Integration of a Multichannel Pulsed-Valve Inlet System to a Linear Quadrupole Ion Trap Mass Spectrometer for the Rapid Consecutive Introduction of Nine Reagents for Diagnostic Ion/Molecule Reactions.

Gas-phase ion/molecule reactions have been used extensively for the structural elucidation of organic compounds in tandem mass spectrometry. Reagents for ion/molecule reactions can be introduced into a mass spectrometer via a continuous flow apparatus or through a pulsed inlet system. However, most of these approaches enable the use of only a single reagent at a time. In this work, a multichannel pulsed-valve inlet system was developed for the rapid consecutive introduction of up to nine different reagents or reagent systems into a linear quadrupole ion trap mass spectrometer for diagnostic gas-phase ion/molecule reactions. Automated triggering of the pulsed valves enabled these experiments to be performed on the high-performance liquid chromatography (HPLC) time scale. This enables high-throughput screening of several functionalities in analytes as they elute from an HPLC column.

Adjacent pulsed nanoelectrospray ionization emitters for the alternating generation of ions of opposite polarity.

An approach that allows for adjacent closely spaced nanoelectrospray ionization (nESI) emitters to be pulsed alternately to generate ions of opposite polarity for transmission through a common interface is described. The potential difference between two or more nESI emitters in close proximity is minimized by applying the same polarity to both emitters at any given point in time but with the magnitude of only the active emitter's potential being sufficiently high to sustain a stable spray. The reduced difference in potential between emitters allows the distance between emitters to be decreased to within a few millimeters so that compromises imposed by the use of multiple emitters for the generation of ions from distinct solutions using a common atmosphere interface are minimized.

Evaluation of a potential tigecycline-warfarin drug interaction.

STUDY OBJECTIVE: To evaluate the potential for a clinically significant drug interaction between tigecycline and warfarin by using pharmacokinetic and anticoagulant assessments. DESIGN: Open-label, nonrandomized study. SETTING: Inpatient clinical pharmacology unit. SUBJECTS: Nineteen healthy male volunteers were enrolled; eight completed all study assessments. INTERVENTION: All subjects received a single oral dose of warfarin 25 mg (day 1). Seven days later (day 8), they received a 100-mg loading dose of intravenous tigecycline, followed by 50 mg every 12 hours for eight additional doses. On day 12, they received another single oral dose of warfarin 25-mg with their last dose of tigecycline. MEASUREMENTS AND MAIN RESULTS: Serum tigecycline and plasma R- and S-warfarin concentrations were determined by high-performance liquid chromatography with tandem mass spectroscopy. Pharmacokinetic parameters were calculated by using noncompartmental methods and analyzed by the two 1-sided tests equivalence procedure. Pharmacodynamic analyses were based on anticoagulant parameters derived from international normalized ratios of prothrombin times. Tigecycline peak concentration, trough concentration, area under the concentration-time curve (AUC) from 0-12 hrs, and clearance were not affected by single-dose warfarin. In contrast, R- and S-warfarin AUC from time zero extrapolated to infinity was increased by 68% and 29%, respectively, and clearance was decreased by 40% and 23%, respectively, when warfarin was administered after eight doses of tigecycline. Nevertheless, tigecycline did not alter the anticoagulant effects of warfarin, which is consistent with a mechanism based only on increased warfarin protein binding. CONCLUSION: These results suggest that a dosage adjustment of either drug is not necessary during coadministration of tigecycline and warfarin. However, consistent with good medical practice, the anticoagulant activity of warfarin should be monitored during coadministration with tigecycline.

Pharmacokinetics of sirolimus (rapamycin) in subjects with severe hepatic impairment.

Nine subjects with severe hepatic impairment (Child-Pugh grade C) and 9 healthy matched control subjects were given a single 15-mg dose of sirolimus by oral solution. Increases (P < or = .002) in mean whole-blood sirolimus t(1/2) (168%), AUC(0-infinity) (210%), and MRT(oral) (261%), together with a decrease (P = .001) in CL/F (-67%), were observed in subjects with severe hepatic impairment compared with healthy matched controls. Sirolimus pharmacokinetic data in Child-Pugh grade A (n = 13, mild) and B (n = 5, moderate) subjects from a previous identically designed study were available for an inter-study comparison. Overall, mean t(1/2), weight-normalized AUC, and MRT(oral) increased steadily, whereas mean CL/F decreased steadily, with increasing degrees of hepatic impairment. CL/F showed large intersubject variabilities within subject types and extensive overlap among the subject types. The results of this study suggest that an initial sirolimus dose reduction of approximately 60% is appropriate in patients with acute severe hepatic impairment; this should be followed by further dose adjustment, based on therapeutic drug monitoring, until the trough concentrations have stabilized at sirolimus levels existing prior to the onset of acute liver failure.

