Abstract SY21-03: Shutting off cancer's fountain of youth: Targeting metabolism to block repair and reverse immortality

Abstract

Abstract The altered metabolism of cancer cells, first recognized by Warburg as aerobic glycolysis, has often been ascribed to answering increased needs for macromolecular biosynthesis to support rapid tumor growth. Recent discoveries that mutations leading to accumulation of fumarate, succinate or 2-hydroxyglutarate (2-HG) may drive cancer via targeting chromatin and DNA modifying enzymes have forced a reevaluation. Indeed, multiple metabolic intermediates can modulate chromatin modifiers, suggesting that altered metabolism may drive epigenetic changes enabling carcinogenesis. Our work has implicated a distinct mechanism, whereby metabolic reprogramming and altered chromatin promote genomic instability and cellular immortality, two hallmarks of cancer. This work began with efforts to target the Warburg effect with small-molecule glucose uptake inhibitors. While targeting glycolysis failed to affect cell proliferation on its own, treated cells were sensitized to radiation, displaying a loss of DNA repair and increased cellular senescence. Working downstream, we found much of this effect was mediated by the hexosamine biosynthetic pathway (HBP) which brings together glucose and glutamine to produce N-acetyl glucosamine (GlcNAc) to support protein glycosylation. Thousands of nuclear, cytoplasmic and mitochondrial protein serine and threonine residues can be modified by O-GlcNAc transferase (OGT) and the resulting O-GlcNAcylation is linked to stress and DNA damage tolerance. Our data implicated the polycomb repressive complexes PRC1 and 2 and their catalytic subunits BMI1 and EZH2 as key OGT targets. Small molecule inhibitors and/or RNAi targeting the HBP, OGT, BMI1 or EZH2 similarly delayed DSB repair and induced senescence after irradiation. In light of others' work linking the histone H3 K27 methyltransferase activity of EZH2 to 53BP1 binding at DSBs to drive repair by non-homologous end-joining (NHEJ), these studies suggest a pathway from the Warburg effect to O-GlcNAcylation and rapid, error-prone DSB repair. Working downstream to validate this mechanism, we discovered that glycolysis and the HBP balanced an antagonistic effect of the TCA cycle on DSB repair. Excess α-ketoglutarate (2-OG) serves as a cofactor for diverse dioxygenases, including the lysine demethylases (KDMs) that antagonize histone methyltransferases such as EZH2. Activating the TCA cycle or providing exogenous 2-OG blocked DSB repair after irradiation and increased senescence while inhibiting all 2-OG-dependent dioxygenases or only the KDM enzymes that remove histone modifications by EZH2 at H3 K27 or G9A at H3 K9 restored DSB repair. This led us to examine the effects of treating cells with the oncometabolite 2-HG, known to mediate its effects by serving as a 2-OG-dependent dioxygenase inhibitor. While prior attention has largely focused on TET DNA demethylases as 2-HG targets and epigenetic mechanisms for carcinogenesis, we observed 2-HG blocking KDM enzymes to accelerate error-prone NHEJ repair, suggesting a role in genomic instability in IDH mutant cells. Toward translation, we have examined the effects of blocking O-GlcNAcylation in vivo as a means to sensitize tumors to radiation. Animals bearing tumors expressing shRNA targeting OGT or treated with OGT inhibitors and then irradiated displayed decreased protein O-GlcNAcylation, a delay in DSB repair, greater cell death, delayed cell recovery and increased senescence. In turn, when animals were treated with GlcNAc or with shRNA or inhibitors targeting O-GlcNAcase, their tumors displayed increased O-GlcNAcylation and dramatic radiation resistance, indicating accelerated DSB repair in vivo. These results implicate OGT and O-GlcNAcyation as a new mechanism by which the Warburg effect may mediate resistance to radiation therapy. Further, our studies point to dietary interventions and/or drugs targeting OGT as a route to improving outcomes from radiotherapy in the clinic. Taken together, an implication of this work is that cancer metabolism may help drive carcinogenesis by shifting repair to NHEJ and away from HR, leading to genomic instability, as well as by overcoming barriers to transformation such as accelerated senescence to promote cell immortality. Our studies argue for renewed efforts to explore cancer metabolism as a target for prevention and therapy, with a focus on leveraging crosstalk with pathways regulating cellular responses to DNA damage. Citation Format: Elena Efimova, Yue Liu, Julian Lutze, Tamica Collins, Amy Flor, Jacqueline Brinkman, Don Wolfgeher, Sara Warrington, Stephen J. Kron. Shutting off cancer's fountain of youth: Targeting metabolism to block repair and reverse immortality [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr SY21-03.