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.

A 9-wk docosahexaenoic acid-enriched supplementation improves endurance exercise capacity and skeletal muscle mitochondrial function in adult rats.

Decline in skeletal muscle mass and function starts during adulthood. Among the causes, modifications of the mitochondrial function could be of major importance. Polyunsaturated fatty (ω-3) acids have been shown to play a role in intracellular functions. We hypothesize that docosahexaenoic acid (DHA) supplementation could improve muscle mitochondrial function that could contribute to limit the early consequences of aging on adult muscle. Twelve-month-old male Wistar rats were fed a low-polyunsaturated fat diet and were given DHA (DHA group) or placebo (control group) for 9 wk. Rats from the DHA group showed a higher endurance capacity (+56%, P < 0.05) compared with control animals. Permeabilized myofibers from soleus muscle showed higher O2 consumptions (P < 0.05) in the DHA group compared with the control group, with glutamate-malate as substrates, both in basal conditions (i.e., state 2) and under maximal conditions (i.e., state 3, using ADP), along with a higher apparent Km for ADP (P < 0.05). Calcium retention capacity of isolated mitochondria was lower in DHA group compared with the control group (P < 0.05). Phospho-AMPK/AMPK ratio and PPARδ mRNA content were higher in the DHA group compared with the control group (P < 0.05). Results showed that DHA enhanced endurance capacity in adult animals, a beneficial effect potentially resulting from improvement in mitochondrial function, as suggested by our results on permeabilized fibers. DHA supplementation could be of potential interest for the muscle function in adults and for fighting the decline in exercise tolerance with age that could imply energy-sensing pathway, as suggested by changes in phospho-AMPK/AMPK ratio.

Disturbance of mitochondrial energy homeostasis caused by the metabolites accumulating in LCHAD and MTP deficiencies in rat brain.

AIMS: We investigated the in vitro effects of 3-hydroxydodecanoic (3HDA), 3-hydroxytetradecanoic (3HTA) and 3-hydroxypalmitic (3HPA) acids, which accumulate in tissues of patients affected by mitochondrial trifunctional protein (MTP) and isolated long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) deficiencies, on various parameters of energy homeostasis in mitochondrial preparations from brain of young rats. MAIN METHODS: We measured the respiratory parameters state 4, state 3, respiratory control ratio (RCR) and ADP/O ratio by the rate of oxygen consumption, as well as the mitochondrial membrane potential and the matrix NAD(P)H levels in the presence of the fatty acids. KEY FINDINGS: We found that 3HDA, 3HTA and 3HPA markedly increased state 4 respiration and diminished the RCR using glutamate plus malate or succinate as substrates. 3HTA and 3HPA also diminished the mitochondrial membrane potential and the matrix NAD(P)H levels. In addition, 3HTA decreased state 3 respiration using glutamate/malate, but not pyruvate/malate or succinate as substrates. Our data indicate that the long-chain 3-hydroxy fatty acids that accumulate in LCHAD/MTP deficiencies act as uncouplers of oxidative phosphorylation, while 3HTA also behaves as a metabolic inhibitor. SIGNIFICANCE: It is presumed that impairment of brain energy homeostasis caused by these endogenous accumulating compounds may contribute at least in part to the neuropathology of LCHAD/MTP deficiencies.

Conjugated linoleic acid and cardiac health: oxidative stress and energetic metabolism in standard and sucrose-rich diets.

Studies on conjugated linoleic acid ingestion and its effect on cardiac tissue are necessary for the safe utilization of this compound as supplement for weight loss. Male Wistar 24-rats were divided into four groups (n=6):(C)given standard chow, water and 0.5 ml saline, twice a week by gavage; (C-CLA)receiving standard chow, water and 0.5 ml of conjugated linoleic acid, twice a week, by gavage; (S)given standard chow, saline by gavage, and 30% sucrose in its drinking water; (S-CLA)receiving standard chow, 30% sucrose in its drinking water and conjugated linoleic acid. After 42 days of treatment S rats had obesity with increased abdominal-circumference, dyslipidemia, oxidative stress and myocardial lower citrate synthase(CS) and higher lactate dehydrogenase(LDH) activities than C. Conjugated linoleic acid had no effects on morphometric parameters in C-CLA, as compared to C, but normalized morphometric parameters comparing S-CLA with S. There was a negative correlation between abdominal adiposity and resting metabolic rate. Conjugated linoleic acid effect, enhancing fasting-VO(2)/surface area, postprandial-carbohydrate oxidation and serum lipid hydroperoxide resembled to that of the S group. Conjugated linoleic acid induced cardiac oxidative stress in both fed conditions, and triacylglycerol accumulation in S-CLA rats. Conjugated linoleic acid depressed myocardial LDH comparing C-CLA with C, and beta-hydroxyacyl-coenzyme-A dehydrogenase/CS ratio, comparing S-CLA with S. In conclusion, dietary conjugated linoleic acid supplementation for weight loss can have long-term effects on cardiac health. Conjugated linoleic acid, isomers c9, t11 and t10, c12c9,t11" and "t10,c12" were changed to "c9, t11" and "t10, c12", respectively. Please check if appropriate.--> presented undesirable pro-oxidant effect and induced metabolic changes in cardiac tissue. Nevertheless, despite its effect on abdominal adiposity in sucrose-rich diet condition, conjugated linoleic acid may be disadvantageous because it can lead to oxidative stress and dyslipidemic profile.

Increase of aerobic capacity by submaximal training and high-fat diets.

Forty male Long-Evans rats were used in an 8-week experiment designed to study the possibilities of using fats to increase aerobic work capacity and to elucidate the mechanisms of aerobic adaptation. The animals were divided into four groups: group SD comprised control rats kept on a standard diet and sedentary regimen; group HFD were given a high-fat diet (78 en.% of fats) and no training; group SDT consumed standard diet and were trained submaximally; group HFDT was on a high-fat diet and submaximal training. Submaximal running endurance (SRE) and maximum oxygen uptake (VO2max) tests were carried out in all groups. To clarify some of the mechanisms of aerobic adaptation we measured the glycogen content in m. gastrocnemius and the activity of key enzymes for the carbohydrate and fat metabolism, citrate synthase (CS), 3-hydroxiacetyl-CoA-dehydrogenase (3-HAD), and carnitine palmitoyl transferase (CPT) in muscle homogenates. It was found that a high-fat diet (HFD) improved the submaximal running endurance and if combined with submaximal training increased its effect several-fold. HFD increased the maximum oxygen uptake, training made it even greater, but the effect on VO2max the greatest when the two were used in combination. CS activity in m. soleus in HFD-fed rats was 23% greater than that in group SD, while 3-HAD activity was 27% higher than that in group SD. CPT activity in group HFDT was twice as great as that of the control group. Under the influence of HFD the muscle fibres underwent a shift of the energy substrate from carbohydrates to fats which was proved by the economical muscle glycogen depletion during physical exercise.

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.