Physiologically-based pharmacokinetic model for alectinib, ruxolitinib, and panobinostat in the presence of cancer, renal impairment, and hepatic impairment.

Renal (RIP) and hepatic (HIP) impairments are prevalent conditions in cancer patients. They can cause changes in gastric emptying time, albumin levels, hematocrit, glomerular filtration rate, hepatic functional volume, blood flow rates, and metabolic activity that can modify drug pharmacokinetics. Performing clinical studies in such populations has ethical and practical issues. Using predictive physiologically-based pharmacokinetic (PBPK) models in the evaluation of the PK of alectinib, ruxolitinib, and panobinostat exposures in the presence of cancer, RIP, and HIP can help in using optimal doses with lower toxicity in these populations. Verified PBPK models were customized under scrutiny to account for the pathophysiological changes induced in these diseases. The PBPK model-predicted plasma exposures in patients with different health conditions within average 2-fold error. The PBPK model predicted an area under the curve ratio (AUCR) of 1, and 1.8, for ruxolitinib and panobinostat, respectively, in the presence of severe RIP. On the other hand, the severe HIP was associated with AUCR of 1.4, 2.9, and 1.8 for alectinib, ruxolitinib, and panobinostat, respectively, in agreement with the observed AUCR. Moreover, the PBPK model predicted that alectinib therapeutic cerebrospinal fluid levels are achieved in patients with non-small cell lung cancer, moderate HIP, and severe HIP at 1-, 1.5-, and 1.8-fold that of healthy subjects. The customized PBPK models showed promising ethical alternatives for simulating clinical studies in patients with cancer, RIP, and HIP. More work is needed to quantify other pathophysiological changes induced by simultaneous affliction by cancer and RIP or HIP.

Efficacy of a single session in-series hemoperfusion and hemodialysis in the management of carprofen overdose in two dogs.

OBJECTIVE: To describe the efficacy of in-series hemoperfusion and hemodialysis in 2 dogs with carprofen overdose. CASE SUMMARY: This report describes the treatment of 2 dogs following accidental carprofen overdoses who underwent a single in-series hemoperfusion and hemodialysis session. Serial serum carprofen concentrations were measured before, during, and after the session. The first patient's session lasted 5 hours, with the largest decrease in serum carprofen concentrations occurring during the first hour of treatment. The carprofen clearance during the following 4 hours of treatment decreased substantially compared to the first hour and was not different from the patient's intrinsic clearance of carprofen after the session was completed. Based on the findings from the first case, the second patient was treated with a 1 hour single hemoperfusion and hemodialysis session. Our results support the hypothesis that carprofen is not effectively removed by conventional hemodialysis and the efficacy of hemoperfusion is short lived due to rapid saturation of the charcoal filter. Once filter saturation occurs, the extracorporeal session is no longer efficacious. Using in-series hemoperfusion and hemodialysis is of benefit to correct the side effects seen with hemoperfusion alone, and hourly charcoal filter replacement may extend the efficacy of treatment in removing carprofen. NEW OR UNIQUE INFORMATION PROVIDED: This is the first published report of in-series hemoperfusion and hemodialysis being used to treat carprofen overdose in a dog. In these 2 cases, the intrinsic clearances of the patients were shown to be equivalent to that of standard hemodialysis alone, indicating that hemodialysis does not produce any advantage in carprofen clearance. In this limited report, we suggest that the efficacy of hemoperfusion in removing carprofen is short-lived, and extending the treatment beyond the first hour does not produce any therapeutic benefit. In order to extend the efficacy of hemoperfusion, hourly replacement of the charcoal filter should be considered.

Biofluid sampler: A new gateway for mail-in-analysis of whole blood samples.

