Influences of Organic Cation Transporter Polymorphisms on the Population Pharmacokinetics of Metformin in Healthy Subjects

This study investigated the effects of genetic polymorphisms in organic cation transporter (OCT) genes, such as OCT1-3, OCTN1, MATE1, and MATE2-K, on metformin pharmacokinetics. Of particular interest was the influence of genetic polymorphisms as covariates on the variability in the population pharmacokinetics (PPK) of metformin using nonlinear mixed effects modeling (NONMEM). In a retrospective data analysis, data on subjects from five independent metformin bioequivalence studies that used the same protocol were assembled and compared with 96 healthy control subjects who were administered a single oral 500 mg dose of metformin. Genetic polymorphisms of OCT2-808 G > T and OCTN1-917C > T had a significant (P < 0.05) effect on metformin pharmacokinetics, yielding a higher peak concentration with a larger area under the serum time–concentration curve. The values obtained were 102 ± 34.5 L/h for apparent oral clearance (CL/F), 447 ± 214 L for volume of distribution (V_d/F), and 3.1 ± 0.9 h for terminal half-life (mean ± SD) by non-compartmental analysis. The NONMEM method gives similar results. The metformin serum levels were obtained by setting the one-compartment model to a first-order absorption and lag time. In the PPK model, the effects of OCT2-808 G > T and OCTN1-917C > T variants on the CL/F were significant (P < 0.001 and P < 0.05, respectively). Thus, genetic variants of OCTN1-917C > T, along with OCT2-808 G > T genetic polymorphisms, could be useful in titrating the optimal metformin dose.

Electronic supplementary material

The online version of this article (doi:10.1208/s12248-013-9460-z) contains supplementary material, which is available to authorized users.KEY WORDS: genetic polymorphism, metformin, OCTs, population pharmacokinetics     

Elucidation of Arctigenin Pharmacokinetics After Intravenous and Oral Administrations in Rats: Integration of In Vitro and In Vivo Findings via Semi-mechanistic Pharmacokinetic Modeling

Although arctigenin (AR) has attracted substantial research interests due to its promising and diverse therapeutic effects, studies regarding its biotransformation were limited. The current study aims to provide information regarding the pharmacokinetic properties of AR via various in vitro and in vivo experiments as well as semi-mechanistic pharmacokinetic modeling. Our in vitro rat microsome incubation studies revealed that glucuronidation was the main intestinal and liver metabolic pathway of AR, which occurred with V_max, K_m, and Cl_int of 47.5 ± 3.4 nmol/min/mg, 204 ± 22 μM, and 233 ± 9 μl/min/mg with intestinal microsomes and 2.92 ± 0.07 nmol/min/mg, 22.7 ± 1.2 μM, and 129 ± 4 μl/min/mg with liver microsomes, respectively. In addition, demethylation and hydrolysis of AR occurred with liver microsomes but not with intestinal microsomes. In vitro incubation of AR and its metabolites in intestinal content demonstrated that glucuronides of AR excreted in bile could be further hydrolyzed back to the parent compound, suggesting its potential enterohepatic circulation. Furthermore, rapid formation followed by fast elimination of arctigenic acid (AA) and arctigenin-4′-O-glucuronide (AG) was observed after both intravenous (IV) and oral administrations of AR in rats. Linear pharmacokinetics was observed at three different doses for AR, AA, and AG after IV administration of AR (0.48–2.4 mg/kg, r² > 0.99). Finally, an integrated semi-mechanistic pharmacokinetic model using in vitro enzyme kinetic and in vivo pharmacokinetic parameters was successfully developed to describe plasma concentrations of AR, AA, and AG after both IV and oral administration of AR at all tested doses.

Electronic supplementary material

The online version of this article (doi:10.1208/s12248-014-9664-x) contains supplementary material, which is available to authorized users.KEY WORDS: arctigenic acid, arctigenin, arctigenin-4′-O-glucuronide, pharmacokinetics, semi-mechanistic pharmacokinetic modeling     

Long-term Alcohol Consumption Increases Pro-Matrix Metalloproteinase-9 Levels via Oxidative Stress

