Function of P2X7 in healthy and diseased skin

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P2X7 is a purinergic receptor which responds to high extracellular ATP by opening a fast ligand-gatedcation channel or on prologued stimulation opening a pore permeable to large organic cations. Pore opening can initiate cellular apoptosis (1). P2X7 expression drives cancer cell growth and proliferation (2,3). Biosceptre has identified a novel partially non-functional form of P2X7 found to be in the many cancers which acts as a calcium channel without forming a large pore, promoting cell invasion and driving metastasis (4,5,6). Conformational changes on non-funtional P2X7 expose new epitopes which can be targeted for cancer therapy. Biosceptre has developed antibodies specific for this form of P2X7 which form the basis of a topical therapeutic that has shown indications of efficacy in treating SCC and BCC. The academic laboratory has focused on studying molecular mechanisms that underly the biology of. They have considerable expertise in working with both normal and human disease tissue with access to a large range of normal and disease cell lines avoiding animal modelsThe focus of this studentship is to examine the expression of both functional and non-functional P2X7 in normal human skin and keratinocyte derived cell lines including SCC. To examine the effects of siRNA and antibody mediated blockade on downstream signalling function in normal and SCC keratinocytes with particular emphasis on apoptotic pathways and investigating how this can be exploited as a therapeutic strategy to treat cutaneous cancer. In particular we wish to understand how targeting conformational changes on non-funtioning P2X7 can promote cell death through activation of apoptosis, receptor endocytosis and antibody dependent cellular cytotoxicity. This project aligns with the BBSRC strategy of Biosciences for Health; Biotechnology for Health and 3Rs. The student willCharacterise P2X7 expression using qPCR, Biosceptre antibodies and commercially available antibodies on a keratinocyte and SCC cell lines and tissue derived from both normal and patient SCCs..Investigate the expression of both functional and non functional isoforms of P2X7 in normal keratinocyte and SCC cell lines and their effect on signalling pathways, calcium second messenger generation, activation of specific kinases and transcription factors including CREB..The role of P2X7 in normal keratinocyte and SCC cell survival will be established using specific siRNA tools, alongside characterisation of the downstream consequences of ligand binding.. Model the response of keratinocytes and SCC cell lines to the effects of high extracellular ATP and in particular how high ATP concentration, a key component of the tumour microenvironment influence P2X7 expression and signalling..Pharmacological characterisation of the P2X7 receptor and its interactions with antibodies and siRNA that target P2X7 and establish the potential for Biosceptre therapeutics to modulate cell signal transduction 2 through P2X7 with regards targeting P2X7 to attenuate SCC in appropriate disease models and establish the pathways by which reagents targeting P2X7 affect SCC cellsReferences 1 Surprenant et al. Science 1996 May 3; 272(5262):735-8. 2 Jelassi et al. Oncogene 2011 May 5;30(18):2108-22. 3 Adinolfi et al. Cancer Res 2012 Jun 15;72(12):2957-69. 4 Slater et al. Breast Cancer Res Treat 2004 Jan;83(1):1-10. 5 Slater et al. Histopathology 2004 Mar;44(3):206-15. 6 Slater and Barden. Histopathology 2005 Aug;47 (2):170-8