Augmentation of anaplerosis, or replenishment of carbon lost during intermediary metabolic transitions, is desirable in energy metabolism defects. Triheptanoin, the triglyceride of 7-carbon heptanoic acid, is anaplerotic via direct oxidation or 5-carbon ketone body generation. In this context, triheptanoin can be used to treat Glucose transporter type 1 deficiency encephalopathy (G1D). An oral triheptanoin dose of 1 g/Kg/day supplies near 35% of the total caloric intake and impacted epilepsy and cognition in G1D. This provided the motivation to establish a maximum, potentially greater dose. Using a 3 + 3 dose-finding approach useful in oncology, we studied three age groups: 4–6, 6.8–10 and 11–16 years old. This allowed us to arrive at a maximum tolerated dose of 45% of daily caloric intake for each group. Safety was ascertained via analytical blood measures. One dose-limiting toxicity, occurring in 1 of 6 subjects, was encountered in the middle age group in the context of frequently reduced gastrointestinal tolerance for all groups. Ketonemia following triheptanoin was determined in another group of G1D subjects. In them, β-ketopentanoate and β-hydroxypentanoate concentrations were robustly but variably increased. These results enable the rigorous clinical investigation of triheptanoin in G1D by providing dosing and initial tolerability, safety and ketonemic potential. ClinicalTrials.gov registration : NCT03041363, first registration 02/02/2017.

Background: The treatment of long-chain mitochondrial p-oxidation disorders (LC-FOD) with a low fat-high carbohydrate diet, a diet rich in medium-even-chain triglycerides (MCT), or a combination of both has been associated with high morbidity and mortality for decades. The pathological tableau appears to be caused by energy deficiency resulting from reduced availability of citric acid cycle (CAC) intermediates required for optimal oxidation of acetylCoA. This hypothesis was investigated by diet therapy with carnitine and anaplerotic triheptanoin (TH).Methods: Fifty-two documented LC-FOD patients were studied in this investigation (age range: birth to 51 years). Safety monitoring included serial quantitative measurements of routine blood chemistries, blood levels of carnitine and acylcarnitines, and urinary organic acids.Results: The average frequency of serious clinical complications were reduced from similar to 60% with conventional diet therapy to 10% with TH and carnitine treatment and mortality decreased from similar to 65% with conventional diet therapy to 3.8%. Carnitine supplementation was uncomplicated.Conclusion: The energy deficiency in LC-FOD patients was corrected safely and more effectively with the triheptanoin diet and carnitine supplement than with conventional diet therapy. Safe intervention in neonates and infants will permit earlier intervention following pre-natal diagnosis or diagnosis by expanded newborn screening. (C) 2015 Elsevier Inc. All rights reserved.

Rett syndrome (RTT) is an autism spectrum disorder (ASD) caused by mutations in the X-linked MECP2 gene that encodes methyl-CpG binding protein 2 (MeCP2). Symptoms range in severity and include psychomotor disabilities, seizures, ataxia, and intellectual disability. Symptom onset is between 6-18 months of age, a critical period of brain development that is highly energy-dependent. Notably, patients with RTT have evidence of mitochondrial dysfunction, as well as abnormal levels of the adipokines leptin and adiponectin, suggesting overall metabolic imbalance. We hypothesized that one contributor to RTT symptoms is energy deficiency due to defective nutrient substrate utilization by the TCA cycle. This energy deficit would lead to a metabolic imbalance, but would be treatable by providing anaplerotic substrates to the TCA cycle to enhance energy production. We show that dietary therapy with triheptanoin significantly increased longevity and improved motor function and social interaction in male mice hemizygous for Mecp2 knockout. Anaplerotic therapy in Mecp2 knockout mice also improved indicators of impaired substrate utilization, decreased adiposity, increased glucose tolerance and insulin sensitivity, decreased serum leptin and insulin, and improved mitochondrial morphology in skeletal muscle. Untargeted metabolomics of liver and skeletal muscle revealed increases in levels of TCA cycle intermediates with triheptanoin diet, as well as normalizations of glucose and fatty acid biochemical pathways consistent with the improved metabolic phenotype in Mecp2 knockout mice on triheptanoin. These results suggest that an approach using dietary supplementation with anaplerotic substrate is effective in improving symptoms and metabolic health in RTT.

