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The safety, tolerance, and pharmacokinetics of a small unilamellar liposomal formulation of amphotericin B (AmBisome) administered for empirical antifungal therapy were evaluated for 36 persistently febrile neutropenic adults receiving cancer chemotherapy and bone marrow transplantation. The protocol was an open-label, sequential-dose-escalation, multidose pharmacokinetic study which enrolled a total of 8 to 12 patients in each of the four dosage cohorts. Each cohort received daily doses of either 1.0, 2.5, 5.0, or 7.5 mg of amphotericin B in the form of AmBisome/kg of body weight. The study population consisted of patients between the ages of 13 and 80 years with neutropenia (absolute neutrophil count, 3 ) who were eligible to receive empirical antifungal therapy. Patients were monitored for safety and tolerance by frequent laboratory examinations and the monitoring of infusion-related reactions. Efficacy was assessed by monitoring for the development of invasive fungal infection. The pharmacokinetic parameters of AmBisome were measured as those of amphotericin B by high-performance liquid chromatography. Noncompartmental methods were used to calculate pharmacokinetic parameters. AmBisome administered as a 1-h infusion in this population was well tolerated and was seldom associated with infusion-related toxicity. Infusion-related side effects occurred in 15 (5%) of all 331 infusions, and only two patients (5%) required premedication. Serum creatinine, potassium, and magnesium levels were not significantly changed from baseline in any of the dosage cohorts, and there was no net increase in serum transaminase levels. AmBisome followed a nonlinear dosage relationship that was consistent with reticuloendothelial uptake and redistribution. There were no breakthrough fungal infections during empirical therapy with AmBisome. AmBisome administered to febrile neutropenic patients in this study was well tolerated, was seldom associated with infusion-related toxicity, was characterized by nonlinear saturation kinetics, and was effective in preventing breakthrough fungal infections.

Background: Tacrolimus is a potent immunosuppressant used in organ transplant recipients; an ointment formulation is being developed as a therapeutic agent for atopic dermatitis. Objective: Our purpose was to define the pharmacokinetics and evaluate tacrolimus 0.3% ointment as therapy for moderate to severe atopic dermatitis. Methods: Thirty-nine patients, 5 to 75 years of age, received 14 applications over 8 days. Serial blood samples were collected on days 1 and 8, with predose samples collected on days 2 through 7. Overall response and signs/symptoms were rated daily on days 1 through 11. Incidence of adverse events and laboratory profile were determined. Results: Mean area under the curve (0.9 to 42.5 ng ·hr/ml) was highly variable and appeared to be related to size of application area. No systemic accumulation of tacrolimus was observed. Comparison to historical intravenous data indicates that absolute bioavailability of topical tacrolimus was less than 0.5%. Ninety-five percent of patients showed at least good improvement. All adverse events were transient. Burning was the most common application site adverse event and vasodilatation ("flushing/warmth") was the most common nonapplication site adverse event. No drug-related changes in laboratory profile were observed. Conclusion: The results of this study suggest that tacrolimus 0.3% ointment may be a safe and effective therapy for atopic dermatitis. (J Am Acad Dermatol 1998;38:69-76.)

AmBisome (ABLP) is a unilamellar liposomal preparation of amphotericin B that has demonstrated an improved safety profile compared to conventional amphotericin B. Single- and multiple-dose pharmacokinetics were determined by using noncompartmental methods for rats administered ABLP at 1, 3, 9, and 20 mg/kg/day. The toxicological profile was evaluated following 30 consecutive days of intravenous ABLP administration. Mean plasma amphotericin B concentrations reached 500 and 380 μg/ml (males and females, respectively) following 30 days of ABLP administration at 20 mg/kg. The overall apparent half-life was 11.2 ± 4.5 h (males) or 8.7 ± 2.2 h (females), and the overall clearance (CL) was 9.4 ± 5.5 ml/h/kg (males) or 10.2 ± 4.1 ml/h/kg (females). ABLP appears to undergo saturable disposition, resulting in a non-dose-proportional amphotericin B area under the curve and a lower CL at higher doses. Histopathological examination revealed dose-related transitional-cell hyperplasia in the transitional epithelium of the urinary tract (kidney, ureters, and urinary bladder) and moderate hepatocellular necrosis at the 20-mg/kg/day dose. Administration of ABLP in doses of up to 20 mg/kg/day resulted in 100-fold higher plasma amphotericin B concentrations, with significantly lower toxicity than that reported with conventional amphotericin B therapy.

