Exploiting metabolic susceptibilities in glioblastoma via glycolytic inhibition and ketogenic therapy.

Abstract

e13558 Background: Glioblastoma (GBM) remains one of the most lethal primary brain tumors in children and adults. Despite enormous efforts to elucidate the genetic and epigenetic drivers of this disease, the prognosis for patients diagnosed with GBM remains dismal. Because tumor cell metabolism differs greatly from that of normal non-cancerous cells, it is possible to develop therapies which more effectively target the cancer cell while sparing normal cells. Growing in popularity is the ketogenic diet, which is a high fat, very low carbohydrate diet resulting in the production of ketone bodies, acetoacetate (AA) and β-hydroxybutyrate (βHB) to generate ATP. Methods: Analysis conducted by open-access GBM patient database, mts assay, Western blot, neurosphere assay, and TEM. Results: Enzymes required for ketone metabolism (BDH1 and OXCT1) were significantly downregulated in GBM while glycolytic enzymes were significantly upregulated (HK2, HK1, SLC2A3, NAMPT, G6PD). GBM stem cell (GSC) markers (CD44, STAT3) positively correlated with glycolytic enzymes. Ultrastructural analysis of GSCs indicated that about half of the mitochondria were missing cristae, highly suggestive of an increased glycolytic dependency. Treatment of patient-derived GSC lines as well as non-stem cell lines SJGBM2 (pediatric) and U87 (adult) resulted in a dose-dependent decrease in viability in response to the glycolytic inhibitor 2-deoxy-D-glucose (2-DG). When cells were exposed to ketone bodies, AA but not βHB induced a dose-dependent decrease in cell viability with 10 mM reducing viability ranging from 20-80% of non-treated controls. Western blot analysis demonstrated robust caspase activation and PARP cleavage in response to AA. Furthermore, AA significantly reduced GSC neurosphere formation at 2.5 mM suggesting inhibition of GSC self-renewal pathways. Combined treatment of low dose 2-DG (50 μM) with increasing concentrations of AA resulted in more cell death than either treatment. The effect was more than additive at the low concentrations of AA (1- 5 mM) suggesting synergy. Conclusions: Glycolytic inhibition in conjunction with the ketogenic diet may be a promising therapeutic route for this difficult-to-treat cancer.