Daniel Rafael Peter Sauter:
Pancreatic ductal adenocarcinoma (PDAC) has one of the worst survival rates of all cancers with >95% of the affected dying from it. Despite of intensive efforts to develop new therapeutic strategies, only few drugs (e.g. gemcitabine, erlotinib) are currently approved for treatment, all exhibit only moderate survival benefits. Therefore, novel targets are urgently needed.
Cl- and K+ channels play integral roles in the regulation of critical cellular processes such as proliferation, apoptosis and migration. These processes are commonly perturbed in tumor cells. A phenotypic hallmark of cancer is its altered pH and Ca2+ homeostasis; a fact that is of particular relevance to tumor biology as many ion channels are regulated by these stimuli. Thus, the aim of this thesis was to elucidate the role of Ca2+-activated and pH – regulated channels in PDAC progression.
Patch-clamp recordings from several pancreatic cancer cell lines (Capan-1, BxPC-3 and AsPC-1) revealed Ca2+-activated Cl- current that showed the biophysical signature of ANO1 (TMEM16A). In line with this, application of ANO1-specific inhibitors and transient gene silencing of ANO1 abrogated this current. Using scratch wound healing assay, we identified ANO1 as a crucial player in cell migration. Surprisingly, no involvement of ANO1 in proliferation was found. We found an abundant functional expression of the intermediate conductance Ca2+-activated K+ channel (KCa3.1) in BxPC-3 and Mia PaCa-2 cell lines employing qPCR, Western blotting and patch-clamp techniques. Results of a transwell migration assay indicate involvement of KCa3.1 in promoting migration. Using a planar high throughput patch-clamp system we identified a pH-regulated K+ current in BxPC-3 cells that followed biophysical and pharmacological characteristics as described for K2P channels. Moreover, when exposed to BL1249, a channel activator of the K2P subunit TREK-1, both cell migration and proliferation was inhibited. In summary, we found that Ca2+-activated currents are pivotal for cellular migration and that pancreatic cancer cells exhibit pH-regulated K+ current that may drive tumor progression.