Date of Award

Spring 2005

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Committee Director

Christopher J. Osgood

Committee Member

James H. Yuan

Committee Member

Lloyd Wolfinbarger

Committee Member

Mark S. Elliot

Abstract

Arglabin-DMA, an analog of farnesyl pyrophosphate (FPP), reportedly inhibits farnesyltransferase (FTase) directly by competitively blocking the binding of Ras protein and its posttranslational modification, as suggested in previous studies. But, the mechanisms by which Arglabin-DMA inhibits tumor growth in vivo and in vitro are still relatively poorly characterized. To determine the mechanism by which this drug inhibits tumor growth, the effects of Arglabin-DMA in two human colon tumor cell lines (mutant K-ras HCT 116 and wild-type ras HT-29) were explored on cell proliferation, apoptosis, and cell cycle kinetics in vitro. In cell viability studies, we showed that Arglabin-DMA had striking morphological and physiological effects on the two human colon tumor cell lines, possibly more so than those of other anticancer drugs. Also, Arglabin-DMA exhibited less harm to normal cells (Hs27) which retained their potential for cell growth. An add-back experiment showed that Arglabin-DMA had no effect on the isoprenoid biosynthetic pathway. The drug not only affects the mutant K-ras human colon tumor cell line, but also the wild-type ras human colon tumor cell line. It may therefore inhibit one or more non-Ras proteins to exert its antitumor effects. Gel electrophoresis, TUNEL assay, Annexin V assay, apoptosis dye-uptake assay, and morphological criteria were used to characterize apoptosis. Adherent cells and freely floating detached cells in Arglabin-DMA treatment were treated as two distinct populations We demonstrated that the detached cells caused by Arglabin-DMA exposure exhibited increased apoptosis in a p53-independent manner. Cell cycle effects were studied using flow cytometry. After Arglabin-DMA was added, the proportion of the two human colon tumor cells in G2/M phase increased, indicating a block in either G2 or M phase. We conclude that Arglabin-DMA has specific cytotoxic effects in two human colon tumor cell lines, and less cytotoxicity to normal cells. It induces arrest at the G2/M phase of the cell cycle. After treatment with Arglabin-DMA, rounded and detached cells enter apoptosis. This mechanism may be analogous to "anoikis," which is the induction of apoptosis in response to loss of cell contact. The utility of this drug in combating cancer remains an attractive, though complex possibility.

DOI

10.25777/ahqz-v755

ISBN

9781109719895

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