Extended access oxycodone self-administration and neurotransmitter receptor gene expression in the dorsal striatum of adult C57BL/6 J mice

Rationale

Although non-medical use of oxycodone continues to be a growing problem in the United States, there are no animal studies examining the effects of long-term oxycodone self-administration (SA).

Objectives

The current study was designed to examine chronic oxycodone SA by mice (14 days), in novel extended (4 h) SA sessions and its effect on selective striatal neurotransmitter receptor mRNA expression.

Methods

Adult male C57/BL6J mice were either allowed to self-administer oxycodone (0.25 mg/kg/infusion, FR1) or served as yoked-saline controls in an extended access paradigm. Mice self-administered oxycodone for 4 h/day for 14 consecutive days. Comparison groups with 14-days exposure to 1-h SA sessions were also studied. Within 1 h of the last extended SA session, mice were sacrificed, dorsal striatum was isolated and selective neurotransmitter receptor mRNA levels were examined.

Results

The oxycodone groups poked the active hole significantly more times than the yoked controls. The number of nose pokes at the active hole rose over the 14 days in the oxycodone group with extended access. The expression of 13 neurotransmitter receptor mRNAs was significantly altered in the dorsal striatum, including the gamma-aminobutyric acid (GABA) A receptor beta 2 subunit (Gabrb2) showing experiment-wise significant decrease, as a result of extended oxycodone SA.

Conclusion

C57BL/6 J mice escalated the amount of oxycodone self-administered across 14 consecutive daily extended sessions, but not 1-h sessions. Decreases in Gabrb2 mRNA levels may underlie escalation of oxycodone intake in the extended access SA sessions.     

Nonclinical pharmacokinetics and metabolism of EPZ‐5676, a novel DOT1L histone methyltransferase inhibitor

(2R,3R,4S,5R)‐2‐(6‐Amino‐9H‐purin‐9‐yl)‐5‐((((1r,3S)‐3‐(2‐(5‐(tert‐butyl)‐1H‐benzo[d]imidazol‐2‐yl)ethyl)cyclobutyl)(isopropyl)amino)methyl)tetrahydrofuran‐3,4‐diol (EPZ‐5676) is a novel DOT1L histone methyltransferase inhibitor currently in clinical development for the treatment of MLL‐rearranged leukemias. This report describes the preclinical pharmacokinetics and metabolism of EPZ‐5676, an aminonucleoside analog with exquisite target potency and selectivity that has shown robust and durable tumor growth inhibition in preclinical models. The in vivo pharmacokinetics in mouse, rat and dog were characterized following i.v. and p.o. administration; EPZ‐5676 had moderate to high clearance, low oral bioavailability with a steady‐state volume of distribution 2–3 fold higher than total body water. EPZ‐5676 showed biexponential kinetics following i.v. administration, giving rise to a terminal elimination half‐life (t_1/2) of 1.1, 3.7 and 13.6 h in mouse, rat and dog, respectively. The corresponding in vitro ADME parameters were also studied and utilized for in vitro–in vivo extrapolation purposes. There was good agreement between the microsomal clearance and the in vivo clearance implicating hepatic oxidative metabolism as the predominant elimination route in preclinical species. Furthermore, low renal clearance was observed in mouse, approximating to f_u‐corrected glomerular filtration rate (GFR) and thus passive glomerular filtration. The metabolic pathways across species were studied in liver microsomes in which EPZ‐5676 was metabolized to three monohydroxylated metabolites (M1, M3 and M5), one N‐dealkylated product (M4) as well as an N‐oxide (M6). Copyright © 2014 John Wiley & Sons, Ltd.     

