Cisplatin-induced apoptosis in human malignant testicular germ cell lines depends on MEK/ERK activation

Abstract

Testicular germ cell tumours (TGCT) represent the most common malignancies in young males. Whereas in 1970s, the survival rate in patients with metastatic testicular tumours was only 5%, these days, 80% of the patients treated by modern chemotherapy will survive their disease. The drug that revolutionised the cure rate for patients with metastatic testicular tumours was cisdiamminedichloroplatinum ( cisplatin, CDDP). In vitro experiments on neoplastic germ cell lines showed that their exquisite sensitivity to CDDP could be attributed to p53-dependent and -independent pathways. Applying cDNA macroarray, semiquantitative RT-PCR and Western blot analyses, blocking experiments, caspase activity assays, and morphological methods, we sought here to define the p53-independent pathway(s) involved in the CDDP-induced apoptosis. For this purpose, we used the human TGCT cell line NCCIT, the mutated p53 of which is known to remain inactive during the course of CDDP-induced apoptosis. Our experiments showed that within hours of CDDP application, two prototype members of the 'mitogen-activated protein kinase' ( MAPK) family, designated 'MAPK ERK kinase' (MEK) and 'extracellular signal-regulated kinase' ( ERK), were dually phosphorylated and caspase-3 became active. Functional assays using MEK inhibitors demonstrated that the phosphorylation of MEK and ERK was required for the activation of caspase-3 as the executing caspase. Interestingly, experiments with the human malignant germ cell line NTERA, which is known to possess wild-type p53, revealed the same results. Thus, our data suggest that CDDP mediates its p53-independent apoptosis-inducing effect on the malignant human testicular germ cells - at least partially - through activation of the MEK - ERK signalling pathway.