The current study reports the development of a novel biofluid sampler (BFS) which is capable of sampling and sample preparation of whole blood without converting it into plasma or serum. The sampler can retain a whole blood sample from 10 to 1000 µL. Although the device shares the same working principle of dried blood spot (DBS) cards, it eliminates most of the technological shortcomings of DBS cards such as low maximum sample volume (~50 µL), sample inhomogeneity due to haematocrit, and poor physical adsorption driven analyte retention by incorporating sol-gel derived high efficiency, multi-functional sorbents on cellulose fabric substrate. The performance of BFS was tested via "Mail-in-Analysis" using three non-steroidal anti-inflammatory drugs (NSAIDs, ketoprofen, carprofen and diclofenac) as the test compounds. Human whole blood samples were fortified with the test compounds and sampled on conventional DBS cards and biofluid samplers (BFSs) in the USA. After drying the blood samples at room temperature, the samples were shipped to Italy for chromatographic analysis. The analytes were back-extracted from the DBS cards and BFSs using methanol and subsequently analysed using a short Symmetry C18 column (75 × 4.6 mm, 3.5 µm). Acetonitrile (ACN) and PBS (30 mM; pH = 2.5) were used as the mobile phases and the elution was performed under isocratic conditions. Compared to the classical dried blood spot cards (DBS), BFSs offer better performance in retaining the selected NSAIDs under conventional postal shipment. By substantially expanding the sampling capacity, eliminating most of the shortcomings of classical DBS cards and exploiting the better materials properties of sol-gel based functional sorbents, BFSs offer a new and profoundly simplified approach for whole blood sampling and analysis and is expected to change the current practice of blood analysis, allowing accurate quantitative analyses either in a local laboratory (on site) or using mail-in-analysis (off site) without compromising the quality of bioanalytical data.

Determination of carbazole and halogenated carbazoles in human serum samples using GC-MS/MS.

Carbazole and halogenated carbazoles have been widely detected throughout the environment in soil, river deposits, and lake sediments. Human exposure to these compounds may occur through inhalation, drinking water, dietary intake and/or skin contact, and exposure levels in the body may be evaluated by measuring them in serum or blood. This paper reports the method development and validation for the analysis of carbazole and 11 halogenated carbazoles in human blood and/or serum samples. A small sample size of 100 µL of blood or serum was employed for the analysis. The samples were prepared through salting-out liquid-liquid extraction (LLE) by using hexane/ethyl acetate (4:1, v/v) as the extraction solvent and aqueous MgSO4 (37.5 wt%) as the salting-out regent, respectively. Sample analysis was performed using gas-chromatography (GC) coupled with a tandem mass spectrometer (MS/MS) in an electron impact (EI) mode. The developed method demonstrated low detection limits in the range of 0.02-0.27 ng/mL, intra-day accuracy ranging from 81.2% to 125%, and inter-day accuracy from 91.0% to 117%. The intra- and inter-day precisions, calculated by relative standard deviations (RSDs), were in the ranges of 1.0-16.0% and 1.8-16.4%, respectively. The developed method was applied to the analysis of 50 human serum samples collected from pregnant women in Southern California in 2012. Low concentrations of carbazole were measured in 18 samples, while halogenated carbazoles were not detected in any of the samples.

Quantification of afatinib, alectinib, crizotinib and osimertinib in human plasma by liquid chromatography/triple-quadrupole mass spectrometry; focusing on the stability of osimertinib.

The development and full validation of a sensitive and selective ultra-performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) method are described for the simultaneous analysis of afatinib, alectinib, crizotinib and osimertinib in human lithium heparinized plasma. Afatinib-d6, crizotinib-d5 and erlotinib-d6 were used as internal standards. Given osimertinib's instability in plasma and whole blood at ambient temperature, samples should be solely processed on ice (T = 0 °C). Chromatographic separation was obtained on an Acquity UPLC ® BEH C18; 2.1 × 50 mm, 1.7 µm column, which was eluted with 0.400 mL/minute flow on a linear gradient, consisting of 10 mM ammonium formate (pH 4.5) and acetonitrile. Calibration curves for all compounds were linear for concentration ranges of 1.00 to 100 ng/mL for afatinib and 10.0 to 1000 ng/mL for alectinib, crizotinib and osimertinib, herewith validating the lower limits of quantification at 1.00 ng/mL for afatinib and 10.0 ng/mL for alectinib, crizotinib and osimertinib. Within-run and between-run precision measurements fell within 10.2%, with accuracy ranging from 89.2 to 110%.

