Jones, A.W.E.G.; (2011) Altered cellular signalling and metabolism in cisplatin cytotoxicity and chemoresistance. Doctoral thesis , UCL (University College London).

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Abstract

Cisplatin (cDDP) is a potent chemotherapeutic agent used in the management of a range of tumours, however its clinical efficacy is limited by tumour cell chemoresistance. Whilst the major target of the drug is considered to be nuclear DNA, cDDP has been shown to interact with other organelles, notably the ER and mitochondria, although the importance of these interactions is not fully understood. The contribution of such extranuclear effects is here addressed through comparative studies of two lung adenocarcinoma cell lines with differing sensitivity towards cDDP. Exposure to cDDP induced proliferative arrest and caspasedependent apoptosis in A549-WT cells, without requirement for caspase-8, calpains, ROS or Ca2+. Two-fold resistance towards cDDP-induced apoptosis was measured in A549-CR cells, generated through long term exposure to the drug. A549-CR cells showed marked inhibition of Ca2+ signalling from the ER due to a 50% reduction of IP3R function. However, cDDP treatment did not alter cytosolic Ca2+ levels nor deplete the ER Ca2+ store. Further, neither chelation of intracellular Ca2+ nor pharmacological inhibition of IP3R Ca2+ channel opening inhibited cDDP-induced A549-WT cell death, thus under the present experimental conditions Ca2+ flux through the IP3R does not modulate cDDP sensitivity. Basal and maximal cellular O2 consumption in A549-WT cells significantly increased following cDDP treatment, but was unaltered in A549-CR cells. Sequencing of mtDNA identified a novel point mutation in A549-CR mtDNA causing a 50% reduction of complex I activity. Mutations of mtDNA and reduced respiratory chain activity might inhibit apoptosis. The abundance of complex I was increased by 50%, potentially an example of pro-survival mitochondrial-nucleus compensatory retrograde signalling. Thus, key differences in mitochondrial function and response to cDDP which may alter A549 cell sensitivity to cDDP have been identified for further investigation.

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