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Several preclinical studies support the use of the therapeutic ketogenic diet (KD) and/or caloric restriction (CR) as an adjuvant to cancer therapy. This data not only suggests that this approach alters multiple hallmarks of cancer growth, but that it may also enhance other therapeutic modalities. We have demonstrated that the unrestricted KD greatly increases survival in a mouse model of malignant glioma when administered in combination with radiation, yet the underlying mechanisms are not fully elucidated. Mounting evidence suggests that the abnormal epigenetic landscape in tumors may, among other things, impact radiosensitivity. Specifically, many cancers exhibit increased histone deacetylase (HDAC) activity which contributes to the epigenetic milieu found in tumors and alters DNA damage repair. It was recently demonstrated that the ketone body β-hydroxybutyrate (BHB) inhibits HDAC activity in normal mouse tissue. As the use of HDAC inhibitors as anti-cancer agents has generated great interest, we examined the effect of BHB on HDAC activity and radiosensitivity in the context of malignant glioma. We found that BHB radiosensitizes mouse glioma, human glioma and human glioma stem-like cells in vitro. We also demonstrate that BHB inhibits HDAC activity in a dose dependent manner and alters key components of DNA damage repair. Taken together this data opens up another avenue for understanding the mechanisms underlying the KD and its impact on radiation therapy, ultimately helping us better understand how to implement it for clinical use in oncology. Citation Format: Eric C. Woolf, Alex P. Rossi, Helena B. Silva-Nichols, Kara D. Gardner, Nelofer Syed, Adrienne C. Scheck. β-hydroxybutyrate inhibits histone deacetylase activity and radiosensitizes malignant glioma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1125A. doi:10.1158/1538-7445.AM2017-1125A

Glioblastoma multiforme (GBM) is an aggressive primary brain tumor with a 5 year survival rate of 25% in children and less than 10% in adults. Improvement in the prognosis of GBM patients requires the development of new therapeutic approaches. One emerging strategy is to target aberrant cell metabolism, a trait shared by virtually all tumor cells. The ketogenic diet (KD), a high fat, low carbohydrate and protein metabolic therapy has been shown to prolong survival in animal glioma models, and when used in conjunction with radiation cured 9 of 11 mice of their implanted tumors. We have also shown that the KD alters hypoxia, angiogenesis, and other hallmarks of glioma progression. To elucidate the underlying mechanisms through which ketones exert their effects on glioma, we are doing analyses of the effect of β-hydroxybutyrate (βHB), the most prevalent ketone body synthesized during ketosis, on glioma cells in vitro. We found that βHB both alone and in conjunction with radiation significantly inhibits proliferation of human and mouse glioma cells and human glioma stem cells (GSC). Alterations in passage through the cell cycle may affect proliferation, and cells are also more sensitive to radiation in the G2/M cell cycle phase. We therefore analyzed the effect of βHB on cell cycle distribution of cells treated with βHB and/or radiation. An analysis of cell cycle status by flow cytometry demonstrated that treating the GSC line L0 with 5mM βHB in combination with 4 Gy of radiation significantly increased the number of cells in G2/M cell cycle arrest. In GL261-Luc2 mouse glioma cells, 5mM βHB alone significantly enhanced G2/M cell cycle arrest, which could lead to radiosensitization. Currently we are analyzing proteins involved in cell cycle progression and apoptosis to better understand βHB mediated changes in growth and radioresistance. In summary, these data provide insight to the radiosensitization and anti-proliferative mechanisms of βHB and may hold implications for the use of the KD in the treatment of GBM. Citation Format: Helena B. Silva-Nichols, Alex P. Rossi, Eric C. Woolf, Marshall J. Fairres, Loic P. Deleyrolle, Brent A. Reynolds, Adrienne C. Scheck. Radiosensitization of glioma cells by the ketone body β-hydroxybutyrate is associated with enhanced cell cycle arrest in the G2/M phase. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1670.

No abstract

Glioblastoma (GB) is the most common primary malignant brain tumor with a dismal median survival of 1.5 years. To address the need for novel therapeutics for the treatment of GB, investigations into high fat, low carbohydrate ketogenic diets (KD) have been explored as potential adjuvant therapeutic modalities. Preclinical studies of the KD in a mouse model of malignant glioma demonstrated that the KD extended median survival when used alone and substantially potentiated both radiotherapy and chemotherapy. However, the underlying mechanisms by which the KD exerts its therapeutic benefits remain poorly understood. A key result of the KD is the production of the ketone bodies β-hydroxybutyrate (βHB), acetoacetate, and acetone. Of these, BHB is the most prevalent ketone body produced. In order to elucidate the underlying molecular mechanisms of the KD, we have examined the interactions between BHB and the mouse glioma cell line GL261-Luc2. Previously, we demonstrated the ability of βHB to recapitulate the in vivo effects of the KD in tumor cells in an in vitro model. In the present study, we demonstrate that the ketone βHB is able to significantly down regulate the master transcription factor c-Myc, which is intimately involved in cell cycle progression, apoptosis, proliferation, DNA damage and repair, and metabolism. Overexpression of c-Myc is tied to a variety of human malignancies. In GB, upregulation of c-Myc signaling has been connected to an increase in tumor glucose metabolism. Furthermore, downregulating c-Myc expression has been demonstrated to confer sensitivity to tumors. We show that doses of both 5mM and 10mM βHB significantly reduced the expression of c-Myc in the GL261-Luc2 cell line when evaluated by both immunocytochemistry and western blot. Downregulation of c-Myc may, in part, be a mechanism by which βHB and thus the KD exert their radiosensitizing and antiproliferative effects on tumors. We intend to further investigate the role of c-Myc in the KD by investigating downstream targets of c-Myc as well as potential regulators of c-Myc transcription that are affected by BHB. Citation Format: Alex P. Rossi, Helena B. Silva-Nichols, Eric C. Woolf, Adrienne C. Scheck. The ketone body β-hydroxybutyrate down regulates c-Myc signaling in a malignant glioma model. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1022.