Simplified LC-MS/MS method for quantification of IG-105, a novel tubulin ligand, and its application to the pharmacokinetic study in rats at the anticancer effective dose.

IG-105, N-(2, 6-dimethoxypyridine-3-yl)-9-methylcarbazole-3-sulfonamide, a novel carbazole sulfonamide, shows a potent anticancer activity in a variety of human tumor cells in vitro and in vivo. In the present study, a rapid and convenient liquid chromatography/tandem mass spectrometry (LC-MS/MS) method was developed and applied to the pharmacokinetic study of IG-105 in rats. Chromatographic separation was accomplished on a C18 column using an isocratic mobile phase of acetonitrile-water-acetic acid (56:44:0.2, v/v/v). The ion transitions of IG-105 and combretastatin A4 (internal standard) in selected reaction monitoring mode were m/z 398→154 and m/z 317→286, respectively. The assay exhibited good linearity over the range of 2-512 ng/mL. Intra- and inter-day precisions were within 8.2 %, and the accuracies ranged from -6.0 to 3.7 %. The extraction recoveries were higher than 90 %, and the matrix effects were negligible. All quality control samples were stable at different storage conditions. The validated LC-MS/MS method was successfully applied to a preclinical pharmacokinetic study of IG-105 in rats after a single oral dose of 100, 250, or 1000 mg/kg which showed tumor growth inhibition activity. The absorption of IG-105 was proved to be rapid but saturated to a certain extent into the blood circulation, from where it was distributed and eliminated gradually.

Bioanalytical liquid chromatography-tandem mass spectrometric assay for the quantification of the ALK inhibitors alectinib, brigatinib and lorlatinib in plasma and mouse tissue homogenates.

Several second and third generation ALK inhibitors have been introduced in recent years. A bioanalytical assay for simultaneous quantification of alectinib, brigatinib, and lorlatinib was developed and validated for human plasma. The method was also partially validated for diluted mouse plasma and tissue homogenates of brain, liver, kidney, and spleen. Samples (40 µl) were pretreated in a 96-well plate by protein precipitation with acetonitrile containing the internal standard [2H8]-alectinib. After chromatographic separation on an ethylene bridged octadecyl silica column by gradient elution at 600 µl/min using 1% (v/v) formic acid (in water) and acetonitrile, compounds were ionized by a turbo electrospray and monitored by selected reaction monitoring on a triple quadrupole mass spectrometer. Validation was performed in a 2-2000 ng/ml concentration range for alectinib and lorlatinib and a 4-4000 ng/ml range for brigatinib. Precisions (within-day and between-day) were in the range 2.2-15.0% and accuracies were in between 87.2 and 110.2% for all matrices and levels. Compounds were sufficiently stable under most investigated conditions. Results of a pilot pharmacokinetic and tissue distribution study for brigatinib in mice are reported. Finally, successful incurred samples reanalysis of tissue homogenate samples containing brigatinib and lorlatinib is presented. Lorlatinib homogenate samples were also successfully reanalyzed using a second independent assay (cross-validation).

Amorphous solid dispersions of carvedilol along with pH-modifiers improved pharmacokinetic properties under hypochlorhydoria.

Carvedilol (CAR) belongs to biopharmaceutics classification system class-II drugs, with poor aqueous solubility and pH-dependent solubility. The present study aimed to develop a novel amorphous solid dispersion (ASD) of CAR with acidic counter ions for pH modifications in microenvironment to improve the pharmacokinetic properties under hypochlorhydric conditions. CAR-ASD was prepared by freeze-drying in combination with counter ions and hydroxypropyl cellulose, and their physicochemical properties including dissolution behavior, storage stability, and photostability were characterized. Pharmacokinetic studies were carried out after oral administration of CAR samples in both normal and omeprazole-treated (30 mg/kg, p.o.) rats as a hypochlorhydria model. Among the tested six counter ions, citric acid (CA) was found to be a preferable pH-modifier of CAR with respect to the dissolution profile and photostability (both potency and colorimetric evaluation). In CAR-ASD formulation with 50% loading of CA (CAR-ASD/CA50), amorphization of CAR was observed during the preparation process. After the oral administration of crystalline CAR in rats under hypochlorhydric condition, there was a 34.4% reduction in the systemic exposure of CAR compared with that in normal rats. However, orally-dosed CAR-ASD/CA50 resulted in limited alterations of pharmacokinetic behavior between normal and omeprazole-treated rats. From these findings, addition of CA as pH-modifier in CAR-ASD might provide consistent pharmacokinetic behavior of CAR even under hypochlorhydric conditions.

