Evan D. Kharasch

Pharmacokinetics and pharmacodynamics of oral oxycodone in healthy human subjects: Role of circulating active metabolites

In vitro experiments suggest that circulating metabolites of oxycodone are opioid receptor agonists. Clinical and animal studies to date have failed to demonstrate a significant contribution of the O‐demethylated metabolite oxymorphone toward the clinical effects of the parent drug, but the role of other putative circulating active metabolites in oxycodone pharmacodynamics remains to be examined.

Identification of Cytochrome P450 2E1 as the Predominant Enzyme Catalyzing Human Liver Microsomal Defluorination of Sevoflurane, Isoflurane, and Methoxyflurane

BackgroundRenal and hepatic toxicity of the fluorinated ether volatile anesthetics is caused by biotransformation to toxic metabolites. Metabolism also contributes significantly to the elimination pharmacokinetics of some volatile agents. Although innumerable studies have explored anesthetic metabolism in animals, there is little information on human volatile anesthetic metabolism with respect to comparative rates or the identity of the enzymes responsible for defluorination. The first purpose of this investigation was to compare the metabolism of the fluorinated ether anesthetics by human liver microsomes. The second purpose was to test the hypothesis that cytochrome P450 2E1 is the specific P450 isoform responsible for volatile anesthetic defluorination in humans. MethodsMicrosomes were prepared from human livers. Anesthetic metabolism in microsomal incubations was measured by fluoride production. The strategy for evaluating the role of P450 2E1 in anesthetic defluorination involved three approaches: for a series of 12 human livers, correlation of microsomal defluorination rate with microsomal P450 2E1 content (measured by Western blot analysis), correlation of defluorination rate with microsomal P450 2E1 catalytic activity using marker substrates (paranitrophenol hydroxylation and chlorzoxazone 6-hydroxylation), and chemical inhibition by P450 isoform-selective Inhibitors. ResultsThe rank order of anesthetic metabolism, assessed by fluoride production at saturating substrate concentrations, was methoxyflurane > sevoflurane > enflurane > isoflurane > desflurane > O. There was a significant linear correlation of sevoflurane and methoxyflurane defluorination with antigenic P450 2E1 content (r = 0.98 and r = 0.72, respectively), but not with either P450 1A2 or P450 3A3/4. Comparison of anesthetic defluorination with either paranitrophenol or chlorzoxazone hydroxylation showed a significant correlation for sevoflurane (r = 0.93, r = 0.95) and methoxyflurane (r = 0.78, r = 0.66). Sevoflurane defluorination was also highly correlated with that of enflurane (r = 0.93), which is known to be metabolized by human P450 2E1. Diethyldithiocarbamate, a selective inhibitor of P450 2E1, produced a concentration-dependent inhibition of sevoflurane, methoxyflurane, and isoflurane defluorination. No other isoform-selective inhibitor diminished the defluorination of sevoflurane, whereas methoxyflurane defluorination was inhibited by the selective P450 inhibitors furafylline (P45O 1A2), sulfaphenazole (P450 2C9/10), and quinidine (P450 2D6) but to a much lesser extent than by diethyldithiocarbamate. ConclusionsThese results demonstrate that cytochrome P450 2E1 is the principal, if not sole human liver microsomal enzyme catalyzing the defluorination of sevoflurane. P450 2E1 is the principal, but not exclusive enzyme responsible for the metabolism of methoxyflurane, which also appears to be catalyzed by P450s 1A2, 2C9/10, and 2D6. The data also suggest that P450 2E1 is responsible for a significant fraction of isoflurane metabolism. Identification of P450 2E1 as the major anesthetic metabolizing enzyme in humans provides a mechanistic understanding of clinical fluorinated ether anesthetic metabolism and toxicity.

Contribution of CYP2E1 and CYP3A to acetaminophen reactive metabolite formation

CYP2E1, 1A2, and 3A4 have all been implicated in the formation of N‐acetyl‐p‐benzoquinone imine (NAPQI), the reactive intermediate of acetaminophen (INN, paracetamol), in studies in human liver microsomes and complementary deoxyribonucleic acid–expressed enzymes. However, recent pharmacokinetic evidence in humans has shown that the involvement of CYP1A2 is negligible in vivo. The purpose of this study was to evaluate the respective roles of CYP2E1 and 3A4 in vivo.

Psychedelic effects of ketamine in healthy volunteers. Relationship to steady-state plasma concentrations

