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Prospective antiviral molecule (2E)-N-methyl-2-[(4-oxo-4H-chromen-3-yl)methylidene]-hydrazinecarbothioamide has been probed using Fourier transform infrared (FTIR), FT-Raman and quantum chemical computations. The geometry equilibrium and natural bond orbital analysis have been carried out with density functional theory employing Becke, 3-parameter, Lee-Yang-Parr method with the 6-311G++(d,p) basis set. The vibrational assignments pertaining to different modes of vibrations have been augmented by normal coordinate analysis, force constant and potential energy distributions. Drug likeness and oral activity have been carried out based on Lipinski's rule of five. The inhibiting potency of 2(2E)-methyl-2-[(4-oxo-4H-chromen-3-yl)methylidene]-hydrazinecarbothioamide has been investigated by docking simulation against SARS-CoV-2 protein. The optimized geometry shows a planar structure between the chromone and the side chain. Differences in the geometries due to the substitution of the electronegative atom and intermolecular contacts due to the chromone and hydrazinecarbothioamide were analyzed. NBO analysis confirms the presence of two strong stable hydrogen bonded NH⋯O intermolecular interactions and two weak hydrogen bonded CH⋯O interactions. The red shift in NH stretching frequency exposed from IR substantiates the formation of NH⋯O intermolecular hydrogen bond and the blue shift in CH stretching frequency substantiates the formation of CH⋯O intermolecular hydrogen bond. Drug likeness, absorption, distribution, metabolism, excretion and toxicity property gives an idea about the pharmacokinetic properties of the title molecule. The binding energy of the nonbonding interaction with Histidine 41 and Cysteine 145, present a clear view that 2(2E)-methyl-2-[(4-oxo-4H-chromen-3-yl)methylidene]-hydrazinecarbothioamide can irreversibly interact with SARS-CoV-2 protease., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020. Published by Elsevier B.V.)

Objective: We developed and validated a simple, convenient and reproducible method for simultaneous estimation of six proton-pump inhibitors (PPIs), omeprazole (OPZ), esomeprazole (EOPZ), lansoprazole (LPZ), pantoprazole (PPZ), rabeprazole (RPZ) and ilaprazole (IPZ) in pharmaceutical dosage forms by a single marker. Meanwhile, the stability of the cited PPIs in 0.9% sodium chloride injection stored in polypropylene syringes up to 48 hours for continuous infusion use was investigated., Materials and Methods: The chromatographic separation was achieved on an InterSustain ® C 18 column (150 × 4.6 mm, 5 μm). The isocratic mobile phase made up of 0.05 M potassium dihydrogen phosphate buffer (pH 4.0): acetonitrile (65:35, v/v) was pumped through the column at a temperature maintained at 30°C and a flow rate of 1.0 mL/min. The relative retention time, UV spectral similarity and relative correction factors between OPZ and the other five PPIs were calculated and investigated using the quantitative analysis of multi-components with a single marker (QAMS) method. The stability study examined physical parameters, pH values and drug concentrations of the PPIs mixtures., Results: Under these conditions, all cited PPIs were separated simultaneously at a retention time of 6.0, 7.3, 7.3, 9.9, 12.5 and 13.9 min for RPZ, OPZ, EOPZ, IPZ, PPZ and LPZ, respectively, with a total run time less than 20.0 min. Comparative analysis results indicated that there were no significant differences observed between the QAMS method and the external standard method. The percentage of initial concentration of each PPI gradually decreased during the storage time., Conclusion: The proposed method, which is selective, economical and accurate, was applied successfully for determination of the cited PPIs in their respective pharmaceutical dosage forms. Admixtures of OPZ, EOPZ, PPZ, IPZ in 0.9% sodium chloride injection were stable for 24 hours and LPZ, RPZ in 0.9% sodium chloride injection were stable for 8 hours in polypropylene syringes., Competing Interests: The authors declare that there is no conflict of interests regarding the publication of this paper., (© 2020 Chen et al.)

