Population pharmacokinetics of tirilazad: effects of weight, gender, concomitant phenytoin, and subarachnoid hemorrhage.

PURPOSE: Data collected during Phase I and II in the development of tirilazad were pooled and analyzed using nonlinear mixed effects models to assess covariates which might affect tirilazad pharmacokinetics. METHODS: Four single dose and five multiple dose studies in normal volunteers were combined with two multiple dose studies performed in patients with subarachnoid hemorrhage (SAH) to identify factors related to intersubject variability in clearance (CL) and central compartment volume (Vc). Data from 253 subjects, which consisted of 7,219 tirilazad concentrations, were analyzed. The effects of weight, gender, patient versus volunteer status, and phenytoin use were evaluated. RESULTS: Relative to male volunteers not receiving concomitant phenytoin, significant effects on clearance included: a 46% increase in volunteers receiving phenytoin, and an 82% increase in clearance associated with SAH patients (all of whom received phenytoin). Significant effects on Vc were: a 26% increase for female volunteers not receiving phenytoin, a 12% decrease for volunteers receiving concomitant phenytoin, a 152% increase for male SAH patients, and a 270% increase for female SAH patients. Incorporating patient covariate effects substantially reduced the interindividual variability (from 27.9% to 24.7% for clearance and from 48.2% to 37.5% for Vc). Residual variability was estimated at 66% coefficient of variation (CV) in SAH patients and at 22-48% CV over the range of predicted concentrations in normal volunteers. CONCLUSIONS: The most important factors affecting tirilazad pharmacokinetics are the administration of phenytoin (increased CL) and SAH (increased Vc and residual variability). The effect of gender on tirilazad pharmacokinetics was modest.

Hormonal effects on tirilazad clearance in women: assessment of the role of CYP3A.

This study assessed whether the previously reported difference in tirilazad clearance between pre- and postmenopausal women is reversed by hormone replacement and whether this observation can be explained by differences in CYP3A4 activity. Ten healthy women from each group were enrolled: premenopausal (ages 18-35), postmenopausal (ages 50-70), postmenopausal receiving estrogen, and postmenopausal women receiving estrogen and progestin. Volunteers received 0.0145 mg/kg midazolam and 3.0 mg/kg tirilazad mesylate intravenously on separate days. Plasma tirilazad and midazolam were measured by HPLC/dual mass spectrophotometry (MS/MS) assays. Tirilazad clearance was significantly higher in premenopausal women (0.51 +/- 0.09 L/hr/kg) than in postmenopausal groups (0.34 +/- 0.07, 0.32 +/- 0.06, and 0.36 +/- 0.08 L/hr/kg, respectively) (p = 0.0001). Midazolam clearance (0.64 +/- 0.12 L/hr/kg) was significantly higher in premenopausal women compared to postmenopausal groups (0.47 +/- 0.11, 0.49 +/- 0.11, and 0.53 +/- 0.19 L/hr/kg, respectively) (p = 0.037). Tirilazad clearance was weakly correlated with midazolam clearance (r2 = 0.129, p = 0.02). Tirilazad clearance is faster in premenopausal women than in postmenopausal women, but the effect of menopause on clearance is not reversed by hormone replacement. Tirilazad clearance in these women is weakly related to midazolam clearance, a marker of CYP3A activity.

Induction of tirilazad clearance by phenytoin.