Investigating the contribution of CYP2J2 to ritonavir metabolism in vitro and in vivo

Ritonavir, an HIV protease inhibitor, is successfully used for the prevention and treatment of HIV infections. Ritonavir pharmacokinetics are complicated by inhibition, induction and pharmacogenetics of cytochrome P450 (CYP) enzymes mediating its clearance. This investigation revealed that CYP2J2, along with CYP3A4/5 and CYP2D6, efficiently metabolizes ritonavir, and to a CYP2J2-specific (minor) metabolite. Chemical inhibition of ritonavir metabolism, clearance, K_I/k_inact and abundance of CYP2J2 in liver microsomes were evaluated and then applied to an in vitro–in vivo static scaling model to estimate the contribution of each isozyme, as a function of CYP abundance, activity, and genotype. Disposition of the CYP2J2-specific metabolite was also evaluated in vivo. In plasma, metabolite abundance was well above previously reported levels with circulating concentrations measured at 2 μM for the main hydroxylisopropyl metabolite. Ritonavir and metabolite plasma profiles were simulated using Simcyp^®. A modest (2–6%) contribution of CYP2J2 to ritonavir clearance is predicted which increases to more than 20% in subjects carrying CYP2D6 poor metabolizer polymorphisms and CYP3A4 irreversible inhibition. These results indicate that minor drug metabolizing enzymes could become quantitatively important in RTV clearance if main metabolic pathways are impeded.     

Differential responses to JNJ‐37822681, a specific and fast dissociating dopamine D2 receptor antagonist,in cynomolgus monkey and Sprague–Dawley rat general toxicology studies: clinical observations,prolactin levels,mammary histopathology findings and toxicokinetics

JNJ‐37822681 is a potent, specific and fast dissociating dopamine D₂ receptor antagonist intended for the treatment of schizophrenia. Its nonclinical toxicological profile was investigated in a series of general repeat dose toxicity studies in cynomolgus monkeys and Sprague–Dawley rats. The maximum duration of treatment was 9 and 6 months, respectively. Interspecies differences were noted in the response to JNJ‐37822681 in terms of extrapyramidal (EPS)‐like clinical signs and prolactin‐mediated tissue changes in the mammary gland. Monkeys showed severe EPS‐like clinical signs such as abnormal posture, abnormal eye movements and hallucination‐like behavior at relatively low exposures compared to those associated with EPS in patients with schizophrenia. The high sensitivity of the monkey to JNJ‐37822681‐induced EPS‐like signs was unexpected based on the fast dissociating properties of the compound. Rats, however, were not prone to EPS. Elevated serum prolactin levels were found in rats and monkeys. While rats showed slight to moderate prolactin‐related tissue changes upon histopathological examination in all studies, which among others affected the mammary gland, only minor mammary gland tissue changes were noted in monkeys. Prolactin levels were only slightly increased in patients with schizophrenia receiving relatively high dose levels of JNJ‐37822681. The monkey toxicology studies did not provide an exposure‐based safety margin, while in rats adverse effects were only noted at exposures considerably higher than those achieved at efficacious plasma concentrations in the clinic. Overall, the available data suggest that the cynomolgus monkey showed better predictivity towards the nature of JNJ‐37822681‐associated adverse events in humans than the Sprague–Dawley rat. Copyright © 2013 John Wiley & Sons, Ltd.     

Surgery‐induced changes in rat IL‐1β and acetylcholine metabolism: role of physostigmine

Pharmacological enhancement of cholinergic activity following administration of physostigmine is known to induce protective effects generally. However, it is unclear whether the effect of physostigmine on inflammation and acetylcholine (ACh) metabolism is related to different types of surgical intervention or anaesthesia alone. To investigate this, rats were subjected to partial liver resection (PLR) or sham surgery, with a control group receiving anaesthesia alone. Half of each treatment group received a single intra‐operative dose of physostigmine (0.04 mg/kg); the others received placebo. Acetylcholinesterase (AChE) activity and plasma and brain concentrations of interleukin (IL)‐1β and ACh were determined. Both PLR and sham operation induced a time‐dependent increase in plasma concentrations of IL‐1β compared with rats receiving anaesthesia alone (3.9‐ and 4.8‐fold increases, respectively). In the brain, IL‐1β concentrations had increased approximately twofold after surgery compared with the control group. Blood AChE was transiently decreased after surgery. Brain AChE activity increased 1.3‐fold (P = 0.014) only after PLR; consequently, cerebral ACh concentrations were significantly reduced. Physostigmine administration significantly reduced IL‐1β and AChE levels. Cerebral ACh concentrations were markedly increased from 544 ± 122 ng/mg protein following placebo administration to 654 ± 93 ng/mg protein after physostigmine administrations (P < 0.001). We conclude that a single dose of physostigmine intra‐operatively has a sustained anti‐inflammatory effect (up to 120 min after injection) that is especially pronounced under the conditions of PLR surgery. In addition to its protective peripheral action, physostigmine exerts a neuroprotective action by increasing levels of the neurotransmitter ACh.     

