Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.712875
Title: Studies of ion channel mechanisms in pancreatic ductal adenocarcinoma (PDAC)
Author: Bonito, Benedetta
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2016
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Abstract:
Pancreatic Ductal Adenocarcinoma (PDAC) is the most common type of pancreatic neoplasia, accounting for more than 90% of all pancreatic cancer cases. Despite improvements in early diagnosis and treatment, the 5-year survival rate has remained unvaried for many decades. The aim of this PhD project was to elucidate the roles of two specific ion channels in the pathophysiology of PDAC: voltage-gated sodium channel (VGSC) and the Ca2+- activated K+ channel KCa3.1. Human pancreatic cell lines of different metastatic potential were used: HPDE (human pancreatic ductal epithelial) cells, Panc-1, MiaPaCa-2, BxPC-3 and Capan-1 cells. VGSCs, typically expressed in neuronal, ‘excitable’ cells, were also found earlier to be functional in several different carcinomas, where they are linked to promotion of the metastatic potential. KCa3.1, which serves a diversity of physiological roles such as cell-volume regulation in erythrocytes and migration in macrophages and microglia, may also promote tumour invasiveness as shown for some tumours including gliomas end endometrial cancer. Initially, the expression of different VGSC subtypes (Nav1.1 - Nav1.7) was screened in PDAC cell lines and abundant levels of mRNA and protein were found. However, whole-cell patch clamp recordings detected a small inward current (~ 30 pA/pF) only in the 10% of MiaPaCa-2 cells, one of the most aggressive cell lines available. Based on the PCR, western blot and patch clamp results obtained, we hypothesized that epidermal growth factor receptor (EGFR) might suppress VGSC expression to protect the Na+/H+ exchanger (NHE1), which has been proposed to serve a ’backbone’ function in driving PDAC metastasis by promoting extracellular acidosis. Indeed, when the EGFR pathway was blocked pharmacologically (by AG1478 10 μM) an increased inward Na+ current was detected by whole-cell patch clamp. Further experiments showed that a combination treatment of AG1478 and TTX (tetrodotoxin) and cariporide (an NHE1 inhibitor) and TTX could reduce cell migration further than treatments alone. This suggested that inhibition of the EGF-EGFR signalling cascade increases VGSC activity and this promoted metastatic cell behaviours. We also found that KCa3.1 channel was overexpressed at mRNA, protein and functional levels in PDAC compared to control cells . We demonstrated both pharmacologically and by gene - silencing approach that this channel regulated the migration and proliferation of MiaPaCa-2 cell line, revealing its potential as another target for therapy, as it has been also suggested in other carcinomas (i.e. endometrial cancer). These results are discussed in their respective chapters. The Thesis ends with a General Discussion chapter, where recent findings on the pathophysiology of PDAC are further highlighted in relation to the findings of the thesis.
Supervisor: Djamgoz, Mustafa Sponsor: Marie Curie (ITN) Network
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.712875  DOI: Not available
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