Matrix metalloproteinases (MMPs) play an important role in alcoholic liver disease. In this study, we evaluated the relationship between pro MMP-9 (pMMP-9) and oxidative stress in plasma of rat exposed to chronic alcohol consumption. Twenty four rats were divided into four groups. Rats in the control group (n = 6) were subjected to physiologic saline by intragastric (i.g.) route. Group Ethanol (n = 6) was given 1 ml of 80% ethanol (v/v) in distilled water through i.g. route. Group Vitamin E (Vit E), (n = 6) was given vitamin E (100 mg kg⁻¹ day⁻¹) by intra peritonealy. Group Vitamin E + Ethanol (n = 6) was given vitamin E 2 h before the administration of ethanol. At the end of 4 weeks, blood samples were taken and plasma malondialdehyde (MDA), protein carbonyls (PCs), aspartate aminotransferase (AST), tumor necrosis factor-α (TNF-α) and pMMP-9 levels were measured. Chronic ethanol administration increased the AST, MDA, PCs, TNF-α and pMMP-9 levels when compared to those in control group (p < 0.05, p < 0.01, p < 0.01, p < 0.05, p < 0.05, respectively). Vitamin E treatment was found to decrease lipid peroxidation and protein oxidation (p < 0.01, p < 0.01, respectively). Also TNF-α and pMMP-9 levels returned to normal by vitamin E treatment. Within all subjects, there was positive correlation between pMMP-9 levels and MDA, PCs levels (p = 0.045, r = 0.454; p = 0.004, r = 0.574, respectively). We conclude that since antioxidant supplementation decreases the alcohol-induced pMMP-9 levels, oxidative stress could be one of the mediators of the generation of MMP-9.     

Modeling Testosterone Circadian Rhythm in Hypogonadal Males: Effect of Age and Circannual Variations

The objective of this study was to characterize the baseline circadian rhythm of testosterone levels in hypogonadal men. A total of 859 baseline profiles of testosterone from hypogonadal men were included in this analysis. The circadian rhythm of the testosterone was described by a stretched cosine function. Model parameters were estimated using NONMEM^® 7.3. The effect of different covariates on the testosterone levels was investigated. Model evaluation was performed using non-parametric bootstrap and predictive checks. A stretched cosine function deeply improved the data goodness of fit compared to the standard trigonometric function (p < 0.001; ΔOFV = −204). The effect of the age and the semester, defined as winter and spring versus summer and fall, were significantly associated with the baseline levels of testosterone (p < 0.001, ΔOFV = −15.6, and p < 0.001, ΔOFV = −47.0). Model evaluation procedures such as diagnostic plots, visual predictive check, and non-parametric bootstrap evidenced that the proposed stretched cosine function was able to model the time course of the diurnal testosterone levels in hypogonadal males with accuracy and precision. The circadian rhythm of the testosterone levels was better predicted by the proposed stretched cosine function than a standard cosine function. Testosterone levels decreased by 5.74 ng/dL (2.4%) every 10 years and were 19.3 ng/dL (8.1%) higher during winter and spring compared to summer and fall.KEY WORDS: circadian rhythm, NONMEM^®, stretched cosine function, testosterone     

Population Pharmacokinetics of Telapristone (CDB-4124) and its Active Monodemethylated Metabolite CDB-4453, with a Mixture Model for Total Clearance

Telapristone is a selective progesterone antagonist that is being developed for the long-term treatment of symptoms associated with endometriosis and uterine fibroids. The population pharmacokinetics of telapristone (CDB-4124) and CDB-4453 was investigated using nonlinear mixed-effects modeling. Data from two clinical studies (n = 32) were included in the analysis. A two-compartment (parent) one compartment (metabolite) mixture model (with two populations for apparent clearance) with first-order absorption and elimination adequately described the pharmacokinetics of telapristone and CDB-4453. Telapristone was rapidly absorbed with an absorption rate constant (Ka) of 1.26 h⁻¹. Moderate renal impairment resulted in a 74% decrease in Ka. The population estimates for oral clearance (CL/F) for the two populations were 11.6 and 3.34 L/h, respectively, with 25% of the subjects being allocated to the high-clearance group. Apparent volume of distribution for the central compartment (V2/F) was 37.4 L, apparent inter-compartmental clearance (Q/F) was 21.9 L/h, and apparent peripheral volume of distribution for the parent (V4/F) was 120 L. The ratio of the fraction of telapristone converted to CDB-4453 to the distribution volume of CDB-4453 (Fmet_est) was 0.20/L. Apparent volume of distribution of the metabolite compartment (V3/F) was fixed to 1 L and apparent clearance of the metabolite (CLM/F) was 2.43 L/h. A two-compartment parent-metabolite model adequately described the pharmacokinetics of telapristone and CDB-4453. The clearance of telapristone was separated into two populations and could be the result of metabolism via polymorphic CYP3A5.KEY WORDS: CDB-4453, mixture model, parent-metabolite, population pharmacokinetics, telapristone (CDB-4124)     

