Abstract 2076: Inhibiting glycolysis and HIF1α: a novel approach to cancer therapy.
Authors: Grandjean Geoffrey V. , Kingston John , Morrow John Kenneth , Zhang Shuxing , Powis Garth 
Published: August 2014

Cancer cells satisfy their increased need for energy under conditions of normal oxygen tension by undergoing a high rate of glycolysis followed by lactic acid fermentation, a process known as the Warburg Effect, rather than oxidative phosphorylation as observed in normal cells. Unlike proliferating cancer cells, the internal environment of most solid tumors is hypoxic due to an inadequate or disorganized blood supply. This results in tumors generating ATP through anaerobic respiration, a process many times less efficient than oxidative phosphorylation. The importance of anaerobic glycolysis for cancer cell energy generation makes it an attractive target for inhibiting cancer growth. However, agents currently in use for regulating energy production in tumors are relatively ineffective analogs of glucose or intermediates of the glycolytic pathway.

We performed a high throughput siRNA screen using an siRNA library targeting all known open reading frames of the human genome (Dharmacon Inc.) to identify genes that when inhibited would block hypoxia inducible factor-1 alpha (HIF-1α) transcription factor luciferase reporter activity. A significant number of genes selected and validated were members of the glycolysis pathway. It is known that many glycolysis enzymes are induced by HIF-1α providing a feed forward loop between glycolysis and HIF-1α. We have addressed two questions: (1) what is the biological mechanism by which these two signaling systems regulate one another?; and (2) can specific inhibitors of glycolysis be developed with the added effect of inhibiting HIF-1α? Our mechanistic studies have shown that of the 40 enzymes of glycolysis involving 11 enzymatic steps and various enzyme isoforms, siRNA knockdown of Aldolase A (ALDOA) produces the maximum inhibition of HIF-1α activity. Further work indicated that ALDOA siRNA inhibition of HIF-1α is mediated through AMPK, a sensor of low ATP levels in the cell, since siRNA knockdown of AMPK results in rescue of HIF-1α activity. The effect of ALDOA inhibition is not mediated through a reduction in HIF-1α protein suggesting that, contrary to the canonical AMPK pathway, mTOR and its regulation of translation is not involved. Instead, p300 is phosphorylated by AMPK thus inhibiting its function as a co-activator of HIF-1α.

A structure based drug design approach was employed for lead identification and optimization using the crystal structure of ALDOA (4ALD) and GOLD virtual screening platform. Three compounds have been identified as having effects on the modulation of ALDOA activity in a protein based biochemical assay as well as inhibitory effects on glycolysis itself using the XF96 Analyzer from Seahorse Bioscience. Here, we present an overview of ALDOA target identification, its mechanism as a regulator of HIF-1α activity and virtual drug design methodologies used in the search for novel pharmacological probes which act as dual inhibitors of energy production and the HIF-1α survival pathway.

Citation Format: Geoffrey V. Grandjean, John Kingston, John Kenneth Morrow, Shuxing Zhang, Garth Powis. Inhibiting glycolysis and HIF1α: a novel approach to cancer therapy. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2076. doi:10.1158/1538-7445.AM2013-2076