Study On The Anticancer Activity And Underlying Mechanism Of A Cytotoxic Peptide LVTX-8

Cancer is one of the life-threatening diseases in the world.Lung cancer is the primary type of cancer responsible for patient death globally.The incidence and mortality of lung cancer is high in China,and it showed an upward trend year by year.The common disadvantage of traditional chemotherapeutic drugs is side effects and drug resistance.Therefore,it is urgent to develop a new type of anticancer drug which is safe,and efficient.Spider toxin is a mixture of active substance containing multiple biological toxins.Most of the spider toxins are polypeptide toxins with different structures and functions.Some of spider toxins which were proved to target ion channel have been a useful tool for investigating ion channel.A few spider toxins which need further studies have showed remarkable anti-cancer activity both in vitro and in vivo.Our laboratory has deep foundations in studying spider toxins.The single components of polypeptide toxin in many spider toxins have been separated,purified and identified.We have constructed c DNA library from many spiders including Lycosa vittate.We have found a cytotoxic peptide LVTX-8 in the c DNA library of Lycosa vittate based on the existing abundant toxins resources in our laboratory to do the further researched on anti-cancer activity and mechanism.LVTX-8 is anα-helix,amphipathic,and cytotoxic anti-cancer peptide（ACP）with the ability to penetrate membranes.Its molecular mass is 2849 Da.After solid-phase peptide chemical synthesis,HPLC separation,mass spectrometry analysis and identification,we have obtained LVTX-8 with high purity.The cytotoxic activity of LVTX-8 against non-small cell lung cancer cell lines（A549 and H460）is high with IC₅₀ value 5μM through the CCK-8 assay.In order to further explore the anti-cancer activity of this cytotoxic peptide and its mechanism,colony formation experiments,Transwell anti-migration experiments and anti-invasion experiments were used to detect the effect of LVTX-8 on non-small cell lung cancer cell lines in vitro.Experiments showed that 5μM LVTX-8 can significantly inhibit the proliferation,migration and invasion of non-small cell lung cancer cell lines in vitro（P<0.05）.Due to the strong cytotoxic activity against cancer cells in vitro,we carried out animal experiments for further investigation in vivo.The results of the xenografts tumor model of nude mice showed the tumors of nude mice treated using LVTX-8 with dose of 10mg/Kg every other day are smaller and the survival time of nude mice are longer after subcutaneous injection.Then,TUNEL experiment（Terminal Deoxynucleotidyl Transferase（Td T）-mediated d UTP Nick-End Labeling（TUNEL）assays）was used to detect the dissected tumor.Compared with the control group（injected with PBS）,there are more apoptotic cells in the tumor tissue,which is consistent with the results in vitro measured by flow cytometry.Therefore,LVTX-8 can promote the apoptosis of cancer cells in a concentration-dependent manner.Lung metastasis model of nude mice were conducted by injecting non-small cell lung cancer cells through the tail vein.And the results showed that 10 mg/kg LVTX-8 injected into nude mice every other day can significantly inhibit the migration and metastasis of lung cancer.In this study,the cytotoxic peptide LVTX-8 was found to have good anti-cancer activity in vivo and in vitro,and it also can inhibit tumor migration,metastasis and invasion both in vivo and in vitro.In addition,we found that LVTX-8 may promote tumor cell apoptosis and inhibit tumor cell growth,migration and invasion by activating genes in the p53 pathway through transcriptomics analysis.Our research provides a vital research paradigm and reference for the investigation of anti-cancer activity and mechanism of similar cytotoxic peptides.