BACKGROUND AND OBJECTIVE Acute repetitive seizures (ARS) are a debilitating part of episodic seizure activity that can sometimes progress to status epilepticus. Currently approved treatment that can be administered by non-medical personnel to patients with ARS is a diazepam rectal gel. While effective, rectal administration can be difficult, inconvenient and objectionable. A diazepam autoinjector has been developed to deliver diazepam via an intramuscular (IM) injection. This study evaluated the dose proportionality of the diazepam autoinjector and the consequent diazepam bioavailability relative to an equivalent dose of diazepam administered rectally as a commercial gel. METHODS This was a phase I, randomized, open-label, two-part, single-dose, crossover, single-centre pharmacokinetic study in 48 healthy young adult (aged 18-40 years) male and female subjects. Part I of the study (n = 24) evaluated the dose proportionality of three strengths of the diazepam autoinjector (5, 10 and 15 mg) administered into the mid-outer thigh via a deep IM injection. Part II (n = 24) assessed the relative bioavailability of the diazepam 10 mg autoinjector versus the diazepam 10 mg rectal gel. Parts I and II were run concurrently. Each subject completed screening up to 30 days prior to three (Part I) or two (Part II) dosing periods. Serial blood sampling for plasma diazepam and desmethyldiazepam (metabolite) concentrations, vital signs and adverse event (AE) assessments were performed at prespecified times. Treatments were separated by a 14-day washout period. RESULTS In Part I, dose proportionality was demonstrated for the diazepam autoinjector at 5, 10 and 15 mg doses by increases in mean maximum plasma concentration (C(max)), mean area under the plasma concentration-time curve (AUC) from time zero to infinity (AUC(∞)), and mean AUC from time zero to time of last measurable concentration (AUC(last)). The median time to reach C(max)(t(max)) was consistent at 1 hour for each dose. In Part II of the study, IM administration via diazepam autoinjector (10 mg) resulted in plasma concentrations of both diazepam and desmethyldiazepam that were slightly higher and less variable than those observed following administration of diazepam rectal gel (10 mg). The geometric mean ratio (diazepam autoinjector/diazepam rectal gel) and 90% confidence intervals for diazepam C(max) and AUC(last) were 0.94 (0.84, 1.05) and 1.14 (1.08, 1.21), respectively, indicating that the overall bioavailability of the diazepam autoinjector was approximately 14% higher than that of diazepam rectal gel. Both treatments were generally well tolerated. Although the incidence of treatment-emergent AEs was higher with diazepam autoinjector compared with diazepam rectal gel (21.7% vs 13.6%), the difference can be attributed to injection site pain. Injection site pain also correlated with the diazepam autoinjector dose administered in Part I: 5 mg (4.3%), 10 mg (21.7%) and 15 mg (27.3%). However, no patients discontinued the trial due to injection site pain. No other AEs correlated with dose, and there was no evidence of respiratory depression with either administration. CONCLUSION Results of the present study indicated that diazepam can be safely and reliably administered IM using a diazepam autoinjector.

1. The primary metabolism of diazepam was studied in human liver microsomes in order to investigate the kinetics and to identify the cytochrome P450 (CYP) isoforms responsible for the formation of the main diazepam metabolites, temazepam and N-desmethyldiazepam. 2. The formation kinetics of both metabolites were atypical and consistent with the occurrence of substrate activation. A sigmoid Vmax model equivalent to the Hill equation was used to fit the data. The degree of sigmoidicity was greater for temazepam formation than for N-desmethyldiazepam formation, so that the ratio of desmethyldiazepam:temazepam formation increased as the substrate (diazepam) concentration decreased. 3. alpha-Naphthoflavone activated both reactions but with a greater effect on temazepam formation than on N-desmethyldiazepam formation. In the presence of 25 microM alpha-naphthoflavone the kinetics for both pathways were approximated by Michaelis-Menten kinetics. 4. Studies with a series of CYP isoform selective inhibitors and with an inhibitory anti-CYP2C antibody indicated that temazepam formation was carried out mainly by CYP3A isoforms, whereas the formation of N-desmethyldiazepam was mediated by both CYP3A isoforms and S-mephenytoin hydroxylase.

