Transient transfection of serum-free suspension F2N78 cell culture for efficient production of human therapeutic protein

초록

Transient transfection can be used for the production of recombinant proteins over a short period (1–14 days) by direct DNA transfer into single-cell suspension cultures. This process has been developed mainly with Chinese hamster ovary (CHO) and human embryonic kidney (HEK293) cells for the production and regulatory approval of therapeutic proteins. Recombinant HEK293 cells constitutively expressing Epstein-Barr virus nuclear antigen-1 (EBNA-1) has been responsible for high levels of plasmid amplification. Because of this enhanced productivity, HEK293 EBNA-1 (293E) is now the most widely used cell line for large-scale transient transfection. Another reason for the wide application of these cells is the easy growth in serum-free conditions. Several improved clones are commercially available for research as well as industry. However, 293E cells are not able to express EBNA-1 gene, so that they cause difficulties on gene expression and engineering of vector DNA. A human hybrid cell line named F2N78 was developed by somatic fusion of HEK293 and Namalwa (Burkitt’s lymphoma) cells. F2N78 cell line inherited many advantageous phenotypes such as ease to suspension, high transfection efficacy, Epstein-Barr virus (EBV) genome insertion in chromosome, and human-specific glycosylation. Because of EBV genome insertion in chromosome, autonomous replication of oriP vector maintained for long time. For that reason, this cell line can be used for the transient transfection for the production of therapeutic proteins. In this study, we compared the potential of F2N78 cells for transient transfection with CHO-K1 and HEK293 cells in serum-free suspension cultures. Through transfection using polycationic polymer polyethylenimine (PEI) at high cell density, human IgG could be produced within 4 days. The constructs used in this study were based on the oriP vector.