The present Special Issue is focused on developing and applying several sensors, biosensor devices, and actuators for the analysis of drugs, foods, and nutraceuticals. Some applications concern classical topics, such as clostridium determination in dairy products, flavouring material in foods like ethylvanillin, or the antioxidant properties of fruit juices, while other applications are more innovative, such as food safety analysis, artificial human senses (electronic nose, or tongue) development, or ethanol determination in pharmaceutical drugs, or forensic purposes using catalytic fuel cell; and lastly, new studies devoted to intelligent food packaging. Therefore, this Special Issue should interest both specialists in the sector and readers who are simply curious, or are simply interested in innovations in the field of food and drug analysis.

The emergence of Mycobacterium tuberculosis strains resistant to current first-line antibiotic regimens constitutes a major global health threat. New treatments against multidrug-resistant tuberculosis (MDR-TB) are thus eagerly needed in particular in countries with a high MDR-TB prevalence. In this context, macozinone (PBTZ169), a promising drug candidate with an unique mode of action and highly potent in vitro tuberculocidal properties against MDR Mycobacterium strains, has now reached the clinical phase and has been notably tested in healthy male volunteers in Switzerland. To that endeavor, a multiplex UHPLC-MS/MS method has been developed for the sensitive and accurate human plasma levels determination of PBTZ169 along with five metabolites retaining in vitro anti-TB activity. Plasma protein precipitation with methanol was carried out as a simplified sample clean-up procedure followed by direct injection of the undiluted supernatant for the bioanalysis of the six analytes within 5 min, using 1.8 μm reversed-phase chromatography coupled to triple quadrupole mass spectrometry employing electrospray ionization in the positive mode. Stable isotopically-labelled PBTZ169 was used as internal standard (ISTD), while metabolites could be reliably quantified using two unlabeled chemical analogues selected as ISTD from a large in-house analogous compounds library. The overall methodology was fully validated according to current recommendations (FDA, EMEA) for bioanalytical methods, which include selectivity, carryover, qualitative and quantitative matrix effect, extraction recovery, process efficiency, trueness, precision, accuracy profiles, method and instrument detection limits, integrity to dilution, anticoagulant comparison and short- and long-term stabilities. Stability studies on the reduced metabolite H2-PBTZ169 have shown no significant impact on the actual PBTZ169 concentrations determined with the proposed assay. This simplified, rapid, sensitive and robust methodology has been applied to the bioanalysis of human plasma samples collected within the frame of a phase I clinical study in healthy volunteers receiving PBTZ169., Competing Interests: The authors have declared that no competing interests exist.

Nilotinib is a broad-based tyrosine kinase inhibitor with the highest affinity to inhibit Abelson (c-Abl) and discoidin domain receptors (DDR1/2). Preclinical evidence indicates that Nilotinib reduces the level of brain alpha-synuclein and attenuates inflammation in models of Parkinson's disease (PD). We previously showed that Nilotinib penetrates the blood-brain barrier (BBB) and potentially improves clinical outcomes in individuals with PD and dementia with Lewy bodies (DLB). We performed a physiologically based population pharmacokinetic/pharmacodynamic (popPK/PD) study to determine the effects of Nilotinib in a cohort of 75 PD participants. Participants were randomized (1:1:1:1:1) into five groups (n = 15) and received open-label random single dose (RSD) 150:200:300:400 mg Nilotinib vs placebo. Plasma and cerebrospinal fluid (CSF) were collected at 1, 2, 3, and 4 hours after Nilotinib administration. The results show that Nilotinib enters the brain in a dose-independent manner and 200 mg Nilotinib increases the level of 3,4-Dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), suggesting alteration to dopamine metabolism. Nilotinib significantly reduces plasma total alpha-synuclein and appears to reduce CSF oligomeric: total alpha-synuclein ratio. Furthermore, Nilotinib significantly increases the CSF level of triggering receptors on myeloid cells (TREM)-2, suggesting an anti-inflammatory effect. Taken together, 200 mg Nilotinib appears to be an optimal single dose that concurrently reduces inflammation and engages surrogate disease biomarkers, including dopamine metabolism and alpha-synuclein., Competing Interests: Charbel Moussa is an inventor of several U.S. and International Georgetown University patents to use Nilotinib and other tyrosine kinase inhibitors as a treatment for neurodegenerative diseases. No other authors declare any conflict of interests with this study.