Single oral 10 mg doses of diazepam and demethyldiazepam were given on different occasions to 16 healthy subjects. The subjects included four poor hydroxylators of debrisoquin and three poor hydroxylators of mephenytoin. There was a correlation between the total plasma clearance of diazepam and demethyldiazepam (rs = 0.83; p less than 0.01). There was no relationship between benzodiazepine disposition and debrisoquin hydroxylation. Poor hydroxylators of mephenytoin had less than half the plasma clearance of both diazepam (p = 0.0008) and demethyldiazepam (p = 0.0001) compared with extensive hydroxylators of mephenytoin. The plasma half-lives were longer in poor hydroxylators than they were in extensive hydroxylators of mephenytoin for both diazepam (88.3 +/- SD 17.2 and 40.8 +/- 14.0 hours; p = 0.0002) and demethyldiazepam (127.8 +/- 23.0 and 59.0 +/- 16.8 hours; p = 0.0001). There was no significant difference in volume of distribution of the benzodiazepines between the phenotypes. This study shows that the metabolism of both diazepam (mainly demethylation) and demethyldiazepam (mainly hydroxylation) is related to the mephenytoin, but not to the debrisoquin, hydroxylation phenotype.

Eleven healthy volunteers received a single intravenous dose of diazepam (0.15 mg/kg), midazolam (0.1 mg/kg), and placebo by 1-minute infusion in a double-blind, three-way crossover study. Plasma concentrations were measured during 24 hours after dosage, and the electroencephalographic (EEG) power spectrum was simultaneously computed by fast-Fourier transform to determine the percentage of total EEG amplitude occurring in the 13 to 30 Hz range. Both diazepam and midazolam had large volumes of distribution (1.2 and 2.3 L/kg, respectively), but diazepam's half-life was considerably longer (33 versus 2.8 hours) and its metabolic clearance lower (0.5 versus 11.0 ml/min kg) than those of midazolam. EEG changes were maximal at the end of the diazepam infusion and 5 to 10 minutes after midazolam infusion. Percent 13 to 30 Hz activity remained significantly above baseline until 5 hours for diazepam but only until 2 hours for midazolam. For both drugs, EEG effects were indistinguishable from baseline by 6 to 8 hours, suggesting that distribution contributes importantly to terminating pharmacodynamic action. The relationship of EEG change to plasma drug concentration indicated an apparent EC50 value of 269 ng/ml for diazepam as opposed to 35 ng/ml for midazolam. However, Emax values were similar for both drugs (+19.4% and +21.3%, respectively).

PURPOSE. The objective of this study was to develop a model to predict the extent to which bile salts can enhance the solubility of a drug, based on the physicochemical properties of the compound. The ability to predict bile salt solubilization of poorly soluble drugs would be a key component in determining which drugs will exhibit fed vs. fasted differences in drug absorption. METHODS. A correlation between the logarithm of the octanol/water partition coefficient [log P] of six steroidal compounds and their solubilities in the presence of various concentrations of sodium taurocholate at 37 degrees C, log [SR]= 2.234 + 0.606 log [P](r2 = 0.987) where SR is the ratio of the stabilization capacity of the bile salt to the solubilization capacity of water for the drug, was used to predict the solubility of the compounds in presence of sodium taurocholate were then measured. RESULTS. The predicted solubilities were within 10% of the experimentally observed solubilities for griseofulvin, cyclosporin A and pentazocine. The model overpredicted the solubility of phenytoin and diazepam in 15 mM sodium taurocholate solution by a factor of 1.33 and 1.62 respectively. CONCLUSIONS. The expected increase in solubility as a function of bile salt concentration can be estimated on the basis of the partition coefficient and aqueous solubility of the compound.

