Protein tyrosine phosphatase 1B modulates GSK3ß/Nrf2 and IGFIR signaling pathways in acetaminophen-induced hepatotoxicity.

Acute hepatic failure secondary to acetaminophen (APAP) poisoning is associated with high mortality. Protein tyrosine phosphatase 1B (PTP1B) is a negative regulator of tyrosine kinase growth factor signaling. In the liver, this pathway confers protection against injury. However, the involvement of PTP1B in the intracellular networks activated by APAP is unknown. We have assessed PTP1B expression in APAP-induced liver failure in humans and its role in the molecular mechanisms that regulate the balance between cell death and survival in human and mouse hepatocytes, as well as in a mouse model of APAP-induced hepatotoxicity. PTP1B expression was increased in human liver tissue removed during liver transplant from patients for APAP overdose. PTP1B was upregulated by APAP in primary human and mouse hepatocytes together with the activation of c-jun (NH2) terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38 MAPK), resulting in cell death. Conversely, Akt phosphorylation and the antiapoptotic Bcl2 family members BclxL and Mcl1 were decreased. PTP1B deficiency in mouse protects hepatocytes against APAP-induced cell death, preventing glutathione depletion, reactive oxygen species (ROS) generation and activation of JNK and p38 MAPK. APAP-treated PTP1B(-/-) hepatocytes showed enhanced antioxidant defense through the glycogen synthase kinase 3 (GSK3)ß/Src kinase family (SKF) axis, delaying tyrosine phosphorylation of the transcription factor nuclear factor-erythroid 2-related factor (Nrf2) and its nuclear exclusion, ubiquitination and degradation. Insulin-like growth factor-I receptor-mediated signaling decreased in APAP-treated wild-type hepatocytes, but was maintained in PTP1B(-/-) cells or in wild-type hepatocytes with reduced PTP1B levels by RNA interference. Likewise, both signaling cascades were modulated in mice, resulting in less severe APAP hepatotoxicity in PTP1B(-/-) mice. Our results demonstrated that PTP1B is a central player of the mechanisms triggered by APAP in hepatotoxicity, suggesting a novel therapeutic target against APAP-induced liver failure.

Acetaminophen-associated hepatic injury: evaluation of acetaminophen protein adducts in children and adolescents with acetaminophen overdose.

Acetaminophen protein adducts (APAP adducts) were quantified in 157 adolescents and children presenting at eight pediatric hospitals with the chief complaint of APAP overdose. Two of the patients required liver transplantation, whereas all the others recovered spontaneously. Peak APAP adducts correlated with peak hepatic transaminase values, time-to-treatment with N-acetylcysteine (NAC), and risk determination per the Rumack-Matthews nomogram. A population pharmacokinetic analysis (NONMEM) was performed with post hoc empiric Bayesian estimates determined for the elimination rate constants (k(e)), elimination half-lives (t(1/2)), and maximum concentration of adducts (C(max)) of the subjects. The mean (+/-SD)k(e) and half-life were 0.486 +/- 0.084 days(-1) and 1.47+/- 0.30 days, respectively, and the C(max) was 1.2 (+/-2.92) nmol/ml serum. The model-derived, predicted adduct value at 48 h (Adduct 48) correlated with adductC(max), adduct T(max), Rumack-Matthews risk determination, peak aspartate aminotransferase (AST), and peak alanine aminotransferase (ALT). The pharmacokinetics and clinical correlates of APAP adducts in pediatric and adolescent patients with APAP overdose support the need for a further examination of the role of APAP adducts as clinically relevant and specific biomarkers of APAP toxicity.

Intermediate syndrome after malathion ingestion despite continuous infusion of pralidoxime.

CASE REPORT: A 33-year-old female ingested an unknown quantity of malathion in a suicide attempt. Cholinergic signs consistent with severe organ, phosphate intoxication developed and were treated within 6 hours of ingestion. Intravenous atropine and a continuous infusion of pralidoxime (400 mg/h) were administered. Prolonged depression of plasma and red blood cell cholinesterases were documented. Despite an initial clinical improvement and the presence of plasma pralidoxime concentrations exceeding 4 microg/mL, the patient developed profound motor paralysis consistent with the diagnosis of Intermediate Syndrome. In addition to the dose and frequency of pralidoxime administration, other factors including persistence of organophosphate in the body, the chemical structure of the ingested organophosphate, and the time elapsed between ingestion and treatment may limit the effectiveness of pralidoxime as an antidote in organophosphate ingestions. This case study suggests that these factors should be taken into account in assessing the risk of Intermediate Syndrome after intentional organophosphate ingestions.