Tirilazad is a membrane lipid peroxidation inhibitor being studied for the management of subarachnoid hemorrhage; phenytoin is used for seizure prophylaxis in the same disorder. The induction of tirilazad clearance by phenytoin was assessed in 12 volunteers (6 male, 6 female). Subjects received phenytoin orally every 8 h for 7 days (200 mg for nine doses and 100 mg for 13 doses) in one phase of a crossover study. In both study phases, 1.5 mg kg-1 tirilazad mesylate was administered by i.v. infusion every 6 h for 29 doses. Tirilazad mesylate and U-89678 (an active metabolite) in plasma were quantified by HPLC. After the final dose, tirilazad clearance was increased by 91.8% in subjects receiving phenytoin + tirilazad versus tirilazad alone. AUC0-6 for U-89678 after the last tirilazad dose was reduced by 93.1% by concomitant phenytoin. These effects were statistically significant. The time course of induction was consistent with that of phenytoin's effect on the ratio of urinary 6 beta-hydroxycortisol to cortisol, a measure of hepatic CYP3A activity. The results show that phenytoin induces metabolism of tirilazad and U-89678 in healthy subjects and that, under these conditions, tirilazad clearance approaches liver blood flow.

Pharmacokinetics of tirilazad in healthy male subjects at doses above 6 mg/kg/day.

The dose proportionality of tirilazad pharmacokinetics at dosages above 6.0 mg/kg/day were assessed in 18 healthy male volunteers between the ages of 19 and 46 years. Subjects were randomized to receive either 1.5 mg/kg, 3.0 mg/kg, or 4.0 mg/kg tirilazad mesylate every 6 hours for 29 doses (daily doses of 6.0, 12.0, and 16.0 mg/kg/day for 7 days). Each drug dose was administered intravenously over 10 minutes. Plasma tirilazad, U-89678, and U-87999 (active reduced metabolites) were quantified by HPLC. Two subjects in the high dose group withdrew before the end of the study. Following the first dose of tirilazad, dose-corrected pharmacokinetic parameters for all 3 compounds did not differ significantly among dose groups. After the final tirilazad the mean half-life of tirilazad was approximately 80 hours. Mean apparent tirilazad clearance did not differ significantly among groups. Mean U-89678 AUC0-6 following the last tirilazad dose did not differ significantly between the 6.0 and 12.0 mg/kg/day doses, but the value for the 16.0 mg/kg dose was higher than values from both lower doses (p = 0.044 and 0.056, respectively). Similar results were obtained for U-87999. The dose effects observed for the pharmacokinetics of these 2 metabolites may have been a function of intersubject variability. When combined with previous data concerning the dose proportionally of tirilazad pharmacokinetics at doses less than 6.0 mg/kg/day, the data from the present study suggest that the pharmacokinetics of tirilazad are approximately linear over a dosage range of 1.0-16.0 mg/kg/day. Due to the inability to assess the plasma protein binding of tirilazad and its reduced metabolites, the clinical significance of the departure from linearity of the pharmacokinetics of U-89678 and U-87999 cannot be directly assessed. Further study at higher doses will be needed to address this issue.

Comparison of the pharmacokinetics of tirilazad mesylate in healthy volunteers and stable subjects with mild liver cirrhosis.

OBJECTIVE: The pharmacokinetics of tirilazad mesylate and an active reduced metabolite, U89678, were studied in 7 volunteers with mild cirrhosis of the liver, and seven age, sex, weight and smoking status matched healthy normal volunteers. Subjects received a single intravenous infusion of 2.0 mg.kg-1 tirilazad mesylate over 10 min. RESULTS: Mean tirilazad AUCzero-infinity was 8.83 mumol h.l-1 and 18.6 mumol h.l-1 in healthy volunteers and cirrhotic subjects, respectively. Mean tirilazad clearance in cirrhotics (12.7 l.h-1) was approximately 2.1 fold lower than in healthy volunteers (27.8 l.h-1). The differences were statistically significant. Mean U-89678 AUCzero-infinity in cirrhotic subjects (3.88 mumol h.l-1) was 2.5 fold higher than in healthy controls (1.53 mumol h.l-1), but the difference was marginally significant. CONCLUSION: These results indicate that clearance of both tirilazad mesylate and U89678 is decreased in subjects with hepatic impairment. This observation may be attributed either to decreases in liver blood flow and/or intrinsic clearance. The results of this study thus suggest that increased monitoring and or a reduction in tirilazad dosing may be necessary in patients with hepatic impairment.

