Publication Date

2014-04-30

Availability

Embargoed

Embargo Period

2016-04-29

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PHD)

Department

Molecular Cell and Developmental Biology (Medicine)

Date of Defense

2014-01-09

First Committee Member

Theodore J. Lampidis

Second Committee Member

Vinata Lokeshwar

Third Committee Member

Diana Lopez

Fourth Committee Member

Xiang-Xi Xu

Abstract

Although cisplatin is the drug of choice in treating a variety of different cancers, acquired resistance appears to be a common and serious drawback to its effectiveness in the clinic. Using three pairs of cisplatin-sensitive and resistant cancer cell lines, we have found significantly altered hexokinase (HK) I and II levels in cisplatin-resistant cancer cell lines. Based on our interest in utilizing glucose analogs to probe tumor metabolism, we investigated the role of HKI and HKII in cisplatin-resistant lung and ovarian cancer cell lines and asked whether these resistant cell lines could be targeted with sugar analogs under both normoxic and anaerobic conditions. Furthermore, we sought to determine if alterations in HKs would lead to metabolic reprogramming that could reveal targets in other metabolic pathways to be exploited for therapeutic gain in patients with cisplatin-resistant tumors. In three different cisplatin-resistant cancer cell lines, we observed lower HKII protein than in the cisplatin-sensitive cancer cell lines from which they were derived. In cisplatin-resistant lung cancer cell lines HKI was also lower, and lower HK isoforms resulted in increased sensitivity to the glycolytic inhibitors 2-deoxyglucose (2-DG) and 2-fluorodeoxyglucose (2-FDG) under anaerobic and hypoxic conditions. This increased sensitivity to 2-DG and 2-FDG correlated with a significant decrease in lactate and ATP production in the presence of these glycolytic inhibitors. Knockdown of HKI or HKII individually with siRNA in cisplatin-sensitive lung cancer cell lines lead to a significant increase in 2-FDG induced cell death under anaerobic conditions, however knockdown of both HKI and HKII with a single novel siRNA sequence yielded the greatest increase in cell death. Furthermore, blockage of alternate metabolic pathways, such as fatty acid oxidation and glutaminolysis, resulted in cell death under normal oxygen conditions in cisplatin-resistant lung cancer cell lines. When expanding our findings into a cisplatin-resistant ovarian cancer cell pair, we found decreased HKII expression in cisplatin-resistant A2780cis correlated with increased sensitivity to 2-DG under anaerobic conditions. Overall, our results indicate that lowered hexokinase levels in two cisplatin-resistant lung cancer cell lines leads to increased sensitivity to glycolytic inhibition under anaerobic and hypoxic conditions. This increased sensitivity to glycolytic inhibitors correlates with a decrease in lactate and ATP production in the presence of 2-DG under anaerobic conditions. By lowering HK levels in cisplatin-sensitive cell lines, we observed a significant increase in 2-FDG mediated cell death under anaerobic conditions. Moreover, the cisplatin-resistant lung cancer cells with lower HK were sensitive to either fatty acid ß-oxidation inhibition or glutamine deprivation under normoxia. When expanding the study to include cisplatin-resistant ovarian cancer cell lines, we found lower HKII levels correlated with increased sensitivity to 2-DG under anaerobic conditions. Thus, utilizing different metabolic inhibitors is effective in inducing cell death selectively in cisplatin-resistant cancer cells in vitro.

Keywords

hexokinase; cisplatin resistance; tumor metabolism

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