Chronic Treatment with Naltrexone Prevents Memory Retention Deficits in Rats Poisoned with the Sarin Analog Diisopropylfluorophosphate (DFP) and Treated with Atropine and Pralidoxime

Humans and rats poisoned with sarin develop chronic neurological disabilities that are not prevented with standardized antidotal therapy. We hypothesized that rats poisoned with the sarin analogue diisopropylfluorophosphate (DFP) and resuscitated with atropine and pralidoxime would have long-term memory deficits that were preventable with naltrexone treatment. Long Evans rats (250–275 g) were randomized to: DFP (N = 8): single subcutaneous (SC) injection of DFP (5 mg/kg). Treatment (N = 9): DFP (5 mg/kg) followed by chronic naltrexone (5 mg/kg/day × 12 weeks). Control (N = 12): single SC injection of isopropyl alcohol, (DFP vehicle) followed by chronic naltrexone (5 mg/kg/day). If toxicity developed after injection, antidotal therapy was initiated with atropine (2 mg/kg) and pralidoxime (25 mg/kg) and repeated as needed. After 12 weeks, rats underwent testing for place learning (acquisition) across 5 days of training using the Morris Water Maze. On day 6 a memory retention test was performed. Statistical analysis was performed using IBM SPSS Statistics. Rats receiving DFP rapidly developed toxicity requiring antidotal rescue. No differences in acquisition were seen between the DFP vs. DFP + naltrexone rats. During memory testing, DFP-poisoned rats spent significantly less time (29.4 ± 2.11 versus 38.5 ± 2.5 s, p < 0.05) and traveled less distance (267 ± 24.6 versus 370 ± 27.5 cm, p < 0.05) in the target quadrant compared to the treatment group. Treatment rats performed as well as control rats (p > 0.05) on the test for memory retention. Poisoning with DFP induced impaired memory retention. Deficits were not prevented by acute rescue with atropine and pralidoxime. Chronic naltrexone treatment led to preserved memory after DFP poisoning.     

Highly Stabilized Curcumin Nanoparticles Tested in an In Vitro Blood–Brain Barrier Model and in Alzheimer’s Disease Tg2576 Mice

The therapeutic effects of curcumin in treating Alzheimer’s disease (AD) depend on the ability to penetrate the blood–brain barrier. The latest nanoparticle technology can help to improve the bioavailability of curcumin, which is affected by the final particle size and stability. We developed a stable curcumin nanoparticle formulation to test in vitro and in AD model Tg2576 mice. Flash nanoprecipitation of curcumin, polyethylene glycol-polylactic acid co-block polymer, and polyvinylpyrrolidone in a multi-inlet vortex mixer, followed by freeze drying with β-cyclodextrin, produced dry nanocurcumin with mean particle size <80 nm. Nanocurcumin powder, unformulated curcumin, or placebo was orally administered to Tg2576 mice for 3 months. Before and after treatment, memory was measured by radial arm maze and contextual fear conditioning tests. Nanocurcumin produced significantly (p = 0.04) better cue memory in the contextual fear conditioning test than placebo and tendencies toward better working memory in the radial arm maze test than ordinary curcumin (p = 0.14) or placebo (p = 0.12). Amyloid plaque density, pharmacokinetics, and Madin–Darby canine kidney cell monolayer penetration were measured to further understand in vivo and in vitro mechanisms. Nanocurcumin produced significantly higher curcumin concentration in plasma and six times higher area under the curve and mean residence time in brain than ordinary curcumin. The P_app of curcumin and tetrahydrocurcumin were 1.8 × 10⁻⁶ and 1.6 × 10⁻⁵ cm/s, respectively, for nanocurcumin. Our novel nanocurcumin formulation produced highly stabilized nanoparticles with positive treatment effects in Tg2576 mice.

Electronic supplementary material

The online version of this article (doi:10.1208/s12248-012-9444-4) contains supplementary material, which is available to authorized users.Key words: Alzheimer’s disease, behavior tests, nanocurcumin, oral route, pharmacokinetic     

Species and Gender Differences Affect the Metabolism of Emodin via Glucuronidation