IMPORTANCE Disorders of brain metabolism are multiform in their mechanisms and manifestations, many of which remain insufficiently understood and are thus similarly treated.Glucose transporter type I deficiency (G1D) is commonly associated with seizures and with electrographic spike-waves.The G1D syndrome has long been attributed to energy (ie, adenosine triphosphate synthetic) failure such as that consequent to tricarboxylic acid (TCA) cycle intermediate depletion.Indeed, glucose and other substrates generate TCAs via anaplerosis.However, TCAs are preserved in murine G1D, rendering energy-failure inferences premature and suggesting a different hypothesis, also grounded on our work, that consumption of alternate TCA precursors is stimulated and may be detrimental.Second, common ketogenic diets lead to a therapeutically counterintuitive reduction in blood glucose available to the G1D brain and prove ineffective in one-third of patients.OBJECTIVE To identify the most helpful outcomes for treatment evaluation and to uphold (rather than diminish) blood glucose concentration and stimulate the TCA cycle, including anaplerosis, in G1D using the medium-chain, food-grade triglyceride triheptanoin. DESIGN, SETTING, AND PARTICIPANTSUnsponsored, open-label cases series conducted in an academic setting.Fourteen children and adults with G1D who were not receiving a ketogenic diet were selected on a first-come, first-enrolled basis.INTERVENTION Supplementation of the regular diet with food-grade triheptanoin.MAIN OUTCOMES AND MEASURES First, we show that, regardless of electroencephalographic spike-waves, most seizures are rarely visible, such that perceptions by patients or others are inadequate for treatment evaluation.Thus, we used quantitative electroencephalographic, neuropsychological, blood analytical, and magnetic resonance imaging cerebral metabolic rate measurements.RESULTS One participant (7%) did not manifest spike-waves; however, spike-waves promptly decreased by 70% (P = .001) in the other participants after consumption of triheptanoin.In addition, the neuropsychological performance and cerebral metabolic rate increased in most patients.Eleven patients (78%) had no adverse effects after prolonged use of triheptanoin.Three patients (21%) experienced gastrointestinal symptoms, and 1 (7%) discontinued the use of triheptanoin. CONCLUSIONS AND RELEVANCETriheptanoin can favorably influence cardinal aspects of neural function in G1D.In addition, our outcome measures constitute an important framework for the evaluation of therapies for encephalopathies associated with impaired intermediary metabolism.

Isovaleric acidemia, resulting from isovaleryl-coenzyme A dehydrogenase deficiency, is associated with marked reduction of free carnitine in both plasma and urine. Fast atom bombardment-mass spectrometry, hydrolysis, and gas chromatography/mass spectrometry have unequivocally identified the existence of isovalerylcarnitine, a new metabolite specific for this disorder. Administration of equimolar amounts of glycine or L-carnitine separately with leucine demonstrated that isovaleryl-coenzyme A is removed by supplemental L-carnitine in the form of isovalerylcarnitine as effectively as it is by glycine, in the form of isovalerylglycine. When L-carnitine is given alone, excretion of isovalerylglycine decreases in preference to enhanced excretion of isovalerylcarnitine and hippurate. Treatment with L-carnitine alone has proven effective in preventing further hospitalizations in a patient with this genetic disorder.

A 19-year-old woman with mild myopathic symptoms from age 6 and fasting intolerance presented with a Reye-like syndrome and a myopathy. Investigations disclosed a lipid storage myopathy, type II glutaric acidemia, and carnitine deficiency in skeletal muscle. Riboflavin and carnitine treatment corrected the metabolic abnormalities and she improved clinically. She later died from pulmonary complications secondary to aspiration. Subsequent studies established electron transfer flavoprotein: ubiquinone oxidoreductase (ETF:QO) deficiency (fibroblast ETF:QO activity was 2.9 mU/mg, normal range is 14.1 +/- 3.8 mU/mg) as the cause of her illness. This is the first documented case of ETF:QO diagnosed in an adult.

