Virtual Screening Of PKM2 Activators And Biological Activity Evaluation

Cancer has become a disease for urgent attention.Suppressing tumorigenesis and cancer cells proliferation become the problem that all researchers are eager to completely overcome.In the aerobic conditions,oxidative phosphorylation is the main metabolic way of normal cells,and in the absence of hypoxia is glycolysis(intracellular glucose produces pyruvate which is subsequently reduced to a large number of lactate and releases a small amount of energy).While,cancer cells have a different metabolism which is highly dependent on glycolysis instead of mitochondrial oxidative phosphorylation,regardless of oxygen availability.This aerobic glycolysis process was termed the Warburg effect,which was discovered by the German chemist Otto Warburg in the 1920 s.Based on the specific metabolic characteristics of tumor cells,more antitumor effective targets have been reported.The activation of tetramer formation of PKM2(M2 type pyruvate kinase)can suppress proliferation and growth of tumor cells effectively,which is considered as a very promising antitumor target.In this work,we try to find PKM2 activators based on the structure of PKM2,using the virtual screening based on molecular docking combined with a variety of evaluation methods and cytotoxicity test.At first,the combinations between PKM2 and compounds from four compounds libraries were evaluated by virtual screening based on molecular docking,including Chembrige Database,FDA-approved Drug Library,SPECS Database and ZINC(Natural Production)Database.Then 168 small molecules were hit by virtual screening according to the results of small molecular structure clustering and binding free energy calculation,and 126 compounds among them were finally selected for the next activity evaluation.Then,the cytotoxicity test(MTT method)was conducted,the results showed totally 12 compounds have significant antitumor effect.The IC50 values of 12 compounds were determined for three cancer cell lines,including human hepatoma cell line HepG2,human cervical cancer cell line Hela and human non-small cell lung cancer cell line A549.The results showed that the IC50 values of 12 compounds were all under 100?M,and the IC50 values of the compound S1 and the compound S13 selected from the SPECS database were below 10 ?M.Finally,we used the method of molecular dynamics simulation,combined with the calculation methods of MM-PBSA binding free energy and free energy decomposition to explore the interaction mode and mechanism between theactivators and PKM2.The results indicated that the van der Waals interaction and nonpolar solvation were the main driving forces of the interaction between the small molecule S1 and the target PKM2.The favorable contributions of the binding mode were from the following residues: Phe26 A,Lys311A,Leu394 A,Phe26B,Leu27 B,Met30B,Leu353 B,Ile389B,Tyr390 B,Leu394B and Glu397 B.By further analysis of the interaction mode,we found that residues Phe26 A,Leu394A,Phe26 B and Leu394 B formed a hydrophobic cavity in the active binding sites and formed a strong van der Waals interaction between S1.There were hydrogen bonds between S1 and residues Lys311 A,Tyr390B and Glu397 B,which were responsible to the stabilization the active conformation ofPKM2.The results of this thesis has important theoretical value and practical value for the discovery of PKM2 activator,which has significance of treatment of related cancers.