Gender does not affect the degree of induction of tirilazad clearance by phenobarbital.

OBJECTIVE: Tirilazad mesylate is a membrane lipid peroxidation inhibitor being evaluated for the treatment of patients with subarachnoid haemorrhage (SAH); phenobarbital may be administered to these patients for seizure prophylaxis. Therefore, the effect of phenobarbital on tirilazad mesylate pharmacokinetics was assessed in 15 healthy volunteers (7M, 8F). METHODS: Subjects received 100 mg phenobarbital orally daily for 8 days in one phase of a two-way crossover study. In both phases, 1.5 mg.kg-1 tirilazed mesylate was administered (as a 10 minute IV infusion) every 6 hours for 29 doses. Three weeks separated study phases. Tirilazad mesylate and U-89678 (an active metabolite) in plasma were quantified by HPLC. RESULTS: Phenobarbital had no effect on the first dose pharmacokinetics of tirilazad or U-89678. After the final dose, clearance for tirilazad was increased 25% in males and 29% in females receiving phenobarbital + tirilazad versus tirilazad mesylate alone. These differences were statistically significant, and the degree of induction was not significantly different between genders. AUC(zero)-6 for U-89678 after the last tirilazad mesylate dose was reduced 51% in males and 69% in females. The decreases were statistically significant, and there was no gender by treatment interaction. CONCLUSION: The results show that phenobarbital induces metabolism of tirilazad and U-89678 similarly in both men and women. Lower levels of tirilazad and U-89678 in SAH patients receiving phenobarbital may adversely impact clinical response.

Effect of age and gender on tirilazad pharmacokinetics in humans.

Tirilazad mesylate pharmacokinetics were assessed in 12 young and 12 elderly volunteers (six men and six women per age group). Subjects received single 10-minute intravenous infusions of 1.5 mg/kg and 3.0 mg/kg tirilazad mesylate. Plasma tirilazad mesylate concentrations were determined by HPLC. There were no significant dose effects on clearance, but half-life increased with dose because of assay insensitivity at the lower dose. Mean half-lives were 16.3 +/- 15.5 and 21.4 +/- 12.6 hours for young and elderly subjects, respectively, at the 3.0 mg/kg dose. At the same dose, mean tirilazad mesylate systemic clearance was 0.630 +/- 0.254 and 0.428 +/- 0.090 L/hr/kg, respectively. The decreased clearance in elderly volunteers was primarily attributable to a lower clearance in elderly women relative to young women. The small effect of age on tirilazad clearance is likely to have minimum clinical impact. Tirilazad clearance was approximately 40% higher in young women than in young men. The clinical importance of this observation is unknown.

Protein binding of tirilazad (U-74006) in human, Sprague-Dawley rat, beagle dog and cynomolgus monkey serum.

Determination of the serum protein binding of tirilazad across species was required to predict the pharmacokinetic behavior of this new drug. Equilibrium dialysis and ultrafiltration techniques, commonly used to study serum protein binding, were shown to be unsuitable for tirilazad due to high nonspecific binding and low aqueous solubility, resulting in unbound drug levels that were nondetectable with current analytical methodology. Ultracentrifugation appeared to offer a technique with which unbound tirilazad could be measured; however, after extensive studies, the apparent lipid partitioning behavior of tirilazad into low density and very low density lipoproteins showed that ultracentrifugation was also unsuitable for determination of the true unbound fraction of tirilazad. Fractions of tirilazad measured in the supernatant were highly correlated with total triglycerides and very low density lipoproteins in the sera of all species analyzed. Studies with delipidized human serum yielded a nonsaturable binding isotherm with free fractions of less than 0.6 +/- 0.02% (mean +/- S.D.) over a concentration range of 4.6 to 55.6 micrograms/ml (normal human in vivo range, 0.01-20 micrograms/ml). These data indicated that, as the triglyceride content of the sera increases, portions of tirilazad bound to serum proteins shift into the lipid phase of lipoproteins. What effect this has on the true unbound fraction is unknown and does not seem to be ascertainable with current technology.

