Enhanced sensitivity to cis-diamminedichloro-platinum (II) of a human carcinoma cell line with mutated p53 gene by cyclin-dependent kinase inhibitor p21WAF1 expression

Abstract

p53-mediated induction of p21WAF1, a cyclin-dependent protein kinase inhibitor, is known to protect cancer cells from the cytotoxic effects of anti-cancer drugs or γ-irradiation. Since the p53 gene is frequently inactivated in cancer cells, we examined whether p21WAF1 expression may alter the sensitivity of cancer cells with mutated p53 gene to anti-cancer drugs. Cells of a colon cancer cell line DLD-1 were transfected with p21WAF1 expression vector controlled by a tetracycline-repressable promoter and transfectants were cloned (Dp21-1). p21WAF1 expression induced by removal of tetracycline from culture media repressed cell proliferation and resulted in altered cell shape, suggesting induction of differentiation. Dp21-1 cells with p21WAF1 expression were more sensitive to cis-diamminedichloropiatinum(II) (CDDP) (IC50 value, 10 μM than those without p21WAF1 expression (IC50, 22 μM). Sensitivity to doxorubicin was not different between Dp21-1 cells with and without p21WAF1 expression. DNA ladder formation was observed in Dp21-1 cells treated with CDDP, indicating that the enhanced sensitivity to CDDP involves apoptosis. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of cytosolic protein revealed that subunit protein bands with Mr 55 kDa and 44 kDa were markedly increased in cells with p21WAF1 expression. By immunoblotting, these proteins were identified as c-Jun N-terminal kinase (JNK) 2 and p38 mitogen-activated protein kinase (MAPK) δ, respectively, both of which are believed to be involved in apoptosis induction by CDDP. These results suggest that p21WAF1 may enhance the sensitivity of colon cancer cells with mutated p53 gene to CDDP, possibly through the JNK and p38 MAPK pathways.