The design of a gene expression construct encoding bevacizumab and investigating its impact on colon cancer cell line model

Abstract

Introduction: Angiogenesis-targeted therapy of cancer is considered as a promising strategy for therapeutic management of cancer progression. The most widely used anti-ngiogenesis drug is bevacizumab which binds to VEGFA and prevents its interaction with VEGF receptor leading to suppression of angiogenesis. Despite the remarkable success in development of angiogenesis inhibitory mAbs, their clinical application is limited by the high-cost of mAbs-based regimen which includes multiple doses of mAbs due to their short biological half-life. Antibody gene transfer is an alternative system for production of antibody in vivo. Purpose: In this study, we have developed a gene-based anti-VEGF mAb system which is expected to produce a high concentration of anti-VEGFA mAb upon a single administration in cancer cells. Methods: The construct expressing anit-VEGF mAb was designed and cloned in pCDH lentivirus vector. The lentiviral particles was produiced in HEK-293 T cells. Bevacizumab expression in the transduced cells was confirmed by qRT-PCR and western blot at both the mRNA and protein level, respectively. Functional analysis of anti-angiogenesis bevacizumab produced in HT-29 cancer cells was performed by tube formation assay in a co-culture system included with endothelial cells. Results: The full-length cDNA bevacizumab light and heavy chains joint with T2A sequence were cloned in pCDH lentivirus vector. The lentiviral particles expressing bevacizumab was produced in HEK-293 T cells. Recombinant lentiviral particles (rLV-bev) containing bevacizumab efficiently transduced HEK-293 cells and produced functional bevacizumab mAb. Bevacizumab expression in the transduced cell was assessed by qRT-PCR and western blot at both the mRNA and protein level, respectively. The functionality of the recombinant bevacizumab was confirmed using the tube formation assay in the co-culture system of endothelial cells and HT-29 cells transduced with rLV-bev viral particles. Conclusion: Our results show that tLV-bev gene delivery system can be useful for angiogenesis-targeted therapy of cancer.