Identification of 1- and 3-methylhistidine as biomarkers of skeletal muscle toxicity by nuclear magnetic resonance-based metabolic profiling.

Nuclear magnetic resonance (NMR)-based metabolomic profiling identified urinary 1- and 3-methylhistidine (1- and 3-MH) as potential biomarkers of skeletal muscle toxicity in Sprague-Dawley rats following 7 and 14 daily doses of 0.5 or 1mg/kg cerivastatin. These metabolites were highly correlated to sex-, dose- and time-dependent development of cerivastatin-induced myotoxicity. Subsequently, the distribution and concentration of 1- and 3-MH were quantified in 18 tissues by gas chromatography-mass spectrometry. The methylhistidine isomers were most abundant in skeletal muscle with no fiber or sex differences observed; however, 3-MH was also present in cardiac and smooth muscle. In a second study, rats receiving 14 daily doses of 1mg/kg cerivastatin (a myotoxic dose) had 6- and 2-fold elevations in 1- and 3-MH in urine and had 11- and 3-fold increases in 1- and 3-MH in serum, respectively. Selectivity of these potential biomarkers was tested by dosing rats with the cardiotoxicant isoproterenol (0.5mg/kg), and a 2-fold decrease in urinary 1- and 3-MH was observed and attributed to the anabolic effect on skeletal muscle. These findings indicate that 1- and 3-MH may be useful urine and serum biomarkers of drug-induced skeletal muscle toxicity and hypertrophy in the rat, and further investigation into their use and limitations is warranted.

Modulation of ascorbic acid metabolism by cytochrome P450 induction revealed by metabonomics and transcriptional profiling.

In the present study, NMR-based urinary metabonomic profiles resulting from dosing with widely recognized microsomal enzyme inducers were evaluated in male rats. Wistar or Sprague-Dawley rats were dosed daily by oral gavage with phenobarbital (PB; 100 mg/kg), diallyl sulfide (DAS; 500 mg/kg), the investigational compound DMP-904 (150 mg/kg), or beta-naphthoflavone (BNF; 100 mg/kg) for 4 days, and urine was collected daily for analysis. Compounds known to increase cytochrome P450 2B enzymes, including PB, DAS and DMP-904, increased the urinary excretion of gulonic and ascorbic acid in a time-dependent manner, reaching a maximum following 3-4 days of dosing. In contrast, BNF, an agent that induces primarily Cyp1A enzymes, did not increase gulonic or ascorbic acid excretion, despite inducing Cyp1A1 more than 200-fold. Given the metabonomic results, hepatic transcriptional changes in the regulation of ascorbic acid biosynthesis were determined by RT-PCR. All Cyp2B inducers increased hepatic mRNA levels of aldo-keto reductase 1A1, an enzyme that catalyzes the formation of gulonic acid from glucuronate with concurrent decreased expression of both regucalcin (Rgn), the enzyme responsible for conversion of gulonic acid to gulono-1, 4-lactone and gulonolactone oxidase (Gulo), the rate-limiting enzyme in ascorbate biosynthesis. These effects would be expected to increase levels of gulonic acid. In addition, Cyp2B inducers also increased hepatic expression of enzymes regulating ascorbic acid reutilization including glutaredoxin reductase (Glrx2) and thioredoxin reductase (Txnrd1). In contrast, BNF did not effect hepatic expression of any enzyme regulating gulonic or ascorbic acid biosynthesis. Thus, some microsomal enzyme inducers alter transcriptional regulation of ascorbic acid biosynthesis, and these changes are detected by noninvasive metabonomic profiling. However, not all microsomal enzyme inducers appear to alter ascorbic acid metabolism. Finally, the work illustrates how metabonomic results can direct additional studies to determine the biochemical mechanisms underlying changes in urinary metabolite excretion.