Some substrates of cytochrome P450 (CYP) 3A4, the most abundant CYP in the human liver responsible for the metabolism of many structurally diverse therapeutic agents, do not obey classical Michaelis-Menten kinetics and demonstrate homotropic and/or heterotropic cooperativity. The unusual kinetics and differential effects observed between substrates of this enzyme confound the prediction of drug clearance and drug-drug interactions from in vitro data. We have investigated the hypothesis that CYP3A4 may bind multiple molecules simultaneously using diazepam (DZ) and testosterone (TS). Both substrates showed sigmoidal kinetics in B-lymphoblastoid microsomes containing a recombinant human CYP3A4 and reductase. When analyzed in combination, TS activated the formation of 3-hydroxydiazepam (3HDZ) and N-desmethyldiazepam (NDZ)(maximal activation 374 and 205%, respectively). For 3HDZ, V(max) values remained constant with increasing TS, whereas the S(50) and Hill values decreased, tending to make the data less sigmoidal. Similar trends were observed for the NDZ pathway. DZ inhibited the formation 6beta-hydroxytestosterone (maximal inhibition, 45% of control), causing a decrease in V(max) but no significant change to the S(50) and Hill values, suggesting that DZ may inhibit via a separate effector site. Multisite rate equation models have been derived to explore the analysis of such complex kinetic data and to allow accurate determination of the kinetic parameters for activation and inhibition. The data and models presented are consistent with proposals that CYP3A4 can bind and metabolize multiple substrate molecules simultaneously; they also provide a generic solution for the interpretation of the complex kinetic data derived from CYP3A4 substrates.

An in situ mouse brain perfusion model predictive of passive and carrier-mediated transport across the blood-brain barrier (BBB) was developed and applied to mdr1a P-glycoprotein (Pgp)-deficient mice [mdr1a(-/-)]. Cerebral flow was estimated from diazepam uptake. Physical integrity of the BBB was assessed with sucrose/inulin spaces; functional integrity was assessed with glucose uptake, which was saturable with a Km of approximately 17 mmol/L and Vmax of 310 mmol x 100 g(-1) x min(-1). Brain uptake of a Pgp substrate (colchicine) was significantly enhanced (two- to fourfold) in mdr1a(-/-) mice. These data suggest that the model is applicable to elucidating the effects of efflux transporters, including Pgp, on brain uptake.

The effect of omeprazole treatment on diazepam plasma levels was studied in four slow and six rapid metabolizers of omeprazole. Single intravenous doses of diazepam (0.1 mg/kg) were administered after 1 week of oral treatment with omeprazole (20 mg) and placebo. This was a double-blind crossover study with randomized placebo and omeprazole treatments. Blood was collected up to 120 hours after diazepam dosing (still during one-daily omeprazole and placebo administration) for measurement of diazepam and its major metabolite desmethyldiazepam. The slow metabolizers of omeprazole also metabolized diazepam slowly, exhibiting only half the diazepam plasma clearance of the others. The mean clearance of diazepam was decreased 26% after omeprazole in the rapid metabolizers, whereas the slow group showed no apparent interaction. The mean plasma concentrations of desmethyldiazepam showed a more rapid formation in the rapid compared with the slow metabolizers, which is a logical consequence of the rate of diazepam metabolism.

We studied the genetically determined hydroxylation polymorphism of S-mephenytoin in a Korean population (N = 206) and the pharmacokinetics of diazepam and demethyldiazepam after an oral 8 mg dose of diazepam administered to the nine extensive metabolizers and eight poor metabolizers recruited from the population. The log10 percentage of 4-hydroxymephenytoin excreted in the urine 8 hours after administration showed a bimodal distribution with an antimode of 0.3. The frequency of occurrence of the poor metabolizers was 12.6% in the population. In the panel study of diazepam in relation to the mephenytoin phenotype, there was a significant correlation between the oral clearance of diazepam and log10 urinary excretion of 4-hydroxymephenytoin (rs = 0.777, p less than 0.01). The plasma half-life of diazepam in the poor metabolizers was longer than that in the extensive metabolizers (mean +/- SEM, 91.0 +/- 5.6 and 59.7 +/- 5.4 hours, p less than 0.005), and the poor metabolizers had the lower clearance of diazepam than the extensive metabolizers (9.4 +/- 0.5 and 17.0 +/- 1.4 ml/min, p less than 0.001). In addition, the plasma half-life of demethyldiazepam showed a statistically significant (p less than 0.001) difference between the extensive metabolizers (95.9 +/- 11.3 hours) and poor metabolizers (213.1 +/- 10.7 hours), and correlated with the log10 urinary excretion of 4-hydroxymephenytoin (rs =-0.615, p less than 0.01). The findings indicate that the Korean subjects have a greater incidence of poor metabolizer phenotype of mephenytoin hydroxylation compared with that reported from white subjects and that the metabolism of diazepam and demethyldiazepam is related to the genetically determined mephenytoin hydroxylation polymorphism in Korean subjects.

