Hydrogen bond donor covalent inhibitor as well as preparation method and application thereof

Abstract

The invention discloses a hydrogen bond donor covalent inhibitor as well as a preparation method and application thereof, and belongs to the technical field of medicinal chemistry, the designed and synthesized hydrogen bond donor-covalent inhibitor structurally comprises three parts, namely a hydrogen bond donor, a connecting arm (LINKER) and a covalent warhead, the three parts are like building blocks, can be combined at will and are subjected to modular synthesis, and the structure of the hydrogen bond donor-covalent inhibitor is optimized. Various hydrogen bond donor-covalent inhibitors with novel structures are obtained. A biological activity experiment proves that the hydrogen bond donor-covalent inhibitor designed and synthesized by the invention has relatively good activity in antibacterial and anti-tumor aspects, and is expected to have potential biological activity and application value in anti-inflammatory and anti-virus aspects and the like. Meanwhile, the synthetic route is simple, the raw material source is rich, the preparation cost is low, modular synthesis can be achieved, and the potential application value in the field of various disease treatment drugs is achieved.

Description

Technical Field

[0001] The present invention belongs to the technical field of pharmaceutical chemistry, and relates to the preparation and application of hydrogen bond donor covalent inhibitors, and specifically to a preparation method of a class of hydrogen bond donor-covalent inhibitors and their application in antibacterial, anti-tumor and other aspects. Background Art

[0002] The emergence of kinase inhibitors has made a big step forward in the fight against diseases and opened the curtain of small molecule targeted therapeutic drugs. Imatinib is an excellent representative of them. Traditional drugs such as aspirin and penicillin are also considered to be kinase covalent inhibitors. Therefore, kinase inhibitors are widely used in antibacterial, anti-tumor, and antiviral aspects. For example, Zanubrutinib, independently developed by BeiGene, is an oral Bruton's tyrosine kinase (BTK) small molecule covalent inhibitor, which was successfully launched in 2019. BTK is a key regulator of the B cell receptor (BCR) signaling pathway, involved in the proliferation, transport, chemotaxis and adhesion of B cells, and widely expressed in different types of hematological malignancies. Zanubrutinib has shown good safety and efficacy in a variety of B cell lymphomas. The key to its therapeutic efficacy is the presence of acrylamide covalent warheads, which covalently bind to the sulfhydryl (-SH) on the cysteine ​​residue of BTK. In recent years, the research on covalent inhibitor drugs has ushered in a wave of revival. In May 2021, Amgen launched Sotorasib, an inhibitor with an acrylamide covalent warhead developed for the difficult-to-drugged target KRAS, on the market first. It became the world's first targeted drug for treating KRAS gene mutation cancers and was approved for the treatment of non-small cell lung cancer carrying the KRAS G12C mutation.

[0003]

[0004] At present, most inhibitors that are on the market or in the research stage have similar aminopyrimidine core structures in their chemical structures. As of 2021, there are 71 small molecule kinase inhibitors approved by the US FDA for marketing, of which 34 contain pyrimidine structures, nearly 1 / 2. These small molecule inhibitors containing pyrimidine core structures are similar to the ATP structure and are competitive binders of ATPase. They mainly exert their inhibitory effects by binding to the hinge region of the kinase through hydrogen bonding. However, precisely because of the similarity to the ATP structure, this type of inhibitor has affinity for many kinases in the body, which can easily cause off-target and drug resistance of the inhibitors. Summary of the invention

[0005] In order to eliminate or reduce off-target and drug resistance effects and improve the therapeutic effect, the present invention designs and synthesizes a class of hydrogen bond donor covalent inhibitors that break through the traditional aminopyrimidine structure and have both hydrogen bond donors and covalent warheads, provides its synthesis method, and demonstrates its application in antibacterial, anti-tumor and other aspects.

[0006] In order to achieve the above object, the present invention adopts the following technical solutions:

[0007] The present invention discloses a type of hydrogen bond donor-covalent inhibitor, whose structural formula is as follows Figure 1 As shown:

[0008] wherein the hydrogen bond donor is selected from urea, thiourea, squaramide or oxalic acid diamide;

[0009] LINKER is selected from the conjugates of the carbon end of an amino acid and substituted piperazine, amino-substituted piperidine or amino-substituted pyrrolidine; or selected from pyrrolidine, piperidine, piperazine substituted with a primary / secondary amine group; or diamine;

[0010] The covalent warhead is selected from covalent warheads that donate an acyl group.

[0011] Preferably, the hydrogen bond donor is selected from

[0012] Among them, R1 is selected from (substituted) aryl, such as (substituted) phenyl, (substituted) naphthyl, etc.; (substituted) heteroaryl, such as (substituted) pyrrole, (substituted) pyridine, substituted (pyrimidine), etc.; (substituted) alkyl, such as tert-butyl, isobutyl, etc.; (substituted) cycloalkyl, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, etc.; (substituted) heterocycloalkyl, such as (substituted) piperidine, (substituted) piperazine, (substituted) morpholine, etc.

[0013] More preferably, R1 is selected from any one of the following formulae:

[0014]

[0015] Preferably, LINKER is selected from

[0016] Any one of .

[0017] Preferably, the covalent warhead providing the acyl group comprises an acryloyl, chloroacetyl, cyanoacetyl, cyanocyclopropylenylacetyl, propionyl, nitroacetyl, oxiranyl or sulfonyl fluoride group.

[0018] Preferably, the above-mentioned class of hydrogen bond donor-covalent inhibitors includes specific compounds of the following structures, named ZSK-1 to ZSK-42:

[0019]

[0020]

[0021] The present invention also discloses a method for preparing a type of hydrogen bond donor-covalent inhibitor, comprising the following steps:

[0022] 1) After dissolving compound A and compound B, triethylamine was added at low temperature and stirred to react in the dark to obtain intermediate II;

[0023] 2) adding dichloromethane and trifluoroacetic acid to intermediate II and reacting at room temperature to obtain intermediate III;

[0024] 3) After dissolving compound C and intermediate III, EDC, HOBt and DMAP were added, and triethylamine was added at low temperature for dissociation to obtain intermediate IV;

[0025] 4) adding dichloromethane and trifluoroacetic acid to intermediate IV and reacting at room temperature to obtain intermediate V;

[0026] 5) After dissolving compound D and intermediate V, triethylamine is added and stirred to react to obtain a hydrogen bond donor-covalent inhibitor;

[0027] in:

[0028] The structural formula of the compound A is as follows:

[0029]

[0030] The structural formula of the compound B is as follows:

[0031]

