Apoptosis of haematopoietic cells upon thymidylate synthase inhibition is independent of p53 accumulation and CD95-CD95 ligand interaction

Treatment of haematopoietic BA/F3 cells with the thymidylate synthase inhib itor 5-fluoro-2'-deoxyuridine (FUdR) activated apoptosis through a mechanis m that required continuous protein synthesis and was inhibited by Bcl-2 ove r-expression. Analysis of p53 levels in cells treated with FUdR indicated a marked accumulation of this protein, Accumulation of p53 was also observed in cells over-expressing Bcl-2, In BA/F3 cells transfected with a cDNA cod ing for the human papilloma virus protein E6, p53 accumulation after FUdR t reatment was inhibited markedly. However, apoptosis was induced in both con trol and E6 cells to a similar extent. The role of the CD95/CD95 ligand (CD 95L) system in FUdR-induced apoptosis was also assessed. As determined by r everse transcriptase PCR, BA/F3 expressed a low constitutive level of CD95L mRNA, which decreased following FUdR treatment. Moreover, blocking CD95-CD 95L interactions with antagonistic CD95 monoclonal antibody did not prevent drug-induced apoptosis. Furthermore, analysis of caspase involvement showe d important differences in apoptosis induced by CD95-triggering or FUdR tre atment. In summary, these results suggest that apoptosis induced by thymine less stress in haematopoietic BA/F3 cells occurs by a mechanism that does n ot require accumulation of p53 and which is independent of CD95-CD95L inter actions.