Sensitive and specific LC-MS/MS method for the simultaneous measurements of viramidine and ribavirin in human plasma.

Viramidine is a prodrug of ribavirin. To facilitate pharmacokinetics studies of viramidine in man, a sensitive and specific LC-MS/MS method for the simultaneous analyses of viramidine and ribavirin in human plasma was developed and validated. The method involved the addition of [13C]viramidine and [13C]ribavirin as internal standards, protein precipitation with acetonitrile, HPLC separation, and quantification by MS/MS system using positive electrospray ionization in the multiple reaction monitoring mode (MRM). The precursor-->product ion transitions were monitored at 245-->113, 250-->113, 244-->112, and 249-->112 for ribavirin, [13C]ribavirin, viramidine, and [13C]viramidine, respectively. The calibration curves for viramidine and ribavirin were linear over a concentration range of 1-1000 ng/mL. For both viramidine and ribavirin, the lower limit of quantification (LLOQ) was 1 ng/mL. For viramidine, intra- and inter-day analyses of QC samples at 1, 5, 250, and 1000 ng/mL indicated good precision (%CV between 1.0 and 7.0%) and accuracy (%bias between -4.3 and 5.2%). For ribavirin, intra- and inter-day analyses of QC samples at 1, 5, 250, and 1000 ng/mL indicated similar precision (%CV between 0.8 and 8.3%) and accuracy (%bias between -5.8 and 9.4%). Both viramidine and ribavirin were stable in human plasma stored at room temperature for at least 3 h, 4 degrees C for at least 6 h, and for at least three freeze-thaw cycles. This accurate and highly specific assay provides a useful method for evaluating the pharmacokinetics of viramidine and ribavirin in man following administration of viramidine.

Reversal of cycloheximide-induced memory disruption by AIT-082 (Neotrofin) is modulated by, but not dependent on, adrenal hormones.

RATIONALE: AIT-082 (Neotrofin), a hypoxanthine derivative, has been shown to improve memory in both animals and humans. In animals, adrenal hormones modulate the efficacy of many memory-enhancing compounds, including piracetam and tacrine (Cognex). OBJECTIVE: To investigate the role of adrenal hormones in the memory-enhancing action of AIT-082. METHODS: Plasma levels of adrenal hormones (corticosterone and aldosterone) in mice were significantly reduced by surgical or chemical (aminoglutethimide) adrenalectomy or significantly elevated by oral administration of corticosterone. The effects of these hormone level manipulations on the memory-enhancing activity of AIT-082 and piracetam were evaluated using a cycloheximide-induced amnesia/passive avoidance model. RESULTS: As previously reported by others, the memory enhancing action of piracetam was abolished by adrenalectomy. In contrast, the memory enhancement by 60 mg/kg AIT-082 (IP) was unaffected. However, a sub-threshold dose of AIT-082 (0.1 mg/kg, IP) that did not improve memory in control animals did improve memory in adrenalectomized animals. These data suggested that, similar to piracetam and tacrine, the memory enhancing action of AIT-082 might be inhibited by high levels of adrenal hormones. As expected, corticosterone (30 and 100 mg/kg) inhibited the action of piracetam, however no dose up to 100 mg/kg corticosterone inhibited the activity of AIT-082. CONCLUSIONS: These data suggest that while AIT-082 function is not dependent on adrenal hormones, it is modulated by them. That memory enhancement by AIT-082 was not inhibited by high plasma corticosterone levels may have positive implications for its clinical utility, given that many Alzheimer's disease patients have elevated plasma cortisol levels.

Neotrofin is transported out of brain by a saturable mechanism: possible involvement of multidrug resistance and monocarboxylic acid transporters.

Neotrofin (AIT-082; leteprinim potassium) is transported out of brain by a saturable mechanism and in this study the mechanisms mediating this efflux were evaluated. Intracerebroventricular coadministration of [(14)C]Neotrofin with verapamil, a P-glycoprotein inhibitor, probenecid, an organic anion transporter inhibitor, 3-[[3-[2-(7-chloroquinolin-2-yl)vinyl]phenyl]-(2-dimethylcarbamoylethylsulfanyl)methylsulfanyl] propionic acid (MK571), a multidrug resistance-associated protein inhibitor, and salicylate or benzoate, both monocarboxylic acid transporter substrates, inhibited the efflux of [(14)C]Neotrofin. Additionally, Neotrofin inhibited the efflux of [(3)H]quinidine from brain. Compounds can diffuse from cerebrospinal fluid (CSF) into extracellular fluid of brain parenchyma and thus, efflux of [(14)C]Neotrofin after intracerebroventricular administration may indicate active transport across choroid plexus epithelium, brain capillary endothelium, or both. To determine whether [(14)C]Neotrofin efflux occurs at the brain capillary endothelium, experiments were performed in which [(14)C]Neotrofin was administered intraparenchymally. The t(1/2) for [(14)C]Neotrofin disappearance from brain after intraparenchymal administration was significantly lower than that for [(3)H]sucrose and the efflux of Neotrofin was inhibited by 600-fold excess of unlabeled Neotrofin, verapamil, MK571, and salicylate. Together, these data suggest that a saturable mechanism for the efflux of Neotrofin is located at the blood-brain barrier and possibly the blood-CSF barrier. It is likely that multiple transporters are involved in the efflux of Neotrofin and these may include multidrug resistance and monocarboxylic acid transporters. These data are discussed in detail with respect to the site of transporter expression, the recent identification of numerous multidrug resistance-associated protein and monocarboxylic acid transporter homologs, the existence of other potential brain efflux transporters, and the availability of specific pharmacological agents with which to distinguish these transporters.