[0032] The structural formula of the intermediate II is as follows:

[0033]

[0034] The structural formula of the intermediate III is as follows:

[0035]

[0036] The structural formula of the compound C is as follows:

[0037]

[0038] The structural formula of the intermediate IV is as follows:

[0039]

[0040] The structural formula of the compound D is as follows:

[0041] R1-X;

[0042] The structural formula of the intermediate V is as follows:

[0043]

[0044] The specific structural formula of the hydrogen bond donor-covalent inhibitor is as follows:

[0045]

[0046] in,

[0047] R1 is

[0048] R2 is H, CH3, isopropyl, tert-butyl, phenyl, substituted phenyl, benzyl or aromatic heterocyclic group;

[0049] R3 is

[0050] X is

[0051] Preferably, in step 1), compound A and compound B are dissolved in dry dichloromethane, and then triethylamine is added at low temperature. After the addition is completed, the mixture is returned to room temperature for stirring and reacting in the dark. After the reaction is completed, water and saturated ammonium chloride solution are added in sequence for dispersion treatment, and then extracted, concentrated, separated and purified to obtain intermediate II;

[0052] The usage ratio of compound A, triethylamine and dichloromethane is (1-5) mmol: (5-10) mmol: (10-50) mL.

[0053] Preferably, in step 2), intermediate II is dissolved in dichloromethane, trifluoroacetic acid is added, and the reaction is carried out at room temperature. After the reaction is completed, the intermediate III is concentrated;

[0054] The usage ratio of intermediate II, trifluoroacetic acid and dichloromethane is (1-5) mmol: (6-12) mmol: (10-50) mL.

[0055] Preferably, in step 3), compound C and intermediate III are dissolved in dry dichloromethane, and then EDC, HOBt and DMAP are added in sequence, and then triethylamine is added at low temperature for dissociation, and after the addition is completed, the reaction is restored to room temperature, and after the reaction is completed, a saturated ammonium chloride solution, water, and a saturated sodium chloride solution are added to the obtained reaction solution in sequence for dispersion, and then the intermediate IV is obtained by extraction, concentration, separation and purification;

[0056] Among them, the usage ratio of compound C, intermediate III, EDC, HOBt, DMAP, triethylamine and dichloromethane is (1-5) mmol: (1-5) mmol: (1-10) mmol: (1-10) mmol: (0.1-1) mmol: (5-50) mmol: (10-100) mL.

[0057] Preferably, in step 4), intermediate IV is dissolved in dichloromethane, trifluoroacetic acid is added, and the mixture is reacted at room temperature. After the reaction is completed, the intermediate V is obtained by concentration treatment;

[0058] The usage ratio of intermediate IV, trifluoroacetic acid and dichloromethane is (1-5) mmol: (6-12) mmol: (10-50) mL.

[0059] Preferably, in step 5), compound D and intermediate V are dissolved in dry dichloromethane or methanol, triethylamine is added at low temperature for dissociation, and after the addition is completed, the temperature is restored to room temperature for reaction. After the reaction is completed, the reaction solution is concentrated and separated and purified to obtain the target product hydrogen bond donor covalent inhibitor;

[0060] The usage ratio of compound D, intermediate V, triethylamine and dichloromethane is (1-5) mmol: (1.05-5.10) mmol: (5.50-30.0) mmol: (10-100) mL.

[0061] The invention also discloses the application of the hydrogen bond donor-covalent inhibitor in the preparation of antibacterial drugs.

[0062] Preferably, the antibacterial drug is a drug against Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, Maltophilia, Staphylococcus aureus or Enterococcus faecalis.

[0063] The invention also discloses the application of the hydrogen bond donor-covalent inhibitor in the preparation of anti-tumor drugs.

[0064] Preferably, the tumor includes liver cancer and brain glioma.

[0065] Preferably, the brain glioma includes human astrocytoblastoma, human glioma and human neuroblastoma.

[0066] Compared with the prior art, the present invention has the following beneficial effects:

[0067] The hydrogen bond donor-covalent inhibitor disclosed in the present invention is a type of covalent inhibitor having both a hydrogen bond donor and a covalent warhead. Unlike the prior art, it breaks through the traditional structure of kinase inhibitors. Most existing kinase inhibitors contain an aminopyrimidine nucleus, which is similar to the ATP heterocyclic nucleus and can be recognized by ATPase. They are competitive binders of ATPase. Due to the similarity with the ATP structure, this type of inhibitor has affinity for many kinases in the body, which can easily cause toxicity, off-target and drug resistance of the inhibitor. The hydrogen bond donor-covalent inhibitor designed and synthesized by the present invention does not contain an aminopyrimidine nucleus, so it is not easy to produce toxicity, off-target and drug resistance. The hydrogen bond donor-covalent inhibitor structure designed and synthesized by the present invention includes three parts, namely a hydrogen bond donor, a linker and a covalent warhead. There are many types of hydrogen bond donors, typically including urea, thiourea, squaramide and oxalyl diamide, etc.; the connecting arm selects natural or non-natural amino acids and various nitrogen-containing (substituted) heterocycles through amide bond coupling, or selects primary (secondary) amine groups to replace nitrogen-containing heterocycles, and the covalent warhead selects a covalent warhead that provides an acyl group. These three parts are like building blocks, which can be combined arbitrarily and synthesized modularly to obtain a variety of novel hydrogen bond donor-covalent inhibitors; biological activity experiments have confirmed that the hydrogen bond donor-covalent inhibitor designed and synthesized by the present invention has good activity in antibacterial and antitumor aspects, and is expected to have potential biological activity and application value in anti-inflammatory and antiviral aspects. At the same time, the synthesis route of the present invention is simple, the source of raw materials is abundant, the preparation cost is low, and modular synthesis can be performed, which has potential application value in the field of drugs for the treatment of various diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0068] Figure 1 It is the structural formula diagram of hydrogen bond donor-covalent inhibitor;

[0069] Figure 2 , Figure 3 , Figure 4 and Figure 5 They are the hydrogen spectrum, carbon spectrum, fluorine spectrum and high-resolution mass spectrum of ZSK-1;

[0070] Figure 6 , Figure 7 , Figure 8 and Fig. 9 They are the hydrogen spectrum, carbon spectrum, fluorine spectrum and high-resolution mass spectrum of ZSK-3;

[0071] Fig.10 , Fig.11 , Fig.12 and Fig.13 They are the hydrogen spectrum, carbon spectrum, fluorine spectrum and high-resolution mass spectrum of ZSK-4;