Background Ketamine has been associated with a unique spectrum of subjective “psychedelic” effects in patients emerging from anesthesia. This study quantified these effects of ketamine and related them to steady‐state plasma concentrations. Methods Ketamine or saline was administered in a single‐blinded crossover protocol to 10 psychiatrically healthy volunteers using computer‐assisted continuous infusion. A stepwise series of target plasma concentrations, 0, 50, 100, 150, and 200 ng/ml were maintained for 30 min each. After 20 min at each step, the volunteers completed a visual analog (VAS) rating of 13 symptom scales. Peripheral venous plasma ketamine concentrations were determined after 28 min at each step. One hour after discontinuation of the infusion, a psychological inventory, the hallucinogen rating scale, was completed. Results The relation of mean ketamine plasma concentrations to the target concentrations was highly linear, with a correlation coefficient of R = 0.997 (P = 0.0027). Ketamine produced dose‐related psychedelic effects. The relation between steady‐state ketamine plasma concentration and VAS scores was highly linear for all VAS items, with linear regression coefficients ranging from R = 0.93 to 0.99 (P < 0.024 to P <0.0005). Hallucinogen rating scale scores were similar to those found in a previous study with psychedelic doses of N,N‐dimethyltryptamine, an illicit LSD‐25‐like drug. Conclusions Subanesthetic doses of ketamine produce psychedelic effects in healthy volunteers. The relation between steady‐state venous plasma ketamine concentrations and effects is highly linear between 50 and 200 ng/ml.

Role of hepatic and intestinal cytochrome P450 3A and 2B6 in the metabolism, disposition, and miotic effects of methadone.

The disposition of the long‐acting opioid methadone, used to prevent opiate withdrawal and treat short‐ and long‐lasting pain, is highly variable. Methadone undergoes N‐demethylation to the primary metabolite 2‐ethyl‐1,5‐dimethyl‐3,3‐diphenylpyrrolinium (EDDP), catalyzed in vitro by intestinal, hepatic, and expressed cytochrome P450 (CYP) 3A4. However, the role of CYP3A4 in human methadone disposition in vivo is unclear. This investigation tested the hypothesis that CYP3A induction (or inhibition) would increase (or decrease) methadone metabolism and clearance in humans.

Human Kidney Methoxyflurane and Sevoflurane Metabolism Intrarenal Fluoride Production as a Possible Mechanism of Methoxyflurane Nephrotoxicity

Background Methoxyflurane nephrotoxicity is mediated by cytochrome P450‐catalyzed metabolism to toxic metabolites. It is historically accepted that one of the metabolites, fluoride, is the nephrotoxin, and that methoxyflurane nephrotoxicity is caused by plasma fluoride concentrations in excess of 50 micro Meter. Sevoflurane also is metabolized to fluoride ion, and plasma concentrations may exceed 50 micro Meter, yet sevoflurane nephrotoxicity has not been observed. It is possible that in situ renal metabolism of methoxyflurane, rather than hepatic metabolism, is a critical event leading to nephrotoxicity. We tested whether there was a metabolic basis for this hypothesis by examining the relative rates of methoxyflurane and sevoflurane defluorination by human kidney microsomes. Methods Microsomes and cytosol were prepared from kidneys of organ donors. Methoxyflurane and sevoflurane metabolism were measured with a fluoride‐selective electrode. Human cytochrome P450 isoforms contributing to renal anesthetic metabolism were identified by using isoform‐selective inhibitors and by Western blot analysis of renal P450s in conjunction with metabolism by individual P450s expressed from a human hepatic complementary deoxyribonucleic acid library. Results Sevoflurane and methoxyflurane did undergo defluorination by human kidney microsomes. Fluoride production was dependent on time, reduced nicotinamide adenine dinucleotide phosphate, protein concentration, and anesthetic concentration. In seven human kidneys studied, enzymatic sevoflurane defluorination was minimal, whereas methoxyflurane defluorination rates were substantially greater and exhibited large interindividual variability. Kidney cytosol did not catalyze anesthetic defluorination. Chemical inhibitors of the P450 isoforms 2E1, 2A6, and 3A diminished methoxyflurane and sevoflurane defluorination. Complementary deoxyribonucleic acid‐expressed P450s 2E1, 2A6, and 3A4 catalyzed methoxyflurane and sevoflurane metabolism, in diminishing order of activity. These three P450s catalyzed the defluorination of methoxyflurane three to ten times faster than they did that of sevoflurane. Expressed P450 2B6 also catalyzed methoxyflurane defluorination, but 2B6 appeared not to contribute to renal microsomal methoxyflurane defluorination because the P450 2B6‐selective inhibitor had no effect. Conclusions Human kidney microsomes metabolize methoxyflurane, and to a much lesser extent sevoflurane, to fluoride ion. P450s 2E1 and/or 2A6 and P450 3A are implicated in the defluorination. If intrarenally generated fluoride or other metabolites are nephrotoxic, then renal metabolism may contribute to methoxyflurane nephrotoxicity. The relative paucity of renal sevoflurane defluorination may explain the absence of clinical sevoflurane nephrotoxicity to date, despite plasma fluoride concentrations that may exceed 50 micro Meter.

