APOPTOSIS INDUCED BY MICROTUBULE DISRUPTING DRUGS IN CULTURED HUMAN LYMPHOMA-CELLS - INHIBITORY EFFECTS OF PHORBOL ESTER AND ZINC-SULFATE

The effects of the microtubule disrupting drugs (MDD) vinblastine, vincristine and colchicine on a human lymphoma cell line, BM 13674, were investigated. Twelve hours after administration of vinblastine (10(-3) mg/ml), VinCristine (10(-2) mg/ml) or colchicine (10(-2) mg/ml), cell death with the characteristic morphology of apoptosis was observed in 71.6%, 82.2% and 76.9% of the cells respectively. The mode of death was confirmed as apoptotic by the occurrence of internucleosomal DNA cleavage, which was demonstrated by agarose gel electrophoresis. For the purpose of casting light on the mechanism involved, inhibition tests were performed on apoptosis induced by one of these drugs, vinblastine, using a phorbol ester (PDBu), zinc sulphate and cyclobeximide. PDBu, an activator of protein kinase C, and zinc sulphate, a putative inhibitor of the endonuclease were thought to be responsible for internucleosomal DNA cleavage; both markedly reduced the induction of apoptosis. The protein synthesis inhibitor cyclobeximide, on the other band, bad no inhibitory effect. Moreover, cyclobeximide treatment per se enhanced apoptosis. This suggests that new protein synthesis is not required for the execution of vinblastine-induced apoptosis. Such a finding is in accord with recent reports suggesting that the ''death program'' within many cell types may be primed but unable to proceed due to concomitant production of specific ''apoptotic inhibitors''. It is suggested that phorbol esters prevent vinblastine-induced apoptosis in the BM 13674 cells by activating one or more of these specific ''apoptotic inhibitors'', possibly by means of PKC-mediated phosphorylation.