Resuspend the isolated bacterial cells with PBS buffer and dilute to obtain a concentration of 1×10 per ml. 8 2 μL of the test solution of compound 21 prepared with DMSO was added to 998 μL of the test bacterial suspension (1×10 8 CFU / mL) to a final concentration of 8×MIC, and an equal volume of PBS was added to the control group. Subsequently, the experimental and control groups were placed in a 37°C incubator for incubation for 2 hours. After the incubation, PI and Syto9 dyes were mixed in equal amounts under dark conditions, and then 3 μL was aspirated and added to the bacterial suspension for staining for 15 minutes. After staining, an appropriate amount of bacterial suspension was aspirated to prepare a smear, and the staining of the bacterial cells was observed under a laser confocal microscope. The LIVE / DEAD fluorescence staining experiment was repeated at least twice.

[0229] Syto9 and PI double staining is often used to detect the viability of bacterial cells. Syto9 can penetrate intact and damaged bacterial cell membranes and stain all tested bacterial cells green, while PI can only stain dead tested bacteria red. Different colors indicate different bacterial cell states. Figure 4As shown, the bacterial cells in the control group treated with PBS buffer exhibited distinct green fluorescence, indicating that the majority of cells were alive. In contrast, the S. aureus ATCC 29213 bacterial cells treated with compound 21 exhibited distinct red fluorescence, indicating that the majority of cells were dead. These experimental observations suggest that compound 21 kills bacteria by disrupting the integrity of the bacterial cell membrane.

[0230] Example 30 Evaluation of in vivo antibacterial activity

[0231] The in vivo antibacterial activity of compound 21 was evaluated in a mouse keratitis infection model caused by S. aureus ATCC 29213. This experiment was performed with the approval of the Experimental Animal Center of South China Agricultural University, and all operations were in compliance with the requirements of the policies of the Ministry of Health.

[0232] The female C57 BL / 6 mice used were 6 to 8 weeks old and weighed 18 to 20 grams. Five days before infection, the mice were treated with cyclophosphamide intraperitoneally at a dose of 100 mg / kg every other day to establish an immunosuppressive model. A sufficient amount of the test strain was picked from the MHA plate using a disposable sterile inoculation loop, resuspended in MHB medium, and cultured in a shaking incubator at 37°C to prepare a medium containing 2×10 8 The test bacterial suspension of each colony was prepared and stored at 4 ° C for subsequent corneal infection. The eyeballs of the same side of the mouse were selected, scratched with a sterile needle to cause corneal tissue damage, and the test bacterial suspension was added dropwise at a volume of 20 μL per mouse for infection. On the second day of infection, the mice were randomly divided into a treatment group, a positive control group and a negative control group, with 5 mice in each group. From the third to the fifth day of infection, the eyeballs of the infected mice were treated every two hours within 6 hours at a volume of 20 μL each time. The treatment group was set to contain 0.5% (5 mg / mL) of the test solution of compound 21, the positive control group was set to contain 5% (50 mg / mL) of vancomycin hydrochloride solution, and the negative control group was set to 5% glucose injection. The test solution and the control solution were both prepared with normal saline. After the treatment, the test mice were euthanized by intraperitoneal injection of a lethal dose of anesthetic, and the isolated damaged corneal tissue was soaked with 1 mL of PBS buffer, and the corneal tissue was then crushed to transfer the bacteria therein to PBS buffer. The MHA plate count method was used to count the surviving bacteria. SPSS 25.0 software was used to perform two independent sample t-tests, and P < 0.05 was considered statistically significant.

[0233] The test results are as follows Figure 5As shown, in a mouse corneal infection model induced by the test strain S. aureus ATCC 29213, treatment with compound 21 (0.5%, 5 mg / mL) reduced the viable bacterial load in the infected cornea by 5.51 logs (P < 0.05), while vancomycin (5%, 50 mg / mL) reduced the bacterial load by 6.54 logs (P < 0.05). The therapeutic effects of the two treatments were not statistically different, and the effective concentration of compound 21 was only one-tenth that of vancomycin. These in vivo antibacterial treatment results indicate that compound 21 can be used to treat bacterial keratitis infections caused by Gram-positive bacteria.

[0234] The above-described embodiments merely illustrate several implementations of the present invention, and while their descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the patent. It should be noted that a person skilled in the art would be able to make numerous variations and improvements without departing from the spirit of the present invention, all of which fall within the scope of protection of the present invention. Therefore, the scope of protection of the patent for this invention shall be determined by the appended claims.

Claims

1. A mercaptophenol derivative having a structure represented by formula (I) or a pharmaceutically acceptable salt thereof or a stereoisomer thereof: in, R1 is selected from: 3,3,4,4,5,5,6,6,7,7,8,8,8-tridecafluorooctyl; R2 is selected from: C2-C6 alkyl substituted by R3, C1-C2 alkyl substituted by R4; R3 is selected from: -NHR5, -NR6R7, R4 is selected from: -C(=O)R8; R5 is selected from the group consisting of: methyl, ethyl, propyl, butyl, methyl substituted with R9, ethyl substituted with R9, propyl substituted with R9; R6 and R7 are independently selected from: methyl, ethyl, or R6, R7 together with the nitrogen atom to which they are connected form R 10 substituted or unsubstituted piperidinyl, piperazinyl or tetrahydropyrrolyl; R8 is selected from: -N(H)-R 12 -C(=O)R 11 ; R9 is selected from the group consisting of amino, methylamino, dimethylamino, (aminoethyl)2-amino; R 10 Selected from: hydrogen, methyl, ethyl; R 11 Selected from: C1-C6 alkoxy, amino, cyano-substituted benzylamino, R 12 Selected from: the residue after removing the carbon terminal carboxyl group and the nitrogen terminal amino group from arginine, and the residue after removing the carbon terminal carboxyl group and the nitrogen terminal amino group from a polypeptide formed by 2-4 arginines.

2. The mercaptophenol derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 1, characterized in that: R2 is 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.

3. The mercaptophenol derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to any one of claims 1 to 2, characterized in that: R3 is selected from the group consisting of methylamino, ethylamino, propylamino, butylamino, dimethylamino, diethylamino, 4-methylpiperazinyl, tetrahydropyrrolyl, 4. The mercaptophenol derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to any one of claims 1 to 2, characterized in that: R 11 Selected from: methoxy, ethoxy, propoxy, amino, cyano-substituted benzylamino, 5. The mercaptophenol derivative or a pharmaceutically acceptable salt thereof or a stereoisomer thereof according to claim 4, characterized in that: R8 is selected from the following structures:

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

7. Use of the mercaptophenol derivative according to any one of claims 1 to 6, or a pharmaceutically acceptable salt thereof, or a stereoisomer thereof in the preparation of a medicament for inhibiting bacterial growth, preventing bacterial infection, or treating bacterial infection.

8. The use according to claim 7, characterized in that: The bacteria are Gram-positive bacteria and / or Gram-negative bacteria.

9. The use according to claim 8, characterized in that: The bacteria are Staphylococcus, Enterococcus, Enterobacter and Acinetobacter.

10. The use according to claim 9, 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.

11. An antibacterial drug, characterized in that: The invention is prepared from active ingredients and pharmaceutically acceptable excipients, wherein the active ingredients include the mercaptophenol derivative according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof or a stereoisomer thereof.

Images