Epidural administration of clonidine, a partial alpha 2-adrenergic agonist, provides effective relief for patients with intractable cancer pain. However, clonidine's long plasma elimination half-life suggests a potential for plasma accumulation following repeated doses or constant infusion. Adult male and female cancer patients experiencing severe, intractable pain were administered a continuous epidural infusion (30 micrograms/h) of clonidine hydrochloride for 14 days. Plasma clonidine concentrations were determined for 31 patients using a radioimmunoassay with a limit of quantitation of 0.062 ng/mL. Mean (+/- SD) plasma clonidine concentration and calculated total body clearance after 7 days of infusion (respectively, 2.19 +/- 1.17 ng/mL, n = 24; 279 +/- 184 mL/min, n = 27) were comparable to those following 14 days of infusion (2.50 +/- 1.51 ng/mL, n = 19; 272 +/- 163 mL/min, n = 21). Clonidine does not appear to accumulate in the plasma compartment during prolonged (14-day) continuous epidural infusion in cancer patients.

Objective: We assessed rates of withdrawal-related events following abrupt discontinuation and gradual tapering of clobazam during Phase I–III studies. Background Clobazam, a 1,5-benzodiazepine, is under FDA review for LGS. Design/Methods: Potential withdrawal-related AEs following abrupt CLB discontinuation were assessed in 4 multiple-dosage Phase I trials (dosing 8−34 days). These studies employed therapeutic (20 and 40 mg/day) and supratherapeutic dosages (120 and 160 mg/day). These events were also assessed in Phase II/III studies (dosing ≤18 weeks) and their open-label extension (OLE) (≤5 years). In these studies, gradual tapering over 2–3 weeks was conducted. Withdrawal-related AEs included sedation, somnolence, insomnia, irritability, anxiety, and seizures. AEs must have occurred ≥1 day following last clobazam dose and within 30 days after last dose, or must have been deemed withdrawal symptoms by investigators. Results: Phase I participants received steady-state dosages of 20–160 mg/day, with the vast majority receiving ≥40 mg/day and ∼50% receiving ≥120 mg/day (≤4-times maximum daily dosage in Phase II/III). In contrast, LGS patients received ≤40 mg/day during Phase II/III and ≤80 mg/day in the OLE. 87 patients discontinued clobazam during Phase II/III and tapered drug. None reported withdrawal-related AEs. Abrupt discontinuation led to AEs consistent with withdrawal. 93 AEs were reported for 68 of 207 in Phase I studies. Nearly all AEs occurred after abrupt discontinuation of clobazam ≥40 mg/day, with ∼50% occurring at ≥120 mg/day. The majority of AEs following abrupt discontinuation were mild, and included headache, insomnia, anxiety, and tremor. No status epilepticus occurred during withdrawal/tapering. Conclusions: Withdrawal-related AEs did not occur during 2- to 3-week tapering at the conclusion of Phase II/III studies of clobazam. No withdrawal-related AEs have been observed with long-term therapy through 5 years In Phase I, abrupt withdrawal led to generally mild AEs at rates comparable to those observed for other benzodiazepines. Supported by: Lundbeck Inc. Disclosure: Dr. Tolbert has received personal compensation for activities with Lundbeck Research USA, Inc as an employee. Dr. Harris has received research support from Lundbeck Research USA, Inc. Dr. Bekersky has received personal compensation for activities with Lundbeck Research USA, Inc. as an employee. Dr. Owen has received personal compensation for activities with Lundbeck Research USA, Inc.

