Apoptosis-resistant E1B-19K-expressing NS/0 myeloma cells exhibit increased viability and chimeric antibody productivity under perfusion culture conditions.

Abstract

We have shown previously that recombinant NS/0 myelomas expressing sufficient amounts of E1B-19K were resistant to apoptosis occurring in the late phase of batch culture and under stressful conditions such as cultivation in glutamine-free medium or following heat shock. However, no significant increase in monoclonal antibodies (MAb) was observed during the prolonged stationary phase of these batch cultures. Here, we show that E1B-19K can enhance cell survival and improve MAb productivity in high cell density perfusion culture. Typically, lymphoid cells grown under steady state in perfusion exhibit decreasing viabilities with concomitant accumulation of apoptotic cells. By modulating the ability of these cells to resist to induction of apoptosis in low nutrient environment, a 3-fold decrease in specific death rate from 0.22 day-1 for NS/0 control to 0.07 day-1 for E1B-19K cells was achieved, resulting in a significant improvement in cell viability throughout perfusion. E1B-19K cells at the perfusion plateau phase also exhibited a 3-fold reduction in specific growth rate concomitant with a lower percentage of S and higher percentage of G1 phase cells. This was associated with a 40% decrease in specific oxygen consumption rate, likely related to a reduction in the specific consumption rates of limiting nutrient(s). Expression of E1B-19K consequently had a significant impact on the steady-state viable cell density, allowing maintenance of 11.5 x 10(6) E1B-19K cells/mL versus 5.9 x 10(6) control NS/0 cells/mL for the same amount of fresh medium brought into the system (half a volume per day). Whereas MAb concentrations found in perfusion culture of control NS/0 myelomas were almost 3-fold higher than those found in batch culture; in the case of E1B-19K-expressing myelomas, the MAb concentration in perfusion was more than 7-fold higher than in batch. This was attributable to the 2-fold increase in viable cell plateau and to a 40% increase in the perfusion to batch ratio of specific MAb productivity (2.2-fold for E1B-19K myelomas versus 1.6-fold for NS/0 control).