The aim of the present study was to define the mechanisms responsible for poor bioavailability of emodin by determining its metabolism using in vitro and in situ disposition models of the intestine and liver. Liver microsomes of mice, rats, guinea pigs, dogs, and humans were used along with the rat intestinal perfusion model and the rat intestinal microsomes. In the rat intestine, excretion rates of emodin-3-O-glucuronide were significantly different (p < 0.05) in four regions of the intestine and were higher in males than in females (p < 0.01). Emodin glucuronidation in liver microsomes was species-dependent, and K_m values varied 5.7-fold (3.2–18.2 μM) in males and 2.8-fold (4.6–13.0 μM) in females. The male intrinsic clearance (CL_int) values differed by 5-fold (27.6–138.3 mL h⁻¹ mg⁻¹ protein), and female CL_int values differed by 4.3-fold (24.3–103.5 mL h⁻¹ mg⁻¹ protein). Since CL_int values of emodin glucuronidation were 10-fold higher than that of isoflavones, emodin was considered rapidly glucuronidated. In contrast to the large species-dependent effects on K_m and CL_int values, gender had a smaller effect on these kinetic parameters (2-fold, p < 0.05). Lastly, glucuronidation rates obtained using liver microsomes from various experimental animals of the same gender correlated well with those in human liver microsomes. In conclusion, Rapid metabolism by UDP-glucuronosyltransferase is the major reason why emodin has poor bioavailability. Species and gender affected emodin metabolism to a different degree, and experimental animals are expected to be useful in predicting emodin glucuronidation in humans.Key words: emodin glucuronidation, first-pass metabolism, gender, species, UGT     

Mercury Exposure Among Artisanal Gold Miners in Madre de Dios, Peru: A Cross-sectional Study

Introduction

Exposure to mercury, a toxic metal, occurs primarily from inhaling mercury vapors or consuming methylmercury-contaminated fish. One third of all anthropogenic mercury emissions worldwide are from artisanal gold mining, which uses mercury to extract gold. Although recent reports suggest that the Madre de Dios region in Peru (with >30,000 artisanal miners) has extensive mercury contamination, residents had never been assessed for mercury exposure. Thus, our objective was to quantify mercury exposure among residents of an artisanal mining town in Madre de Dios and to assess risk factors for exposure.

Methods

We conducted a cross-sectional assessment of 103 residents of an artisanal gold mining town in July 2010. Each participant provided a urine and blood sample and completed a questionnaire assessing potential exposures and health outcomes. We calculated geometric mean (GM) urine total mercury and blood methylmercury concentrations and compared log-transformed concentrations between subgroups using linear regression.

Results

One third (34.0 %) of participants were gold miners. All participants had detectable urine total mercury (GM, 5.5 μg/g creatinine; range, 0.7–151 μg/g creatinine) and 91 % had detectable blood methylmercury (GM, 2.7 μg/L; range, 0.6–10 μg/L); 13 participants (13 %) reported having kidney dysfunction or a neurological disorder. Urine total mercury concentrations were higher among people who heated gold–mercury amalgams compared with people who never heated amalgams (p < 0.05); methylmercury concentrations were higher among fish consumers compared with nonfish consumers (p < 0.05).

Conclusion

Our findings suggest that mercury exposure may be widespread in Huaypetue.     

A Novel MDR1 GT1292-3TG (Cys431Leu) Genetic Variation and Its Effect on P-glycoprotein Biologic Functions

P-glycoprotein (P-gp) is a membrane-bound transporter protein that is encoded by the human multidrug resistance gene MDR1 (ABCB1). P-gp recognizes a wide range of xenobiotics, is pivotal in mediating cancer drug resistance, and plays an important role in limiting drug penetration across the blood–brain barrier. MDR1 genetic variation can lead to changes in P-gp function and may have implications on drug pharmacokinetics. We have identified a novel MDR1_GT1292-3TG (Cys431Leu) genetic variation through systematic profiling of subjects with leukemia. The cellular and transport function of this variation was investigated with recombinant human embryonic kidney cells expressing MDR1. Compared with the wild type, MDR1_GT1292-3TG recombinant cells exhibited a lower drug resistance phenotype for a panel of chemotherapeutic agents. When compared with wild type, MDR1_GT1292-3TG recombinant cells exposed exhibited a 75% decrease in IC₅₀ for doxorubicin (162.6 ± 17.4 to 37.9 ± 2.6 nM) and a 50% decrease in IC₅₀ for paclitaxel (155.7 ± 27.5 to 87.7 ± 9.2 nM), vinblastine (128.0 ± 15.9 to 65.9 ± 5.1 nM), and vincristine (593.7 ± 61.8 to 307.3 ± 17.0 nM). The effects of the Cys431Leu variation, due to MDR1_GT1292-3TG nucleotide transition, on P-gp-dependent intracellular substrate accumulation appeared to be substrate dependent where doxorubicin, vinblastine, and paclitaxel exhibit an increased accumulation (p < 0.05), while verapamil and Hoechst33342 exhibit a decreased intracellular concentration compared with wild type (p < 0.05). Collectively, these data suggest MDR1_GT1292-3TG variation of P-gp may reduce drug resistance and that subjects with this genotype undergoing chemotherapy with drugs that are transported by P-gp could potentially be more responsive to therapy than those with MDR1 wild-type genotype.Key words: ABC transporter, drug resistance, genetic variation, MDR1, P-glycoprotein, polymorphism, transporter     

