Document Type

Dissertation

Date of Degree

Spring 2015

Degree Name

PhD (Doctor of Philosophy)

Degree In

Molecular and Cellular Biology

First Advisor

Raymond J. Hohl

Abstract

Schweinfurthins are a family of natural products with significant anti-cancer activities. They were originally identified in the National Cancer Institute (NCI) human 60 cancer cell line screening. The growth inhibition profile of schweinfurthins is distinct from other clinically used anti-cancer agents, indicating that they have a novel mechanism of action or have a previously unrecognized protein target. Previous studies showed that schweinfurthins affect multiple cellular processes in cancer cells. For example, schweinfurthins can alter cytoskeleton organization, induce ER stress and apoptosis, and inhibit the mevalonate pathway. The mevalonate pathway is responsible for the production of isoprenoids and cholesterol, which have been shown to play regulatory roles in the Hedgehog (Hh) signaling pathway. In this study, we found that the Hh signaling pathway in NIH-3T3 and SF-295 cells was inhibited by schweinfurthins. The supplementation of mevalonate and cholesterol partially restored Hh signaling, indicating that schweinfurthins inhibit Hh signaling partially by down-regulating the products from the mevalonate pathway. Interestingly, schweinfurthins in combination with cyclopamine, an inhibitor of the Hh singaling pathway, synergistically decreased cell viability.

In order to better understand the underlying mechanism of the anti-cancer action of schweinfurthins, we attempted to identify the protein target of schweifnurthins. Affinity chromatography was performed to pull down the protein target. We found that schweinfurhtins bound to the M2 isoform of pyruvate kinase (PKM2) and inhibit its pyruvate kinase activity. Knockdown of PKM2 by siRNA increased the sensitivity of SF-295 cells to schweinfurthins. The inhibition of PKM2 by schweinfurthins led to a reduction in the rate of glycolysis in cancer cells. Fructose 1,6-bisphosphate (FBP), an activator of PKM2, could alleviate schweinfurthin-mediated inhibition on PKM2 and glycolysis. Notably, FBP could also partially reverse the reduction of cell viability in the presence of schweinfurthins. Taken together, these studies revealed the mechanism by which schweinfurthins inhibit Hh signaling. In addition, we uncovered PKM2 as a schwienfurthin target and highlighted the importance of glycolysis suppression as a mechanism of the anti-cancer action of schweinfurthins.

Public Abstract

Glioblastoma multiforme (GBM) is the most prevalent and lethal type of brain tumor. Despite advancements in surgery, chemotherapy and radiotherapy, GBM remains incurable with an average survival time of 12 to 15 months. New therapeutic agents are urgently required for developing treatments for glioblastoma patients. Schweinfurthins are a family of naturally occurring compounds discovered from Macaranga schweinfurthii. Schweinfurthins display significant anti-cancer activity towards a broad variety of cancer types including GBM. However, the mechanisms by which schweinfurthins kill cultured GBM cells are largely unknown. Our proposed research aims to uncover the molecular target as well as the mechanism of action of schweinfurthins. We identified PKM2as a target of schweinfurthins and targeting this protein suppressed the glucose metabolism in GBM cells in vitro. We also found that schweinfurthins inhibit the Hedgehog (Hh) signaling pathway. The addition of cholesterol alleviates the inhibitory effects on Hh signaling and the cytotoxic effects of schweinfurthins. GBM cells show elevated glycolysis and lipids biosynthesis through which cancer cells can produce more building blocks for their growth and division. Based on our observations and previous studies, we proposed that schweinfurthins kill GBM cells by inhibiting both glycolysis and lipogenesis simultaneously. In sum, our research has provided new insights into the mechanisms of the anti-cancer activity of schweinfurthins. More importantly, our studies highlight that targeting glucose and lipids metabolism might be exploited as a strategy in the pursuit of a cure for GBM.

Keywords

publicabstract, glioblastoma, glycolysis, PKM2, schweinfurthins, the Hedgehog signaling pathway, the mevalonate pathway

Pages

xiii, 148 pages

Bibliography

Includes bibliographical references (pages 127-148).

Copyright

Copyright 2015 Chaoqun Zheng

Included in

Cell Biology Commons

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