A study of the pharmacokinetics and thromboxane inhibitory activity of a single intramuscular dose of carprofen as a means to establish its potential use as an analgesic drug in white rhinoceros.

The alleviation of pain and prevention of suffering are key aspects of animal welfare. Unfortunately, analgesic drugs are not available for all species. White rhinoceros (Ceratotherium simum), representing one of such species, which survive poaching attempts inflicted with severe facial injuries and gunshot wounds, nonetheless require analgesic support. To improve treatment conditions, this study explored the use of carprofen for the treatment of pain and inflammation in white rhinoceros. The pharmacokinetics of 1 mg/kg intramuscular carprofen was evaluated in six healthy white rhinoceros. The half-life of λz and mean residence time was 105.71 ± 15.67 and 155.01 ± 22.46 hr, respectively. The area under the curve and the maximum carprofen concentration were 904.61 ± 110.78 µg ml-1  hr-1 and 5.77 ± 0.63 µg/ml, respectively. Plasma TXB2 inhibition demonstrated anti-inflammatory properties and indicated that carprofen may be effective for a minimum of 48 hr in most animals. With its long half-life further indicating that a single dose could be effective for several days, we suggest that carprofen may be a useful drug for the treatment of white rhinoceros.

Effect of dronedarone on the pharmacokinetics of carvedilol following oral administration to rats.

Dronedarone is a CYP2D6 inhibitor; therefore, it is prudent to exercise caution when concurrently administering CYP2D6-metabolized ß-blockers because of a lack of published data on potential drug interactions. The aim of this study was to investigate the effect of dronedarone on the pharmacokinetics of orally administered carvedilol in rats. Twenty male Sprague-Dawley rats were randomly divided into two groups and 10mg/kg carvedilol was administered to the rat with or without dronedarone pretreatment in a parallel design. Blood samples were collected before and after 0.25, 0.5, 0.75, 1, 2, 4, 6, 8, 12, and 24h of drug administration. The plasma concentration of carvedilol was determined using LC-MS/MS. The systemic exposure to carvedilol was significantly increased and elimination of carvedilol was significantly decreased in the dronedarone-pretreated rats than in the vehicle-pretreated rats. The one-compartment model with first-order absorption and elimination was sufficient to explain the pharmacokinetic characters after single oral administration of carvedilol to both vehicle-pretreated and dronedarone-pretreated rats. This study suggests that dronedarone inhibits CYP2D6-mediated carvedilol metabolism, and dose adjustment is needed in carvedilol and dronedarone combination therapy. Further studies are needed to clarify the effect of dronedarone on carvedilol and CYP2D6 substrates in clinical use.

Comparative study on stabilizing ability of food protein, non-ionic surfactant and anionic surfactant on BCS type II drug carvedilol loaded nanosuspension: Physicochemical and pharmacokinetic investigation.

Carvedilol (CAR) in its pure state has low aqueous solubility and extremely poor bioavailability which largely limit its clinical application. The aim of the study is to improve the dissolution rate and the bioavailability of CAR via preparing nanosuspensions with different stabilizers. Antisolvent precipitation-ultrasonication technique was used here. Attempts have been made to use food protein- Whey protein isolate (WPI) as a stabilizer in CAR loaded nanosuspension and also to compare its stabilizing potential with conventional nanosuspension stabilizers such as non-ionic linear copolymer-poloxamer 188 (PLX188) and anionic surfactant-sodium dodecyl sulfate (SDS). Optimized nanosuspensions showed narrow size distribution with particle size ranging from 275 to 640nm. Amorphous state of CAR nanocrystals which also improved the solubility by 16-, 25-, 55-fold accordingly was confirmed by powder X-ray diffraction (PXRD) and differential scanning calorimetry (DSC). From scanning electron microscopy (SEM), flaky shape of PLX188 and SDS nanosuspensions could be revealed but WPI nanosuspension was sphere-shaped. Up to 70% dissolution of loaded drug was observed within 15min in phosphate buffer (pH6.8). A pharmacokinetic study in rats indicated that both Cmax and AUC0-36 values of nanosuspensions were estimated to be 2-fold higher than those of reference, suggesting a significant increase in CAR bioavailability.

