Abstract IA-17: The roles of prolyl hydroxylases in metabolism and cancer

Abstract

Abstract Both succinate dehydrogenase (SDH) and fumarate hydratase (FH) are Krebs cycle enzymes that convert succinate to fumarate and fumarate to malate, respectively. SDH is also a functional member (complex II) of the Electron Transport Chain. Surprisingly, although SDH and FH are ‘housekeeping genes’ with key bioenergetic roles, germline mutations in these genes cause cancer. We demonstrated that succinate, a Krebs cycle metabolite (an SDH substrate), functions as an intracellular messenger between mitochondria and the cytosol. Succinate, which accumulates in mitochondria due to the inactivation of SDH, leaks out to the cytosol, where it inhibits the enzymatic machinery of oxygen sensing, mediated by a family of α-ketoglutarate-dependent Prolyl Hydroxylase enzymes (PHD). PHD inhibition by succinate triggers the accumulation and activation of the hypoxia inducible factor (HIF) in the nucleus and the pseudohypoxic response that enhances tumour vascularisation and glycolysis. These and other independent studies collectively suggested that PHD inhibition and pseudohypoxia are important factors that link mitochondrial tumour suppressors and cancer. This observation received further support recently from the fact that PHD2 was genetically identified as a tumour suppressor. The inhibition of PHD by succinate is achieved by a product inhibition mechanism, given that PHD hydroxylates its targets while utilizing α-ketoglutarate as a substrate and producing succinate. We further characterized the nature of succinate-mediated inhibition of PHD and showed that this inhibition is competitive in nature. Therefore, the ratio, rather than the absolute concentrations, of α-ketoglutarate to succinate or fumarate in cells critically affect PHD activity and HIF activity. We designed and synthesised cell-permeating α-ketoglutarate esters that are hydrolyzed in the cytosol by esterases. These compounds support PHD activity thereby lowering HIF levels and activity in SDH deficient cells. Furthermore, we also demonstrated that α-ketoglutarate increased the affinity of PHD to oxygen and reactivate PHD in cells under hypoxia, resulting in destabilization of the HIFα proteins. It is likely that the binding of α-ketoglutarate by PHD ‘primes’ the enzyme for the binding of oxygen. Our study suggests that well-designed α-ketoglutarate derivatives may have therapeutic potential in the treatment of tumors with functional down-regulation of oxidative phosphorylation. Furthermore, the functional consequences of the observations that PHDs are sensors for oxygen and α-ketoglutarate will be discussed. Citation Information: Cancer Res 2009;69(23 Suppl):IA-17.