[0072] Fig.14 , Fig.15 , Fig.16 and Fig.17They are the hydrogen spectrum, carbon spectrum, fluorine spectrum and high-resolution mass spectrum of ZSK-18;

[0073] Fig.18 , Fig.19 , Fig. 20 and Fig.21 They are the hydrogen spectrum, carbon spectrum, fluorine spectrum and high-resolution mass spectrum of ZSK-26;

[0074] Fig. 22 , Fig.23 , Fig.24 and Fig.25 They are the hydrogen spectrum, carbon spectrum, fluorine spectrum and high-resolution mass spectrum of ZSK-31;

[0075] Fig.26 , Fig. 27 , Fig.28 and Fig.29 They are the hydrogen spectrum, carbon spectrum, fluorine spectrum and high-resolution mass spectrum of ZSK-36;

[0076] Fig.30 is the anti-Hep G2 cell activity of ZSK-3 (code Z2 is ZSK-3);

[0077] Fig.31 The results of cell scratch assay of hydrogen bond donor covalent inhibitor against glioma cell migration;

[0078] Fig.32 The experimental results of apoptosis induction of glioma cells by hydrogen bond donor covalent inhibitors;

[0079] Fig.33 Western-Blot (WB) experimental results of hydrogen bond donor covalent inhibitors acting on focal adhesion kinase. DETAILED DESCRIPTION

[0080] In order to enable those skilled in the art to better understand the scheme of the present invention, the technical scheme in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work should fall within the scope of protection of the present invention.

[0081] It should be noted that the terms "first", "second", etc. in the specification and claims of the present invention and the above-mentioned drawings are used to distinguish similar objects, and are not necessarily used to describe a specific order or sequence. It should be understood that the data used in this way can be interchanged where appropriate, so that the embodiments of the present invention described herein can be implemented in an order other than those illustrated or described herein. In addition, the terms "including" and "having" and any variations thereof are intended to cover non-exclusive inclusions, for example, a process, method, system, product or device that includes a series of steps or units is not necessarily limited to those steps or units that are clearly listed, but may include other steps or units that are not clearly listed or inherent to these processes, methods, products or devices.

[0082] The present invention is further described in detail below in conjunction with the accompanying drawings:

[0083] 1. General preparation method of hydrogen bond donor covalent inhibitor disclosed in the present invention

[0084] The reaction route is as follows:

[0085]

[0086] The steps include:

[0087] (1) The compound shown in step 1 is dissolved in dry dichloromethane, and triethylamine is added to react under low temperature and in the dark with stirring to obtain the intermediate II shown in FIG.

[0088] (2) adding dichloromethane and trifluoroacetic acid to the compound shown in step 2 to react at room temperature to obtain the intermediate III shown;

[0089] (3) The compound shown in step 3 was dissolved in dry dichloromethane, EDC, HOBt, and DMAP were added, and triethylamine was added at low temperature to dissociate, thereby obtaining the intermediate IV shown in FIG.

[0090] (4) adding dichloromethane and trifluoroacetic acid to the compound shown in step 4 to react at room temperature to obtain the intermediate V shown;

[0091] (5) The compound shown in step 5 is dissolved in dry dichloromethane, and triethylamine is added and stirred to react to obtain a hydrogen bond donor covalent inhibitor.

[0092] in:

[0093]

[0094] etc.; R2=H, CH3, isopropyl (i-Pr), tert-butyl (t-Bu), phenyl (Ph), substituted phenyl, benzyl (Bn), aromatic heterocyclic group, etc.; wait; wait.

[0095] Preferably, in step (1), the compound is dissolved in dry dichloromethane, and triethylamine is added at low temperature. After the addition is completed, the mixture is returned to room temperature and protected from light for reaction. After the reaction is completed, the reaction solution is dispersed with water and saturated ammonium chloride solution in sequence, extracted with dichloromethane 2-3 times, and the organic layer is concentrated under vacuum and then separated and purified to obtain the intermediate represented by general formula II.

[0096] Further preferably, in step (1), the compound, triethylamine and dichloromethane are added in a ratio of 1 mmol:5 mmol:10 mL.

[0097] Preferably, in step (2), the intermediate of formula II is dissolved in dichloromethane, trifluoroacetic acid is added, and the mixture is reacted at room temperature. After the reaction is completed, the mixture is concentrated under reduced pressure to obtain the intermediate of formula III.

[0098] Further preferably, in step (2), the compound, trifluoroacetic acid and dichloromethane are added in a ratio of 1 mmol:6 mmol:10 mL.

[0099] Preferably, in step (3), the compound is dissolved in dry dichloromethane, and then EDC, HOBt, and DMAP are added in sequence. Triethylamine is added at low temperature for dissociation. After the addition is completed, the reaction is restored to room temperature. After the reaction is completed, the reaction solution is dispersed with saturated ammonium chloride solution, water, and saturated sodium chloride solution in sequence, extracted with dichloromethane 2-3 times, and the organic layer is concentrated under vacuum and then separated and purified to obtain the intermediate represented by general formula IV.

[0100] Further preferably, in step (3), the compound, EDC, HOBt, DMAP, triethylamine, and dichloromethane are added in a ratio of 1 mmol: 1 mmol: 1 mmol: 0.1 mmol: 5 mmol: 10 mL.

[0101] Preferably, in step (4), the intermediate of formula IV is dissolved in dichloromethane, trifluoroacetic acid is added, and the mixture is reacted at room temperature. After the reaction is completed, the mixture is concentrated under reduced pressure to obtain the intermediate of formula V.

[0102] Further preferably, in step (4), the compound, trifluoroacetic acid and dichloromethane are added in a ratio of 1 mmol:6 mmol:10 mL.

[0103] Preferably, in step (5), the compound is dissolved in dry dichloromethane (or methanol), and triethylamine is added at low temperature for dissociation. After the addition is completed, the reaction is resumed at room temperature. After the reaction is completed, the obtained reaction solution is concentrated under vacuum pressure and separated and purified to obtain the hydrogen bond donor covalent inhibitor of general formula I.

[0104] Further preferably, in step (5), the compound, triethylamine and dichloromethane are added in a ratio of 1 mmol:5 mmol:10 mL.