Development of liquid chromatographic methods for enantioseparation and sensitive detection of ß-adrenolytics/ß2-agonists in human plasma using a single enantiomer reagent.

Enantioseparation of four commonly used ß-adrenolytics (bisoprolol, salbutamol, and carvedilol, marketed as racemic mixtures) has been achieved by both TLC and RPHPLC via an indirect approach. A new chiral reagent, (S)-naproxen benzotriazole ester, was synthesized and it was characterized by UV, IR, 1HNMR, elemental analysis and polarimetry. It was used to synthesize diastereomeric derivatives of the three ß-adrenolytics under microwave irradiation. TLC separation of diastereomeric derivatives was achieved which were then isolated by preparative approach; these were characterized and were used as standard reference for determining absolute configuration of diastereomeric derivatives and for establishing validated HPLC method for enantioseparation and sensitive detection of the three ß-adrenolytics in human plasma. Mobile phase in gradient mode containing methanol and aqueous triethylaminephosphate (TEAP) was successful for HPLC separation; conditions with respect to pH, flow rate, and buffer concentration were optimized. The method is capable to accurately quantitate ß-adrenolytics in human plasma with minimal sample clean-up and rapid separation by TLC and RPHPLC. The limit of detection values were 0.97 and 0.87ng/mL for diastereomeric derivatives of (S)- and (R)-bisoprolol, respectively, which are very low in comparison to literature reports.

Alectinib versus crizotinib in patients with ALK-positive non-small-cell lung cancer (J-ALEX): an open-label, randomised phase 3 trial.

BACKGROUND: Alectinib, a potent, highly selective, CNS-active inhibitor of anaplastic lymphoma kinase (ALK), showed promising efficacy and tolerability in the single-arm phase 1/2 AF-001JP trial in Japanese patients with ALK-positive non-small-cell lung cancer. Given those promising results, we did a phase 3 trial to directly compare the efficacy and safety of alectinib and crizotinib. METHODS: J-ALEX was a randomised, open-label, phase 3 trial that recruited ALK inhibitor-naive Japanese patients with ALK-positive non-small-cell lung cancer, who were chemotherapy-naive or had received one previous chemotherapy regimen, from 41 study sites in Japan. Patients were randomly assigned (1:1) via an interactive web response system using a permuted-block method stratified by Eastern Cooperative Oncology Group performance status, treatment line, and disease stage to receive oral alectinib 300 mg twice daily or crizotinib 250 mg twice daily until progressive disease, unacceptable toxicity, death, or withdrawal. The primary endpoint was progression-free survival assessed by an independent review facility. The efficacy analysis was done in the intention-to-treat population, and safety analyses were done in all patients who received at least one dose of the study drug. The study is ongoing and patient recruitment is closed. This study is registered with the Japan Pharmaceutical Information Center (number JapicCTI-132316). FINDINGS: Between Nov 18, 2013, and Aug 4, 2015, 207 patients were recruited and assigned to the alectinib (n=103) or crizotinib (n=104) groups. At data cutoff for the second interim analysis, 24 patients in the alectinib group had discontinued treatment compared with 61 in the crizotinib group, mostly due to lack of efficacy or adverse events. At the second interim analysis (data cutoff date Dec 3, 2015), an independent data monitoring committee determined that the primary endpoint of the study had been met (hazard ratio 0·34 [99·7% CI 0·17-0·71], stratified log-rank p<0·0001) and recommended an immediate release of the data. Median progression-free survival had not yet been reached with alectinib (95% CI 20·3-not estimated) and was 10·2 months (8·2-12·0) with crizotinib. Grade 3 or 4 adverse events occurred at a greater frequency with crizotinib (54 [52%] of 104) than alectinib (27 [26%] of 103). Dose interruptions due to adverse events were also more prevalent with crizotinib (77 [74%] of 104) than with alectinib (30 [29%] of 103), and more patients receiving crizotinib (21 [20%]) than alectinib (nine [9%]) discontinued the study drug because of an adverse event. No adverse events with a fatal outcome occurred in either treatment group. INTERPRETATION: These results provide the first head-to-head comparison of alectinib and crizotinib and have the potential to change the standard of care for the first-line treatment of ALK-positive non-small-cell lung cancer. The dose of alectinib (300 mg twice daily) used in this study is lower than the approved dose in countries other than Japan; however, this limitation is being addressed in the ongoing ALEX study. FUNDING: Chugai Pharmaceutical Co, Ltd.