Bioanalysis plays a key role during the drug discovery process to generate the pharmacokinetic data to facilitate unbiased evaluation of leads, optimized leads and drug candidates. Such pharmacokinetic data are used to enable key decisions in the drug discovery process. The aim of the work is to put forward a new strategy of performing the incurred sample reanalysis for select small molecule novel chemical entities at different stages of drug discovery prior to candidate selection. Three discovery programs representing hits, leads and optimized lead candidates were selected for the incurred sample reanalysis (ISR) analysis. From each discovery program, two novel chemical entities were selected for the ISR analysis. The time points considered for ISR generally varied among the programs; however, samples coinciding with drug absorption, distribution and elimination were considered in the ISR assessment. With the exception of a single ISR value that gave a high deviation (about 63%), the observed ISR values supported the discovery bioanalytical assays. While the individual bioanalytical laboratory can draw an algorithm for selecting novel chemical entities and fixing the acceptance criteria for the ISR data, it is proposed that the percentage difference between ISR vs. original concentration for 67% of the repeat samples is contained within ±30% for discovery bioanalysis., (© 2018 John Wiley & Sons, Ltd.)

This manuscript aims to provide insights and updates on emerging technologies from a throughput and multiplexing perspective and to update readers on changes in previously reported technologies. The technologies discussed range from nascent (ultrasensitive Cira, Intellicyt ® , Dynaxi and Captsure™) to the more established (Ella and SQIDlite™). For the nascent technologies, there was an emphasis on user interviews and reviews, where available, to help provide an unbiased view to our readers. For the Ella, a review of published user data as well as author and other user experiences are summarized. Due to their emergent nature, all the technologies described are applicable in the early drug development stage, may require an upfront investment of capital and may not perform as expected.

In this work, for the first time, a method has been developed for the determination of AZD6118, a candidate drug, in dog plasma samples. The method is based on microextraction by packed sorbent (MEPS) of the drug prior to liquid chromatography-electrospray ionization tandem mass spectrometry assay. Various important factors affecting MEPS performance were optimized, and under the optimized condition, a linear calibration curve in the concentration range of 20-25,000nmolL -1 with a coefficient of determination over 0.99 was obtained. The back-calculated values of the calibration points showed good agreement with the theoretical concentrations (coefficients of variation percent between 0.3-3.8). The lower limit of quantification and limit of detection were 20.0 and 2.9nmolL -1 , respectively. The repeatability and accuracy of the method was evaluated by determination of quality control samples at three concentration levels (low, medium and high) using the developed method, and the results (coefficients of variation values were between 1.9% and 3.2%, relative recoveries ranged between 93.5-102.1%) confirm that a powerful method has been developed for the extraction and determination of the investigated drug in dog plasma., (Copyright © 2016 Elsevier B.V. All rights reserved.)

AZD5423 is a selective glucocorticosteroid receptor modulator developed for the inhaled use in asthma and COPD. This study reports the initial, first-in-man, single and repeat dose-escalating studies in healthy male individuals, including one cohort of male Japanese individuals. Inhaled, nebulized AZD5423 was safe and well tolerated up to and including the highest doses tested for up to 2 weeks of once-daily treatment. Plasma exposure suggested dose-proportional pharmacokinetics and dose-related effects on 24-hr plasma and urine cortisol. There were no or marginal effects on other biomarkers tested (osteocalcin, TRAP5b, DHEA-S and 4β-OH-cholesterol). No clinically relevant differences in safety or pharmacokinetics could be distinguished between the two study populations, although hypothalamus-pituitary-adrenal (HPA) effects appeared to be marginally greater in the Japanese- versus the Caucasian-dominant study population. AZD5423, inhaled via nebulization, can be used in healthy individuals at doses of at least 300 μg for 2 weeks. The effects on the HPA axis reported herein, together with efficacy data reported elsewhere, indicate that benefit-risk ratio may be improved relative to conventional inhaled steroids., (© 2016 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).)

