Caffeine-suppressed ATM pathway leads to decreased p53 phosphorylation and increased programmed cell death in gamma-irradiated leukaemic molt-4 cells

Ionising radiation (ir) is one of the main treatment modalities in oncology. however,we still search for substances which can radio-sensitize tumour cells. in this study we used caffeine,a non-specific ataxia-telangiectasia mutated kinase (atm) inhibitor,and studied its effect on the activation of the proteins involved in cell cycle control and the induction of apoptosis in human t-lymphocyte leukaemic molt-4 cells (p53 wt). we evaluated the expression of the tumour-suppressor p53 (itself and phosphorylated on ser15 and ser392),the cell cycle regulator p21,and the anti-apoptotic protein myeloid cell leukemia 1 (mcl-1). after treatment with 2 mm caffeine,the cells were irradiated by 1 or 3 gy,lysed and the proteins detected by western-blotting. apoptosis was determined by flow-cytometric annexin v/propidium iodine detection. irradiation by 1 or 3 gy induced p53 phosphorylation at ser15 and ser392 after 2 h with maximum after 4 h. adding caffeine significantly inhibited ser15 phosphorylation,which is atm-dependent but surprisingly also ser392 phosphorylation,which is atm-independent,suggesting that caffeine might have another cellular target (protein kinase). similarly,caffeine caused a substantial decrease in p21 in combination with both doses of ir and also mcl-1 was down-regulated. three days after irradiation,caffeine significantly increased induction of apoptosis. the atm/p53 pathway was suppressed by caffeine,which led to increased apoptosis accompanied by a p53-independent decrease in mcl-1. it also caused down-regulation of p21,which possibly contributed to the shortened cell cycle arrest necessary for effective dna repair and thus impeded radio-resistance. caffeine promotes the cytotoxic effect of ionising radiation and provides a possible platform for the development of new anti-cancer therapeutics known as radio-sensitizers.