Effects of Type 2 Diabetes Mellitus in Patients on Treatment With Glibenclamide and Metformin on Carvedilol Enantiomers Metabolism.

Carvedilol is available in clinical practice as a racemate in which (S)-(-)-carvedilol is a ß- and α1 -adrenergic antagonist and (R)-(+)-carvedilol is only an α1 -adrenergic antagonist. Carvedilol is mainly metabolized by glucuronidation, by CYP2D6 to hydroxyphenylcarvedilol (OHC), and by CYP2C9 to O-desmethylcarvedilol (DMC). This study evaluated the pharmacokinetics of carvedilol enantiomers and their metabolites OHC and DMC in healthy volunteers (n = 13) and in type 2 diabetes mellitus patients with good glycemic control (n = 13). The healthy subjects were enrolled to receive either a 25-mg oral single dose of carvedilol alone (no DDI) or carvedilol simultaneously with 5 mg glibenclamide and 500 mg metformin (DDI), whereas type 2 diabetes mellitus patients who were on long-term treatment with glibenclamide (5 mg/8 h) and metformin (500 mg/8 h) were enrolled to receive only a single oral dose of 25 mg carvedilol. The plasma concentrations of the (R)-(+)-carvedilol, (R)-(+)-DMC, and (R)-(+)-OHC were higher than those of (S)-(-)-carvedilol, (S)-(-)-DMC, and (S)-(-)-OHC in all investigated groups. The pharmacokinetics of the carvedilol enantiomers did not differ between the groups. However, the AUC values of the DMC enantiomers were lower in the type 2 diabetes mellitus patients than in the healthy volunteers (DDI and no DDI) [(R)-(+), 6.9, 10.4, 11.9 ng·h/mL; and (S)-(-), 2.4, 4.3, 4.0 ng·h/mL, respectively]. In contrast, the AUC values of the OHC enantiomers were higher in the type 2 diabetes mellitus patients [(R)-(+), 13.9, 6.6, 4.9 ng·h/mL; and (S)-(-), 7.2, 1.5, 1.5 ng·h/mL], which explains the fact that the carvedilol pharmacokinetics was unchanged.

An UPLC-MS/MS method for the quantitation of alectinib in rat plasma.

Currently, crizotinib is the first generation drug, which has been used in the treatment of ALK-rearranged non-small cell lung cancer (NSCLC). However, more and more patients are found in crizotinib-resistance. In the last year, alectinib has been approved for treatment of patients with crizotinib-resistance. In this study, we aim to develop and validate a simple, rapid and sensitive tandem mass spectrometry (UHPLC-MS/MS) method for determination of alectinib in rat plasma. Diazepam was chosen as an internal standard (IS). Protein precipitation by acetonitrile was utilized to prepare plasma samples. Chromatographic separation was achieved on a RRHD Eclipse Plus C18 (2.1×50mm, 1.8µ) column with a gradient mobile phase consisting of acetonitrile and water (containing 0.1% formic acid). The analytes were detected by an electrospray ionization (ESI) source in positive mode. A dynamic multiple reaction monitoring (MRM) method was developed to detect specific precursor and product ions. The target fragment ions were m/z 483.2→396.1 for alectinib and m/z 285.0→192.9 for diazepam (IS). Linear calibration plots were achieved in the range of 1-500ng/ml for alectinib (R2=0.997) in rat plasma. Mean recoveries of alectinib in rat plasma ranged from 84.2% to 92.2%. The intra- and inter-day precision was below 9.3% and accuracy was from -1.4% to 12.1%. No obvious matrix effect was found. This method shows a good performance: accuracy, precision and stability. It has been fully validated and successfully applied to pharmacokinetic study of alectinib.

