IP 3 receptor antagonist, 2‐APB, attenuates cisplatin induced Ca 2+ ‐influx in HeLa‐S3 cells and prevents activation of calpain and induction of apoptosis

Background and purpose: Cisplatin drives specific types of tumour cells to apoptosis. In this study we investigate the involvement of intracellular calcium ([Ca 2+ ] i ) in triggering apoptosis in two different cell lines. As cisplatin is used for the treatment of several forms of cancer we choose HeLa‐S3 and U2‐OS as two examples of tumour cell lines. Experimental approach: Cisplatin (1nM–10μM) was applied to HeLa‐S3 and U2‐OS cells and [Ca 2+ ] i measured with fluo‐4, using laser scanning microscopy. Inositol‐1,4,5‐trisphosphate (IP 3 ) receptors were visualized with immunostaining. Membrane conductances were measured with patch‐clamp techniques. Levels of calpain and caspases were assessed by western blots and apoptotic cells were stained with Hoechst 33342 and counted. Key results: Cisplatin increases [Ca 2+ ] i concentration‐dependently in HeLa‐S3 but not in U2‐OS cells. This elevation of [Ca 2+ ] i depended on extracellular Ca 2+ but was reduced by the IP 3 receptor blocker, 2‐APB. This effect was not due to a Ca 2+ release triggered by Ca 2+ entry. Immunostaining showed IP 3 ‐receptors (type1‐3) at the cellular membrane of HeLa‐S3 cells, but not in U2‐OS cells. Electrophysiological experiments showed an increased membrane conductance with cisplatin only when Ca 2+ was present extracellularly. Increase of [Ca 2+ ] i was related to the activation of calpain but not caspase‐8 and triggered apoptosis in HeLa‐S3 but not in U2‐OS cells. Conclusions and implications: Our observations on the activation of IP 3 ‐receptors, calcium entry and apoptotic rate by cisplatin in specific carcinogenic cells might open new possibilities in the treatment of some forms of cancer. British Journal of Pharmacology (2007) 151 , 1176–1186; doi: 10.1038/sj.bjp.0707335