Microsomal protein recovery and hepatocellularity have been determined and investigated as scaling factors for interrelating clearance by hepatic microsomes, freshly isolated hepatocytes and whole liver from untreated (UT) rats and rats treated with either the cytochrome P450 inducer phenobarbital (PB) or dexamethasone (DEX). Hepatocellularity in UT rats (1.1 x 10(8) hepatocytes/g liver) was not significantly different after either PB or DEX induction (1.1 and 1.3 x 10(8) hepatocytes/g liver, respectively). However the microsomal protein recovery index, which provides a scaling factor that is inversely related to the efficiency of the microsomal preparation procedure, was 47 mg/g liver in both PB and DEX microsomes and differs from UT rats (60 mg/g liver). These contrasting findings are consistent with the interlaboratory trends in the literature, indicating that, although hepatocellularity estimates are in good accord, microsomal recovery can vary 2-fold; this has implications for scaling. The oxidation of diazepam to its three primary metabolites was measured in PB and DEX microsomes and hepatocytes and the scaling factors were applied to these data and previously reported UT data. Marked changes in kinetics occur on induction resulting in a shift in the major pathway. In particular, 3-hydroxylation is induced over 20-fold by DEX. Diazepam CL(int) was determined in vivo after administration of a bolus dose into the hepatic portal vein of UT, PB, and DEX rats; values of 127, 191, and 323 ml/min/SRW (where SRW is a standard rat weight of 250 g), respectively, were obtained. Using these scaling factors, the hepatocyte predictions of CL(int) were excellent (99, 144, and 297 ml/min/SRW for UT, PB, and DEX, respectively), whereas only the DEX prediction (248 ml/min/SRW) was accurate for the microsomal system, with a substantial underprediction for UT and PB (46 and 68 ml/min/SRW, respectively). Evidence is presented for product inhibition, resulting from accumulation of primary metabolites within the microsomal preparation, as the mechanism responsible for this underprediction. These results illustrate that the scaling factor approach is applicable to induced livers in which both cytochrome P450 complement and zonal distribution are altered. These data, together with our previous studies, demonstrate that CL(int) in cells (2.4-297 ml/min/SRW), microsomes (2.7-248 ml/min/SRW), and in vivo (1.5-323 ml/min/SRW) are related in a linear fashion and hence inherently both in vitro systems are of equal value in predicting in vivo CL(int).

To determine the effect of fluoxetine on diazepam's pharmacokinetic and psychomotor responses, single oral doses of 10 mg diazepam were administered to six normal subjects on three occasions, either alone or in combination with 60 mg fluoxetine. Diazepam was given alone, after a single dose of fluoxetine, and after eight daily doses of fluoxetine. Psychometric data showed that fluoxetine had no significant effect on the psychomotor responses to diazepam. However, the pharmacokinetic data indicated a change in diazepam disposition after fluoxetine administration. Diazepam AUC was larger, the half-life was longer, and the plasma clearance was lower after fluoxetine administration, suggesting that fluoxetine inhibited the metabolism of diazepam. The reduced formation of an active metabolite, N-desmethyldiazepam, also suggested that fluoxetine inhibited diazepam's metabolism. The clinical implications of this pharmacokinetic drug-drug interaction are minor because psychomotor responses were unaffected and offsetting changes in the kinetics of diazepam and its metabolite occurred. Dosage modification of either fluoxetine or diazepam is unlikely to be necessary.