Deregulation of choline kinase in pancreatic cancer and its inhibition as a potential therapeutic strategy

Abstract

In this dissertation I describe the role of Choline Kinase Alpha (CHKA) in pancreatic ductal adenocarcinoma (PDAC) and its proposal as a therapeutic target. I show that PDAC cell lines and human PDAC overexpress CHKA. Immunohistochemistry (IHC) of human PDAC samples reveals an intense CHKA staining that is absent from normal pancreatic tissue and chronic pancreatitis. There is lack of correlation between cytoplasmic staining and survival but a positive correlation between nuclear staining and survival in well and moderately-differentiated tumors. In PDAC cells CHKA is important for cell proliferation; genetic approaches and pharmacological inhibition support this hypothesis. PDAC cell lines express high levels of CHKA compared with non-malignant pancreatic cell lines (HPDE/HPNE) and similar to CHKA overexpressing tumors. Genetic CHKA down-regulation, using a specific shRNA, decreases growth rate by more than 50% in all cell lines without morphological changes. I assessed the therapeutic potential of CHKA inhibitor, MN58b. This drug induces apoptosis on several PDAC cells lines depending on their CHKA levels suggesting that it is a predictive marker. MN58b is also effective against Pancreatic Stellate Cells (PSC) one of the main cell types of PDAC stroma that contributes to its chemorresistance. MN58b also synergizes with different chemotherapeutic agents (Gemcitabine, Oxaliplatin and 5-Fluorouracil) commonly used in PDAC treatment. Genetic CHKA down-regulation sensitizes PDAC cell lines to these drugs. Moreover gemcitabine-resistant PDAC cells show enhanced sensitivity to MN58b. Resistance to chemotherapy is a general problem in Oncology. For that reason I went beyond the therapeutic potential of MN58b and generated a MN58b resistant cellular model. Resistant cells show less choline (CHO) uptake, slower proliferation and reduced migration capacities than its counterpart. I describe ABCB-1 and ABCB-4 up-regulation as the mechanism of resistance. These two proteins belong to the multidrug resistance proteins family (MDR), and extrude drugs from inside of the cell. Consistently with this mechanism, blockade of these trans-membrane proteins restores the original MN58bsensitivity. In summary this data suggest that CHKA support PDAC development and its pharmacological inhibition could be a new therapeutic approach.