Pharmacokinetics and pharmacodynamics of bumetanide in critically ill pediatric patients.

This prospective, open-label, clinical trial was conducted to describe the pharmacology of bumetanide in pediatric patients with edema. Nine infants, children, and young adults with edema who were selected for diuretic therapy were studied. After a brief baseline period, each patient received parenteral bumetanide 0.2 mg/kg divided into two equal doses and administered every 12 hours. Urine excretion rate, fractional and total excretion of Na+, Cl-, and K+, creatinine clearance, and plasma and urine concentrations of bumetanide were measured at multiple intervals after drug administration. Bumetanide caused significant increases in the excretion rate of urine and each measured electrolyte. Unexpectedly, creatinine clearance increased dramatically after each dose. Adverse effects, including hypokalemia and hypochloremic metabolic alkalosis, were evident by the end of the treatment period. The plasma pharmacokinetics of bumetanide revealed mean +/- standard deviation values for total clearance and apparent volume of distribution of 3.9 +/- 2.4 mL/min/kg and 0.74 +/- 0.54 L/kg, respectively. Patients excreted an average of 34% of each dose unchanged in the urine over 12 hours. Plasma concentrations of bumetanide accurately predicted several renal effects using a link model with similar pharmacodynamic parameters in each case. Parenteral bumetanide 0.1 mg/kg administered every 12 hours produced significant beneficial and adverse effects in these critically ill pediatric patients with edema. Pharmacokinetic parameters are similar to those previously reported for infants. Plasma concentrations of bumetanide can predict effect-compartment pharmacodynamics.

Pharmacokinetics of famotidine in patients with cystic fibrosis.

Famotidine pharmacokinetics were studied in 13 patients with severe cystic fibrosis (CF) ranging from 10 to 47 years of age and 25 to 72 kg in weight. Patients were randomized to first receive famotidine either 20 mg intravenously or 40 mg orally. Twelve patients were crossed over to the alternate treatment. Repeated blood samples were obtained over 12 hours after intravenous and oral administration and urine was collected over 24 hours for quantitation of famotidine by means of high-performance liquid chromatography (HPLC). A compartment model-dependent approach was used to characterize the disposition of famotidine. From the intravenous data, the mean +/- standard deviation elimination half-life (t1/2) was 2.11 +/- 0.75 hours, the total clearance (Cl) was 0.79 +/- 0.41 L/kg/hr, the renal clearance was 0.57 +/- 0.26 L/kg/hr, the fraction eliminated unchanged in the urine was 83% +/- 16%, and the apparent volume of distribution (Vdss) was 1.33 +/- 0.53 L/kg. The bioavailability determined from comparison of intravenous and oral area under the curve data was 71% +/- 27%. Results of this study support an initial famotidine dose of 20 mg intravenously or 40 mg orally every 12 hours in patients with CF who are older than 9 years of age.

Pharmacokinetics and pharmacodynamics of famotidine in children.

The pharmacokinetics and pharmacodynamics of intravenous famotidine were studied in 12 children (1.1-12.9 years of age; mean weight +/- standard deviation = 27.6 +/- 21.2 kg) who were given the drug for prophylactic management of stress ulceration. After a 0.5-mg/kg infusion of famotidine, timed blood (n = 10) and urine (n = 6) samples and repeated evaluations of intragastric pH (n = 13) were obtained from each subject. Pharmacokinetic parameters were determined from curve fitting of serum concentration data. The mean (+/- SD) maximum serum concentration (Cmax) was 527.6 +/- 281.2 ng/mL, the elimination half-life (t1/2) was 3.2 +/- 3.0 hours, and the apparent steady-state volume of distribution (Vdss) was 2.4 +/- 1.7 L/kg. Plasma clearance (Cl) and renal clearance (ClR) were 0.70 +/- 0.34 L/hr/kg and 0.43 +/- 0.24 L/hr/kg, respectively. Over 24 hours, 73.0 +/- 27.3% of the dose was excreted unchanged in the urine (Fel). Pharmacodynamic analysis of gastric pH data using the sigmoid Emax model predicted that 50% of the maximal effect of famotidine (EC50) occurs at a serum concentration of 26.0 +/- 13.2 ng/mL. Children who did not have an initial intragastric pH < or = 4 did not have a significant response in pH after receiving famotidine. Although Vdss and Cl were higher in these children than those seen in adults, statistically significant relationships between these parameters and age were not observed in the study population. The pharmacodynamics and pharmacokinetics of famotidine in children older than one year of age appear to be similar to those noted in adults.

