Design And Synthesis Of Novel 3-hydroxypyridin-4(1H)-ones-based Hybrids As Pseudomonas Aeruginosa Biofilm Inhibitors

Bacteria resistance to antibiotics has become a global threat,and traditional antibiotics can no longer play a basic role in the treatment of resistant bacteria.Antibiotic resistance caused by biofilm formation is an important reason why chronic bacterial infection is difficult to cure.The chronic infection caused by the biofilm is easy to relapse and is also difficult to cure,so the treatment cost is greatly increased and the treatment time is prolonged.Therefore,the development of new antimicrobial agents for bacterial biofilm is expected to alleviate the worldwide problem of bacterial resistance.In this thesis,based on the 3-hydroxypyridin-4(1H)-one pharmacophore,we designed and synthesized 3-hydroxypyridin-4(1H)-one hydrids as Pseudomonas aeruginosa biofilm inhibitors,which provides a new direction for the subsequent search and development of potential biofilm inhibitors.Objective: There are three common quorum sensing(QS)systems in Pseudomonas aeruginosa: las,rhl and pqs systems,respectively.In the early stage of our research group,the series of hydroxypyrone and hydroxypyridone derivatives have been screened to show potent anti-biofilm activity.Among them,using the bio-isosterism principle,the hydroxypyridone derivatives obtained by replacing the quinoline ring of the pqs system signal molecule PQS(2-heptyl-3-hydroxy-4-quinolone)with the hydroxypyridone structure can interfere with iron uptake and simultaneously inhibit the pqs system,which is a new dual-target Pseudomonas aeruginosa biofilm inhibitor.Therefore,in this thesis we also use hydroxypyridone as the parent skeleton to splice other antibiofilm pharmacophore,hope to find more active Pseudomonas aeruginosa biofilm inhibitors and multi-channel quorum sensing inhibitors.Methods: Using classical medicinal chemistry splicing operations,the pharmacophore of las system inhibitors and pqs system inhibitors are combined with the parent skeleton of hydroxypyridone,to design and synthesize new hydroxypyridone hybrids,with a view to obtaining quorum sensing inhibitors for both las and pqs channels.Firstly,we take kojic acid as the raw material,and obtain the key intermediate at C-6 position via hydroxyl protection,amination(hydroxylation),and Gabriel synthesis reaction,then realizes amide condensation with the pharmacophore((methoxy)phenylamino)of the las system and virulence inhibitors,then deprotects the group to obtain the target products of series I hydroxypyridone hydrids;Secondly,we use kojic acid as the raw material,through reduction,aldol condensation,hydroxyl protection,amination,and Gabriel synthesis reaction to obtain the key intermediate at C-2 position.Similar to series I,series II hydroxypyridinone target products were obtained by amide condensation and deprotection;Series III target products were obtained by nucleophilic substitution reaction between the intermediate of hydroxypyridinone and the pharmacophore of pqs system inhibitor(benzimidazole);Based on isosteric principle,benzimidazole was replaced by benzoxazole and benzothiazole respectively to obtain series IV and V target products;Series VI target products were obtained by splicing hydroxypyridinone skeleton with a single heterocycle.Based on this,six series of hydroxypyridinone hydrids were obtained and their minimum inhibitory concentration(MIC)and biofilm inhibitory activity against Pseudomonas aeruginosa were determined.Results: In this thesis,six series of hydroxypyridinone hydrids were designed and synthesized,and a total of 73 compounds were obtained.All compounds were confirmed by 1H NMR,13 C NMR and HRMS.The results of biological activity showed that the MIC were all greater than 512 μM,and the compounds in series I and II showed weak biofilm inhibitory activity at 10 μM,indicating the effective groups((methoxy)phenylamino)of las system inhibitors are not good for biofilm activity;And the substituted hydroxypyridone derivatives at C-6 position basically show no biofilm inhibitory activity at 10 μM,indicating that C-6-position may be an invalid modification site.Interestingly,in the series of compounds Ⅲ,Ⅳ,Ⅴ and Ⅵ,most of them exhibited biofilm inhibitory activity at 10 μM.The structure-activity relationships of series Ⅲ,Ⅳ,Ⅴ and Ⅵ hydroxypyridinone compounds were summarized: 1.The activity of benzimidazole and benzoxazole skeleton splicing derivatives is better when 5-position is used as the substitution point,however,benzothiazole derivatives have more excellent activity when they take 6-position as the substitution site;2.In general,the substitution of electron withdrawing group is beneficial to enhance the activity,while the substitution of electron donating group is greatly weakened;3.The derivatives substituted by single heterocycle such as imidazole ring,thiazole ring and p-Nitrobenzene ring show better inhibitory activity,indicating that the combination may have synergistic effect;4.When hydroxypyridinone C-6position is the modification site the activity was almost lost.Among them,compound 28h(IC50= 10.57±1.54 μM)showed the best biofilm inhibition activity,and the biofilm inhibition rate was 50.85 ± 1.59% at the concentration of 10 μM,indicating that benzimidazole,a pharmacophore of pqs system inhibitor,may play an anti-biofilm synergistic effect after with the combination of hydroxypyridinone skeleton.Conclusion: Based on the parent skeleton of hydroxypyridone known to act on inhibition of Pseudomonas aeruginosa biofilm,through hybrid pharmacophore method,the 73 hydroxypyridone hybrids were synthesized in order to find new biofilm inhibitors.Fortunately,we discovered that 28 h of the seed compound with anti-biofilm activity,can inhibit the secretion of elastase and pyocyanin,as a quorum sensing inhibitor of the las and pqs dual-pathway,which provides a new idea for the research of Pseudomonas aeruginosa biofilm inhibitors.