Ketogenic Diet with Concurrent Chemoradiation in Head and Neck Squamous Cell Carcinoma: Preclinical and Phase 1 Trial Results.

Ketogenic diets (KD) are high in fat and low in carbohydrates, forcing cells to utilize mitochondrial fatty acid oxidation for energy production. Since cancer cells demonstrate increased mitochondrial oxidative stress relative to normal cells, we hypothesized that a KD may selectively enhance metabolic oxidative stress in head and neck cancer cells, sensitizing them to radiation and platinum-based chemotherapy without causing increased toxicity in surrounding normal tissues. This hypothesis was tested in preclinical murine xenografts and in a phase 1 clinical trial (NCT01975766). In this study, mice bearing human head and neck cancer xenografts (FaDu) were fed either standard mouse chow or KetoCal® KD (90% fat, 8% carbohydrate, 2% protein) and exposed to ionizing radiation. Tumors were harvested from mice to test for glutathione, a biomarker of oxidative stress. In parallel, patients with locally advanced head and neck cancer were enrolled in a phase 1 clinical trial where they consumed KD and received radiation with concurrent platinum-based chemotherapy. Subjects consumed KetoCal KD via percutaneous endoscopic gastrostomy (PEG) tube and were also allowed to orally consume water, sugar-free drinks, and foods approved by a dietitian. Oxidative stress markers including protein carbonyls and total glutathione were assessed in patient blood samples both pre-KD and while consuming the KD. Mice bearing FaDu xenografts that received radiation and KD demonstrated a slight improvement in tumor growth rate and survival compared to mice that received radiation alone; however a variation in responses was seen dependent on the fatty acid composition of the diet. In the phase 1 clinical trial, a total of twelve patients were enrolled in the study. Four patients completed five weeks of the KD as per protocol (with variance in compliance). Eight patients did not tolerate the diet with concurrent radiation and platinum-chemotherapy (5 were patient decision and 3 were removed from study due to toxicity). The median number of days consuming a KD in patients who did not complete the study was 5.5 (range: 2-8 days). Reasons for discontinuation included "stress of diet compliance" (1 patient), grade 2 nausea (3 patients), and grade 3 fatigue (1 patient). Three patients were removed from the trial due to dose-limiting toxicities including: grade 4 hyperuricemia (2 patients) and grade 3 acute pancreatitis (1 patient). Median weight loss was 2.95% for the KD-tolerant group and 7.92% for patients who did not tolerate the diet. In conclusion, the ketogenic diet shows promise as a treatment combined with radiation in preclinical mouse head and neck cancer xenografts. A phase 1 clinical trial evaluating the safety and tolerability of KD demonstrated difficulty with diet compliance when combined with standard-of-care radiation therapy and cisplatin chemotherapy.

Mood stabilizers regulate cytoprotective and mRNA-binding proteins in the brain: long-term effects on cell survival and transcript stability.

Manic depressive illness (MDI) is a common, severe, chronic and often life-threatening illness. Despite well-established genetic diatheses and extensive research, the biochemical abnormalities underlying the predisposition to, and the pathophysiology of, these disorders remain to be clearly established. Despite formidable obstacles in our attempts to understand the underlying neurobiology of this illness, there is currently considerable excitement about the progress that is being made using novel strategies to identify changes in gene expression that may have therapeutic relevance in the long-term treatment of MDI. In this paper, we describe our recent research endeavours utilizing newer technologies, including a concerted series of mRNA RT-PCR studies, which has led to the identification of novel, hitherto completely unexpected targets for the long-term actions of mood stabilizers - the major cytoprotective protein bcl-2, a human mRNA binding (and stabilizing) protein, AUH, and a Rho kinase. These results add to the growing body of data suggesting that mood stabilizers may bring about some of their long-term benefits by enhancing neuroplasticity and cellular resilience. These results are noteworthy since recent morphometric brain imaging and post-mortem studies have demonstrated that MDI is associated with the atrophy and/or loss of neurons and glia. The development of novel treatments which more directly target molecules involved in critical CNS cell survival and cell death pathways have the potential to enhance neuroplasticity and cellular resilience, and thereby modulate the long-term course and trajectory of these devastating illnesses.