Purpose: Whether microdosing studies can be used to evaluate the human pharmacokinetics of new anticancer drugs remains unclear. The disposition of docetaxel in cancer patients is linear in terms of dose proportionality. We examined whether the pharmacokinetics of docetaxel in a clinically relevant therapeutic dose could be predicted from the pharmacokinetics of a microdose of docetaxel in Japanese patients with cancer., Methods: A microdose of docetaxel (100 μg/patient) was given by 5-min intravenous infusion on day 1, followed by a therapeutic dose of docetaxel (60-75 mg m(-2)), given by 1-h intravenous infusion on day 8. Plasma docetaxel was analyzed by liquid chromatography-tandem mass spectrometry. A two-compartment pharmacokinetic model was used to calculate the area under the plasma concentration-time curve from time 0 to infinity (AUC0-inf)., Results: Nine patients received both a microdose and therapeutic dose of docetaxel. The AUC0-inf after microdosing was 3640 ± 1150 ng h L(-1), while that after therapeutic dosing adjusted to 100 mg/patient was 2230 ± 757 µg h L(-1). The ratio of docetaxel clearance in therapeutic dose to that in microdose was 1.8 (P = 0.0041). Plasma α1-acid glycoprotein concentrations negatively correlated with docetaxel clearance at therapeutic dose, whereas the trend was weak at microdose., Conclusion: Docetaxel clearance showed marginal nonlinearity between microdose and therapeutic dose, presumably because of saturation of plasma protein binding; however, the magnitude was within twofold, allowing practically acceptable extrapolation.

PF-05231023, a long-acting fibroblast growth factor 21 (FGF21) analog, was generated by covalently conjugating two engineered [des-His1, Ala129Cys]FGF21 molecules to a nontargeting human IgG1 κ scaffold. The pharmacokinetics (PK) of PF-05231023 after i.v. and s.c. administration was evaluated in rats and monkeys using two enzyme-linked immunosorbent assays with high specificity for biologically relevant intact N termini (NT) and C termini (CT) of FGF21. Intact CT of FGF21 displayed approximately 5-fold faster systemic plasma clearance (CL), an approximately 2-fold lower steady-state volume of distribution, and at least 5-fold lower bioavailability compared with NT. In vitro serum stability studies in monkeys and humans suggested that the principal CL mechanism for PF-05231023 was degradation by serum proteases. Direct scaling of in vitro serum degradation rates for intact CT of FGF21 underestimated in vivo CL 5-fold, 1.4-fold, and 2-fold in rats, monkeys, and humans, respectively. The reduced steady-state volume of distribution and the bioavailability for intact CT relative to NT in rats and monkeys were compatible with proteolytic degradation occurring outside the plasma compartment via an unidentified mechanism. Human CL and PK profiles for intact NT and CT of FGF21 were well predicted using monkey single-species allometric and Dedrick scaling. Physiologically based pharmacokinetic models incorporating serum stability data and an extravascular extraction term based on differential bioavailability of intact NT and CT of FGF21 in monkeys improved accuracy of human PK predictions relative to Dedrick scaling. Mechanistic physiologically based pharmacokinetic models of this nature may be highly valuable for predicting human PK of fusion proteins, synthetically conjugated proteins, and other complex biologics., (Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.)

