Structure-activity Relationships And Mechanism Studies Of Piperlongumine And Optimization Of Keap1-Nrf2 Inhibitors

1.Design,synthesis and mechanism studies of piperlongumine derivativesPiperlongumine is a natural alkaloid isolated from the long pepper with multiple biological activities.It shows potent anticancer effect targeting cancer cells but leaves normal cells intact.Besides,it can enhance the potential therapeutic effect of some antitumor drugs.In recent years,with more and more widely use of chemical biology methods in the target identification of new compounds,activity-based probes(ABPs)have emerged as a new technology which is applied to study the protein function.Multi-functionalized probes can selectively label the proteins under particular status in complex biological specimen,that makes the study aimed at the separation,identification,distribution and function of these active proteins possible.On the basis of previously designed compound PL-51,thirty PL derivatives were synthesized.Most of them exhibited similar in vitro antitumor potency compared with PL-51 and PL.In particular,compound P15 and P18 showed superior cytotoxicity than PL against six tumor cells with IC50 values of nM level.Structure-activity relationships indicated that derivatives with single substituent on benzene-ring were better than derivatives with double and triple substituents.Under mechanistic study of the previously synthesized compound PL-23,it was found that PL-23 was not as same as PL to exert its antiproliferative activity via increasing cellular ROS level in cancer cells and also not targeting common kinases.Small molecule probes were applied to identify the target of PL-23.Active probe ZM76 and two inactive probe ZM77 and ZM78 were obtained and then confirmed the target gel band at the protein molecular weight of 40 kDa.Two potential proteins Annexin A1 and Annexin A2 got the highest score through initial MS analysis.2.Optimization of Keap1-Nrf2 protein-protein interaction inhibitorsKeap1-Nrf2-ARE signaling pathway plays a central role in oxidative stress related diseases such as inflammation and cancer.Nrf2 is an important intracellular transcription factor,negatively regulated by Keap1.Under stressed condition,Nrf2 can escape Keap1-mediated degradation and translocate into nucleus,activating ARE-dependent cytoprotective genes.There are few reports of Keap1-Nrf2 non-peptide molecule inhibitors with high potency indicating the study was in the stage.Fragment-Based Drug Design was applied to further optimize compound S47,a previously found Keap1-Nrf2 inhibitor,to obtain six potent derivatives possessing similar or superior activity compared to S47.In particular,compound AS01 interrupted the two proteins interaction with binding value KD2 of 1.138 ?M,which was six-fold potent over S47.Structure-activity relationships showed that acetoxy compounds were better than propionyloxy compounds.