Absence of an interaction between tigecycline and digoxin in healthy men.

STUDY OBJECTIVE: To evaluate a potential interaction between tigecycline and digoxin using pharmacokinetic and pharmacodynamic assessments. DESIGN: Open-label, three-period, one-sequence crossover study. SETTING: Hospital-affiliated, inpatient clinical pharmacology unit. SUBJECTS: Twenty healthy men. INTERVENTION: Tigecycline 100 mg was administered intravenously as a single dose on day 1 (period 1). Digoxin was administered as a 0.5-mg oral loading dose on day 7, followed by 0.25 mg/day on days 8-14 (period 2). Digoxin 0.25 mg/day was continued on days 15-19; in addition, on day 15, a loading dose of tigecycline 100 mg was administered intravenously, followed by 50 mg every 12 hours starting on the evening of day 15 through the morning of day 19 (period 3). MEASUREMENTS AND MAIN RESULTS: Pharmacokinetic assessments were performed on days 1 and 19 for tigecycline and on days 14 and 19 for digoxin. Electrocardiographic parameters were measured at baseline and on days 1, 14, and 19 to assess digoxin pharmacodynamics. Serum tigecycline concentrations were determined by liquid chromatography with tandem mass spectrometry detection, and plasma and urine digoxin concentrations were determined by radioimmunoassay. Tigecycline area under the concentration-time curve (AUC), AUC from 0-12 hours (AUC(0-12)), weight-normalized clearance, and mean resistance time were not affected by concomitant multiple-dose digoxin administration, but tigecycline half-life was decreased during period 1, apparently due to fewer detectable terminal concentrations in some subjects. Digoxin steady-state AUC(0-24), weight-normalized oral dose clearance, cumulative amount of drug excreted in urine over 24 hours, renal clearance, and QTc (change from baseline) were not affected by multiple-dose tigecycline administration. CONCLUSION: No significant effects of tigecycline on digoxin pharmacokinetics and pharmacodynamics were noted, but a small effect of digoxin on tigecycline pharmacokinetics cannot be ruled out due to design issues with period 1 of the study.

A comparative study of sirolimus tablet versus oral solution for prophylaxis of acute renal allograft rejection.

This multicenter, open-label study compared the efficacy, safety, and pharmacokinetic parameters of sirolimus (rapamycin) tablet and liquid formulations for prevention of efficacy failure. A total of 477 renal allograft recipients were randomly assigned (1:1) to receive either tablet or solution formulations of sirolimus for 12 months, plus cyclosporine (CsA) and steroids. Pharmacokinetic parameters were analyzed based on trough concentrations and 24-hour pharmacokinetic profiles. There were no significant differences in efficacy failure at 3 or 12 months between tablet and solution groups. Graft survival, patient survival, rate of first biopsy-confirmed acute rejection, time to and severity of acute rejection, and laboratory parameters were not significantly different between groups. Mean steady-state sirolimus and CsA pharmacokinetic parameters on days 30 and 90 were not significantly different by formulation, except for longer sirolimus t(max) after tablet administration. Multivariate logistic regression analysis indicated that low sirolimus C(min,TN) and more human leukocyte antigen mismatches were predictors of acute rejection. The tablet and solution formulations of sirolimus demonstrated therapeutic equivalence.

Pharmacokinetics and metabolic disposition of sirolimus in healthy male volunteers after a single oral dose.

