Experimental Study Of LOHP Combining Liposome-mediated Cytosine Deaminase-endostatin Fusion Gene Delivery On The Treatment Of Rectal Cancer

Background:Rectal cancer is one of the popular malignant tumors in our country. A large number of statistical data indicates that the rates of incidence and mortality of the disease in our country have been going up in recent years. The limitation of conventional cancer therapy derives from the imitis on their inability to ablate cancer without excessively damaging the patient. Endostatin, an angiogenesis inhibitor tested in multiple clinical trials, selectively targets neovascular endothelial cells, suppressing tumor growth. To enhance the therapeutic efficacy, we fused endostatin gene with cytosine deaminase gene, which converts a prodrug 5-flucytosine into a cytotoxic 5-fluorouracil.We reasoned that an endostatin-cytosine deaminase fusion protein should possess tumor-targeting property of endostatin to allow selective tumor-killing effect of cytosine deaminase in the presence of 5-FC, in addition to antiangiogenesis activity of endostatin, and thus might produce potent antitumor activity. However, the safety and the transfection efficiency are two very important puzzles faced by gene trepapy. Our research is to slolve these problems.Objective:Aims To investigate the effect of oxaliplatin (LOHP) on liposome-mediated gene delivery .we also want to examine whether liposome-mediated CD-Endostatin gene delivery repeatedly combining LOHP can enhance killing of rectal cancer HR-8348 cells xenografts in nude mouse.Methods (1) The recombinant plasmid containing CD-Endostatin gene was constructed.Prior to liposome transfection, HR-8348 cells were treated with LOHP at doses of IC50. After 36 hours of liposome transfection, green fluorocytes were counted. The transfection efficiency was figured out and compared with each other. (2) The cytotoxicity and therapeutic effects of LOHP combined with CD-Endostatin gene and 5-FC were evaluated by MTT assay. (3) A murine transplant model was established by subcutaneous inculation 1×106 HR-8348 cells into nude mouse. Befor delivered with liposome-mediated CD-Endostatin gene repeatedly ,tumors were treated with LOHP. At the same time, mice were injected i.p.with 500mg/kg/day of 5-FC for 12 days. Three control groups were set up respectively. Ten mice are in each group. Twenty-eight days after implantation, tumor size and weight, inhibition rate,intratumoral microvessel density(MVD) were evaluated respectively after the mice were sacrificed.Results With treatment with IC50 LOHP,the plasmid transfer efficiency in the HR-8348 cancer cells was improved about 18 times. The combination of LOHP and liposome-mediated gene delivery can achieve the approximate transfection efficiency of virus vector. In contrast with the controls, The tumors treated with LOHP combining liposome-mediated cytosine deaminase-endostatin gene grew very slowly and tumors regression reached to 79.69% of the weight of the blank control and 81.5% of the volume of the blank control. The MVD was decreased significantly in the group.There was a synergistic benefit for LOHP combining with liposome-mediated cytosine deaminase-endostatin gene-therapy of having stronger inhibitory effects and this was statistical significance (P<0.01).Conclusion LOHP can improve the transfection efficiency of liposome-mediated gene delivery markedly. The results suggest the combination of CD-Endostatin fusion gene and LOHP therapy lead to pronounced anticancer effect without any notable toxicity.