Dose-dependent reduction of apoptosis in nutrient-limited cultures of NS/0myeloma cells transfected with the E1B-19K adenoviral gene

It is now well documented that apoptosis represents the prevalent mode of d eath in lymph old cultures and occurs spontaneously in late-exponential pha se of batch cultures following nutrient exhaustion. In an attempt to enhanc e the cell survival of these cell lines, we have initially engineered nonpr oducing NS/0 myeloma cells with a vector expressing the adenoviral E1B-19K protein. NS/0 cells transfected with E1B-19K were found to be more resistan t to apoptosis occurring in the late phase of batch culture and under stres sful conditions such as cultivation in glutamine-free medium or following h eat shock. In this study, we have characterised a number of NS/0 subclones constitutively expressing different levels of E1B-19K, as well as several s ubclones in which the expression of E1B-19K was regulated by a tetracycline -controllable gene switch. We have found that a threshold E1B-19K level was required in order to achieve protection against apoptosis. The extent of r esistance against cell death induced by nutrient deprivation in glutamine-f ree medium and in the late phase of batch cultures correlated with the leve l of E1B-19K expression up to an optimal level where further increases in E 1B-19K levels did not result in significant additional protection. To asses s the effects of E1B-19SK on antibody productivity, an apoptosis-resistant NS/0 clone was then transfected with a chimeric antibody construct. Despite their improved viability, the antibody productivity of E1B-19K clones in b atch culture was not significantly improved. Moreover, while the use of E1B -19K considerably delayed cell death, cells eventually died by apoptosis. S urprisingly, E1B-19K had no beneficial effect on the efficiency of fusion o f NS/0 myelomas and splenocytes for the generation of hybridoma cells. Furt hermore, the resulting hybridomas, although expressing E1B-19K at levels co mparable to the myeloma parent, were no longer resistant to apoptosis. This indicates that the ability of E1B-19K to prevent apoptosis is not only dos e-dependent but also seems to be cell-type dependent. (C) 1999 John Wiley & Sons, Inc.