[0105] The specific embodiment is described below:

[0106] Example 1

[0107] The synthetic route of ZSK-1 is as follows:

[0108]

[0109] Specific operation: Take a 100mL dry single-mouth round-bottom flask, put in a stirring magnet, add N-Boc piperazine (1.87g, 10mmol), dissolve it with a mixed solvent of 25mL dichloromethane (DCM) and 5mL triethylamine (Et3N), cool the reaction bottle to 0℃ with an ice bath, and carefully drop a dichloromethane solution (5mL) containing 0.91g acryloyl chloride. React at 0℃ for 30min, warm to room temperature and react for 1.5h. Remove the solvent by rotary evaporation, dissolve the residue with 20mL dichloromethane, slowly add 3.45g trifluoroacetic acid (TFA, 30mmol) at room temperature, stir and react for 3h to obtain the key intermediate A, which is not separated and purified. Remove the solvent and excess trifluoroacetic acid by rotary evaporation, and obtain a dark yellow viscous residue that is dissolved in 10mL dichloromethane for use. Take another 100mL dry single-mouth round-bottom flask, add Boc-L-phenylalanine (Boc-L-Phe-OH, 2.60g, 9.8mmol) and dissolve it with 20mL dichloromethane and 20mL triethylamine, add condensation agent EDC hydrochloride (EDC·HCl, 2.5g, 15mmmol), 1-hydroxybenzotriazole (HOBt, 1.50g, 11mmol), 4-N,N-dimethylpyridine (DMAP, 0.13g, 1.1mmol) to the reaction flask in turn, and stir to react for 15min. Slowly add the dichloromethane solution containing the dark yellow viscous residue to the mixed solution in the 100mL single-mouth round-bottom flask, and stir to react at room temperature for 8h. The solvent was removed by rotary evaporation, and the obtained residue was dissolved in 60 mL of dichloromethane, washed with water and saturated brine in sequence, and the organic phase was dried over anhydrous sodium sulfate. The dichloromethane was removed by rotary evaporation, and the residue was purified by column chromatography (elution system: n-hexane / ethyl acetate = 1:1, v / v) to obtain 2.41 g of the key intermediate B as a white solid for standby use (total yield of three steps: 62%).

[0110] Take 0.195g (0.5mmol) of the key intermediate B and dissolve it in 5mL of dichloromethane, cool it in an ice bath, add 0.17g of trifluoroacetic acid (TFA, 1.5mmol) and react at room temperature for 2h. Cool the reaction solution to 0°C with an ice bath again, slowly add 3mL of triethylamine, stir and react for 15min, then add 70mg (0.5mmol) of 4-fluorophenyl isocyanate in dichloromethane solution (2mL) to the above system, stir and react at room temperature for 8h, evaporate the solvent, and purify the residue by column chromatography (elution system: n-hexane / ethyl acetate = 1:1 to 1:3, v / v) to obtain 0.17g of white solid ZSK-1 (total yield of two steps 80%).

[0111] ZSK-1 test data: white solid; melting point: 94.1-94.7℃; 1 HNMR(400MHz,Chloroform-d)δ7.56(d,J=36.6Hz,1H),7.40–7.00(m,8H),6.91(t,J=8.5Hz,2H),6.43(s,1H),6.28(d,J=16.5Hz,1H ),5.72(dd,J=10.3,1.9Hz,1H),5.25–5.01(m,1H),3.88–3.32(m,5H),3.31–3.06(m,3H),3.05–2.92(m,1H),2.70(d,J=60.1Hz,1H); 13 CNMR(101MHz,Chloroform-d)δ172.4,165.5,158.7(d,J=242.0Hz),155.4,136.0,134.8(d,J=2.7Hz),129. 5,129.0,128.9,127.5,126.8,121.2(d,J=7.6Hz),115.5(d,J=22.6Hz),50.7,45.7,44.9,42.0,41.1,40.3; 19 F NMR(376MHz,Chloroform-d)δ-120.1.HRMS-ESI(m / z):[M+Na]+calcd.for C 23 H 25 N4O3FNa + :447.1808,found:447.1807.

[0112] For the structural identification of ZSK-1, see Figure 2 to Figure 5 .

[0113] Example 2

[0114] The synthetic route of ZSK-3 is as follows:

[0115]

[0116] The synthesis operation of ZSK-3 is similar to that of Example 1, except that the isocyanate in the operation steps of Example 1 is replaced by 3,5-ditrifluoromethylphenyl squaramide monomethyl ester and the solvent is replaced by methanol.

[0117] ZSK-3 test data: light yellow solid; melting point: 147.8-148.2℃; 1 H NMR (400MHz, DMSO-d6) δ10.48(s,1H),8.34(s,1H),8.06(s,2H),7.68(s,1H),7.40–7.16(m,5H),7.01–6.60(m,1 H),6.25–6.04(m,1H),5.76–5.65(m,1H),5.44(d,J=8.1Hz,1H),3.51(d,J=43.4Hz,6H),3.16(d,J=76.5Hz,4H); 13 C NMR(101MHz,DMSO-d6)δ184.2,180.4,168.5(d,J=4.5Hz),164.3,162.6,141.1,135.6,131.4(q,J=32.8Hz), 129.7, 128.4, 127.8 (d, J = 21.3Hz), 127.1 (d, J = 22.6Hz), 124.5, 121.8, 117.9, 114.8, 54.9, 45.8, 44.4, 40.5; 19 F NMR(376MHz,DMSO-d6)δ-61.7.HRMS-ESI(m / z):[M+Na]+calcd.for C 28 H 24 N4O4F6Na + :617.1594,found:617.1597.

[0118] For the structural identification of ZSK-3, see Figure 6 to Figure 9 .

[0119] Example 3

[0120] The synthetic route of ZSK-4 is as follows:

[0121]

[0122] The synthesis operation of ZSK-4 is similar to that of Example 1, except that the isocyanate in the operation steps of Example 1 is replaced by isothiocyanate.

[0123] ZSK-4 test data: light yellow solid; melting point: 103.5-104.4℃; 1 H NMR(400MHz,Chloroform-d)δ8.93(s,1H),8.75–8.41(m,1H),7.88(s,2H),7.39(s,1H),7.33–7.19(m,5H),6.38(s,1H),6. 31–6.10(m,1H),5.75–5.60(m,1H),5.57–5.37(m,1H),3.79–3.34(m,6H),3.28(dd,J=12.8,5.5Hz,1H),3.18–2.71(m,3H); 13 C NMR(101MHz,Chloroform-d)δ180.7,172.8,165.4,139.9,135.0,131.5(q,J=36.6,35.1Hz),1 29.3,129.0(d,J=14.6Hz),128.1,126.7,124.3,122.4,121.6,117.8,55.6,45.9,42.4,39.2; 19 F NMR(376MHz,Chloroform-d)δ-63.1.HRMS-ESI(m / z):[M+Na]+calcd.for C 25 H 24 N4O2F6SNa + :581.1422,found:581.1424.

[0124] For the structural identification of ZSK-4, see Figure 10 to Figure 13 .