GDC-0834, a Bruton's tyrosine kinase inhibitor investigated as a potential treatment of rheumatoid arthritis, was previously reported to be extensively metabolized by amide hydrolysis such that no measurable levels of this compound were detected in human circulation after oral administration. In vitro studies in human liver cytosol determined that GDC-0834 (R)-N-(3-(6-(4-(1,4-dimethyl-3-oxopiperazin-2-yl)phenylamino)-4-methyl-5-oxo- 4,5-dihydropyrazin-2-yl)-2-methylphenyl)-4,5,6,7-tetrahydrobenzo[b] thiophene-2-carboxamide) was rapidly hydrolyzed with a CLint of 0.511 ml/min per milligram of protein. Aldehyde oxidase (AO) and carboxylesterase (CES) were putatively identified as the enzymes responsible after cytosolic fractionation and mass spectrometry-proteomics analysis of the enzymatically active fractions. Results were confirmed by a series of kinetic experiments with inhibitors of AO, CES, and xanthine oxidase (XO), which implicated AO and CES, but not XO, as mediating GDC-0834 amide hydrolysis. Further supporting the interaction between GDC-0834 and AO, GDC-0834 was shown to be a potent reversible inhibitor of six known AO substrates with IC50 values ranging from 0.86 to 1.87 μM. Additionally, in silico modeling studies suggest that GDC-0834 is capable of binding in the active site of AO with the amide bond of GDC-0834 near the molybdenum cofactor (MoCo), orientated in such a way to enable potential nucleophilic attack on the carbonyl of the amide bond by the hydroxyl of MoCo. Together, the in vitro and in silico results suggest the involvement of AO in the amide hydrolysis of GDC-0834., (Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.)

Body surface area (BSA) scaling has been used for prescribing individualized dosages of various drugs and has also been recommended by the U.S. Food and Drug Administration as one method for using data from animal model species to establish safe starting dosages for first-in-human clinical trials. Although BSA conversion equations have been used in certain clinical applications for decades, recent recommendations to use BSA to derive interspecies equivalents for therapeutic dosages of drug and natural products are inappropriate. A thorough review of the literature reveals that BSA conversions are based on antiquated science and have little justification in current translational medicine compared to more advanced allometric and physiologically based pharmacokinetic modeling. Misunderstood and misinterpreted use of BSA conversions may have disastrous consequences, including underdosing leading to abandonment of potentially efficacious investigational drugs, and unexpected deadly adverse events. We aim to demonstrate that recent recommendations for BSA are not appropriate for animal-to-human dosage conversions and use pharmacokinetic data from resveratrol studies to demonstrate how confusion between the "human equivalent dose" and "pharmacologically active dose" can lead to inappropriate dose recommendations. To optimize drug development, future recommendations for interspecies scaling must be scientifically justified using physiologic, pharmacokinetic, and toxicology data rather than simple BSA conversion., (© FASEB.)

Sorafenib (Nexavar®) is currently the only FDA-approved small molecule targeted therapy for advanced hepatocellular carcinoma. The use of structural analogues and derivatives of sorafenib has enabled the elucidation of critical targets and mechanism(s) of cell death for human cancer lines. We previously performed a structure-activity relationship study on a series of sorafenib analogues designed to investigate the inhibition overlap between the major targets of sorafenib Raf-1 kinase and VEGFR-2, and an enzyme shown to be a potent off-target of sorafenib, soluble epoxide hydrolase. In the current work, we present the biological data on our lead sorafenib analogue, t-CUPM, demonstrating that this analogue retains cytotoxicity similar to sorafenib in various human cancer cell lines and strongly inhibits growth in the NCI-60 cell line panel. Co-treatment with the pan-caspase inhibitor, Z-VAD-FMK, failed to rescue the cell viability responses of both sorafenib and t-CUPM, and immunofluorescence microscopy shows similar mitochondrial depolarization and apoptosis-inducing factor release for both compounds. These data suggest that both compounds induce a similar mechanism of caspase-independent apoptosis in hepatoma cells. In addition, t-CUPM displays anti-proliferative effects comparable to sorafenib as seen by a halt in G0/G1 in cell cycle progression. The structural difference between sorafenib and t-CUPM significantly reduces inhibitory spectrum of kinases by this analogue, and pharmacokinetic characterization demonstrates a 20-fold better oral bioavailability of t-CUPM than sorafenib in mice. Thus, t-CUPM may have the potential to reduce the adverse events observed from the multikinase inhibitory properties and the large dosing regimens of sorafenib.