Simultaneous quantification of vortioxetine, carvedilol and its active metabolite 4-hydroxyphenyl carvedilol in rat plasma by UPLC-MS/MS: Application to their pharmacokinetic interaction study.

To establish a rapid and sensitive ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the determination of vortioxetine, carvedilol and its metabolite 4-hydroxyphenyl carvedilol in rat plasma. The analytes and the internal standard (diazepam) were separated on an Acquity UPLC BEH C18 chromatography column (2.1mm×50mm, 1.7µm) using gradient elution with a mobile phase of acetonitrile and 0.1% formic acid in water at a flow rate of 0.4mL/min. The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) mode to monitor the precursor-to-product ion transitions of m/z 299.2→150.1 for vortioxetine, m/z 407.2→100.3 for carvedilol, m/z 423.2→100.1 for 4-hydroxyphenyl carvedilol and m/z 285.2→193.1 for diazepam (IS) using a positive electrospray ionization interface. The method was validated over a concentration range of 0.5-100ng/mL for vortioxetine, 0.5-1000ng/mL for carvedilol and 0.1-50ng/mL for 4-hydroxyphenyl carvedilol. Total time for each chromatograph was 3.0min. The intra- and inter-day precision and accuracy of the quality control samples at low, medium, and high concentration levels exhibited relative standard deviations (RSD)<11.6% and the accuracy values ranged from -12.2% to 11.3%. The analytical method was successfully applied to a pharmacokinetic interaction study of vortioxetine and carvedilol after oral administration vortioxetine and carvedilol in rats. Results suggested that the co-administration of vortioxetine and carvedilol results in a significant drug interaction in rats.

Flutriciclamide (18F-GE180) PET: First-in-Human PET Study of Novel Third-Generation In Vivo Marker of Human Translocator Protein.

Neuroinflammation is associated with neurodegenerative disease. PET radioligands targeting the 18-kDa translocator protein (TSPO) have been used as in vivo markers of neuroinflammation, but there is an urgent need for novel probes with improved signal-to-noise ratio. Flutriciclamide (18F-GE180) is a recently developed third-generation TSPO ligand. In this first study, we evaluated the optimum scan duration and kinetic modeling strategies for 18F-GE180 PET in (older) healthy controls. METHODS: Ten healthy controls, 6 TSPO high-affinity binders, and 4 mixed-affinity binders were recruited. All subjects underwent detailed neuropsychologic tests, MRI, and a 210-min 18F-GE180 dynamic PET/CT scan using metabolite-corrected arterial plasma input function. We evaluated 5 different kinetic models: irreversible and reversible 2-tissue-compartment models, a reversible 1-tissue model, and 2 models with an extra irreversible vascular compartment. The minimal scan duration was established using 210-min scan data. The feasibility of generating parametric maps was also investigated using graphical analysis. RESULTS: 18F-GE180 concentration was higher in plasma than in whole blood during the entire scan duration. The volume of distribution (VT) was 0.17 in high-affinity binders and 0.12 in mixed-affinity binders using the kinetic model. The model that best represented brain 18F-GE180 kinetics across regions was the reversible 2-tissue-compartment model (2TCM4k), and 90 min resulted as the optimum scan length required to obtain stable estimates. Logan graphical analysis with arterial input function gave a VT highly consistent with VT in the kinetic model, which could be used for voxelwise analysis. CONCLUSION: We report for the first time, to our knowledge, the kinetic properties of the novel third-generation TSPO PET ligand 18F-GE180 in humans: 2TCM4k is the optimal method to quantify the brain uptake, 90 min is the optimal scan length, and the Logan approach could be used to generate parametric maps. Although these control subjects have shown relatively low VT, the methodology presented here forms the basis for quantification for future PET studies using 18F-GE180 in different pathologies.