The pharmacokinetics and metabolic disposition of sirolimus (rapamycin, Rapamune), a macrocyclic immunosuppressive agent for the prevention of allograft rejection in organ transplantation, were investigated in 6 healthy male volunteers after a single nominal 40-mg oral dose of the C-radiolabeled drug, with the added aim of assessing the potential role of sirolimus metabolites in the clinical pharmacology of the parent drug. The absorption of parent drug and derived materials was rapid (tmax 1.3 +/- 0.5 hours, mean +/- SD), and the elimination of sirolimus was slow (t(1/2) 60 +/- 10 hours, mean +/- SD) in whole blood. The high whole blood to plasma (B/P) concentration ratio of sirolimus (142 +/- 39) was consistent with its extensive partitioning into formed blood elements. The markedly lower B/P value based on radioactivity (2.7 +/- 0.4) suggested that drug-derived products partitioned into formed blood elements to a much lesser extent. Based on AUC0-144h values, unchanged sirolimus represented an average 35% of total radioactivity in whole blood. Drug-derived products in whole blood were characterized by HPLC, LC/MS, and LC/MS/MS as 41-O-demethyl, 7-O-demethyl, and several hydroxy, dihydroxy, hydroxy-demethyl and didemethyl sirolimus metabolites. The percentage distribution of sirolimus metabolites in whole blood ranged from 3%-10% at 1 hour to 6%-17% at 24 hours after drug administration. Based on their low immunosuppressive activities and relative abundance in whole blood of humans after sirolimus administration, metabolites of sirolimus do not appear to play a major role in the clinical pharmacology of the parent drug. A majority of the administered radioactivity (91.0 +/- 8.0%) was recovered from feces, and only 2.2% +/- 0.9% was renally excreted.

Pharmacokinetics of sirolimus (rapamycin) in subjects with mild to moderate hepatic impairment.

Eighteen adult subjects with mild to moderate hepatic impairment and 18 healthy control subjects were given a single 15-mg dose of sirolimus by oral solution. Mean whole-blood sirolimus weight-normalized oral-dose clearances (CL/F) were significantly decreased (P = .02) in subjects with mild to moderate hepatic impairment by -31.8% and -36.0%, respectively, compared with controls. There were no significant differences in mean sirolimus C(max) and t(max) values among groups. The observed decreases in CL/F may be relevant in renal transplant patients with mild to moderate hepatic impairment, based on the close similarity of sirolimus CL/F in controls and previously studied stable renal transplant patients receiving multiple-dose administration of sirolimus and cyclosporine. There was considerable overlap in the CL/F values of hepatic-impaired subjects and controls, suggesting that whole-blood sirolimus trough concentrations in renal transplant patients exhibiting mild to moderate hepatic impairment be initially monitored to assess the need for dose adjustments.

Safety and pharmacokinetics of ascending single doses of sirolimus (Rapamune, rapamycin) in pediatric patients with stable chronic renal failure undergoing dialysis.

Sirolimus (Rapamune, rapamycin) has been shown to be an effective and safe immunosuppressive drug in adult kidney transplant patients when administered concomitantly with cyclosporine (CsA) and steroids. This study reports on a phase 1 assessment of the drug's tolerance, safety, and pharmacokinetic parameters in pediatric patients. The safety and pharmacokinetic profiles of ascending single doses of sirolimus oral solution were investigated in 32 clinically stable pediatric patients on chronic hemodialysis (n = 26) or peritoneal dialysis (n = 6). Patients were divided into two age groups (5-11 and 12-18 yr), and each patient received either a single dose of sirolimus (1, 3, 9, or 15 mg/m(2)) or placebo. Whole blood and plasma samples were collected from each patient for the determination of sirolimus pharmacokinetic parameters. Safety assessments were based on reports of adverse events and results of scheduled physical examinations, vital sign measurements and clinical laboratory tests. The younger patients (5-11 yr) showed statistically significant increases in whole blood sirolimus t(max) (p < or = 0.05) and weight-normalized CL/F (p<0.05) when compared with older patients (12-18 yr). There were no differences in terminal t(1/2), V(ss)/F, dose-normalized peak concentration (C(max)) and AUC, or the B/P. The whole blood sirolimus mean t(max) and weight-normalized CL/F in younger patients were increased by approximately 41.5% and 30%, respectively. Whole blood sirolimus concentrations exhibited less than proportional increases with ascending doses, which may have been caused by the large inter-subject variability in CL/F, small number of subjects, and a potentially inherent decrease in sirolimus bioavailability in younger pediatric patients. Adverse events occurred in all dose and age groups, with headache and stomach pain being the most frequently observed events. No deaths or serious adverse events were reported, and no patient withdrew from the study because of an adverse event. Based on an inter-study analysis, weight-normalized CL/F in the current population of younger pediatric dialysis patients (5-11 yr, 544 +/- 463 mL/h/kg, n = 7) was increased by 90% (p < or = 0.05) compared with healthy adults (19-36 yr, 287 +/- 111 mL/h/kg, n = 25). These results suggest that younger pediatric patients might require an increased maintenance dose of sirolimus to achieve whole blood exposures similar to those in healthy adults. Sirolimus is well tolerated as a single dose of 1, 3, 9, or 15 mg/m(2).