Purpose: To develop a population pharmacokinetic (PK) model for HM781-36 (poziotinib) and its metabolites in cancer patients., Methods: Blood samples were collected from three phase I studies in which fifty-two patients received oral HM781-36B tablets (0.5-32 mg) once daily for 2 weeks, and another 20 patients received oral HM781-36B tablets (12, 16, 18, 24 mg) in fasting (12 patients) or fed (eight patients) state once daily for 4 weeks. Nonlinear mixed effect modeling was employed to develop the population pharmacokinetic model., Results: HM781-36 PK was ascribed to a two-compartment model and HM781-36-M1/-M2 PK to one-compartment model. HM781-36 oral absorption was characterized by first-order input (absorption rate constant: 1.45 ± 0.23 h⁻¹). The central volume of distribution (185 ± 12.7 L) was influenced significantly by body weight. The absorption rate constant was influenced by food. The typical HM781-36 apparent clearance was 34.5 L/h (29.4 %CV), with an apparent peripheral volume of distribution of 164 L (53.5 %CV). Other covariates did not significantly further explain the PKs of HM781-36., Conclusions: The proposed model suggests that HM781-36 PKs are consistent across most solid tumor types, and that the absorption process of HM781-36 is affected by the fed state before dosing. HM781-36 PKs are not complicated by patient factors, other than body weight.

In this paper, an ex vivo model for the study of the transcorneal permeation of drugs, based on porcine tissues, was evaluated. The setup is characterized by ease of realization, absence of O₂ and CO₂ bubbling and low cost; additionally, the large availability of porcine tissue permits a high throughput. Histological images showed the comparability between porcine and human corneas and confirmed the effectiveness of the isolation procedure. A new de-epithelization procedure based on a thermal approach was also set up to simulate cornea permeability in pathological conditions. The procedure did not affect the integrity of the underlying layers and allowed the characterization of the barrier properties of epithelium and stroma. Six compounds with different physicochemical properties were tested: fluorescein, atenolol, propranolol, diclofenac, ganciclovir and lidocaine. The model highlighted the barrier function played by epithelium toward the diffusion of hydrophilic compounds and the permselectivity with regard to more lipophilic molecules. In particular, positively charged compounds showed a significantly higher transcorneal permeability than negatively charged compounds. The comparability of results with literature data supports the goodness and the robustness of the model, especially taking into account the behavior of fluorescein, which is generally considered a marker of tissue integrity., (© 2014 Wiley Periodicals, Inc. and the American Pharmacists Association.)

