Acylcarnitines: Nomenclature, Biomarkers, Therapeutic Potential, Drug Targets, and Clinical Trials.

Acylcarnitines are fatty acid metabolites that play important roles in many cellular energy metabolism pathways. They have historically been used as important diagnostic markers for inborn errors of fatty acid oxidation and are being intensively studied as markers of energy metabolism, deficits in mitochondrial and peroxisomal ß -oxidation activity, insulin resistance, and physical activity. Acylcarnitines are increasingly being identified as important indicators in metabolic studies of many diseases, including metabolic disorders, cardiovascular diseases, diabetes, depression, neurologic disorders, and certain cancers. The US Food and Drug Administration-approved drug L-carnitine, along with short-chain acylcarnitines (acetylcarnitine and propionylcarnitine), is now widely used as a dietary supplement. In light of their growing importance, we have undertaken an extensive review of acylcarnitines and provided a detailed description of their identity, nomenclature, classification, biochemistry, pathophysiology, supplementary use, potential drug targets, and clinical trials. We also summarize these updates in the Human Metabolome Database, which now includes information on the structures, chemical formulae, chemical/spectral properties, descriptions, and pathways for 1240 acylcarnitines. This work lays a solid foundation for identifying, characterizing, and understanding acylcarnitines in human biosamples. We also discuss the emerging opportunities for using acylcarnitines as biomarkers and as dietary interventions or supplements for many wide-ranging indications. The opportunity to identify new drug targets involved in controlling acylcarnitine levels is also discussed. SIGNIFICANCE STATEMENT: This review provides a comprehensive overview of acylcarnitines, including their nomenclature, structure and biochemistry, and use as disease biomarkers and pharmaceutical agents. We present updated information contained in the Human Metabolome Database website as well as substantial mapping of the known biochemical pathways associated with acylcarnitines, thereby providing a strong foundation for further clarification of their physiological roles.

Antibacterial activity of apramycin at acidic pH warrants wide therapeutic window in the treatment of complicated urinary tract infections and acute pyelonephritis.

BACKGROUND: The clinical-stage drug candidate EBL-1003 (apramycin) represents a distinct new subclass of aminoglycoside antibiotics for the treatment of drug-resistant infections. It has demonstrated best-in-class coverage of resistant isolates, and preclinical efficacy in lung infection models. However, preclinical evidence for its utility in other disease indications has yet to be provided. Here we studied the therapeutic potential of EBL-1003 in the treatment of complicated urinary tract infection and acute pyelonephritis (cUTI/AP). METHODS: A combination of data-base mining, antimicrobial susceptibility testing, time-kill experiments, and four murine infection models was used in a comprehensive assessment of the microbiological coverage and efficacy of EBL-1003 against Gram-negative uropathogens. The pharmacokinetics and renal toxicology of EBL-1003 in rats was studied to assess the therapeutic window of EBL-1003 in the treatment of cUTI/AP. FINDINGS: EBL-1003 demonstrated broad-spectrum activity and rapid multi-log CFU reduction against a phenotypic variety of bacterial uropathogens including aminoglycoside-resistant clinical isolates. The basicity of amines in the apramycin molecule suggested a higher increase in positive charge at urinary pH when compared to gentamicin or amikacin, resulting in sustained drug uptake and bactericidal activity, and consequently in potent efficacy in mouse infection models. Renal pharmacokinetics, biomarkers for toxicity, and kidney histopathology in adult rats all indicated a significantly lower nephrotoxicity of EBL-1003 than of gentamicin. INTERPRETATION: This study provides preclinical proof-of-concept for the efficacy of EBL-1003 in cUTI/AP. Similar efficacy but lower nephrotoxicity of EBL-1003 in comparison to gentamicin may thus translate into a higher safety margin and a wider therapeutic window in the treatment of cUTI/API. FUNDING: A full list of funding bodies that contributed to this study can be found in the Acknowledgements section.