A repeat-dose thorough QT study of inhaled fluticasone furoate/vilanterol combination in healthy subjects

AIMS

This study was designed as a thorough QT (TQT) study to evaluate the effects of fluticasone furoate(FF)/vilanterol (VI) in healthy subjects. Supportive data from a TQT study conducted with FF are also presented.

METHODS

This was a randomized, placebo-and positive-controlled, double-dummy, double-blind, four-way crossover study, in which healthy subjects (n = 85) were randomized to 7 days of once-daily treatment of FF/VI (200/25 or 800/100 μg) or placebo or single-dose oral moxifloxacin (single-blind, 400 mg). In the supportive TQT study, subjects (n = 40) were randomized to single-dose inhaled FF(4000 μg), oral moxifloxacin (400 mg) or placebo.

RESULTS

There was a lack of effect of FF/VI (200/25 μg) on QTcF (Fridericia''s correction); all time-matched mean differences from baseline relative to placebo (0–24 h) were <5 ms, with upper 90% confidence intervals (CI) of <10 ms. At 800/100 μg, FF/VI had no significant clinicaleffect on QTcF except at 30 min postdose when the 90% CI was >10 ms [mean (90% CI), 9.6 ms (7.2, 12.0)]. No effect on QTci (individually corrected) was observed at either strength of FF/VI, with mean time-matched treatment differences <5 ms at all time points [upper 90% CIs <10 ms (0–24 h)]. Assay sensitivity was confirmed; moxifloxacin prolonged QTcF and QTci, with time-matched mean differences from baseline relative toplacebo of >10 ms (1–8 h postdose).

CONCLUSIONS

Repeat once-daily dosing of FF/VI (200/25 μg), which is the highest therapeutic strength used in phase III studies, is not associated with QTc prolongation in healthy subjects. Supratherapeutic strength FF/VI (800/100 μg) demonstrated a small transient effect on QTcF but not on QTci.     

Sewage‐based epidemiology in monitoring the use of new psychoactive substances: Validation and application of an analytical method using LC‐MS/MS

Sewage‐based epidemiology (SBE) employs the analysis of sewage to detect and quantify drug use within a community. While SBE has been applied repeatedly for the estimation of classical illicit drugs, only few studies investigated new psychoactive substances (NPS). These compounds mimic effects of illicit drugs by introducing slight modifications to chemical structures of controlled illicit drugs. We describe the optimization, validation, and application of an analytical method using liquid chromatography coupled to positive electrospray tandem mass spectrometry (LC‐ESI‐MS/MS) for the determination of seven NPS in sewage: methoxetamine (MXE), butylone, ethylone, methylone, methiopropamine (MPA), 4‐methoxymethamphetamine (PMMA), and 4‐methoxyamphetamine (PMA). Sample preparation was performed using solid‐phase extraction (SPE) with Oasis MCX cartridges. The LC separation was done with a HILIC (150 x 3 mm, 5 µm) column which ensured good resolution of the analytes with a total run time of 19 min. The lower limit of quantification (LLOQ) was between 0.5 and 5 ng/L for all compounds. The method was validated by evaluating the following parameters: sensitivity, selectivity, linearity, accuracy, precision, recoveries and matrix effects. The method was applied on sewage samples collected from sewage treatment plants in Belgium and Switzerland in which all investigated compounds were detected, except MPA and PMA. Furthermore, a consistent presence of MXE has been observed in most of the sewage samples at levels higher than LLOQ. Copyright © 2015 John Wiley & Sons, Ltd.