Importance The physiological relevance of acid maltase (acid α-glucosidase, an enzyme that degrades lysosomal glycogen) is well recognized in liver and muscle. In late (adult)–onset acid maltase deficiency (glycogen storage disease type II [GSD II]), glycogen accumulates inside muscular lysosomes in the context of reduced enzymatic activity present not only in muscle, but also throughout the organism. Yet, disease manifestations are commonly attributed to lysosomal disruption and autophagic vesicle buildup inside the myofiber due to a lack of obvious hepatic or broader metabolic dysfunction. However, current therapies primarily focused on reducing glycogen deposition by dietary or enzyme replacement have not been consistently beneficial, providing the motivation for a better understanding of disease mechanisms. Objective To provide a systematic overview of metabolism and methylation capacity using widely available analytical methods by evaluating secondary compromise of (1) the citric acid cycle, (2) methylation capacity, and (3) nutrient sensor interaction in as many as 33 patients with GSD II (ie, not all patients were available for all assessments) treated with only a low-carbohydrate/high-protein, calorie-balanced diet. Design, Setting, and Patients Case series including clinical and analytical characterization in an academic setting involving 33 enzymatically proved adults with GSD II treated only with a low-carbohydrate/high-protein, calorie-balanced diet. Main Outcome and Measure Biochemical analysis of blood and urine samples. Results Patients exhibited evidence for disturbed energy metabolism contributing to a chronic catabolic state and those who were studied further also displayed diminished plasma methylation capacity and elevated levels of insulin-like growth factor type 1 and its carrier protein insulin-like growth factor binding protein 3 (IGFBP-3). Conclusions and Relevance The simplest unifying interpretation of these abnormalities is nutrient sensor disturbance with secondary energy failure leading to a chronic catabolic state. Data also provide the framework for the investigation of potentially beneficial interventions, including methylation supplementation, as adjuncts specifically targeted to ameliorate the systemic metabolic abnormalities of this disorder.

IMPORTANCE The physiological relevance of acid maltase (acid α-glucosidase, an enzyme that degrades lysosomal glycogen) is well recognized in liver and muscle. In late (adult)-onset acid maltase deficiency (glycogen storage disease type II [GSD II]), glycogen accumulates inside muscular lysosomes in the context of reduced enzymatic activity present not only in muscle, but also throughout the organism. Yet, disease manifestations are commonly attributed to lysosomal disruption and autophagic vesicle buildup inside the myofiber due to a lack of obvious hepatic or broader metabolic dysfunction. However, current therapies primarily focused on reducing glycogen deposition by dietary or enzyme replacement have not been consistently beneficial, providing the motivation for a better understanding of disease mechanisms. OBJECTIVE To provide a systematic overview of metabolism and methylation capacity using widely available analytical methods by evaluating secondary compromise of (1) the citric acid cycle, (2) methylation capacity, and (3) nutrient sensor interaction in as many as 33 patients with GSD II (ie, not all patients were available for all assessments) treated with only a low-carbohydrate/high-protein, calorie-balanced diet. DESIGN, SETTING, AND PATIENTS Case series including clinical and analytical characterization in an academic setting involving 33 enzymatically proved adults with GSD II treated only with a low-carbohydrate/high-protein, calorie-balanced diet. MAIN OUTCOME AND MEASURE Biochemical analysis of blood and urine samples. RESULTS Patients exhibited evidence for disturbed energy metabolism contributing to a chronic catabolic state and those who were studied further also displayed diminished plasma methylation capacity and elevated levels of insulin-like growth factor type 1 and its carrier protein insulin-like growth factor binding protein 3 (IGFBP-3). CONCLUSIONS AND RELEVANCE The simplest unifying interpretation of these abnormalities is nutrient sensor disturbance with secondary energy failure leading to a chronic catabolic state. Data also provide the framework for the investigation of potentially beneficial interventions, including methylation supplementation, as adjuncts specifically targeted to ameliorate the systemic metabolic abnormalities of this disorder.

APBD is a rare disorder most often affecting adults of Ashkenazi Jewish origin due to partial deficiency of the glycogen brancher enzyme (GBE). It is characterized by progressive involvement of both the central and peripheral nervous systems and deposition of amylopectin-like polyglucosan bodies. There have been no metabolic derangements that might suggest effective therapy nor have there been any clinical improvements for control of its relentless progression. The APBD patients, in this study, experienced stabilization of disease progression, and limited functional improvement in most patients with dietary triheptanoin. Due to a plateau in clinical improvement, the reduced plasma creatinine and methionine levels prompted evaluation of other plasma methylation intermediates in this complex integrated pathway system: decreased S-adenosylmethionine (SAM) (p<0.002), increased S-adenosylhomocysteine (p<0.001), elevated creatine (p=0.001) and increased free choline (p<0.001). Plasma levels of homocysteine and guanidinoacetate were normal. Impaired metabolism of choline and creatine may relate to the progressive dysmyelination and progressive muscle weakness associated with APBD. The partial deficiency of GBE appears to produce a secondary energy deficit possibly related to inadequate reserves of normal glycogen for efficient degradation to free glucose. Dysfunctional regulation of glycogen synthase (GS) may result in continued synthesis and deposition of polyglucosan bodies. This investigation has demonstrated, for the first time, arrest of clinical deterioration with limited functional recovery with triheptanoin diet therapy and the existence of significant derangement of methylation pathways that, when corrected, may lead to even greater therapeutic benefits.

