INFLUENCE OF BCL-2 OVEREXPRESSION ON THE CERAMIDE PATHWAY IN DAUNORUBICIN-INDUCED APOPTOSIS OF LEUKEMIC-CELLS

We have previously demonstrated that daunorubicin (DNR) induces apopto sis in some leukemic myeloid cell lines. We investigated a potential p rotective role for Bcl-2 in apoptosis induced by DNR in two leukemic c ell lines, one myeloid and one lymphoid, overexpressing the anti-apopt otic gene Bcl-2. Parental cells treated with DNR exhibited classical f eatures of apoptosis 6 h after drug exposure, all the cells being dead after 30-48 h. In contrast, overexpression of Bcl-2 significantly del ayed, but did not prevent the occurrence of DNR-induced apoptosis, wit h no surviving cells 96 h after drug exposure. To elucidate the mechan ism of the protection mediated by Bcl-2, we explored the signaling pat hway which initiates DNR-induced apoptosis. In this report, we show th at, in both the myeloid and lymphoid parental cell lines, DNR triggere d a sphingomyelin (SM) hydrolysis after 10-15 min with a concomitant c eramide generation. Moreover, exogenous ceramide induced DNA fragmenta tion in these cells, with levels similar to those observed with DNR tr eatment. In contrast, Bcl-2 overexpression protected the cells against apoptosis induced by ceramide treatment, without preventing the early SM hydrolysis nor the ceramide generation in these cells. Our results strongly suggest that Bcl-2-mediated protection of DNR-induced apopto sis is effected downstream of the SM-ceramide signaling pathway.