Date of Award

2011

Degree Type

Dissertation

Degree Name

Ph.D.

Department

Chemistry and Biochemistry

First Advisor

Pandey, Siyaram (Chemistry and Biochemistry)

Keywords

Biochemistry.

Rights

CC BY-NC-ND 4.0

Abstract

Cancer is a disease of uncontrolled cell growth and proliferation that is predicted to directly affect one-third of Canadians. Standard chemotherapy, which targets DNA or its replicative machinery, effectively eradicates cancer cells but also causes non-specific toxicity to non-cancerous cells. Pancratistatin is a natural alkaloid isolated from Hymenocallis littoralis found to selectively induce apoptosis (programmed cell death) in numerous human cancer cell lines with an insignificant effect on non-cancerous cells. The major objectives of this study were to: 1) assess the selectivity and efficacy of pancratistatin and synthetic derivatives on patient-obtained and commercially available leukemia models; 2) test the effect of pancratistatin on human colon and prostate cancer cells in vitro and in vivo; and 3) to determine the mechanism of action of pancratistatin. Results suggested that 1 ┬╡M pancratistatin induces apoptosis in leukemia ex vivo with an insignificant effect on non-cancerous peripheral blood mononuclear cells. Apoptosis was monitored by nuclear staining and phosphatidylserine exposure by microscopy and flow cytometry. Structure-activity relationship screening of synthetic derivatives of pancratistatin on leukemia (Jurkat) cells revealed that certain analogs retain activity of the native compound, albeit at higher concentrations. The effects of pancratistatin on colon (HCT116, HT-29) and prostate (LNCaP, DU145) cancer cell lines, expressing either the wild-type or functionally inactive p53 tumor suppressor protein, were studied using cell-based assays and animal models. Pancratistatin treatment caused increased production of reactive oxygen species, collapse of mitochondrial membrane potential, and release of the pro-apoptotic proteins cytochrome c, apoptosis-inducing factor and endonuclease G to the cytosol. Furthermore, pancratistatin induced cell death independent of p53, caspase activation or Bax expression. Importantly, components of the mitochondrial respiratory chain were deemed crucial for pancratistatin activity, as mtDNA-deficient 'Rho-0' cells were resistant to pancratistatin. A significant finding of this study was that pancratistatin reduced growth of human colon and prostate tumor xenografts in immune-compromised mice, was well-tolerated and determined to be non-toxic to vital organs compared to control. In conclusion, pancratistatin is a natural anti-cancer compound that selectively targets cancer cell mitochondria to induce apoptosis and significantly reduces growth of human tumor xenografts.

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