Abstract B009: Sub-oxidant ascorbate sensitizes glioblastoma stem cells to DNA damaging agents by inducing DNA de-methylation and altering the chromatin landscape
Authors: Malloy Maureen , Sudhir Sweta , Khela Harmon , Fagundo-Lugo Maria , Lal Bachchu , Bressler Joseph , Laterra John , Bertoni Hernando Lopez 
Published: December 2022

Abstract
Gliomas are the most common primary brain cancer and glioblastoma (GBM) represents the most aggressive and lethal subtype. The high GBM morbidity and mortality are due to profound genetic, epigenetic, and phenotypic heterogeneity within the tumor that contributes to an almost guaranteed development of therapeutic resistance and tumor recurrence. GBM stem-like cells (GSCs) are tumor cell sub-populations that disproportionately drive tumor propagation, therapy resistance, and tumor recurrence. Epigenetic deregulation is thought to play a fundamental role in the formation and expansion of GSCs and their contributions to oncogenesis. However, the epigenetic reprogramming events that contribute to GSC tumor propagation and therapy resistance remain elusive. DNA methylation (5mC) can be modified to generate hydroxymethylation (5hmC) or reversed through enzymatic reactions coordinated by the Ten-Eleven Translocation (TET) enzymes. Multiple studies show a decrease in both 5hmC levels and TET expression in gliomas and a correlation between loss of 5hmC and increasing glioma grade and poor prognosis. These observations suggest that re-establishing TET activity and 5hmC levels in tumor cells will have tumor suppressive effects. Ascorbate (vitamin C) has gained pre-clinical interest as an attractive antitumor agent for GBM due to its ability to regulate redox status, cross the blood-brain barrier, and enhance TET enzymatic activity; therefore, providing an innovative potential epigenetic therapeutic for GBM. We show that low-dose ascorbate (i.e. sub-pro-oxidant concentrations) enhances TET activity and elevates global 5hmC levels concurrent with decreases in 5mC in GSCs. These changes in DNA methylation correlate with a decrease in self-renewal capacity and increased tumor cell death induced by temozolomide (TMZ) and ionizing radiation (IR). We also present a novel mechanism whereby ascorbate increases the histone mark H3K36me3 by upregulating GSC expression of the histone methyltransferase NSD1 via demethylation of the gene promoter region. We propose that the enhanced susceptibility of ascorbate-treated tumor cells to DNA damage therapies results from increased H3K36me3-induced euchromatin states that increases genomic susceptibility to DNA damage. These findings support the need to better understand the epigenetic, transcriptomic, and chromatin architectural effects of ascorbate in cancer. These results also strongly indicate that ascorbate could serve as an adjunct to standard-of-care GBM treatment by restoring TET tumor-suppressor activity and re-sensitizing GSCs to conventional radio- and chemotherapeutics.
Citation Format: Maureen Malloy, Sweta Sudhir, Harmon Khela, Maria Fagundo-Lugo, Bachchu Lal, Joseph Bressler, John Laterra, Hernando Lopez Bertoni. Sub-oxidant ascorbate sensitizes glioblastoma stem cells to DNA damaging agents by inducing DNA de-methylation and altering the chromatin landscape. [abstract]. In: Proceedings of the AACR Special Conference: Cancer Epigenomics; 2022 Oct 6-8; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2022;82(23 Suppl_2):Abstract nr B009.