OBJECTIVE:To evaluate effects of fats with odd and even numbers of carbon atoms on muscle metabolism in exercising horses with polysaccharide storage myopathy (PSSM).ANIMALS:8 horses with PSSM (6 females and 2 males; mean +/- SD age, 6.3 +/- 3.9 years).PROCEDURES:Isocaloric diets (grain, triheptanoin, corn oil, and high-fat, low-starch [HFLS] feed) were fed for 3 weeks each; horses performed daily treadmill exercise. Grain was fed to establish an exercise target, and HFLS feed was fed as a negative control diet. Daily plasma samples were obtained. For each diet, a 15-minute exercise test was performed, and gluteus medius muscle specimens and blood samples were obtained before and after exercise.RESULTS:Feeding triheptanoin, compared with the corn oil diet, resulted in exercise intolerance; higher plasma creatine kinase (CK) activity and concentrations of C3:0- and C7:0-acylcarnitine and insulin; and lower concentrations of nonesterified fatty acids (NEFA) and C16:0-, C18:1-, and C18:2-acylcarnitine, without changes in concentrations of plasma glucose or resting muscle substrates and metabolites. Feeding grain induced higher CK activity and insulin concentrations and lower NEFA concentrations than did corn oil or HFLS feed. Feeding grain induced higher glucose concentrations than did triheptanoin and corn oil. In muscle, feeding grain resulted in lower glucose-6-phosphate, higher citrate, and higher postexercise lactate concentrations than did the other diets.CONCLUSIONS AND CLINICAL RELEVANCE:Triheptanoin had detrimental effects, reflecting decreased availability of NEFA, increased insulin stimulation of glycogen synthesis, and potential inhibition of lipid oxidation. Long-chain fats are the best dietetic for PSSM.

Pyruvate carboxylase (PC) is a regulated mitochondrial enzyme that catalyzes the conversion of pyruvate to oxaloacetate, a critical transition that replenishes citric acid cycle intermediates and facilitates other biosynthetic reactions that drive anabolism. Its deficiency causes multiorgan metabolic imbalance that predominantly manifests with lactic acidemia and neurological dysfunction at an early age. Three clinical forms of PC deficiency have been identified: an infantile form (Type A), a severe neonatal form (Type B), and a benign form (Type C), all of which exhibit clinical or biochemical correlates of impaired anaplerosis. There is no effective treatment for these patients and most, except those affected by the benign form, die in early life. We review the physiology of this enzyme and dissect the major clinical, biochemical, and genetic aspects of its dysfunction, emphasizing features that distinguish PC deficiency from other causes of lactic acidemia that render PC deficiency potentially treatable using novel interventions capable of enhancing anaplerosis.

We investigated the interrelations between C4 ketogenesis (production of β-hydroxybutyrate + acetoacetate), C5 ketogenesis (production of β-hydroxypentanoate + β-ketopentanoate), and anaplerosis in isolated rat livers perfused with 13C-labeled octanoate, heptanoate, or propionate. Mass isotopomer analysis of C4 and C5 ketone bodies and of related acyl-CoA esters reveal that C4 and C5 ketogenesis share the same pool of acetyl-CoA. Although the uptake of octanoate and heptanoate by the liver are similar, the rate of C5 ketogenesis from heptanoate is much lower than the rate of C4 ketogenesis from octanoate. This results from the channeling of the propionyl moiety of heptanoate into anaplerosis of the citric acid cycle. C5 ketogenesis from propionate is virtually nil because acetoacyl-CoA thiolase does not favor the formation of β-ketopentanoyl-CoA from propionyl-CoA and acetyl-CoA. Anaplerosis and gluconeogenesis from heptanoate are inhibited by octanoate. The data have implications for the design of diets for the treatment of long chain fatty acid oxidation disorders, such as the triheptanoin-based diet.