The aim of the present work was to design a pH-modified solid dispersion (pH(M)-SD) that can improve the dissolution and bioavailability of poorly water-soluble weakly basic GT0918, a developing anti-prostate cancer drug. To select the appropriate acidifiers, a solubility test was carried out first. Solid dispersions (SDs) containing GT0918 and polyvinylpyrrolidone (PVP) were prepared using a solvent evaporation method and were characterized using dissolution studies in different media. The solid states of the SDs were investigated using scanning electron microscopy (SEM), powder X-ray diffraction (PXRD), differential scanning calorimetry (DSC) and Fourier transformed infrared spectroscopy (FTIR). The in vivo pharmacokinetics of the pH(M)-SDs tablets were also studied in beagle dogs compared to the conventional tablets. The optimized pH(M)-SD (GT0918/PVP/citric acid, 1:2:2 weight ratio) exhibited a significant improvement in the dissolution behavior compared to both the physical mixture and the binary SDs. Solid-state characterization revealed that the amorphous formation of GT0918 in the SDs and the strong H-bonding were only found in the pH(M)-SDs containing citric acid. Furthermore, the GT0918-loaded pH(M)-SD tablets showed a higher AUC and a lower tmax compared to the conventional tablets. Accordingly, the pH(M)-SD might be an efficient route for enhancing the dissolution and bioavailability of poorly water-soluble GT0918., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The prediction of brain extracellular fluid (ECF) concentrations in human is a potentially valuable asset during drug development as it can provide the pharmacokinetic input for pharmacokinetic-pharmacodynamic models. This study aimed to compare two translational modelling approaches that can be applied at the preclinical stage of development in order to simulate human brain ECF concentrations. A population-PBPK model of the central nervous system was developed based on brain microdialysis data, and the model parameters were translated to their corresponding human values to simulate ECF and brain tissue concentration profiles. In parallel, the PBPK modelling software Simcyp was used to simulate human brain tissue concentrations, via the bottom-up prediction of brain tissue distribution using two different sets of mechanistic tissue composition-based equations. The population-PBPK and bottom-up approaches gave similar predictions of total brain concentrations in both rat and human, while only the population-PBPK model was capable of accurately simulating the rat ECF concentrations. The choice of PBPK model must therefore depend on the purpose of the modelling exercise, the in vitro and in vivo data available and knowledge of the mechanisms governing the membrane permeability and distribution of the drug., (Copyright © 2014 John Wiley & Sons, Ltd.)

LY2605541 is a novel basal insulin analog with a prolonged duration of action. Two Phase I studies assessed LY2605541 pharmacokinetics (PK), glucodynamics (GD), and tolerability in healthy subjects. In Study 1, 33 subjects received single subcutaneous (SC) doses of LY2605541 (0.01-2.22 U/kg) and insulin glargine (0.5-0.8 U/kg) followed by euglycemic clamp for up to 24-36 hours. In Study 2, absolute bioavailability of SC LY2605541 was assessed in 8 subjects by comparing dose normalized area under concentration versus time curve of SC against IV administration. Time-to-maximum plasma concentration (medians) and geometric means for half-life (t½ ) and apparent clearance, respectively, ranged from 18.0 to 42.0 hours, 24.4-45.5 hours, and 1.8-2.8 L/h for SC LY2605541, versus 10.0-12.0 hours, 12.2-14.9 hours, and 51.4-65.2 L/h for SC insulin glargine. LY2605541 glucose infusion rate (GIR) profiles were sustained for ≥36 hours versus glargine GIR profiles, which waned at 24 hours. After IV administration, LY2605541's geometric mean t½ was 2.3 hours. LY2605541 intra-subject variability (CV%) was <18% for PK and <32% for GD parameters. The most common adverse events were related to study procedures and were mild-moderate in severity. These results established a well-tolerated baseline dose for LY2605541 with a relatively flat PK profile and low intra-subject variability., (© 2014, The American College of Clinical Pharmacology.)

The identification of the no observed adverse effect level (NOAEL) is the key regulatory outcome of toxicity studies. With the introduction of "omics" technologies into toxicological research, the question arises as to how sensitive these technologies are relative to classical regulatory toxicity parameters. BASF SE and metanomics developed the in vivo metabolome database MetaMap®Tox containing metabolome data for more than 500 reference compounds. For several years metabolome analysis has been routinely performed in regulatory toxicity studies (REACH mandated testing or new compound development), mostly in the context of 28 day studies in rats (OECD 407 guideline). For those chemicals for which a toxicological NOAEL level was obtained at either high or mid-dose level, we evaluated the associated metabolome to investigate the sensitivity of metabolomics versus classical toxicology with respect to the NOAEL. For the definition of a metabolomics NOAEL the ECETOC criteria (ECETOC, 2007) were used. In this context we evaluated 104 cases. Comparable sensitivity was noted in 75% of the cases, increased sensitivity of metabolomics in 8%, and decreased sensitivity in 18% of the cases. In conclusion, these data suggest that metabolomics profiling has a similar sensitivity to the classical toxicological study (e.g. OECD 407) design., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

