P53-INDUCED APOPTOSIS IN THE HUMAN T-ALL CELL-LINE CCRF-CEM

The tumor suppressor p53 has been implicated in apoptosis induction an d is mutated in human T-ALL CCRF-CEM cells. To investigate possible co nsequences of wild-type p53 loss, we reconstituted CEM-C7H2, a subclon e of CCRF-CEM, with a temperature-sensitive p53 allele (p53ts). Stably transfected lines expressed high levels of p53ts and shift to the per missive temperature (32 degrees C) caused rapid induction of p53-regul ated genes, such as p21(CIP1/WAF1), mdm-2 and bax. This was followed b y extensive apoptosis within 24 h to 36 h, supporting the notion that mutational p53 inactivation contributed to the malignant phenotype. p5 3-dependent apoptosis was preceded by digestion of poly(ADP-ribose) po lymerase, a typical target of interleukin-1 beta-converting enzyme (IC E)-like proteases/caspases, and was markedly resistant to the ICE/casp ase-1 and FLICE/caspase-8 inhibitor acetyl-Tyr-Val-Afa-Asp.chloromethy lketone (YVAD), but sensitive to the CPP32/caspase-3 inhibitor nzyloxy carbonyl-Asp-Glu-Val-Asp.fluoromethylketone (DEVD) and benzyloxycarbon yl-Val-Ala-Asp.fluoromethylketone (zVAD), a caspase inhibitor with bro ader specificity. This indicated an essential involvement of caspases, but argued against a significant role of ICE/caspase-1 or FLICE/caspa se-8. Actinomycin D or cycloheximide prevented cell death, suggesting that, in this system, p53-induced apoptosis depends upon macromolecule biosynthesis. Introduction of functional p53 into CEM cells enhanced their sensitivity to the DNA-damaging agent doxorubicin, but not to th e tubulin-active compound vincristine. Thus, mutational p53 inactivati on in ALL might entail relative resistance to DNA-damaging, but not to tubulin-destabilizing, chemotherapy.