[0125] Example 4

[0126] The synthetic route of ZSK-18 is as follows:

[0127]

[0128] The synthesis operation of ZSK-18 is similar to that of Example 1, except that the isocyanate in the operation steps of Example 1 is replaced by 4-trifluoromethylphenyl squaramide monomethyl ester and the solvent is replaced by methanol.

[0129] ZSK-18 test data: white solid; melting point: 174.8-175.9℃; 1 H NMR (400MHz, DMSO-d6) δ10.22(s,1H),8.34(d,J=8.3Hz,1H),7.73–7.57(m,4H),7.35–7.20(m,5H),6.76(dd,J=16.1,10.2Hz,1H),6.12 (dd,J=16.6,2.3Hz,1H),5.71(dd,J=10.4,2.4Hz,1H),5.45(q,J=7.5Hz,1H),3.74–3.35(m,6H),3.28–3.18(m,1H),3.13–2.98(m,3H); 13 C NMR (101MHz, DMSO-d6) δ184.0,180.1,168.9,168.2,165.2,163.2,141.9,135.3,129.5,128.5(d,J=12.3Hz ),127.2(d,J=16.0Hz),126.6,125.6,123.1,122.8(d,J=4.4Hz),118.1,54.0,45.0,44.4,41.7,40.9,40.1; 19 F NMR(376MHz,DMSO-d6)δ-60.1.HRMS-ESI(m / z):[M+Na]+calcd.for C 27 H 25 N4O4F3Na + :549.1726,found:549.1730.

[0130] The relevant spectra for the structural identification of ZSK-18 can be found in Figure 14 to Figure 17 .

[0131] Example 5

[0132] The synthetic route of ZSK-26 is as follows:

[0133]

[0134] The synthesis operation of ZSK-26 is similar to that of Example 1. After obtaining the key intermediate A, the isocyanate in the operation steps of Example 1 is replaced with 3,5-ditrifluoromethylphenyl squaramide monomethyl ester, which is directly reacted with the key intermediate A, and the solvent is replaced with methanol.

[0135] The test data of ZSK-26 is: light yellow solid; melting point: 130.6-131.2℃; 1H NMR (400MHz, DMSO-d6) δ10.21(s,1H),7.95(s,2H),7.68(s,1H),6.86(dd,J= 16.7,10.4Hz,1H),6.32–6.04(m,1H),5.90–5.61(m,1H),3.95–3.66(m,8H); 13 C NMR (101MHz, DMSO-d6) δ185.9,181.2,168.6,164.4,161.8,141.0,130.6(q,J=32.9 ,32.4Hz),127.9(d,J=7.9Hz),124.6,121.9(d,J=3.5Hz),119.4,114.8,44.9,41.2; 19 F NMR(376MHz,DMSO-d6)δ-61.7.HRMS-ESI(m / z):[M+Na]+calcd.for C 19 H 15 N3O3F6Na + :470.0915,found:470.0915.

[0136] The relevant maps for the structural identification of ZSK-26 can be found in Figure 18 to Figure 21 .

[0137] Example 6

[0138] The synthetic route of ZSK-31 is as follows:

[0139]

[0140] The synthesis operation of ZSK-31 is similar to that of Example 1, except that the acryloyl chloride in the operation steps of Example 1 is replaced by chloroacetyl chloride, the isocyanate is replaced by 4-trifluoromethylphenyl squaramide monomethyl ester, and the solvent is replaced by methanol.

[0141] ZSK-31 test data: white solid; melting point: 161.1-162.2℃; 1 H NMR (400MHz, DMSO-d6) δ10.18(d,J=9.3Hz,1H),8.29(t,J=8.0Hz,1H),7.78–7.52(m,4H),7.39–7.15(m ,5H),5.44(s,1H),4.38(q,J=6.5,5.8Hz,2H),3.63–3.38(m,5H),3.31–3.17(m,2H),3.11–2.94(m,3H); 13C NMR (101MHz, DMSO-d6) δ184.3,180.2,168.6,168.4,164.8,163.1,142.5,135.7,129. 6(d,J=2.1Hz),128.4,127.0,126.7(d,J=3.2Hz),117.9,53.8,44.7,41.8,41.4,40.4; 19 F NMR(376MHz,DMSO-d6)δ-60.2HRMS-ESI(m / z):[M+Na]+calcd.for C 26 H 24 N4O4F3ClNa + :571.1336,found:571.1343.

[0142] The relevant maps for the structural identification of ZSK-31 can be found in Figure 22 to Figure 25 .

[0143] Example 7

[0144] The synthetic route of ZSK-35 is as follows:

[0145]

[0146]

[0147] The synthesis operation of ZSK-35 is similar to that of Example 1, except that the acryloyl chloride in the operation steps of Example 1 is replaced by cyanoacetic acid, the isocyanate is replaced by 4-trifluoromethylphenyl squaramide monomethyl ester, and the solvent is replaced by methanol.

[0148] ZSK-35 test data: light yellow solid; melting point: 161.8-162.6℃; 1 H NMR(400MHz,DMSO-d6)δ11.09–10.55(m,1H),8.49(s,1H),8.09(s,2H),7.66(s,1H),7.36–7.19(m,5H),5 .42(q,J=6.2Hz,1H),4.05(d,J=3.0Hz,2H),3.70–3.40(m,5H),3.21(d,J=30.2Hz,2H),3.13–2.95(m,3H); 13C NMR (101MHz, DMSO-d6) δ184.2, 180.4, 168.5 (d, J = 5.7Hz), 162.6, 161.7, 141.2, 135.7, 131.4 (q, J = 33.0Hz), 129.6 (d, J = 5.6 Hz),128.4,127.1(d,J=23.6Hz),124.5,121.8,119.1,117.9,116.0,114.7,53.9,45.1–44.3(m),41.6–40.8(m),40.3,24.8; 19 F NMR(376MHz,DMSO-d6)δ-61.8.HRMS-ESI(m / z):[M+Na]+calcd.for C 28 H 23 N5O4F6Na + :630.1552,found:630.1561.

