Defense Date

2009

Document Type

Dissertation

Degree Name

Doctor of Philosophy

Department

Pharmacology & Toxicology

First Advisor

Paul Ph.D. Dent

Abstract

The first study examined the interaction between low doses of the multi-kinase inhibitor sorafenib and the histone deacetylase inhibitor vorinostat in colon cancer cells. Sorafenib and vorinostat synergized to kill HCT116 and SW480 cells. In SW480 cells, sorafenib+vorinostat toxicity correlated with CD95 activation and CD95-stimulated autophagy. Drug lethality in SW480 cells was blocked by knock down of CD95. In SW620 cells that are patient matched to SW480 cells, sorafenib+vorinostat toxicity was significantly reduced that correlated with a lack of CD95 activation and lower expression of ceramide synthase 6 (LASS6). Overexpression of LASS6 in SW620 cells enhanced drug-induced CD95 activation and tumor cell killing, whereas knock down of LASS6 in SW480 cells suppressed CD95 activation. In HCT116 cells, sorafenib+vorinostat did not increase CD95 plasma membrane levels, weakly induced caspase 8 association with CD95, and knock down of CD95 enhanced drug lethality. In HCT116 cells sorafenib+vorinostat treatment caused CD95-dependent autophagy that was a protective signal. Thus, treatment of tumor cells with sorafenib+vorinostat activates CD95 that promotes viability via autophagy or degrades survival via extrinsic or intrinsic pathways. Drug-induced activation of the de novo ceramide synthesis pathway plays a key role in CD95 activation. The second project explores the mechanism by which the combination of 17AAG, an hsp90 inhibitor, and PD184352, a MEK1/2 inhibitor alters survival in colon cancer cells. 17AAG and PD184352 synergized to kill HCT116 and SW480 cells. In HCT116 cells drug-exposure increased CD95 plasma membrane levels In SW620 cells, 17AAG and PD184352 toxicity was significantly reduced that correlated with a lack of CD95 activation and lower expression LASS6. Overexpression of LASS6 in SW620 cells enhanced drug-induced CD95 activation and tumor cell killing. In Mia Paca2 cells, a pancreatic cell line, inhibition of caspase 8 or overexpression of c-FLIP-s suppressed cell killing by PD184352 and 17AAG exposure. Drug lethality in Mia Paca2 cells was blocked by knock down of CD95. Additionally, overexpression of Bcl-xL or knockdown of caspase 9 decreased cell killing in 17AAG and PD184352 combination treatment. Thus, 17AAG+PD184352 exposure activates the extrinsic and intrinsic apoptotic pathways to kill Mia Paca2 cells. This document was created in Microsoft Word 2000.

Rights

© The Author

Is Part Of

VCU University Archives

Is Part Of

VCU Theses and Dissertations

Date of Submission

May 2009

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