Strategies for enhancing transient gene expression system of chinese hamster ovary cells in serum-free suspension culture

Abstract

Recombinant proteins are increasingly important in fundamental research and for clinical applications. As many of these recombinant proteins are of human or animal origin, cultivated mammalian cells are the host of choice to ensure their functional folding and proper posttranslational modifications. Large-scale transient trans-fection of human embryonic kidney 293 or Chinese hamster ovary (CHO) cells is now an established technolo-gy that can be used in the production of hundreds of milligram to gram quantities of a recombinant protein. Despite the time-consuming and labor-intensive process of stable cell line generation, stable CHO cell lines are most widely used as the production platform for therapeutic proteins in the pharmaceutical industry because they provide high yields and validated quality. However, in the early development phases, a number of therapeutic protein candidates should be available in a timely manner and in sufficient quantity. In mamma-lian cells, transient gene expression (TGE) rather than stable expression is well-suited to meet this requirement. In an effort to improve TGE in CHO cells, the combinatorial engineering of polyoma virus large T-antigen (PyLT) and Bcl-xL in CHO DG 44 cells was performed. The developed cell line (CHOP-off-Bcl-xL), which constitutively expresses PyLT and inducibly expresses Bcl-xL, is capable of episomal replication with the use of the DNA expression vector encoding PyOri, EBNA-1, and OriP (pWP-Ang-EBNA/OriP-PyOri) and it is apoptosis-resistant. When the recombinant antibody was transiently expressed, this combinatorial engineering in the CHO cells resulted in a more than two-fold increase in the product titer in various culture conditions such as batch, fed-batch, and cultures with sodium butyrate additions. Controlled proliferation is the method for enhancing the recombinant protein production in mammali-an cells by increase cell specific productivity. To improve transient gene expression (TGE) in Chines...