Induction of enoyl-CoA hydratase by LD50 exposure to perfluorocarboxylic acids detected by two-dimensional electrophoresis.

The effect of in vivo exposure to perfluoro-n-octanoic and perfluoro-n-decanoic acids was examined in the rat liver by two-dimensional electrophoresis (2DE). Using nonequilibrium pH-gradient electrophoresis in the first dimension separation, proteins associated with the mitochondrial/peroxisomal cell fraction were observed and immunologically identified. Conspicuous inductions in peroxisomal enoyl-CoA hydratase and other proteins of the peroxisomal beta-oxidative pathway were observed following single-dose exposure to each compound. The abundance of the tentatively-identified mitochondrial equivalent, crotonase, was not altered by these intoxications. These results confirm previous observations of perfluorocarboxylic acid toxicity and support the use of 2D protein-pattern alterations in biomarker research. The ability to identify this type of alteration via 2DE, in association with specific toxic effects by chemically related compounds, may provide new and additional markers for chemical-induced tissue damage.

Effect of hypolipidemic drugs gemfibrozil, ciprofibrate, and clofibric acid on peroxisomal beta-oxidation in primary cultures of rainbow trout hepatocytes.

Primary cultures of hepatocytes were established from sexually mature male rainbow trout (Oncorhyncus mykiss) and treated with the hypolipidemic drugs gemfibrozil (0.25-1.25 mM), clofibric acid (2.25-3.00 mM), or ciprofibrate (0.25-1.00 mM). Significant dose-related increases in peroxisomal fatty acyl-CoA oxidase (FACO) were seen after exposure for 48 hr to clofibric acid (P < 0.01) and ciprofibrate (P < 0.05) but not gemfibrozil (P = 0.08). Strong correlation was obtained between increased acyl-CoA oxidase activity and the relative amount of peroxisomal bifunctional enzyme (PBE), further supporting evidence of a proliferative effect. These preliminary studies demonstrate that peroxisomal beta-oxidation can be induced in vitro in a primary rainbow trout hepatocyte system.

Comparative effects of dehydroepiandrosterone and related steroids on peroxisome proliferation in rat liver.

Dehydroepiandrosterone (DHEA) is known to induce peroxisome proliferation and peroxisomal enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase (PBE) mRNA in the rat liver. We have compared the effects of 6 intermediate metabolites of DHEA on the induction of peroxisome proliferation and PBE mRNA. Administration of epiandrosterone, etiocholanolone, androstenedione, estrone or estradiol for 2 weeks in the diet at 0.45% concentration to adult male F-344 rats failed to induce significant increases in peroxisome proliferation and PBE mRNA when compared to the parent compound DHEA. Dietary administration of 5-androstene-3 beta,17 beta-diol (ADIOL) for 2 weeks at 0.45% concentration caused an increase in PBE mRNA and peroxisome proliferation but to a lesser extent than DHEA. Following a single intragastric dose of DHEA an increase in PBE mRNA level was observed in the liver at 1 hr and continued to 16 hrs., but not with its metabolites. These results strongly suggest that DHEA or possibly another yet to be identified metabolite might be responsible for peroxisome proliferation.

Regulation of rat hepatic peroxisomal enoyl-CoA hydratase-3-hydroxyacyl-CoA dehydrogenase bifunctional enzyme by thyroid hormone.

Rat hepatic t protein that is negatively regulated by thyroid hormone in nuclear globulin extract was characterized by the antibodies. The following evidence indicated that t protein is a peroxisomal enoyl-CoA hydratase-3-hydroxyacyl-CoA dehydrogenase bifunctional enzyme (bifunctional enzyme). 1. Both proteins had an identical molecular size, and were immunologically indistinguishable from each other. 2. The t protein was abundant in mitochondrial fraction which contained abundant peroxisomes. 3. The amount of the t protein was increased by a peroxisomal proliferator. 4. The activity of the peroxisomal bifunctional enzyme corresponded to the t protein in CM-Sephadex column chromatography. The amount of peroxisomal bifunctional enzyme was increased by thyroidectomy and decreased by 3,5,3'- triiodo-L-thyronine treatment in the whole homogenate of rat liver. These results indicate that the levels of peroxisomal bifunctional enzyme were regulated by thyroid hormone in vivo.