Bioanalysis of alectinib and metabolite M4 in human plasma, cross-validation and impact on PK assessment.

BACKGROUND: Alectinib is a novel anaplastic lymphoma kinase (ALK) inhibitor for treatment of patients with ALK-positive non-small-cell lung cancer who have progressed on or are intolerant to crizotinib. To support clinical development, concentrations of alectinib and metabolite M4 were determined in plasma from patients and healthy subjects. METHODS: LC-MS/MS methods were developed and validated in two different laboratories: Chugai used separate assays for alectinib and M4 in a pivotal Phase I/II study while Roche established a simultaneous assay for both analytes for another pivotal study and all other studies. CONCLUSION: Cross-validation assessment revealed a bias between the two bioanalytical laboratories, which was confirmed with the clinical PK data between both pivotal studies using the different bioanalytical methods.

Sorptive thin film microextraction followed by direct solid state spectrofluorimetry: A simple, rapid and sensitive method for determination of carvedilol in human plasma.

A poly acrylate-ethylene glycol (PA-EG) thin film is introduced for the first time as a novel polar sorbent for sorptive extraction method coupled directly to solid-state spectrofluorimetry without the necessity of a desorption step. The structure, polarity, fluorescence property and extraction performance of the developed thin film were investigated systematically. Carvedilol was used as the model analyte to evaluate the proposed method. The entire procedure involved one-step extraction of carvedilol from plasma using PA-EG thin film sorptive phase without protein precipitation. Extraction variables were studied in order to establish the best experimental conditions. Optimum extraction conditions were the followings: stirring speed of 1000 rpm, pH of 6.8, extraction temperature of 60 °C, and extraction time of 60 min. Under optimal conditions, extraction of carvedilol was carried out in spiked human plasma; and the linear range of calibration curve was 15-300 ng mL(-1) with regression coefficient of 0.998. Limit of detection (LOD) for the method was 4.5 ng mL(-1). The intra- and inter-day accuracy and precision of the proposed method were evaluated in plasma sample spiked with three concentration levels of carvedilol; yielding a recovery of 91-112% and relative standard deviation of less than 8%, respectively. The established procedure was successfully applied for quantification of carvedilol in plasma sample of a volunteer patient. The developed PA-EG thin film sorptive phase followed by solid-state spectrofluorimetric method provides a simple, rapid and sensitive approach for the analysis of carvedilol in human plasma.

Postoperative Analgesia Due to Sustained-Release Buprenorphine, Sustained-Release Meloxicam, and Carprofen Gel in a Model of Incisional Pain in Rats (Rattus norvegicus).

Postoperative analgesia in laboratory rats is complicated by the frequent handling associated with common analgesic dosing requirements. Here, we evaluated sustained-release buprenorphine (Bup-SR), sustained-release meloxicam (Melox-SR), and carprofen gel (CG) as refinements for postoperative analgesia. The aim of this study was to investigate whether postoperative administration of Bup-SR, Melox-SR, or CG effectively controls behavioral mechanical and thermal hypersensitivity in a rat model of incisional pain. Rats were randomly assigned to 1 of 5 treatment groups: saline, 1 mL/kg SC BID; buprenorphine HCl (Bup HCl), 0.05 mg/kg SC BID; Bup-SR, 1.2 mg/kg SC once; Melox-SR, 4 mg/kg SC once; and CG, 2 oz PO daily. Mechanical and thermal hypersensitivity were tested daily from day-1 through 4. Bup HCl and Bup-SR attenuated mechanical and thermal hypersensitivity on days 1 through 4. Melox-SR and CG attenuated mechanical hypersensitivity-but not thermal hypersensitivity-on days 1 through 4. Plasma concentrations, measured by using UPLC with mass spectrometry, were consistent between both buprenorphine formulations. Gross pathologic examination revealed no signs of toxicity in any group. These findings suggest that postoperative administration of Bup HCl and Bup-SR-but not Melox-SR or CG-effectively attenuates mechanical and thermal hypersensitivity in a rat model of incisional pain.