GROWTH-FACTOR AND BCL-2 MEDIATED SURVIVAL DURING ABORTIVE PROLIFERATION OF HYBRIDOMA CELL-LINE

Cultures of the CRL-1606 hybridoma (ATCC) have been reported to underg o continuous proliferation with simultaneous death during nutrient lim ited fed-batch fermentations, The bcl-2 proto-oncogene has been shown to prevent cell death under a variety of otherwise death inducing cond itions. We were interested in elucidating the nature of the massive de ath observed in cultures of CRL-1606, specifically with respect to the possible environmental causes, and the ability of overexpressed human bcl-2 (hbcl-2) to mitigate cell death. Abortive proliferation, or con tinuous proliferation in the presence of continuous death, could be in duced in serum free cultures of CRL-1606 through the withdrawal of ins ulin provided the culture was competent for cell proliferation. Cultur e competency for proliferation was found to be solely determined by th e presence of cell culture nutrients. Abortive proliferation was defec tive in cultures transfected with hbcl-2 and the enhanced viability ob served resulted from an increased viable cell population and at the ex pense of the nonviable cell population normally found in untransfected cultures. Abortive proliferation was also observed in serum containin g cultures upon serum shiftdowns. Like the insulin-supplemented serum free culture system, hbcl-2 transfected cultures exhibited defects in the abortive proliferation process. These results suggest that the mas sive death observed during nutrient-limited fed-batch fermentation ori ginate, in part, from growth or survival factor limitations. Hence, ap proaches to design cell culture media that account for the cell's prol iferation requirements without accounting for the cell's survival requ irements may represent. a cell death sentence. Given the transformed n ature of the hybridomas, we conclude that the abortive proliferation o f CRL-1606 is a consequence of inappropriate cell cycle entry in a sur vival factor limited environment. (C) 1998 John Wiley & Sons, Inc.