Inhibition Studies Of Hydroxamic Acids And Thiosemicarbazones On Metallo-β-lactamase

β-Lactamases,especially metallo-β-lactamases（MβLs）mediated bacterial resistance,have posed a serious threat to global public health security.MβLs hydrolyze almost all current antibiotics in clinical practice,but there is still no an effective method for it.The current methods to combat this super-resistance are the development of new antibacterial drugs or the development of inhibitors of MβLs,which can be used in combination with antibiotics to kill resistant bacteria.Based on this idea,this paper has carried out the following three parts of work:Antibiotic resistance has become an important problem that plagues countries all over the world.There are many ways for bacteria to resist antibiotics,the most important of which is to produce hydrolytic enzymes to inactivate antibiotics.In particular,the bacterial resistance caused by metalβ-lactamases（MβLs）has caused serious public health threats,such as NDM-1,VIM-2,etc.These enzymes can hydrolyze mostβ-lactamases.Amide antibiotics,including penicillins,cephalosporins,and carbapenems.The current strategy for dealing with antibiotic resistance is to develop new antibacterial drugs,and the other is to develop inhibitors of target proteins that cause antibiotic resistance to inactivate drug-resistant bacteria through their cooperation with antibiotics.Based on this idea,this paper has carried out the following three parts of work:1.Nineteen new hydroxamic acid derivatives were successfully designed and synthesized.Evaluation of biological activity revealed that,these hydroxamic acid derivatives had significant broad-spectrum inhibition on MβLs Imi S,L1,VIM-2 and IMP-1,and their activity reached the micromolar level,with IC₅₀ value in the range of 0.6-9.4,1.3-27.4,5.4-43.7 and 5.2-49.7μM;Hydroxamic acid synergized with cefazolin and meropenem to increase the sterilization activity of these antibiotics on 2-32-fold;mechanism characterization showed that,compound 17 reversibly and competitively inhibited L1 with K_i was determined to be 2.5μM.Animal experiments exhibited that,it can cooperate with cefazolin to significantly reduce the infection of L1-producing E.coli in the spleen and liver of mice.The docking study suggested that,it was tightly bound to the Zn（II）of VIM-2 and Cph A through the oxygen atom of the sulfonamide group,but bound to the Zn（II）of L1through the oxygen atom of the hydroxamic acid group.These studies indicate that hydroxamic acid is an effective scaffold for the development of MβLs inhibitors.2.A total of twenty-six thiosemicarbazones were designed.Evaluation of biological activity revealed that,the obtained molecules effectively inhibited NDM-1 with IC₅₀ in the range of0.038-34.7μM（except 1e,2e,and 3d）,and 1c is the most potent inhibitor(IC₅₀=0.038μM).The structure-activity relationship revealed that the diaryl-substitutes improved inhibitory activities of the inhibitors.The determination of MIC showed,the thiosemicarbazones exhibited synergistic antimycobacterial actions against E.coli-NDM-1,resulted a 2-512-fold reduction in MIC of meropenem,while 1c restored 16-256-,16-,and 2-fold activity of the antibiotic on clinical isolates ECs,K.pneumonia and P.aeruginosa harboring NDM-1,respectively.Animal experiments showed that 1c had a synergistic antibacterial ability with meropenem,reduced the bacterial load clinical isolate EC08 in the spleen and liver.This work provided a highly promising scaffold for the development of NDM-1 inhibitors.A novel vancomycin derivative3.A dipyridyl-substituted thiosemicarbazone（DpC）was synthesized.Evaluation of biological activity indicated that,DpC is the broad-spectrum inhibitor of MβLs（NDM-1,VIM-2,IMP-1,Imi S,L1）,with an IC₅₀ value in the range of 0.021-1.08μM.Mechanism studies showed that,it reversibly and competitively inhibited NDM-1 with the K_i value of10.2 n M.The determination of MIC value showed that,DpC had broad-spectrum antibacterial effect on clinical isolate K.pneumonia,CRE,VRE,CRPA and MRSA,with MIC value in the range of 16-32μg/m L,and exhibited synergistic antibacterial effect with meropenem on MβLs-producing bacteria,resulting in a 2-16-,2-8-,and 8-fold reduction in MIC of meropenem against EC-MβLs,EC01-EC24,K.pneumo nia,respectively.Moreover,Animal experiments showed that,DpC significantly reduced the infection of the clinically resistant bacteria K.pneumoniae in the spleen and liver of mice.In this work,DpC was identified to be a potent scaffold for the development of broad-spectrum inhibitors of MβLs.