Study On The Anti-melanoma Activity And Target Proteins Identification Of Dehydrocostus Lactone

In recent years,the incidence of melanoma has shown an increasing trend.However,there is still a lack of effective drugs for the treatment of melanoma.As the source of new drug development,natural compounds have important research value.Dehydrocostus lactone is an effective active ingredient of Saussurea lappa(Decne.)C.B.Clarke which has great inhibitory effects on various cancers such as breast cancer and pharyngeal cancer.However,there are few researches on melanoma,and its specific target and mechanism are still unclear.The development of chemical proteomics based on probe and mass spectrometry provides a powerful technical support to elucidate the target proteins and mechanism of action of natural drugs.Chemical proteomics can realize the detection and identification of trace proteins,which can effectively screen out the target proteins of drugs.Therefore,the purpose of this study is to research the anti-melanoma activity of dehydrocostus lactone and identify the target proteins interacting with it.In A375 cell lines,the activity of dehydrocostus lactone was detected from the aspects of cell proliferation,cell cycle,apoptosis and migration,and the expression of related proteins,as well as the changes of intracellular MMP and ROS.Then,a dehydrocostus lactone probe modified with terminal alkynyl benzophenone was designed and synthesized,and it was characterized by mass spectrometry,nuclear magnetic resonance spectroscopy and anti-melanoma activity.The probe was used to indirectly link the target proteins to the biotin through click chemical reaction,and the biotin-streptavidin system was used to catch the proteins.Finally,the peptides were labeled with a novel iSIPL2-180/181 labeling reagents,and the target proteins were identified by LC-MS/MS.The results showed that dehydrocostus lactone inhibited the proliferation of A375 cells.The IC50 of dehydrocostus lactone was 3.886 ± 0.596 μM after 24 h of treatment.Dehydrocostus lactone could increase intracellular ROS content,decrease MMP,induce apoptosis and G2/M phase cell cycle arrest,and up-regulate the expression of p53,caspase-8,cleaved caspase-3 and cleaved PARP,as well as down-regulate Cyclin B1 and CDK1 in A3 75 cells.A total of 69 target proteins that may interact directly with the molecule were identified by using the chemical probe.These target proteins are involved in the regulation of multiple pathways such as cell metabolism,survival and death,including the PI3K-Akt signaling pathway.In addition,the most influential proteins on the antimelanoma activity of dehydrocostus lactone were screened out,such as ENO1,HSPA8,HSP90 and so on.In conclusion,dehydrocostus lactone showed great anti-melanoma activity in this study,and it has multiple protein targets.In this study,the anti-melanoma activity of dehydrocostus lactone was studied for the first time,and its possible target proteins were identified,which laid a research foundation for elucidating its mechanism of action,and also provided a theoretical basis for its future use as a clinical anti-tumor drug candidate.