[0149] Example 8

[0150] The synthetic route of ZSK-36 is as follows:

[0151]

[0152] The key intermediate D (0.80 g, 2 mmol) obtained in Example 7 was suspended in 20.0 mL of toluene, and 1.0 mL of pyrrolidine and 170 uL (2.1 mmol) of cyclopropylcarboxaldehyde were added in sequence, and the mixture was heated and stirred in an oil bath at 80 ° C for 3 h. The reaction process was monitored by thin layer chromatography (TLC). After the reaction was completed, the solvent was removed by rotary evaporation, and the residue was dissolved in 100 mL of ethyl acetate, washed with water and saturated brine in sequence, and the organic phase was dried over anhydrous sodium sulfate. The ethyl acetate was removed by rotary evaporation, and the residue was purified by column chromatography (elution system: n-hexane / ethyl acetate = 1:2, v / v) to obtain 0.77 g of the key intermediate E as a white solid for standby use (yield 85%). Take the above key intermediate E 0.23 g (0.5 mmol) and dissolve it in 5 mL of dichloromethane, cool it in an ice bath, add 0.17 g of trifluoroacetic acid (TFA, 1.5 mmol) and react at room temperature for 2 h. The reaction solution was cooled to 0°C with an ice bath again, 3 mL of triethylamine was slowly added dropwise, and the reaction was stirred for 15 min. A methanol solution (2 mL) containing 0.17 g (0.51 mmol) of 3,5-ditrifluoromethylphenyl squaramide monomethyl ester was added to the above system, and the reaction was stirred at room temperature for 30 h. The solvent was evaporated and the residue was purified by column chromatography (elution system: n-hexane / ethyl acetate = 1:3 to 1:5, v / v) to obtain 0.26 g of off-white solid ZSK-36 (total yield of two steps 75%).

[0153] ZSK-36 test data: white solid; melting point: 143.4-144.8℃; 1 H NMR (400MHz, DMSO-d6) δ10.93(s,1H),8.53(d,J=37.2Hz,1H),8.10(s,2H),7.64(s,1H),7.38–7.19(m,5H),6.70–6.37(m, 1H),5.42(t,J=6.9Hz,1H),3.72–3.43(m,6H),3.26–2.94(m,4H),2.06–1.80(m,1H),1.29–1.15(m,2H),0.99–0.79(m,2H); 13 CNMR(101MHz,DMSO-d6)δ184.2,180.4,168.6(d,J=14.3Hz),164.1,162.7,161.9,141.3,135.7,131.3(q,J=32.9Hz),129.7,1 28.4,127.1(d,J=28.3Hz),124.5,121.8,119.1,117.8,116.7,115.6,114.6,105.9,105.2,54.1,48.5,44.7,41.2,15.4,10.0; 19 F NMR(376MHz,DMSO-d6)δ-61.8.HRMS-ESI(m / z):[M+H]+calcd.for C 32 H 28 N5O4F6 + :660.2040,found:660.2038.

[0154] The relevant spectra for the structural identification of ZSK-36 can be found in Figure 26 to Figure 29 .

[0155] Example 9

[0156] The synthetic route of ZSK-40 is as follows:

[0157]

[0158] Take a 100 mL dry single-necked round-bottom flask, put in a stirring magnetic bar, add piperazine (1.30 g, 15 mmol) and 1.16 g (3 mmol) Boc-protected L-phenylalanine (Boc-L-Phe-OH), dissolve with 25 mL dichloromethane (DCM), add condensation agent EDC hydrochloride (EDC·HCl, 0.6 g, 3.3 mmmol), 1-hydroxybenzotriazole (HOBt, 0.33 g, 3.3 mmol), 4-N,N-dimethylpyridine (DMAP, 36 mg, 0.3 mmol) into the reaction flask in turn, and stir at room temperature for 8 h. The solvent was removed by rotary evaporation, and the residue was dissolved in 50 mL of dichloromethane, washed with water and saturated brine in turn, and the organic phase was dried over anhydrous sodium sulfate, and dichloromethane was removed by rotary evaporation. The residue was purified by column chromatography (elution system: n-hexane / ethyl acetate = 1:1-1:3, v / v) to obtain 0.92 g of the key intermediate F as a pale yellow solid for standby use (yield 90%). 0.66 g (2 mmol) of the key intermediate F was weighed into a 100 mL single-mouth round-bottom reaction bottle, dissolved in 20 mL of dichloromethane, cooled to 0°C in an ice bath, and a dichloromethane solution (5 mL) containing 0.22 g of epoxypropionic acid chloride (2.1 mmol) was carefully added dropwise, and the reaction was carried out at 0°C for 30 min, and then heated to room temperature for 1.5 h. The solvent was removed by rotary evaporation, and the residue was dissolved in 50 mL of dichloromethane, washed with water and saturated brine in turn, and the organic phase was dried over anhydrous sodium sulfate, and dichloromethane was removed by rotary evaporation. The residue was purified by column chromatography (elution system: n-hexane / ethyl acetate = 2:1 to 1:1, v / v) to obtain 0.61 g of the key intermediate G as a white solid for standby use (yield 76%). 0.21 g of the key intermediate G (0.5 mmol) was dissolved in 10 mL of dichloromethane, and 0.30 g of trifluoroacetic acid (TFA, 2.5 mmol) was slowly added at room temperature, and the reaction was stirred for 2 h, and the solvent and excess trifluoroacetic acid were removed by rotary evaporation to obtain a dark yellow viscous residue, which was not further separated and purified. The dark yellow viscous residue obtained above was dissolved with 10 mL of methanol, the reaction solution was cooled to 0°C in an ice bath, 1.5 mL of triethylamine was slowly added dropwise, and the reaction was stirred for 15 min. Then, a methanol solution (5 mL) containing 0.14 g (0.52 mmol) of 4-trifluoromethylphenyl squaramide monomethyl ester was added to the above system, and the reaction was stirred at room temperature for 30 h. The solvent was evaporated and the obtained residue was purified by column chromatography (elution system: n-hexane / ethyl acetate = 1:1 to 1:5, v / v) to obtain 0.24 g of white solid ZSK-40 (two-step yield 86%).

[0159] ZSK-40 test data: white solid; melting point: 103.5-105.6℃; 1H NMR(400MHz, DMSO-d6)δ10.87(s,1H),8.47(d,J=15.6Hz,1H),7.56–7.42(m,2H),7.34–7.20(m,5H),6.78– 6.64(m,2H),4.48(t,J=7.2Hz,1H),3.64–3.32(m,6H),3.52(s,1H),3.18–2.82(m,4H),2.10–1.86(m,2H); 13 C NMR(101MHz,DMSO-d6)δ183.0,179.6,166.8,164.2,162.8,160.6,139.4,136.8,132.2,13 0.1,128.2,127.3,123.8,120.5,118.4,117.2,116.8,116.6,58.7,56.8,50.4,47.6,37.5; 19 F NMR(376MHz,DMSO-d6)δ-64.3.HRMS-ESI(m / z):[M+H]+calcd.for C 27 H 26 F3N4O5 + :543.1850,found:543.1856.

