HUMAN LYMPHOBLASTOID CELL-LINES EXPRESSING MUTANT P53 EXHIBIT DECREASED SENSITIVITY TO CISPLATIN-INDUCED CYTOTOXICITY

Human lymphoblastoid cells were transfected with expression vectors co ntaining p53 cDNA mutated at either codon 135 or 246. The cells were s ubjected to cisplatin treatment or gamma-radiation and observed for ch anges in the cell cycle arrest and apoptosis, We found that compared t o the parental cell line, cells overexpressing mutant p53 (either 246( val) or 135(ser)) exhibited decreased apoptosis in response to gamma-r adiation or cisplatin as measured by: propidium iodide (PI) staining o f the cellular DNA (cell cycle analysis) and decrease in PARP (poly AD P-ribose polymerase) cleavage as detected by Western blotting. Interes tingly the cells expressing mutant p53(135(ser)) protein were less res istant to cisplatin-induced apoptosis than the p53(246(val))-bearing c ell line. A significant decrease in the G1/S arrest assayed by bromode oxyuridine and PI staining (cell cycle/proliferation assay) was also o bserved in response to irradiation and cisplatin in cell lines express ing either of the mutant p53 constructs. A lower basal level and reduc ed magnitude of protein induction of the cell cycle inhibitor p21/Waf1 was seen both after cisplatin and gamma-radiation treatment in the mu tant p53 expressing lymphoblastoid variant when compared to the wild t ype p53 parental cell line but induction of the p53 regulator MDM2 was comparable in both. No increase in basal levels of Bc12 protein in wi ld type or mutant p53 expressing cells was observed in response to cis platin or irradiation. Unexpectedly, following cisplatin treatment we observed an increase in mutant and wild type p53 RNA steady state leve ls in addition to increased levels of p53 protein. These results sugge st that irradiation or cisplatin treatment may not only stabilize wild type p53 protein but also may increase the steady state p53 RNA level s. Finally these results indicate that both irradiation and cisplatin should be used with caution in the treatment of lymphoid tumors bearin g mutations of p53.