Metformin decreases bacterial trimethylamine production and trimethylamine N-oxide levels in db/db mice.

The current study aimed to explore whether metformin, the most widely prescribed oral medication for the treatment of type 2 diabetes, alters plasma levels of cardiometabolic disease-related metabolite trimethylamine N-oxide (TMAO) in db/db mice with type 2 diabetes. TMAO plasma concentration was up to 13.2-fold higher in db/db mice when compared to control mice, while in db/db mice fed choline-enriched diet, that mimics meat and dairy product intake, TMAO plasma level was increased 16.8-times. Metformin (250 mg/kg/day) significantly decreased TMAO concentration by up to twofold in both standard and choline-supplemented diet-fed db/db mice plasma. In vitro, metformin significantly decreased the bacterial production rate of trimethylamine (TMA), the precursor of TMAO, from choline up to 3.25-fold in K. pneumoniae and up to 26-fold in P. Mirabilis, while significantly slowing the growth of P. Mirabilis only. Metformin did not affect the expression of genes encoding subunits of bacterial choline-TMA-lyase microcompartment, the activity of the enzyme itself and choline uptake, suggesting that more complex regulation beyond the choline-TMA-lyase is present. To conclude, the TMAO decreasing effect of metformin could be an additional mechanism behind the clinically observed cardiovascular benefits of the drug.

Long-chain acylcarnitines determine ischaemia/reperfusion-induced damage in heart mitochondria.

The accumulation of long-chain fatty acids (FAs) and their CoA and carnitine esters is observed in the ischaemic myocardium after acute ischaemia/reperfusion. The aim of the present study was to identify harmful FA intermediates and their detrimental mechanisms of action in mitochondria and the ischaemic myocardium. In the present study, we found that the long-chain acyl-CoA and acylcarnitine content is increased in mitochondria isolated from an ischaemic area of the myocardium. In analysing the FA derivative content, we discovered that long-chain acylcarnitines, but not acyl-CoAs, accumulate at concentrations that are harmful to mitochondria. Acylcarnitine accumulation in the mitochondrial intermembrane space is a result of increased carnitine palmitoyltransferase 1 (CPT1) and decreased carnitine palmitoyltransferase 2 (CPT2) activity in ischaemic myocardium and it leads to inhibition of oxidative phosphorylation, which in turn induces mitochondrial membrane hyperpolarization and stimulates the production of reactive oxygen species (ROS) in cardiac mitochondria. Thanks to protection mediated by acyl-CoA-binding protein (ACBP), the heart is much better guarded against the damaging effects of acyl-CoAs than against acylcarnitines. Supplementation of perfusion buffer with palmitoylcarnitine (PC) before occlusion resulted in a 2-fold increase in the acylcarnitine content of the heart and increased the infarct size (IS) by 33%. A pharmacologically induced decrease in the mitochondrial acylcarnitine content reduced the IS by 44%. Long-chain acylcarnitines are harmful FA intermediates, accumulating in ischaemic heart mitochondria and inducing inhibition of oxidative phosphorylation. Therefore, decreasing the acylcarnitine content via cardioprotective drugs may represent a novel treatment strategy.

Suppression of intestinal microbiota-dependent production of pro-atherogenic trimethylamine N-oxide by shifting L-carnitine microbial degradation.