Bispectral Index monitoring during sedation with sevoflurane, midazolam, and propofol

Background Bispectral Index (BIS) has been used to measure sedation depth. Ideally, to guide anesthetic management, range of BIS scores at different sedation levels should not overlap, and BIS should be independent of drug used. This study assessed ability of BIS to predict sedation depth between sevoflurane, propofol, and midazolam. Quality of recovery was also compared. Methods Patients undergoing surgery with local or regional anesthesia and sedation were randomized to sevoflurane (n = 23), midazolam (n = 21), or propofol (n = 22). Sedation was titrated to Observers’s Assessment of Alertness–Sedation score of 3 (responds slowly to voice). BIS and Observers’s Assessment of Alertness–Sedation were measured every 5 min. BIS prediction probability (PK) was compared between drugs. Recovery was assessed by BIS and Digit Symbol Substitution and memory tests. Results Bispectral Index of responders to voice was significantly different from nonresponders (86 ± 10 vs. 74 ± 14, mean ± SD;P < 0.001) However, wide variability and overlap in BIS were observed (25th–75th percentile, responders vs. non-responders: 79–96 vs. 65–83). BIS of responders was different for sevoflurane versus propofol and midazolam. BIS was a better predictor of propofol sedation than sevoflurane or midazolam (PK = 0.87 ± 0.11, 0.76 ± 0.01, and 0.69 ± 0.02, respectively;P < 0.05). At 10 min after the procedure, 76, 48, and 24% of sevoflurane, propofol, midazolam patients, respectively, returned to baseline Digit Symbol Substitution scores (P < 0.05). Excitement–disinhibition occurred in 70, 36, and 5% of sevoflurane, propofol, and midazolam patients, respectively (P < 0.05). Conclusion Individual BIS scores demonstrate significant variability, making it difficult to predict sedation depth. The relation between BIS and sedation depth may not be independent of anesthetic agent. Quality of recovery was similar between drugs, but excitement occurred frequently with sevoflurane.

Gender differences in drug effects: implications for anesthesiologists

Background:  The gender aspect in pharmacokinetics and pharmacodynamics of anesthetics has attracted little attention. Knowledge of previous work is required to decide if gender‐based differences in clinical practice is justified, and to determine the need for research.

assessment of Low-flow Sevoflurane and Isoflurane Effects on Renal Function Using Sensitive Markers of Tubular Toxicity

Background: Carbon dioxide absorbents degrade sevoflurane, particularly at low gas flow rates, to fluoromethyl‐2,2‐difluoro‐1‐(trifluoromethyl)vinyl ether (compound A). Compound A causes renal proximal tubular injury in rats but has had no effect on blood urea nitrogen (BUN) or creatinine concentrations in patients. This investigation compared the effects of low‐flow sevoflurane and isoflurane on renal tubular function in surgical patients using conventional (BUN and creatinine) and finer indices of renal injury, specifically those biomarkers sensitive for compound A toxicity in rats (glucosuria, proteinuria, and enzymuria [N‐acetyl‐beta‐D‐glucosaminidase (NAG) and alpha‐glutathione‐S‐transferase (alpha GST)]). Methods: Consenting patients with normal preoperative renal function at two institutions were randomized to receive sevoflurane (n = 36) or isoflurane (n = 37) in oxygen and air. Total gas flow was 1 l/min, opioid doses were minimized, and barium hydroxide lime was used to maximize anesthetic degradation. Inspiratory and expiratory compound A concentrations were quantified every 30–60 min. Blood and urine were obtained before and 24–72 h after anesthesia for laboratory evaluation. Results: Sevoflurane and isoflurane groups were similar with respect to age, weight, sex, American Society of Anesthesiologists status, anesthetic duration (3.7 or 3.9 h), and anesthetic exposure (3.6 or 3 minimum alveolar concentration [MAC]‐hour). Maximum inspired compound A concentration (mean +/‐ standard deviation) was 27 +/‐ 13 ppm (range, 10–67 ppm). Areas under the inspired and expired compound A concentration versus time curves (AUC) were 79 +/‐ 54‐ppm‐h (range, 10–223 ppm‐h) and 53 +/‐ 40 ppm‐h (range, 6–159 ppm‐h), respectively. There was no significant difference between anesthetic groups in postoperative serum creatinine or BUN, or urinary excretion of protein, glucose, NAG, proximal tubular alpha GST, or distal tubular pi GST. There was no significant correlation between compound A exposure (AUC) and protein, glucose, NAG, alpha GST, or pi GST excretion. Postoperative alanine and aspartate aminotransferase concentrations were not different between the anesthetic groups, and there were no significant correlations between compound A exposure and alanine or aspartate aminotransferase concentrations. Conclusions: The renal tubular and hepatic effects of low‐flow sevoflurane and isoflurane were similar as assessed using both conventional measures of hepatic and renal function and more sensitive biochemical markers of renal tubular cell necrosis. Moderate duration low‐flow sevoflurane anesthesia, during which compound A formation occurs, appears to be as safe as low‐flow isoflurane anesthesia.

Role of P‐glycoprotein in the intestinal absorption and clinical effects of morphine

There is considerable and unexplained individual variability in the morphine dose‐effect relationship. The efflux pump P‐glycoprotein regulates brain access and intestinal absorption of numerous drugs. Morphine is a P‐glycoprotein substrate in vitro, and P‐glycoprotein affects morphine brain access and pharmacodynamics in animals. However, the role of P‐glycoprotein in human morphine disposition and clinical effects is unknown. This investigation tested the hypothesis that plasma concentrations and clinical effects of oral and intravenous morphine are greater after inhibition of intestinal and brain P‐glycoprotein, with the P‐glycoprotein inhibitor quinidine used as an in vivo probe.