A sensitive and selective high-performance liquid chromatographic (HPLC) method was developed for the determination of 1,3-di(4-imidazolino-2-methoxyphenoxy)propane (DMP) in rat, dog and human plasma (50-5000 ng/ml) and urine (0.1-10 micrograms/ml). DMP and DMPent (dimethoxyimidizolinopentamidine, the internal standard), are extracted from alkanized plasma with n-butyl chloride-n-butanol (9:1, v/v). The organic phase is dried under nitrogen, reconstituted in mobile phase, and washed with hexane. Separation is achieved by ion-pair chromatography on a Zorbax Rx C8 column with fluorescence detection. The analysis of pooled plasma (80, 400, and 4000 ng/ml) and urine controls (0.3, 1.6, and 8 micrograms/ml) demonstrated excellent precision and accuracy over a three-day period. The recovery of DMP is90% from rat, dog, and human plasma and85% from rat and human urine, and 60-70% from dog urine. The limit of quantitation (LOQ) of the assay is 50 ng/ml in rat, dog and human plasma. Using the high-sensitivity assay, the limit of quantitation was decreased to 5, 2 and 0.6 ng/ml in rat, dog and human plasma, respectively. The LOQ of the assay is 0.1 microgram/ml in rat, dog and human urine. The assay was used to determine plasma and urine concentrations of DMP in pharmacokinetic studies in rat and dog.

(1979). Influence of Gut Microflora on Bioavailability. Drug Metabolism Reviews: Vol. 9, No. 2, pp. 259-279.

Six adult female subjects who use saccharin-containing products in their diet were asked to take divided equal doses of saccharin every 6 hr to maintain their average daily intake for 3 days. At the end of this period, each subject took a single dose that was equal to one divided dose. Saccharin concentrations in plasma and urine samples were used to assess the kinetic profile. Saccharin absorption was rapid with maximum concentrations in plasma in 0.5 to 1.0 hr. Maximum plasma concentrations and areas under the plasma concentration-time curves were proportional to dose. Renal clearance exceeded glomerular filtration rate in all cases and approximated renal plasma flow when corrected for the saccharin free fraction in plasma. Mean elimination half-life was 7.5 hr and mean apparent volume of distribution was 264 l. The kinetic parameters indicate that saccharin is distributed as a function of lean rather than total body mass (suggesting that saccharin does not distribute into body fat). This observation, together with data from studies in animals, suggests that there may be one or more high-retention compartments for saccharin. Clinical Pharmacology and Therapeutics (1981) 30, 558–563; doi:10.1038/clpt.1981.203

A rapid, sensitive and selective high-performance liquid chromatographic (HPLC) assay was developed for the determination of (E)-1,2,3,4-tetrahydro-1,1,4,4-tetramethyl-6-(1-methyl-2- phenylethenyl)naphthalene (I) and its phenolic metabolite, (E)-4-[2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)-2- methylethenyl]phenol (II) in plasma. The assay for both compounds involves precipitation of the plasma proteins with acetonitrile, followed by extraction of the entire mixture into methyl tert.-butyl ether and subsequent analysis by reversed-phase HPLC. The overall recovery of I and II was 96.9 +/- 5.3 and 96.2 +/- 5.8% for dog plasma, 80.0 +/- 4.0 and 93.5 +/- 5.0% for human plasma and 97.4 +/- 2.9 and 97.6 +/- 9.6% for rat plasma, respectively. The sensitivity limit is 20 ng/ml of plasma for both compounds I and II using UV detection at 280 nm. The HPLC assay was used in studies in the dog and in the rat.

(1983). Use of the Isolated Perfused Kidney as a Tool in Drug Disposition Studies. Drug Metabolism Reviews: Vol. 14, No. 5, pp. 931-960.

