Study On Mechanism Of Sponge Derived Agelasine B Against Non-small Cell Lung Cancer

Objective:Screen the active compounds from natural products derived from sponge and study its mechanism.Contents and methods:Lung cancer is currently the world’s highest incidence and mortality of malignant tumors,and more than 80%were non-small cell lung cancer（NSCLC）,the five year survival rate is only about 15%.In recent years,significant progress has been made in the development of drugs for NSCLC from the cytotoxic drugs to low toxicity and targeted drugs,but there are still problems such as drug resistance and insensitivity.Overexpression of cyclin dependent kinase（CDK）in tumor tissues has been considered as an important target for anticancer drug development.At present,there is a class of CDK inhibitor such as palbociclib approved for clinical use,and nearly 20 CDK inhibitors including flavopiridol,roscovitine,dinaciclib,palbociclib were in clinical research.In this study,we isolated and screened a kind of alkaloid Agelasine B（AB）from Xisha sponge which has significant anti-lung cancer activity.Two human NSCLC cell lines PC9 and A549 were selected as the research object,and the cell biology and molecular biology techniques were used to elucidate the mechanism of natural product AB against NSCLC.Results:1.The natural product AB with anti-lung cancer activity and non-toxicity to normal cells was successfully isolated and screened.2.AB could significantly diminish the proliferative activities and markedly reduce the colony formation rate of NSCLC cells in a time-dependent and dose-dependent manner.3.AB could induce cell cycle arrest in G1 phase and apoptosis in NSCLC cells.4.AB could significantly increase the level ofγ-H₂AX expression,a marker of DNA double-strand breaks,and activate the release of intracellular ROS.5.Preliminary study of mechanism demonstrated AB downregulate the level of CDK2 gene and protein expression,and downregulate the expression of Cyclin E1 protein and Rb phosphorylation level.AB could activate ATR-p53-p21signaling pathway and inhibit DNA repair pathway Chk1-cdc25C.Inhibition of Cyclin E1,p-Rb and the activation of p21,p-p53 by AB were eliminated after knockdown expression of CDK2.Conclusion:These results suggest that AB can increase intracellular ROS release and induce DNA damage.AB causes cell cycle arrest,inhibits proliferation and induces apoptosis by downregulating CDK2 and activating ATR-p53-p21 pathway and inhibiting Chk1-cdc25C pathway.This study provides a theoretical and experimental basis for the development of AB as a lead compound for lung cancer.