Disposition of tirilazad (U74006F), a 21-aminosteroid, in the plasma, heart, brain, and liver of the rat.

Tirilazad mesylate (Freedox, Upjohn) is the first agent of a new class of compounds called lazaroids currently under clinical investigation for its potential beneficial effects following neurotrauma. Tirilazad's therapeutic effect is due to its ability to inhibit iron-dependent lipid peroxidation. This study was conducted in Sprague-Dawley rats to determine the basic pharmacokinetics and distribution of tirilazad into the brain, heart, and liver. Rats (N = 50) were killed in groups of five at 0, 10, 20, and 40 min, and at 1.5, 2, 3, 4, 6, and 8 hr after intravenous administration of 10 mg/kg of tirilazad mesylate. Tirilazad was assayed in plasma, heart, liver, and brain tissue by HPLC. Using the mean concentrations at each time point, pharmacokinetic parameters were estimated using standard noncompartmental techniques and statistical moment theory. Tirilazad was rapidly eliminated from the plasma with a half-life of 2.4 hr and clearance of 6.1 ml/min. The volume of distribution at steady-state was large, 4.8 liters/kg. The concentrations of tirilazad were highest in the liver and heart and lowest in the plasma and brain. Elimination from tissues paralleled elimination from plasma with half-lives of 1.9, 1.6, and 1.5 hr in the brain, heart, and liver, respectively. Tirilazad appears to be a highly extracted, hepatically eliminated drug, suggesting its clearance is dependent on liver blood flow, and alterations in plasma protein binding are unlikely to affect its clearance but may affect the fraction unbound.

Pretreatment with U74006F improves neurologic outcome following complete cerebral ischemia in dogs.

We examined the 21-aminosteroid U74006F, a potent inhibitor of lipid peroxidation, for potential neuroprotective effects in a canine model of complete cerebral ischemia. Two 1.5-mg/kg boluses were administered to six dogs, the first bolus 15 minutes prior to a 12-minute episode of complete cerebral ischemia and the second bolus after 11 minutes of ischemia, 1 minute prior to reperfusion. Using this dosage regimen, plasma U74006F levels of greater than 0.3 microgram/ml were maintained for up to an hour postischemia. An additional six animals received equal volumes of the citrate vehicle solution. At 24 and 48 hours postischemia, the dogs were neurologically evaluated by an observer blinded as to treatment selection. All six U74006F-treated animals had a normal neurologic outcome at 48 hours postischemia, while the citrate vehicle-treated animals all suffered moderate to severe neurologic deficits. The difference in outcome was significant at both 24 and 48 hours (p less than 0.005). Although U74006F is a 21-aminosteroid, it is not reported to possess glucocorticoid activity. This is supported by the present finding that no changes in plasma glucose concentration were observed following administration of the drug. The systemic vitamin E levels of citrate vehicle-treated animals decreased significantly (from 4.10 +/- 0.46 micrograms/ml to 2.95 +/- 0.38 micrograms/ml, p less than 0.05), whereas the vitamin E levels in U74006F-treated animals did not decrease significantly. These results suggest that U74006F may be of benefit in improving neurologic outcome when administered prior to an episode of complete cerebral ischemia.(ABSTRACT TRUNCATED AT 250 WORDS)

Plasma compatibility of injectables: comparison of intravenous U-74006F, a 21-aminosteroid antioxidant, with Dilantin brand of parenteral phenytoin.