Sodium/hydrogen exchange inhibition with cariporide reduces leukocyte adhesion via P-selectin suppression during inflammation

Background and purpose:

The Na⁺/H⁺ exchange (NHE) inhibitor cariporide is known to ameliorate ischaemia/reperfusion (I/R) injury by reduction of cytosolic Ca²⁺ overload. Leukocyte activation and infiltration also mediates I/R injury but whether cariporide reduces I/R injury by affecting leukocyte activation is unknown. We studied the effect of cariporide on thrombin and I/R induced leukocyte activation and infiltration models and examined P-selectin expression as a potential mechanism for any identified effects.

Experimental approach:

An in vivo rat mesenteric microcirculation microscopy model was used with stimulation by thrombin (0.5 μ ml⁻¹) superfusion or ischaemia (by haemorrhagic shock for 60 min) and reperfusion (90 min).

Key results:

Treatment with cariporide (10 mg kg⁻¹ i.v.) significantly reduced leukocyte rolling, adhesion and extravasation after thrombin exposure. Similarly, cariporide reduced leukocyte rolling (54±6.2 to 2.4±1.0 cells min⁻¹, P<0.01), adherence (6.3±1.9 to 1.2±0.4 cells 100 μm⁻¹, P<0.01) and extravasation (9.1±2.1 to 2.4±1.1 cells per 20 × 100 μm perivascular space, P<0.05), following haemorrhagic shock induced systemic ischaemia and reperfusion. The cell adhesion molecule P-selectin showed a profound decrease in endothelial expression following cariporide administration in both thrombin and I/R stimulated groups (35.4±3.2 vs 14.2±4.1% P-selectin positive cells per tissue section, P<0.01).

Conclusions and implications:

The NHE inhibitor cariporide is known to limit reperfusion injury by controlling Ca²⁺ overload but these data are novel evidence for a vasculoprotective effect of NHE inhibition at all levels of leukocyte activation, an effect which is likely to be mediated at least in part by a reduction of P-selectin expression.     

In Vitro Lipolysis Data Does Not Adequately Predict the In Vivo Performance of Lipid-Based Drug Delivery Systems Containing Fenofibrate

The present study investigated the utility of in vitro lipolysis performance indicators drug solubilization and maximum supersaturation ratio (SR^M) for their predictive use for the in vivo performance in a minipig model. The commercial Lipanthyl formulation and a series of LbDDS based on identical self-nanoemulsifying drug delivery systems (SNEDDS) containing 200 mg of fenofibrate, either dissolved or suspended, were subjected to combined gastric (pH 2) and intestinal (pH 6.5) in vitro lipolysis. Based on the solubilization profiles and SRM the rank-order SNEDDS (75% drug load) > super-SNEDDS (150% drug load, dissolved) = SNEDDS suspension (150% drug load, partially suspended) > Lipanthyl was established, with an increased likelihood of drug precipitation above SR^M > 3. The in vitro performance, however, was not reproduced in vivo in a minipig model as the mean plasma concentration over time curves of all LbDDS were comparable, independent of the initial physical state of the drug. There was no correlation between the area under the solubilization-time curves (AUC_{in vitro}) of the intestinal step and the AUC_{in vivo}. The study suggests careful interpretation of in vitro performance criteria and revision of LbDDS optimization towards increased solubilization.KEY WORDS: in vitro ipolysis, in vitro/in vivo correlation, lipids, SNEDDS suspensions, super-SNEDDS, supersaturation     

Biodistribution, pharmacokinetics and metabolism of interleukin-1 receptor antagonist (IL-1RA) using [¹⁸F]-IL1RA and PET imaging in rats

BACKGROUND AND PURPOSE

Positron emission tomography (PET) has the potential to improve our understanding of the preclinical pharmacokinetics and metabolism of therapeutic agents, and is easily translated to clinical studies in humans. However, studies involving proteins radiolabelled with clinically relevant PET isotopes are currently limited. Here we illustrate the potential of PET imaging in a preclinical study of the biodistribution and metabolism of ¹⁸F-labelled IL-1 receptor antagonist ([¹⁸F]IL-1RA) using a novel [¹⁸F]-radiolabelling technique.

