Mcl-1 Overexpression Leads to Higher Viabilities and Increased Production of Humanized Monoclonal Antibody in Chinese Hamster Ovary Cells

摘要

Bioreactor stresses,including nutrient deprivation,shear stress,and byproduct accumulation can cause apoptosis,leading to lower recombinant protein yields and increased costs in downstream processing. Although cell engineering strategies utilizing the overexpression of antiapoptotic Bcl-2 family proteins such as Bcl-2 and Bcl-x_L potently inhibit apoptosis,no studies have examined the use of the Bcl-2 family protein,Mcl-1,in commercial mammalian cell culture processes. Here,we overexpress both the wild type Mcl-1 protein and a Mcl-1 mutant protein that is not degraded by the proteasome in a serum-free Chinese hamster ovary (CHO) cell line producing a therapeutic antibody. The expression of Mcl-1 led to increased viabilities in fed-batch culture,with cell lines expressing the Mcl-1 mutant maintaining ~90% viability after 14 days when compared with 65% for control cells. In addition to enhanced culture viability,Mcl-1-expressing cell lines were isolated that consistently showed increases in antibody production of 20-35% when compared with control cultures. The quality of the antibody product was not affected in the Mcl-1-expressing cell lines,and Mcl-1-expressing cells exhibited 3-fold lower caspase-3 activation when compared with the control cell lines. Altogether,the expression of Mcl-1 represents a promising alternative cell engineering strategy to delay apoptosis and increase recombinant protein production in CHO cells.