Synthesis And Preliminary Evaluation Of Celastrol Analogues

ObjectivesA series of Celastrol analogues were synthesized. The antiproliferative activity of these analogues was studied with C6 and PC 12 cells. Their protective effect against oxidative stress was investigated on neuronal PC12 cells. To understand the neuroprotective mechanism of action, we determined the expression of Hsp70 proteins by Western blotting. Our purpose of the present work was to obtain new compounds with higher activity and less toxicity. Analysis of structure-activity relationship of the new compounds provides new information for future study.Methods1. Synthesize a series of Celastrol analogues.2. Characterize structures by 1H NMR and ESIMS.3. Antiproliferative Assay:Cells (C6, PC12) were grown to confluence in a humidified atmosphere (37℃,5% CO2). Compounds were added and cells were returned to the incubator for 48 h. Cell viability was measured by the MTT method. IC50 values were calculated from three independent experiments using SPSS.4. Cytoprotective effect against l-BHP-induced PC 12 cells damage:PC 12 cells were cultured. Drugs at different concentrations were added, and the cells were incubated at 37℃for 1 h. BHP (200μM) was then added, and the cells were incubated for another 24 h at 37℃under 5% CO2. Cell viability was measured by the MTT assay.5. Neuroprotective mechanism of action:Western blotting for Hsp70 was performed following standard procedure. Expression of Hsp70 protein was compared with Celastrol.Results1. New analogues Cel 1-13 were synthesized, and the structures of all new compounds were identified by 1H NMR and ESIMS.2. Analogues Cel 1-7 and Cel-13 showed better antitumor activity on PC 12 and C6 cells than DDP. Cel-1 and Cel-6 have considerable activity compare to the natural product.3. Cel-6, Cel-7 and Cel-13 inhibit t-BHP-induced PC 12 cell damage. Cel-13 was more effective in protecting cells from t-BHP-induced cell damage than its parent Celastrol. Both Celastrol and Cel-13 up-regulated the chaperone protein Hsp70 at the tested concentrations.ConclusionsCel 1-7 and Cel-13 were active on PC 12 and C6 cells. Cel-6, Cel-7 and Cel-13 blocked t-BHP-induced PC 12 cell damage at the tested concentrations. Celastrol and Cel-13 up-regulated the chaperone protein Hsp70. These results suggest that the protective effect and the Hsp70 protein expression were not well-correlated for Celastrol and Cel-13. It remains to be uncovered if other mechanisms of action were in play for their neuroprotective effects.