EXPERIMENTAL APPROACH

IL-1RA was radiolabelled by reductive amination on lysine moieties with [¹⁸F]fluoroacetaldehyde. Sprague-Dawley rats were injected intravenously with [¹⁸F]IL-1RA and imaged with a PET camera for 2 h. For the study of IL-1RA metabolites by ex vivoγ-counting of samples, rats were killed 20 min, 1 h or 2 h after injection of [¹⁸F]IL-1RA.

KEY RESULTS

[¹⁸F]IL-1RA distribution into the major organs of interest was as follows: kidneys >> liver > lungs >> brain. In lungs and liver, [¹⁸F]IL-1RA uptake peaked within 1 min post-injection then decreased rapidly to reach a plateau from 10 min post-injection. In the brain, the uptake exhibited slower pharmacokinetics with a smaller post-injection peak and a plateau from 6 min onward. IL-1RA was rapidly metabolized and these metabolites represented ∼40% of total activity in plasma and ∼80% in urine, 20 min after injection.

CONCLUSIONS AND IMPLICATIONS

Preclinical PET imaging is a feasible method of assessing the biodistribution of new biological compounds of therapeutic interest rapidly. The biodistribution of [¹⁸F]IL-1RA reported here is in agreement with an earlier study suggesting low uptake in the normal brain, with rapid metabolism and excretion via the kidneys.     

Risk Factors for Complications of Drug-Induced Seizures

The purpose of this study is to determine clinical factors associated with complications of drug-induced seizures. This prospective observational study was conducted at an American Association of Poison Control Centers-certified regional poison control center (PCC) over a 1-year period. All consecutive cases reported to a PCC involving seizures were forwarded to investigators, who obtained standardized information including the specific drug or medication exposure, dose, reason for exposure, vital signs, laboratory data, treatment, and outcome. Patients were monitored by daily telephone follow-up until death or discharge. Subjects were excluded if the seizure was deemed to be unrelated to exposure. Odds ratios were used to analyze variables for associations with admission to the hospital for >72 h, endotracheal intubation, status epilepticus, anoxic brain injury, or death. One hundred twenty-one cases met inclusion criteria. Sixty-three (52%) were male, and the mean age was 30 (SD14) years. Common exposures included: antidepressants (33%), stimulants (15%), and anticholinergics (10%). One hundred and three (85%) of the exposures were intentional, of which 74 were suicide attempts and 16 were drug abuse or misuse. Forty-nine (40%) patients required endotracheal intubation, 12(10%) had status epilepticus, 50(41%) were hospitalized for more than 72 h, and one patient died. Median hospital stay was 3 days. Variables significantly associated with complications included stimulant exposure (odds ratios, OR = 11 [95% confidence intervals (CI) 1.9–52]), suicide attempt (OR = 2.2 [95% CI 1.02–4.7]), initial hypotension (OR = 11.2 [95% CI 1.4–89.3]), admission glucose >130 mg/dL (OR = 5.4 [95% CI 1.6–18.1]), and admission HCO₃ < 20 mEq/L (OR = 4.0 [95% CI 1.4–11.3]). Significant clinical factors associated with complications of drug-related seizures include stimulant exposure, suicide attempt, initial hypotension, and admission acidosis or hyperglycemia.     

Investigation of the Influence of FcRn on the Distribution of IgG to the Brain

It has been suggested that the neonatal Fc receptor (FcRn) is a primary determinant of the distribution of IgG to the brain. In the present report, ¹²⁵I-labeled 7E3, a monoclonal IgG1 antibody, was injected intravenously to groups of FcRn-deficient mice and C57BL/6J control mice. Sub-groups of three mice were sacrificed at several time points. Blood and brain tissue were harvested and radioactivity was assessed. Antibody concentrations in brain were corrected for residual blood using ⁵¹Cr-labeled red blood cells. Data were analyzed via WinNonlin, and areas under plasma and tissue concentration vs. time curves (AUCs) were assessed via the Bailer method. The apparent plasma elimination half-life and clearance of 7E3 were 13.61 ± 0.61 days and 6.5 ± 0.10 ml/day/kg in control mice and 0.70 ± 0.05 days and 63.5 ± 2.7 ml/day/kg in the knockout mice. Plasma and brain AUCs (0–10 days) were found to be 3,338.7 ± 50.4 and 7.46 ± 0.5 nM day in control animals and 781.2 ± 16.6 and 1.65 ± 0.1 nM day in FcRn-deficient animals. There was no significant difference between brain-to-plasma AUC ratios in control and FcRn-deficient mice (0.0022 ± 0.00015 vs. 0.0021 ± 0.00011, p = 0.3347). Two-way analysis of variance showed no significant differences, at any time point, between brain-to-plasma concentration ratios determined from control and knockout animals. The results suggest that FcRn does not contribute significantly to the “blood–brain barrier” for IgG in mice, and the data suggest that FcRn is not responsible for the low exposure of IgG in the brain relative to plasma.Key words: antibody, blood brain barrier, FcRn, IgG, pharmacokinetics     

