Anti-tumor Effects Of The Intrabody Against CDK4 On Liver Cancer

Liver cancer,especially hepatocellular carcinoma(HCC),is a malignant tumor with high morbidity and mortality in the world.It is a serious harm to human life and health,and currently lacks effective treatment methods for liver cancer.China has always been a high-risk area of liver cancer,and the incidence of liver cancer is also increasing year by year.Therefore,it is of great scientific and social significance to explore effective prevention and treatment strategies for liver cancer.Uncontrolled cell cycle regulation is one of the important causes of malignant proliferation of tumor cells.CDK4 is the first activated cyclin-dependent kinase that enters the proliferative phase and is the rate-limiting factor in G1/S transition during cell cycle progression.Overexpression of CDK4 occurs in many types of human cancers,including hepatocellular carcinoma.Intrabodies are antibodies that are expressed in cells and act on intracellular components.They can target intracellular specific antigens,knock out important target protein functions in cells.Intrabody,as a new class of molecules,has been showing great potential and application prospects in anti-tumor gene therapy.In this study,the experiments by cell lines as well as at the animal models have demonstrated the anti-tumor effects of intrabody targeting the CDK4(NLS-AK)on liver cancer.RT-PCR,western blot,and indirect immunofluorescence experiments demonstrated the expression and localization of anti-CDK4 intrabodies in HepG2 cells.Co-immunoprecipitation experiments showed that anti-CDK4 intrabodies could specifically bind to endogenous CDK4 in HepG2 cells.MTT assay showed that NLS-AK could inhibit the growth and proliferation of HepG2 cells.Flow cytometric analysis showed that the expression of NLS-AK in HepG2 induced cell cycle G1 arrest and cell apoptosis.Wound healing assay showed that NLS-AK expression could significantly inhibit the migration ability of HepG2 cells.We established a treatment model for allograft tumors of liver cancer,and found that NLS-AK could effectively inhibit the growth of hepatocarcinoma transplanted in nude mice,and it showed a good effect on inhibiting the development of liver cancer.This study provides experimental evidence for clinical treatment of liver cancer by targeting CDK4 intrabody.