Retroviral Vector Mediated Humane Ribonuclease Inhibitor Inhibites B16 Melanoma Growth In Vivo

BACKGROUND: Ribonuclease Inhibitor (RI) is an acidic glycoprotein with a molecular weight of around 50 KDa. One interesting function of RI is its antiangiogenesis effect against Angiogenin (Ang). Ang expresses highly and promotes angiogenesis in colon, liver tumor,etc. RI may inhibit angiogenesis through blocking on b-FGF pass way. Many experiments proved that angiogenesis is necessary to the tumor's growth and metastasis. Folkman et al figured out that antiangiogenesis could block the growth of tumors, and with little side effect. Antiangiogenesis therapy offers relative specialty on tumors from normal tissues and reduces tumor cells' tolerance to antitumor medication. The positive relationship of inhibition on tumor and blood plasma level of RI was confirmed by many researchers. Because of RI's short half life in blood plasma, it is difficult to administrate RI in vein with high efficiency as antitumor medication. In vivo gene therapy strategy might be a promising way to circumvent the puzzle. Nowadays, most of gene therapies are carried out with retroviral vectors. Retroviral vector pLNCX in this thesis was reconstructed from Moloney's Murine Leukemia Virus and Moloney's Murine Sarcoma Virus. Once transducted in packaging cell line PA317, retroviral vectors pLNCX are packaged. Thus, replication-incompetent retroviruses with target gene sequence subcloned are produced.AIM: To investigate the antitumor efficiency of in vivo gene delivery approach by pLNCX-based vector mediated human ribonuclease inhibitor (hRI) cDNA sequence. To assess safety of this gene therapy system to peripheral blood and hematopoietic system. METHODS: After packaged in package cell line PA317, recombinated retrovirus bearing hRI cDNA sequence are concentrated and purified. Controlled by virgin retroviral vector and PBS, the virions were injected intravenously into C57BL/6J mice bearing B16 melanomas that were inoculated subcutaneously. The expression of target gene in mice organs and tumors, the antiangiogenesis effect on melanoma growth and the impact of viral vector to hemopoietic system were examined. RESULTS: The transfected hRI gene expression product exerted antitumor growth and antiangiogenesis effect without examinable side effect on hemopoietic system. It is also found that hRI expressed highly in fibroblasts of tumor's fibrous capsule.CONCLUSION: Intravenous in vivo transfection of retroviral vector bearing hRI sequence suppressed the growth of B16 melanoma inoculated in C57BL mice through antiangiogenesis mechanism.