Blood Levels of Methemoglobin in Patients with Aluminum Phosphide Poisoning and its Correlation with Patient’s Outcome

Although methemoglobinemia following aluminum phosphide (AlP) intoxication has been reported, probable effect of blood level of methemoglobin (Met-Hb) on outcome of AlP-poisoned patients has not yet been investigated. This study aimed to evaluate blood levels of methemoglobin in patients with AP intoxication and its correlation with patient’s outcome. This prospective study was carried out at the Loghman–Hakim poison hospital from April 2009 to August 2009. All patients aged >12 years who had ingested AlP and were admitted at the hospital were enrolled in the study. Using the co-oximetry, blood Met-Hb level was measured at the time of admission and 24 h later if the patient survived. Forty-eight patients with AlP intoxication including 24 males were evaluated. Mean age of the patients was 25.5 ± 9.5 years. There was significant association between blood level of Met-Hb at the time of admission and mortality (2.4% ± 7.1% in survivors versus 15.2% ± 13.5% in non-survivors, P < 0.001). The same association was found at the 2nd day of admission (2.9% ± 8.2% in survivors versus 26.5% ± 19.9% in non-survivors, P = 0.02). The present study found an association between blood level of Met-Hb and mortality in patients with AlP intoxication. Effect of administration of vitamin C and methylene blue on outcome of patients with AlP intoxication should be investigated in future studies.     

Integrated Pharmacokinetic-Driven Approach to Screen Candidate Anticancer Drugs for Brain Tumor Chemotherapy

The goal of the study was to develop an effective screening strategy to select new agents for brain tumor chemotherapy from a series of low molecular weight anticancer agents [ON123x] by the combined use of in silico, in vitro cytotoxicity, and in vitro ADME profiling studies. The results of these studies were cast into a pipeline of tier 1 and tier 2 procedures that resulted in the identification of ON123300 as the lead compound. Of the 154 ON123xx compounds, 13 met tier 1 screening criteria based on physicochemical properties [i.e., MW < 450 Da, predicted log P between 2 and 3.5] and in vitro glioma cell cytotoxicity [i.e., IC50 < 10 μM] and were further tested in tier 2 assays. The tier 2 profiling studies consisted of metabolic stability, MDCK-MDR1 cell permeability and plasma and brain protein binding that were combined to globally assess whether favorable pharmacokinetic properties and brain penetration could be achieved in vivo. In vivo cassette dosing studies were conducted in mice for 12 compounds that permitted examination of in vitro/in vivo relationships that confirmed the suitability of the in vitro assays. A parameter derived from the in vitro assays accurately predicted the extent of drug accumulation in the brain based on the area under the drug concentration–time curve in brain measured in the cassette dosing study (r² = 0.920). Overall, the current studies demonstrated the value of an integrated pharmacokinetic-driven approach to identify potentially efficacious agents for brain tumor chemotherapy.

Electronic supplementary material

The online version of this article (doi:10.1208/s12248-012-9428-4) contains supplementary material, which is available to authorized users.KEY WORDS: brain tumor, CNS, drug development, pharmacokinetics, preclinical     

An Electrically Tight In Vitro Blood–Brain Barrier Model Displays Net Brain-to-Blood Efflux of Substrates for the ABC Transporters,P-gp,Bcrp and Mrp-1

