Abstract 4448: Identification of a small molecule inhibitor of aldolase A for the targeting of hypoxic cancer cells

Abstract

Abstract Cancer cells are critically dependent on glycolysis. Currently, there is no effective pharmacotherapy that exploits this metabolic vulnerability. We have uncovered a feed forward cycle of anaerobic glycolysis and hypoxia-inducible factor-1 (HIF-1). Glycolysis under hypoxic conditions normally maintains high ATP levels, which is driven by transcriptional activity of HIF-1. Glycolysis inhibition results in a cellular energy crisis, i.e. an increased AMP:ATP ratio, leading to AMPK-mediated phosphorylation of the HIF-1 co-activator p300/CBP. This prevents HIF-1 activity, although HIF-1 protein levels are unchanged, abrogating the feed forward cycle. We identified fructose-1,6-bisphosphate aldolase A (ALDOA) as a top glycolytic enzyme target for inhibiting hypoxic cancer cell glycolysis and HIF-1 activity. Our aim was to identify small molecule inhibitors for ALDOA that may yield compounds for further preclinical development. To this end, a primary high-throughput screen (HTS) was performed with 65,936 compounds using a fluorescence-based NADH oxidation aldolase (cell-free) assay. 640 compounds were further tested in a cherry-pick confirmation screen, from which 112 hits underwent concentration-response curve validation in cell-free assays. From these, 4 hits were further tested in cell-based assays by using colorectal, breast and pancreatic cancer cell lines. A lead compound that showed micromolar potency in inhibiting ALDOA, induced cell death under hypoxic conditions with IC50 values ranging between 2 - 8 μM in the cancer cell lines tested. It also inhibited extracellular flux in a real-time glycolysis assay, inhibited extracellular lactate production under hypoxic conditions as a measure of glycolysis, and blocked hypoxia responsive element (HRE) HIF-1 reporter activity but not HIF-1 protein levels. No effect of the lead compound was observed on mitochondrial respiration. Thus, this compound provides a valuable chemical probe for inhibiting glycolysis in cancer cells. Furthermore, the work provides proof-of-concept that targeting glycolysis with a small molecule inhibitor exerts potent antitumor effects in vitro and is currently being tested in preclinical models as a first-in-class oncological agent. Citation Format: Petrus R. de Jong, Geoffrey V. Grandjean, Ashwini K. Devkota, Eun Jeong Cho, Kevin N. Dalby, Garth Powis. Identification of a small molecule inhibitor of aldolase A for the targeting of hypoxic cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4448. doi:10.1158/1538-7445.AM2015-4448