An electron-capture gas-liquid chromatographic assay was developed for the determination of clonazepam (as its methyl derivative) and of flunitrazepam and its N-desmethyl metabolite in blood. The method was used to measure blood levels in man following single oral 2-mg doses of clonazepam and flunitrazepam, and has a sensitivity limit of 0.5 to 1.0 ng of compound per milliliter of blood.

The pharmacokinetics of saccharin were studied in two human subjects. One male and one female subject ingested a single 100 mg oral dose of saccharin. Absorption was fast and the elimination half-lives were 1.2 and 6.6 hours for the male and female subject, respectively. Approximately 85% of the oral dose was recovered in the urine as intact saccharin and renal clearance approximated renal plasma flow when corrected for the unbound fraction of saccharin in plasma. Saccharin elimination appears to be related to previous saccharin ingestion, suggesting a high-retention compartment.

A randomized, four-way cross-over study was conducted in eight healthy male volunteers to determine the relative and absolute bioavailability of prednisone (PN) and prednisolone (PL). PN and PL were administered as single, oral 10-mg tablet doses and as 10-mg zero-order 0.5-hour intravenous infusions. Comparable mean PN and PL maximum plasma concentrations (Cmax), times for Cmax, areas under the plasma concentration-time curves (AUC), and apparent elimination rate constants between tablet treatments demonstrated that PN and PL tablets were bioequivalent. Absolute bioavailability (F) determinations based on plasma PL concentrations were independent of which IV treatment was used as reference and indicated complete systemic availability of PL from both PN and PL tablets. However, F based on plasma PN data was contradictory. Using IV PN as reference, approximately 70% systemic availability was observed from both tablets, whereas using IV PL as reference, systemic availability was greater than unity. PN and PL are model compounds that exemplify the difficulties involved in accurately determining the relative and absolute bioavailability of substances that undergo reversible metabolism.

A sensitive, rapid, and specific high pressure liquid chromatographic (HPLC) assay was developed for the determination of salicylic (SA) and salicyluric (SU) acids in plasma and urine. The compounds are extracted into ethyl ether at acid pH, evaporated, and reconstituted prior to instrumental separation. Overall recovery of both compounds is 90 ± 5%, and the sensitivity limits are 150 ng of SU and 300 ng SA per ml of biological fluid. The assay was used for the determination of both compounds in plasma and urine of man following oral doses of 40 mg/kg of sodium salicylate.

The spectrofluorodensitometric assay for the determination of flurazepam and its major metabolites in blood involves selective extraction of flurazepam and its major metabolites into ether and hydrolysis in acid to their respective benzophenones, which are cyclized in dimethylformamide-potassium carbonate to their respective 9-acridanone derivatives and separated by TLC. The fluorescence of the 9-acridanones is measured directly on the chromatoplate, using a scanning TLC analyzer in the reflectance mode. The limits of detection of the assay are 0.5-1.0 ng of each compound/ml of blood using a 4-ml specimen/analysis.

The isolated perfused rat kidney was used to study the pharmacokinetic-pharmacodynamic relation of bumetanide and furosemide. Diuresis, as indicated by the concomitant increase in urine volume and fractional excretion of sodium, was produced with both drugs. The action of furosemide was dependent on a high clearance resulting from combined glomerular filtration and tubular secretion. The site of furosemide action was the luminal side of the nephron and a large amount of drug was required in the tubular lumen to produce diuresis. A bidirectional transport of bumetanide was indicated. Although tubular secretion of bumetanide was demonstrated, the action of bumetanide was not dependent on secretion and the highest response was achieved when bumetanide was filtered and partially reabsorbed. The lack of dependency on secretion to produce a response may be indicative of bumetanide reaching its site of action from the luminal as well as the vascular side of the nephron. In the isolated perfused rat kidney, although both drugs had the same pharmacodynamic endpoint, each drug had a different pharmacokinetic profile that characterized its response.

