Anti-tumor Activity Screening And Mechanism Exploration Of Lucidum Meroterpenoids

Objective: Screening series of compounds which own great anti-tumor cell activity of lucidum meroterpenoids and researching the mechanism of compound with strong anti-tumor activity obtained by screen.Researching the absorption,distribution,metabolism and excretion in vivo with the help of pharmacokinetic study,thus providing possible experimental basis for the research and development of new anti-tumor drugs.Methods: CCK8 combined with the scratch methods were used to detect cytotoxicity and cell migration inhibition respectively,thus screening small molecule compound LZ28,which owns great anti-tumor activity of human esophageal cancer KYSE30 cell line.Furthermore,the influence of LZ28 on cell apoptosis,cell cycle,ROS production was analyzed,using flow cytometry,and the protein expression of autophagy marker LC3 B and P62 was detected by western blot.GFP-RFP-LC3 B double fluorescent transfection assay was used to investigate the effect of compound LZ28 on autophagy flow.Observing cell morphology by means of transmission electron microscopy,we found that LZ28 can inhibit autophagy.What’s more,LZ28 can enhence the sensitivity of cisplatin,which is one of the clinical anticancer drugs.The bioactivity of small molecule probes and prototype compound was decteded by CCK8 method.Affinity proteomics identifies and enriches target proteins of LZ28.Specific bands of small molecule probes were anayzed by western blot and silver staining.Fluorescence gel electrophoresis was used to verify the binding force between target proteins and LZ28.The function of target proteins was dected by CCK8 method.We detected the pharmacokinetic parameters of the LZ28 by liquid-mass spectrometry.Results: LZ28 significantly inhibites the growth and migration of esophageal cancer cell KYSE30 and induces non-apoptotic cell death.It also inhibits the production of ROS,blocks the cell cycle in G2 phase,up-regulates the protein expression of LC3 B and P62 and inhibits autophagy.It enhances the sensitivity of anticancer drug cisplatin(CDDP).The small molecule probe SZU086 has similar cytotoxicity to the prototype compound LZ28.The molecule weights of specific bands of small molecular probes were mainly distributed in the range of 25-35 、35-45 、 40-55 k Da.The target protein STING specifically binds to SZU086.Pharmacokinetic experiments show that the viscera clearance rate of LZ28 is more higher,the half-life period is around 4.6 h,and the bioavailability is 9.17%.Conculsion: Lucidum meroterpenoids have good anti-tumor activity and can inhibit the growth and migration of esophageal cancer cells.The compound LZ28 may exert its anti-tumor effects through cell cycle arrest and autophagy inhibition.LZ28 has good solubility and tolerance,but low bioavailability in vivo.The functional target protein of LZ28 still needs to be further searched and verified.