[0160] Example 10

[0161] The synthetic route of ZSK-41 is as follows:

[0162]

[0163] The synthesis operation of ZSK-41 is similar to that of Example 9, except that the piperazine in the operation steps of Example 9 is replaced by N-sulfonyl fluoride piperazine hydrochloride.

[0164] ZSK-41 test data: white solid; melting point: 121.8-123.2℃; 1 H NMR(400MHz,DMSO-d6)δ10.82(s,1H),8.74(s,1H),8.06(s,2H),7.72(s,2H),7.28–7.16(m,5 H),6.74–6.68(m,1H),3.78(m,1H),3.68–3.60(m,2H),3.26–2.94(m,4H),2.76–2.63(m,4H); 13C NMR (101MHz, DMSO-d6) δ186.0,182.2,170.8,142.8,138.6,136.4,130.2,129.8,128.4,127.6,123.7,121.8,116.8,60.2,48.9,45.5,40.2; 19 FNMR(376MHz,DMSO-d6)δ-64.6,-43.2.HRMS-ESI(m / z):[M+H]+calcd.for C 24 H 23 F4N4O5S + :555.1320,found:555.1324.

[0165] Embodiment 11

[0166] The synthetic route of ZSK-42 is as follows:

[0167]

[0168] ZSK-42 test data: white solid; melting point: 135.3-136.6℃; 1 H NMR(400MHz,DMSO-d6)δ10.84(s,1H),8.65(s,1H),7.78(s,2H),7.65(s,2H),7.32–7.1 8(m,5H),5.45(s,2H),4.06(m,1H),3.65–3.54(m,4H),3.43–3.32(m,4H),3.25(m,2H); 13 C NMR(101MHz,DMSO-d6)δ183.4,179.6,170.3,168.4,144.2,142.8,136.4,135.2,130.5,13 0.1,129.2,128.3,126.7,118.3,117.2,116.8,116.2,113.3,76.6,59.2,50.4,49.3,39.8; 19 F NMR(376MHz,DMSO-d6)δ-62.2.HRMS-ESI(m / z):[M+H]+calcd.for C 26 H 25 F3N5O6 + :560.1751,found:560.1748.

[0169] 2. Activity test experiments and related results

[0170] 1. Antibacterial activity test of hydrogen bond donor covalent inhibitors

[0171] The antibacterial activity of the above hydrogen bond donor covalent inhibitor samples was tested according to the standard operating procedures of the minimum inhibitory concentration method (MIC method). Escherichia coli (E.coli), Klebsiella pneumoniae (K.Pneumoniae), Pseudomonas aeruginosa (P.aeruginosa), Acinetobacter baumannii, Ps.maltophilia, Staphylococcus aureus (S.aureus), and Enterococcus faecalis were selected as active test bacteria, and ZSK-1 to ZSK-42 were selected for general screening. The experiment confirmed that the hydrogen bond donor covalent inhibitor containing a square amide structure has good antibacterial ability, among which ZSK-3 has an IC 50 It is 1~2uM.

[0172] 2. Antitumor activity test of hydrogen bond donor covalent inhibitors

[0173] The anti-tumor activity of the above hydrogen bond donor covalent inhibitor samples was tested according to the standard operating procedures of the MTT method.

[0174] Human liver cancer cells (Hep G2) were selected for anti-tumor activity testing, and ZSK-1 to ZSK-42 were selected for general screening. The results are as follows Fig.30 As shown in the experiment, it was confirmed that the hydrogen bond donor covalent inhibitor containing square amide structure had moderate anti-Hep G2 activity, among which the IC 50 It is 25~50uM.

[0175] Human astrocytic glioblastoma cells (human malignant glioma cells, U-87MG), human glioma cells (U251), and human neuroblastoma cells (SH SY5Y) were selected for anti-tumor activity testing, and ZSK-1 to ZSK-42 were selected for general screening. The experiment confirmed that the hydrogen bond donor covalent inhibitors containing amide structure have good anti-glioma activity, among which ZSK-31 has an IC of 0.05 for U-87MG. 50 The results of Western Blot (immunoblotting) showed that the hydrogen bond donor covalent inhibitor containing amide structure acted on focal adhesion kinase (FAK), inhibiting the autophosphorylation of FAK, thereby inhibiting the proliferation and migration of tumor cells.

[0176] Fig.31The results of the cell scratch test of hydrogen bond donor-covalent inhibitors against glioma cell migration are shown. Human glioma cells (U251) were selected as the research object, and the control group (control), positive control group (TMZ, using temozolomide TMZ, a drug for clinical treatment of glioma, as the control), and drug administration group (using synthetic hydrogen bond donor-covalent inhibitors ZSK-2, ZSK-3, ZSK-6, and ZSK-10 as drugs) were set up to conduct cell scratch experiments. The results showed that compared with the positive control drug TMZ, ZSK-2, ZSK-3, ZSK-6, and ZSK-10 all had a certain ability to inhibit U251 cell migration, among which ZSK-3 containing a square amide structure had the strongest ability to inhibit U251 cell migration.

[0177] Fig.32 The effect of hydrogen bond donor-covalent inhibitor ZSK-3 on apoptosis of brain glioma cells is shown. Human glioma cells (U251) were selected as the research object, and a control group (TAE226 is a commonly used control drug for anti-glioma) and a drug group (synthesized hydrogen bond donor-covalent inhibitor ZSK-3 is used as a drug) were set up to conduct cell apoptosis experiments. The results showed that compared with the positive control drug TAE226, ZSK-3 has better biological activity than the positive control drug TAE226, and can induce a large number of U251 cells to undergo apoptosis at an early stage at a lower concentration (16uM).

[0178] Fig.33 The effect of hydrogen bond donor-covalent inhibitors on focal adhesion kinase (FAK) autophosphorylation is shown. A control group (control), a positive control group (TAE226 is a commonly used control drug for anti-glioma), and a drug-treated group (with synthetic hydrogen bond donor-covalent inhibitors ZSK-31 and ZSK-18 as drugs) were set up for WB experiments. The results showed that compared with the positive control drug TAE226, both ZSK-31 and ZSK-18 could significantly affect focal adhesion kinase (FAK) autophosphorylation, confirming that ZSK-31 and ZSK-18 target FAK and exert anti-glioma effects by inhibiting FAK autophosphorylation.

[0179] The above experiments confirmed that the synthesized hydrogen bond donor covalent inhibitor has application value in antibacterial and anti-tumor aspects.

