Comparison of Three Different Methods of Transfection for the Production of Recombinant Adenovirus Expressing Human Carcinoembryonic Antigen Gene.

Abstract

Adenoviral vectors (AdVs) are widely used as a gene delivery vehicle and vaccine design due to their genetic stability, transfer capacity of large genes, production at high titers, and remarkable efficacy of transduction. One of the most important applications of AdVs is in cancer immunotherapy. Tumor-associated antigens are overexpressed in cancer cells; however, they cannot induce immune responses sufficiently. Therefore, the immune system must be stimulated against these antigens to kill the cancer cells. This study described the construction steps of a recombinant AdV expressing human carcinoembryonic antigen (CEA) gene. Furthermore, in order to achieve a high titer of the virus, an efficient transfection was required. Three various transfection reagents were compared to achieve the best method of transfection. Carcinoembryonic antigen was cloned into the pAdV and transfected into the A293 cells using three different reagents, including polyethylenimine (PEI), calcium phosphate, and DMRIE-C. The PEI had the highest transfection efficiency, which was selected for the transfection of the recombinant plasmid. It has low toxicity for cells and is suitable for large-scale transfection. The virus produced in this study can be applied as a vaccine in cancer immunotherapy for stimulating the immune system against CEA-expressing tumors.