A high-pressure liquid chromatographic method was developed to measure nonradioactive iothalamate in serum and urine for use in estimating glomerular filtration rate (GFR). This method was used to study the renal handling of cibenzoline, an experimental antiarrhythmic drug. The mean cibenzoline clearance was 3.5 +/- 2.5 (SD) times the glomerular filtration rate. The clearance of non-protein-bound cibenzoline was seven times GFR, indicating excretion by the renal tubular secretory pathway for organic bases. This drug, at the doses used, did not lower creatinine clearance, indicating that the effect of basic drugs competing with creatinine for the base secretory pathway appears to be dose and drug dependent.

The production of bile salts by rat liver is regulated by a feedback mechanism, but it is not known which enzyme controls endogenous bile acid synthesis. In order to demonstrate the biochemical site of this control mechanism, bile fistula rats were infused intravenously with (14)C-labeled bile acid precursors, and bile acid biosynthesis was inhibited as required by intraduodenal infusion of sodium taurocholate. The infusion of taurocholate (11-14 mg/100 g of rat per hr) inhibited the incorporation of acetate-1-(14)C, mevalonolactone-2-(14)C, and cholesterol-4-(14)C into bile acids by approximately 90%. In contrast, the incorporation of 7alpha-hydroxycholesterol-4-(14)C into bile acids was reduced by less than 10% during taurocholate infusion. These results indicate that the regulation of bile acid biosynthesis is exerted via cholesterol 7alpha-hydroxylase provided that hepatic cholesterol synthesis is adequate.

An improved method for the chemical synthesis of HMG-CoA is described. The procedure is designed to prevent the formation of 3-acetoxy-HMG-CoA.

Isoniazid was administered orally to two slow acetylator subjects, and plasma and saliva samples were assayed for intact drug. Saliva/plasma concentration ratios tended to be scattered about the value of 1. The results indicate that salivary level measurements may provide a useful approach for acetylation phenotyping. However, because of instability of isoniazid in frozen specimens, analyses of drug levels had to be initiated within 1 hr from the time of collection. Protein binding studies indicated that INH is not bound to plasma proteins to any appreciable extent.

This report presents data to demonstrate the clinical utility of a previously published electron-capture GLC assay in the measurement of blood levels of 7-chloro-1,3-dihydro-5-(2'-chlorophenyl)-2 H -1,4-benzodiazepin-2-one in humans following a single oral 4-mg dose. It also describes a differential pulse polarographic assay for the quantitation of its major urinary metabolite, lorazepam, in humans. This assay has greater sensitivity and specificity than the previously published Bratton–Marshall spectrophotometric assay.

A rapid, sensitive and selective high-performance liquid chromatographic (HPLC) assay was developed for the determination of 6-(2-chlorophenyl)-4-hydroxy-4H-imidazo[1,5-a]-[1,4]benzodiazepine- 3-carboxamide (I) and its benzophenone carboxylic acid metabolite, 1-[2-(2-chlorobenzoyl)phenyl]-4-(aminocarbonyl)-1H-imidazole-5-car boxylic acid (II) in plasma. The assay involves the extraction of both compounds into benzene from buffered plasma (pH 5.4) and subsequent analysis by reversed-phase HPLC. The overall recovery of I and II is 98.3 +/- 9.4 and 59.7 +/- 15.7% for dog plasma, 86.0 +/- 14.7 and 52.8 +/- 15.1% for rat plasma and 98.1 +/- 9.3 and 66.9 +/- 18.0% for human plasma, respectively. The sensitivity limit of the assay for I and II is 20.0 and 40.0 ng/ml of plasma using ultraviolet detection at 254 nm. The assay was used in studies in dog and rat.

Clonazepam (1 mg/kg) and antipyrine (0.1 mg/kg) were administered simultaneously by intravenous bolus injection to three dogs. After 2 weeks of chronic phenobarbital administration, the studies were repeated. Plasma concentration-time curves in all studies were biexponential. Phenobarbital administration increased total plasma clearances of clonazepam and antipyrine by 102% and 98.5%, respectively. Volumes of distribution were not altered, and consequently reductions in terminal exponential half-lives observed after phenobarbital were attributed to increases in clearances.