The 21-aminosteroid antioxidant U-74006F (1) is being developed as an iv injectable agent for the treatment of human CNS trauma and ischemia. Because of its poor water solubility, the plasma compatibility of the parenteral formulation of 1 was evaluated using three models: (I) static solubility, (II) aggregometric, and (III) dynamic flow. The flow model was designed to mimic an iv infusion into the human antecubital vein, which was assumed to have plasma flow of 10 mL/min. Dilantin (phenytoin), the positive control, produced a precipitate in all three models from a 10% (v/v) mixture with human plasma, which approximates the in vivo ratio when the drug is infused at the recommended rate of 1 mL/min. Approximately 39% of the phenytoin dose in the flow model was retained on a downstream 3-microns filter as crystals. In comparison, the parenteral formulation of 1 produced minimal precipitate in models I and II from 40% mixtures with plasma, but higher percentages produced unstable suspensions with time-dependent precipitation. The percentage of the dose of the parenteral formulation of 1 retained on the filter in the flow model was 0.5% or less at infusion rates as high as 10 mL/min and 3% at 19 mL/min. At the 10-mL/min infusion rate, the mass of 1 retained on the filter per minute was less than 1% of the mass of phenytoin retained at the 1-mL/min infusion rate for Dilantin. The acceptable plasma compatibility of the parenteral formulation of 1 appears to be related to the solubilizing effects of plasma protein binding and pH suppression by the citric acid vehicle.

Effect of the aminosteroid U74006F after cardiopulmonary arrest in dogs.

The oxygen free radical-induced lipid peroxidative reactions that occur during resuscitation from normothermic cardiac arrest may contribute to the degree of neurologic dysfunction sustained. A blinded, randomized experimental trial was performed to determine whether U74006F, a potent inhibitor of lipid peroxidation, reduces morbidity and 24-hour mortality after 10 minutes of normothermic cardiopulmonary arrest; ventricular fibrillation was induced by electrical stimulation in 24 open-chest, halothane-anesthetized dogs, and circulation was reestablished by direct cardiac compressions, administration of a standardized drug regime, and internal countershocks. When spontaneous circulation was restored, a bolus injection of 1.5 mg/kg U74006F (n = 12) or 25 mM citrate vehicle (n = 12) was infused intravenously in 15 minutes and an infusion was continued at 0.125 mg/kg/hr for the next 12 hours. In the drug-treated group, plasma U74006F concentration averaged 0.13 microgram/ml between 3 and 12 hours after cardiac arrest. By 24 hours after arrest, 10 of 12 (83%) vehicle-treated dogs had died but only four of 12 (33%) U74006F-treated dogs had died (p = 0.017). U74006F-treated dogs survived significantly longer (mean +/- SEM 22 +/- 1 hr) than vehicle-treated dogs (18 +/- 1 hr), with significantly better neurologic function 1, 2, and 24 hours after arrest. Plasma fatty acid hydroperoxide concentrations 12 hours after arrest were 88 +/- 81 pmol/ml in U74006F-treated and 241 +/- 49 pmol/ml in vehicle-treated dogs (p less than 0.05). Vitamin E concentrations were significantly higher in the plasma of U74006F-treated dogs 2, 3, and 6 hours after arrest compared with vehicle-treated dogs.(ABSTRACT TRUNCATED AT 250 WORDS)

High-performance liquid chromatographic determination of a 21-aminosteroid antioxidant in plasma.

A novel 21-aminosteroid, 21-[4-(2,6-di-1-pyrrolidinyl-4-pyrimidinyl)-1- piperazinyl]-16 alpha-methylpregna-1,4,9(11)-triene-3,20-dione monomethanesulfonate (I), is under development for the treatment of central nervous system injury in humans. This report describes a reversed-phase high-performance liquid chromatographic (RP-HPLC) method using ultraviolet detection at 254 nm for the determination of I in plasma with low nanogram per milliliter sensitivity. Plasma was deproteinated by mixing with acetonitrile and centrifuging, and I was extracted from the supernatant with disposable bonded-phase columns. The extracts were chromatographed on an octylsilane bonded-phase HPLC column with an acetonitrile-water mobile phase containing triethylammonium acetate, pH5. Quantification was accomplished by peak-height ratio analysis using an analogue of I as the assay internal standard. The method was suitable for the determination of I following a 30 mg/kg intraperitoneal dose in the rat.