[0180] The above contents are only for explaining the technical idea of ​​the present invention and cannot be used to limit the protection scope of the present invention. Any changes made on the basis of the technical solution in accordance with the technical idea proposed by the present invention shall fall within the protection scope of the claims of the present invention.

Claims

1. A class of hydrogen bond donor-covalent inhibitors, characterized in that Its structural formula is shown in Figure 1: wherein the hydrogen bond donor is selected from urea, thiourea, squaramide or oxalic acid diamide; LINKER is selected from the conjugate of the carbon end of an amino acid and substituted piperazine, amino-substituted piperidine or amino-substituted pyrrolidine; or selected from pyrrolidine, piperidine, piperazine substituted with a primary / secondary amine group; or diamine; The covalent warhead is selected from covalent warheads that donate an acyl group.

2. A class of hydrogen bond donor-covalent inhibitors according to claim 1, characterized in that: The hydrogen bond donor is selected from Wherein, R1 is selected from substituted aryl, substituted heteroaryl, substituted alkyl, substituted cycloalkyl, and substituted heterocycloalkyl.

3. A class of hydrogen bond donor-covalent inhibitors according to claim 2, characterized in that: The substituted aryl includes substituted phenyl and substituted naphthyl; the substituted heteroaryl includes substituted pyrrole, substituted pyridine and substituted pyrimidine; the substituted alkyl includes tert-butyl and isobutyl; the substituted cycloalkyl includes cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl and cyclooctyl; the substituted heterocycloalkyl includes substituted piperidine, substituted piperazine and substituted morpholine.

4. A class of hydrogen bond donor-covalent inhibitors according to claim 3, characterized in that: R1 is selected from any one of the following formulae:

5. A class of hydrogen bond donor-covalent inhibitors according to claim 1, characterized in that: LINKER is selected from Any one of .

6. A class of hydrogen bond donor-covalent inhibitors according to claim 1, characterized in that: Covalent warheads providing acyl groups include acryloyl, chloroacetyl, cyanoacetyl, cyanocyclopropylenylacetyl, propionyl, nitroacetyl, oxiranyl, or sulfonyl fluoride.

7. A hydrogen bond donor-covalent inhibitor according to any one of claims 1 to 6, characterized in that: Specific compounds including the following structures are designated as ZSK-1 to ZSK-42:

8. The method for preparing a type of hydrogen bond donor-covalent inhibitor according to any one of claims 1 to 7, characterized in that: The following steps are involved: 1) After dissolving compound A and compound B, triethylamine was added at low temperature and stirred to react in the dark to obtain intermediate II; 2) adding dichloromethane and trifluoroacetic acid to intermediate II and reacting at room temperature to obtain intermediate III; 3) After dissolving compound C and intermediate III, EDC, HOBt and DMAP were added, and triethylamine was added at low temperature for dissociation to obtain intermediate IV; 4) adding dichloromethane and trifluoroacetic acid to intermediate IV and reacting at room temperature to obtain intermediate V; 5) After dissolving compound D and intermediate V, triethylamine is added and stirred to react to obtain a hydrogen bond donor-covalent inhibitor; in: The structural formula of the compound A is as follows: The structural formula of the compound B is as follows: The structural formula of the intermediate II is as follows: The structural formula of the intermediate III is as follows: The structural formula of the compound C is as follows: The structural formula of the intermediate IV is as follows: The structural formula of the compound D is as follows: R1-X; The structural formula of the intermediate V is as follows: The specific structural formula of the hydrogen bond donor-covalent inhibitor is as follows: in, R1 is R2 is H, CH3, isopropyl, tert-butyl, phenyl, substituted phenyl, benzyl or aromatic heterocyclic group; R3 is X is Preferably, in step 1), compound A and compound B are dissolved in dry dichloromethane, and then triethylamine is added at low temperature. After the addition is completed, the mixture is returned to room temperature for light-shielded stirring reaction. After the reaction is completed, water and saturated ammonium chloride solution are added in sequence for dispersion treatment, and then the intermediate II is obtained through extraction, concentration, separation and purification; wherein the amount ratio of compound A, triethylamine and dichloromethane is (1-5) mmol: (5-10) mmol: (10-50) mL; Preferably, in step 2), after the intermediate II is dissolved in dichloromethane, trifluoroacetic acid is added, reacted at room temperature, and concentrated after the reaction to obtain the intermediate III; wherein the amount ratio of the intermediate II, trifluoroacetic acid and dichloromethane is (1-5) mmol: (6-12) mmol: (10-50) mL; Preferably, in step 3), after dissolving compound C and intermediate III in dry dichloromethane, EDC, HOBt and DMAP are added in sequence, and then triethylamine is added at low temperature for dissociation. After the addition is completed, the reaction is restored to room temperature. After the reaction is completed, the reaction solution is sequentially added with saturated ammonium chloride solution, water and saturated sodium chloride solution for dispersion, and then extracted, concentrated, separated and purified to obtain intermediate IV; wherein the amount ratio of compound C, intermediate III, EDC, HOBt, DMAP, triethylamine and dichloromethane is (1-5) mmol: (1-5) mmol: (1-10) mmol: (1-10) mmol: (0.1-1) mmol: (5-50) mmol: (10-100) mL; Preferably, in step 4), after the intermediate IV is dissolved in dichloromethane, trifluoroacetic acid is added, and the reaction is carried out at room temperature. After the reaction is completed, the intermediate V is concentrated and treated; wherein the amount ratio of the intermediate IV, trifluoroacetic acid and dichloromethane is (1-5) mmol: (6-12) mmol: (10-50) mL; Preferably, in step 5), compound D and intermediate V are dissolved in dry dichloromethane or methanol, triethylamine is added at low temperature for dissociation, and after the addition is completed, the temperature is restored to room temperature for reaction. After the reaction is completed, the reaction solution is concentrated and separated and purified to obtain the target product hydrogen bond donor covalent inhibitor; wherein the amount ratio of compound D, intermediate V, triethylamine and dichloromethane is (1-5) mmol: (1.05-5.10) mmol: (5.50-30.0) mmol: (10-100) mL.

9. Use of the hydrogen bond donor-covalent inhibitor according to any one of claims 1 to 7 in the preparation of antibacterial drugs, characterized in that: The antibacterial drug is a drug against Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, Maltophilia, Staphylococcus aureus or Enterococcus faecalis.

10. Use of the hydrogen bond donor-covalent inhibitor according to any one of claims 1 to 7 in the preparation of anti-tumor drugs, characterized in that: The tumors include liver cancer and brain glioma; Further, it includes human brain astroglioma, human glioma and human neuroblastoma.

Images