Efflux transporters of the ATP-binding cassette superfamily including breast cancer resistance protein (Bcrp/Abcg2), P-glycoprotein (P-gp/Abcb1) and multidrug resistance-associated proteins (Mrp’s/Abcc’s) are expressed in the blood–brain barrier (BBB). The aim of this study was to investigate if a bovine endothelial/rat astrocyte in vitro BBB co-culture model displayed polarized transport of known efflux transporter substrates. The co-culture model displayed low mannitol permeabilities of 0.95 ± 0.1 · 10⁻⁶ cm·s⁻¹ and high transendothelial electrical resistances of 1,177 ± 101 Ω·cm². Bidirectional transport studies with ³H-digoxin, ³H-estrone-3-sulphate and ³H-etoposide revealed polarized transport favouring the brain-to-blood direction for all substrates. Steady state efflux ratios of 2.5 ± 0.2 for digoxin, 4.4 ± 0.5 for estrone-3-sulphate and 2.4 ± 0.1 for etoposide were observed. These were reduced to 1.1 ± 0.08, 1.4 ± 0.2 and 1.5 ± 0.1, by addition of verapamil (digoxin), Ko143 (estrone-3-sulphate) or zosuquidar + reversan (etoposide), respectively. Brain-to-blood permeability of all substrates was investigated in the presence of the efflux transporter inhibitors verapamil, Ko143, zosuquidar, reversan and MK 571 alone or in combinations. Digoxin was mainly transported via P-gp, estrone-3-sulphate via Bcrp and Mrp’s and etoposide via P-gp and Mrp’s. The expression of P-gp, Bcrp and Mrp-1 was confirmed using immunocytochemistry. The findings indicate that P-gp, Bcrp and at least one isoform of Mrp are functionally expressed in our bovine/rat co-culture model and that the model is suitable for investigations of small molecule transport.KEY WORDS: blood–brain barrier, breast cancer resistance protein, multidrug resistance-associated protein, p-glycoprotein, polarized small molecule transport     

Pharmacokinetic Evaluation of Intranasally Administered Vinyl Polymer-Coated Lorazepam Microparticles in Rabbits

The intranasal (IN) administration of lorazepam is desirable in order to maximize speed of onset and minimise carry-over sedation; however, this benzodiazepine is prone to chemical hydrolysis and poor airway retention, and thus, innovative epithelial presentation is required. The aim of this study was to understand how the in situ self-assembly of a mucoretentive delivery system, formed by the dissolution of vinyl polymer-coated microparticles in the nasal mucosa, would influence lorazepam pharmacokinetics (PK). IN administration of the uncoated lorazepam powder (particle size, 6.7 ± 0.1 μm) generated a biphasic PK profile, which was indicative of sequential intranasal and oral absorption (n = 6; dose, 5 mg/kg). Coating the drug with the vinyl polymer, MP1 (9.9 ± 0.5 μm with 38.8 ± 14.0%, w/w lorazepam) and MP2 (10.7 ± 0.1 μm with 47.0 ± 1.0%, w/w lorazepam), allowed rapid systemic absorption (MP1, T_max 14.2 ± 4.9 min; MP2, T_max 9.3 ± 3.8 min) in rabbits and modified the PK profiles in a manner that suggested successful nasal retention. The poly(vinyl pyrrolidone)-rich MP2 system provided the best comparative bioavailability, it prolonged the early-phase nasal drug absorption and minimised drug mucociliary clearance, which correlated well with the intermolecular hydrogen-bond-driven vinyl polymer interactions observed in vitro.KEY WORDS: intranasal, lorazepam, microparticles, pharmacokinetics, poly(vinyl alcohol)     

Influence of rosuvastatin on the NAD(P)H oxidase activity in the retina and electroretinographic response of spontaneously hypertensive rats

Background and purpose:

Retinal complications may be encountered during the development of hypertension as a response to oxidative stress. Statins may reduce the risk of developing hypertension and ocular diseases. We evaluate the effects of rosuvastatin (ROSU) on retinal functionality and oxidative stress levels in normotensive and spontaneously hypertensive rats (SHR).

Experimental approach:

Wistar Kyoto (WKY) and SHR were treated for 3 weeks with rosuvastatin (10 mg kg⁻¹ day⁻¹). Electroretinograms (ERG) were recorded before and after rosuvastatin treatment. Reactive oxygen species (ROS) were determined in the retina with dihydroethidium staining and NAD(P)H oxidase activity was evaluated.

Key results:

Retinal ganglion cell ROS and retinal NAD(P)H oxidase activity were higher in SHR than in WKY rats, respectively (17.1±1.1 vs 10.2±1.2 AU, P<0.01; 38095±8900 vs 14081±5820 RLU mg⁻¹; P<0.05). The ERG b-wave amplitude in SHR was significantly lower than that in WKY rats. Rosuvastatin reduced SBP in SHR but did not change plasma lipid levels. Rosuvastatin treatment in SHR significantly decreased ROS levels (11.2±1.3, P<0.01), NAD(P)H activity in retinal ganglion cells (9889±4290; P<0.05), and increased retinal plasmalogen content in SHR, but did not modify the ERG response.

Conclusions and implications:

Rosuvastatin, beyond lowering cholesterol levels, was able to lower ROS in the retina induced by hypertension, but without improving retinal function in SHR. These findings point to a complex relationship between ROS in the pathogenesis of retinal disease and hypertension.