AIMS: Trimethylamine-N-oxide (TMAO) is produced in host liver from trimethylamine (TMA). TMAO and TMA share common dietary quaternary amine precursors, carnitine and choline, which are metabolized by the intestinal microbiota. TMAO recently has been linked to the pathogenesis of atherosclerosis and severity of cardiovascular diseases. We examined the effects of anti-atherosclerotic compound meldonium, an aza-analogue of carnitine bioprecursor gamma-butyrobetaine (GBB), on the availability of TMA and TMAO. MAIN METHODS: Wistar rats received L-carnitine, GBB or choline alone or in combination with meldonium. Plasma, urine and rat small intestine perfusate samples were assayed for L-carnitine, GBB, choline and TMAO using UPLC-MS/MS. Meldonium effects on TMA production by intestinal bacteria from L-carnitine and choline were tested. KEY FINDINGS: Treatment with meldonium significantly decreased intestinal microbiota-dependent production of TMA/TMAO from L-carnitine, but not from choline. 24hours after the administration of meldonium, the urinary excretion of TMAO was 3.6 times lower in the combination group than in the L-carnitine-alone group. In addition, the administration of meldonium together with L-carnitine significantly increased GBB concentration in blood plasma and in isolated rat small intestine perfusate. Meldonium did not influence bacterial growth and bacterial uptake of L-carnitine, but TMA production by the intestinal microbiota bacteria K. pneumoniae was significantly decreased. SIGNIFICANCE: We have shown for the first time that TMA/TMAO production from quaternary amines could be decreased by targeting bacterial TMA-production. In addition, the production of pro-atherogenic TMAO can be suppressed by shifting the microbial degradation pattern of supplemental/dietary quaternary amines.

Magnesium nitrate attenuates blood pressure rise in SHR rats.

The administration of magnesium supplements and nitrates/nitrites decreases arterial blood pressure and attenuates the development of hypertension-induced complications. This study was performed to examine the effects of treatment with magnesium nitrate on the development of hypertension and its complications in spontaneously hypertensive (SHR) rats. Male SHR rats with persistent hypertension at the age of 12-13 weeks were allocated to two groups according to their arterial blood pressure. Rats from the control group received purified water, while the experimental animals from the second group received magnesium nitrate dissolved in purified water at a dose of 50 mg/kg. After four weeks of treatment, blood pressure was measured, the anatomical and functional parameters of the heart were recorded using an ultrasonograph, vascular reactivity was assayed in organ bath experiments and the cardioprotective effects of magnesium nitrate administration was assayed in an ex vivo experimental heart infarction model. Treatment with magnesium nitrate significantly increased the nitrate concentration in the plasma (from 62 ± 8 µmol/l to 111 ± 8 µmol/L), and attenuated the increase in the arterial blood pressure. In the control and magnesium nitrate groups, the blood pressure rose by 21 ± 3 mmHg and 6 ± 4 mmHg, respectively. The administration of magnesium nitrate had no effect on the altered vasoreactivity, heart function or the size of the heart infarction. In conclusion, our results demonstrate that magnesium nitrate effectively attenuates the rise in arterial blood pressure. However, a longer period of administration or earlier onset of treatment might be needed to delay the development of complications due to hypertension.

Effects of long-term mildronate treatment on cardiac and liver functions in rats.

Mildronate is a cardioprotective drug that improves cardiac function during ischaemia and functions by lowering l-carnitine concentration in body tissues and modulating myocardial energy metabolism. The aim of the present study was to characterise cardiovascular function and liver condition after long-term mildronate treatment in rats. In addition, changes in the plasma lipid profile, along with changes in the concentration of mildronate, l-carnitine and gamma-butyrobetaine were monitored in the rat tissues. Wistar rats were perorally treated daily with a mildronate dose of either 100, 200 or 400 mg/kg for 4, 8 or 12 weeks. The l-carnitine-lowering effect of mildronate was dose-dependent. However, the carnitine levels reached a plateau after about four weeks of treatment. During the additional weeks of treatment, the carnitine levels were not considerably changed. The obtained results provide evidence that even a high dose of mildronate does not alter cardiovascular parameters and the function of isolated rat hearts. Furthermore, the histological evaluation of liver tissue cryosections and measurement of biochemical markers of hepatic toxicity showed that all the measured values were within the normal reference range. Our results provide evidence that long-term mildronate administration induces significant changes in carnitine homeostasis, but it is not associated with cardiac impairment or disturbances in liver function.