GC-MS determination of amphetamine and methamphetamine in human urine for 12 hours following oral administration of dextro-methamphetamine: lack of evidence supporting the established forensic guidelines for methamphetamine confirmation.

Ten human volunteers, naive to amphetamines and divided into two groups of five each, were given an oral dose of 30 mg/70 kg D-methamphetamine in one of two different paradigms: the initial dose at 0930 h or the initial dose at 2130 h. One week later, each subject was crossed over with regard to time but given the same dose. A total of 214 urine specimens were collected either prior to dosing or at each micturition for a 12-h period post dose. Specimens were analyzed on a blind basis for methamphetamine and one of its metabolites, amphetamine, by gas chromatography-mass spectrometry (GC-MS) using coinjection of extracted sample and pentafluoropropionic anhydride and selected-ion monitoring. Approximately 20% of the D-methamphetamine was recovered unchanged from the urine specimens, and 2% was recovered as amphetamine. The mean urine methamphetamine concentration in both groups reached a maximum within 4-6 h and declined thereafter. A residual amount of methamphetamine was found in some predose specimens at the crossover evaluation, reflecting that methamphetamine may be detected in urine for up to 7 days. The amphetamine concentration reached a plateau by 4-6 h. This observation coupled with the finding that all subjects excreted approximately 2% of the methamphetamine dose as amphetamine suggested a saturable process for its biotransformation. Concentrations of both methamphetamine and amphetamine tended to be higher, but were not significantly different, for night administration. Methamphetamine concentrations were consistently greater than the 500-ng/mL cutoff in most post-dosing specimens, whereas amphetamine concentrations generally did not achieve the 200-ng/mL cutoff specified by the Substance Abuse and Mental Health Services Administration (SAMHSA) guidelines for GC-MS confirmation of methamphetamine. Some specimens containing methamphetamine had no amphetamine metabolite. The current guidelines would have resulted in 90.2% of the specimens containing methamphetamine being ruled negative by confirmation following either night or day administration, whereas one subject following the initial day administration and another following night crossover administration would have been judged positive at most time intervals. These findings suggest that the current SAMHSA guidelines select for individual metabolic variations and that GC-MS confirmation of methamphetamine will result in most occasional users being ruled negative following an oral dose of methamphetamine while some will be ruled positive.

Serum sickness-like reactions to cefaclor: role of hepatic metabolism and individual susceptibility.

In an effort to explain the increased incidence of serum sickness-like reactions (SSLR) in patients receiving cefaclor, we used an in vitro murine microsomal system as a surrogate for in vivo hepatic drug biotransformation. Lymphocytes from three groups of subjects were exposed to a nonselective mixture of cefaclor metabolites. After an 18-hour incubation of lymphocytes with these metabolites, cells were examined for viability by trypan blue exclusion. The subject groups consisted of patients with a previous history of SSLR after cefaclor therapy (n = 19), patients who experienced adverse reactions to cefaclor suggestive of immediate hypersensitivity (n = 11), and control subjects who had previously tolerated at least two courses of cefaclor therapy without adverse effect (n = 9). Additionally, immediate family members of six subjects with cefaclor-associated SSLR were studied. Lymphocyte killing was 100% greater than baseline (i.e., a non-drug-containing control) in subjects with SSLR compared with those with immediate hypersensitivity reactions (4% cell death above baseline; p < 0.001) and nonaffected control subjects (6% cell death above baseline; p < 0.001). Family studies were consistent with a pattern of maternal inheritance; five of six mothers who had not received cefaclor had a positive (i.e., > or = 35% cell death above baseline) in vitro cytotoxic response. Other studies confirmed the requirement for biotransformation of the parent drug to elicit cell death, demonstrated specificity of the reaction to cefaclor, illustrated a lack of cross-reactivity to cephalexin in subjects with SSLR to cefaclor, and verified the reproducibility of the reaction over time in an affected subject. Our findings indicate that cefaclor associated SSLR may be a unique adverse drug reaction that requires biotransformation of the parent drug and may result from inherited defects in the metabolism of reactive intermediates. Furthermore, this condition can be retrospectively confirmed with an in vitro lymphocyte-based cytotoxicity assay.