Exposure-response relationships and drug interactions of sirolimus.

Sirolimus (rapamycin, RAPAMUNE, RAPA) is an immunosuppressive agent used for the prophylaxis of renal allograft rejection and exhibits an immunosuppressive mechanism that is distinct from that for cyclosporine and tacrolimus. The purpose of this manuscript is to discuss the exposure-response relationships and drug interactions of sirolimus. The various factors affecting sirolimus whole blood exposure included first-pass extraction, formulation, food, demographics, liver disease, assay method, and interacting drugs. Clinically significant effects caused by food, pediatric age, hepatic impairment, and interacting drugs require recommendations for the safe and efficacious use of sirolimus in renal allograft patients. An exposure-response model based on multivariate logistic regression was developed using the interstudy data from 1832 renal allograft patients. The analysis revealed an increased probability of acute rejection for sirolimus troughs <5 ng/mL, cyclosporine troughs <150 ng/mL, human leukocyte antigen (HLA) mismatches > or =4, and females. The outcomes suggested that individualization of sirolimus doses immediately after transplantation, based on HLA mismatch and sex, would likely decrease the probability of acute rejections in renal allograft recipients who receive concomitant sirolimus, cyclosporine (full-dose), and corticosteroid therapy. Sirolimus is a substrate for both Cytochrome P450 3A (CYP3A) and P-glycoprotein (P-gp) and undergoes extensive first-pass extraction. Drugs that are known to inhibit or induce these proteins may potentially affect sirolimus whole blood exposure. In healthy volunteers, cyclosporine, diltiazem, erythromycin, ketoconazole, and verapamil significantly increased sirolimus whole blood exposure, and rifampin significantly decreased sirolimus exposure. However, sirolimus whole blood exposure was not affected by acyclovir, atorvastatin, digoxin, ethinyl estradiol/norgestrel, glyburide, nifedipine, or tacrolimus. Among the 15 drugs studied, sirolimus significantly increased the exposures of only erythromycin and S-(-)verapamil.

Pharmacokinetic interactions between sirolimus and microemulsion cyclosporine when orally administered jointly and 4 hours apart in healthy volunteers.

Sirolimus (RAPA) and cyclosporine (CsA) are immunosuppressive compounds that are being used concomitantly in renal transplant patients. Both drugs are dosed orally, have common intestinal and hepatic metabolism and intestinal transport mechanisms, and thus offer potential for pharmacokinetic drug interactions. A single-dose, open-label, four-period, four-treatment, randomized crossover study was completed in 15 male and 6 female volunteers. Each subject received a 10-mg oral dose of RAPA alone (Rapamune Oral Solution), a 300-mg oral dose of CsA alone (3 x 100-mg Neoral Soft Gelatin Capsules), RAPA and CsA jointly, and CsA followed by RAPA delayed by 4 hours. Blood samples were collected for either 144 hours (RAPA) or 48 hours (CsA) and analyzed by either liquid chromatography/tandem mass spectrometry (RAPA) or radioimmunoassay (CsA). RAPA bioavailability was markedly increased by CsA when given jointly, with Cmax,tmax, and AUC being increased 116%, 92%, and 230%, respectively. However, when RAPA was administered 4 hours after CsA, increases in RAPA Cmax, tmax, and AUC were only 37%, 58%, and 80%, respectively. CsA did not affect t1/2 or mean residence time (MRT) by either mode of combined administration. RAPA did not significantly affect CsA bioavailability after either joint or delayed combined administrations. It was concluded that CsA markedly increases the bioavailability of RAPA, which may be attributed to a large intestinal and hepatic first-pass effect, rather than altered elimination. RAPA did not affect the bioavailability of CsA.