A sensitive and specific electron-capture GLC assay was developed for the determination of bromazepam (I), 7-bromo-1,3-dihydro-5-(2-pyridyl)-2H-1,4-benzodiazepin-2-one, in blood. The overall recovery of I from blood is 85 ± 5.0%, and the sensitivity limit of detection is 5–10 ng I/ml blood. In the determination of the urinary excretion, the major urinary metabolites, 3-hydroxybromazepam (II) and 2-amino-3-hydroxy-5-bromobenzoylpyridine (V), are selectively extracted into ether from urine buffered to pH 9.0 after incubation with glucuronidase—sulfatase, whereas I and 2-amino-5-bromobenzoylpyridine (IV) are directly extractable from urine into ether at pH 9.0. The residues of the respective ether extracts are dissolved in 1.0 M phosphate buffer (pH 5.5) and analyzed by differential pulse polarography, which yields two distinct peaks for the benzodiazepin-2-one and the benzoylpyridine component in each fraction. The overall recovery of I and IV is 80 ± 5.0% with sensitivity limits of 100 and 50 ng/5 ml urine, respectively; the recovery of II and V is about 45 ± 5.0% with sensitivity limits of 100 ng each/5 ml urine analyzed. Blood levels of bromazepam and the urinary excretion of the unchanged drug and its major metabolites were determined in humans following the administration of single 12-mg oral doses.

This chapter deals largely with the action of thyroid hormones on bile acid metabolism, because few definitive studies with other hormones (e.g., pituitary, adrenal, gonadal) have been carried out so far.

The hepatic biosynthesis of bile salts in the rat has been shown to be controlled homeostatically by the quantity of bile salt returning to the liver via the portal circulation. The feedback mechanism was demonstrated in two kinds of experiments. In the first, rats with bile fistulas were infused intraduodenally with sodium taurocholate 12 hr after surgery. If the rate of infusion was greater than 10 mg per 100 g rat per hr, the increase in bile acid output normally observed in bile fistula rats was prevented. In the second type of experiment, the rats were infused with taurocholate 48-72 hr after biliary diversion, when bile acid output had reached a maximal value. Provided the rate of infusion exceeded 10 mg per 100 g rat per hr, bile acid secretion returned to the low levels observed in intact rats. Previous attempts to demonstrate the feedback control have been unsuccessful because too little bile salt was infused. The taurocholate pool of the experimental animals was measured as approximately 15 mg per 100 g rat; it was calculated from this and the above results that this pool circulated 10-13 times daily.

The tolerance and pharmacokinetics (PK) of tacrolimus (T) by the addition of mycophenolate mofetil (MMF) in stable kidney transplant patients (6/group) on long-term tacrolimus-based therapy were investigated. Patients received combination T and MMF therapy at three MMF doses: 1, 1.5, and 2 g/day administered twice daily. A 12-hour blood PK profile for T was obtained prior to MMF dosing; concomitant 12-hour profiles for T, mycophenolic acid (MPA), and mycophenolic acid glucuronide (MPAG) were obtained after 2 weeks of administration. Tolerance was monitored through 3 months. The intra- and intergroup PK of T were variable. The mean AUC0-12 of T for each group was increased after 2 weeks of concomitant MMF administration, but the increase was not statistically significant. Both drugs were well tolerated. Gastrointestinal events were of interest as such have been attributed to both T and MMF. Events reported were diarrhea, nausea, dyspepsia, and vomiting. Other common adverse events were headache, hypomagnesemia, and tremors. Most were mild, although a few were considered to be moderate. There was no apparent relationship between the incidence of any adverse event and MMF treatment group. In the present study, the coadministration of T and MMF did not significantly alter T pharmacokinetics.