Summary Pharmacological ascorbate has been proposed as a potential anti-cancer agent when combined with radiation and chemotherapy. The anti-cancer effects of ascorbate are hypothesized to involve the autoxidation of ascorbate leading to increased steady-state levels of H2O2; however, the mechanism(s) for cancer cell-selective toxicity remain unknown. The current study shows that alterations in cancer cell mitochondrial oxidative metabolism resulting in increased levels of O2•− and H2O2 are capable of disrupting intracellular iron metabolism thereby selectively sensitizing non-small cell lung cancer (NSCLC) and glioblastoma (GBM) cells to ascorbate through pro-oxidant chemistry involving redox active labile iron and H2O2. In addition, preclinical studies and clinical trials demonstrate the feasibility, selective toxicity, tolerability, and potential efficacy of pharmacological ascorbate in GBM and NSCLC therapy.

Cancer cells, relative to normal cells, demonstrate significant alterations in metabolism that are proposed to result in increased steady-state levels of mitochondrial-derived reactive oxygen species (ROS) such as O2•−and H2O2. It has also been proposed that cancer cells increase glucose and hydroperoxide metabolism to compensate for increased levels of ROS. Given this theoretical construct, it is reasonable to propose that forcing cancer cells to use mitochondrial oxidative metabolism by feeding ketogenic diets that are high in fats and low in glucose and other carbohydrates, would selectively cause metabolic oxidative stress in cancer versus normal cells. Increased metabolic oxidative stress in cancer cells would in turn be predicted to selectively sensitize cancer cells to conventional radiation and chemotherapies. This review summarizes the evidence supporting the hypothesis that ketogenic diets may be safely used as an adjuvant therapy to conventional radiation and chemotherapies and discusses the proposed mechanisms by which ketogenic diets may enhance cancer cell therapeutic responses.

SUMMARY Pharmacological ascorbate has been proposed as a potential anti-cancer agent when combined with radiation and chemotherapy. The anti-cancer effects of ascorbate are hypothesized to involve the autoxidation of ascorbate leading to increased steady-state levels of H2O2; however, the mechanism(s) for cancer cell-selective toxicity remain unknown. The current study shows that alterations in cancer cell mitochondrial oxidative metabolism resulting in increased levels of O2.− and H2O2 are capable of disrupting intracellular iron metabolism, thereby selectively sensitizing non-small-cell lung cancer (NSCLC) and glioblastoma (GBM) cells to ascorbate through pro-oxidant chemistry involving redox-active labile iron and H2O2. In addition, preclinical studies and clinical trials demonstrate the feasibility, selective toxicity, tolerability, and potential efficacy of pharmacological ascorbate in GBM and NSCLC therapy.

The toxicity of pharmacological ascorbate is mediated by the generation of H2O2 via the oxidation of ascorbate. Since pancreatic cancer cells are sensitive to H2O2 generated by ascorbate they would also be expected to become sensitized to agents that increase oxidative damage such as ionizing radiation. The current study demonstrates that pharmacological ascorbate enhances the cytotoxic effects of ionizing radiation as seen by decreased cell viability and clonogenic survival in all pancreatic cancer cell lines examined, but not in non-tumorigenic pancreatic ductal epithelial cells. Ascorbate radiosensitization was associated with an increase in oxidative stress-induced DNA damage, which was reversed by catalase. In mice with established heterotopic and orthotopic pancreatic tumor xenografts, pharmacological ascorbate combined with ionizing radiation decreased tumor growth and increased survival, without damaging the gastrointestinal tract or increasing systemic changes in parameters indicative of oxidative stress. Our results demonstrate the potential clinical utility of pharmacological ascorbate as a radiosensitizer in the treatment of pancreatic cancer.

Purpose Ketogenic diets (KDs) are high in fat and low in carbohydrates as well as protein which forces cells to rely on lipid oxidation and mitochondrial respiration rather than glycolysis for energy metabolism. Cancer cells (relative to normal cells) are believed to exist in a state of chronic oxidative stress mediated by mitochondrial metabolism. The current study tests the hypothesis that KDs enhance radio-chemo-therapy responses in lung cancer xenografts by enhancing oxidative stress. Experimental Design Mice bearing NCI-H292 and A549 lung cancer xenografts were fed a KD (KetoCal® 4:1 fats: proteins+carbohydrates) and treated with either conventionally fractionated (1.8-2 Gy) or hypofractionated (6 Gy) radiation as well as conventionally fractionated radiation combined with carboplatin. Mice weights and tumor size were monitored. Tumors were assessed for immuno-reactive 4-hydroxy-2-nonenal-(4HNE) modified proteins as a marker of oxidative stress as well as PCNA and γH2AX as indices of proliferation and DNA damage, respectively. Results The KD combined with radiation resulted in slower tumor growth in both NCI-H292 and A549 xenografts (p<0.05), relative to radiation alone. The KD also slowed tumor growth when combined with carboplatin and radiation, relative to control. Tumors from animals fed a KD in combination with radiation demonstrated increases in oxidative damage mediated by lipid peroxidation as determined by 4HNE-modified proteins as well as decreased proliferation as assessed by decreased immunoreactive PCNA. Conclusions These results show that a KD enhances radio-chemo-therapy responses in lung cancer xenografts by a mechanism that may involve increased oxidative stress.