In the present paper, BA99 and AC88 were used as model compounds for intraperitoneal (i.p.) administration to Sprague-Dawley rats. A major problem for the compounds, like many others newly developed pharmaceutical drugs, is the poor solubility in water. To solve solubility related problems, development of nanosuspensions is an attractive alternative. Both compounds are suitable for nanosuspensions, using the milling approach. After 2 weeks in freezer, the nanoparticles aggregated to form particles in the 400-2000 nm interval. However, following a 20 s ultrasonication step, the original particle sizes (about 200 nm) were obtained. Adding 5% mannitol before the samples were frozen abolished aggregation. It is also possible to freeze-dry the nanosuspension in the presence of 5% mannitol and re-disperse the formulation in water. Nanosuspensions of both compounds were injected i.p. to rats at 5 and 500 µmoL/kg. At the low dose, also a microsuspension was administered. I.p. administration resulted in overall improved C(max) for both AC88 and BA99 compared to s.c. and oral administration. I.p. is the preferred route of administration of tolerable drugs when a fast onset of action is desired and when a significant first passage metabolism occurs. The net charge of the active molecule appeared to affect the absorption kinetics. In the present work, the neutral molecule was favored over the negatively charged one.

Aims: This clinical trial assessed whether a potent, selective GPR109A agonist, GSK256073, could, through inhibition of lipolysis, acutely improve glucose homeostasis in subjects with type 2 diabetes mellitus., Methods: Thirty-nine diabetic subjects were enrolled in the randomized, single-blind, placebo-controlled, three-period crossover trial. Each subject received placebo and two of four regimens of GSK256073 for 2 days. GSK256073 was dosed 5 mg every 12 h before breakfast and supper (BID), 10 mg every 24 h before breakfast (QD), 25 mg BID and 50 mg QD., Results: The change from baseline weighted mean glucose concentration for an interval from 24 to 48 h after the initial drug dose was significantly reduced for all GSK256073 regimens, reaching a maximum of -0.87 mmol/l (-1.20, -0.52) with the 25 mg BID dose. Sustained suppression of non-esterified fatty acid (NEFA) and glycerol concentrations was observed with all GSK256073 doses throughout the 48-h dosing period. Serum insulin and C-peptide concentrations fell in concert with glucose concentrations and calculated HOMA-IR scores decreased 27-47%, consistent with insulin sensitization. No marked differences were evident between either 10 and 50 mg total daily doses or QD versus BID dosing., Conclusions: Administration of a GPR109A agonist for 2 days significantly decreased serum NEFA and glucose concentrations in diabetic subjects. Glucose improvements were associated with decreased insulin concentrations and measures of enhanced insulin sensitivity. Improved glucose control occurred with GSK256073 doses that were generally safe and not associated with events of flushing or gastrointestinal disturbances., (© 2013 John Wiley & Sons Ltd.)

In recent years, the development of microfluidic chip separation devices coupled to MS has dramatically increased for high-throughput bioanalysis. In this review, advances in different types of microfluidic chip separation devices, such as electrophoresis- and LC-based microchips, as well as 2D design of microfluidic chip-based separation devices will be discussed. In addition, the utilization of chip-based separation devices coupled to MS for analyzing peptides/proteins, glycans, drug metabolites and biomarkers for various bioanalytical applications will be evaluated.

Aim: The investigations reported here aimed to evaluate the incremental benefit for dose finding by concentration-response analysis versus dose-response analysis., Methods: Trials were simulated using an Emax model for a range of scenarios of drug properties, trial design options and target response levels. The simulated data were analysed by concentration-response and dose-response modelling; a dose was then chosen to target a specific response level in a confirmatory trial. The two approaches were compared in terms of the quality of model parameter estimation and the success rate for the confirmatory trial., Results: While the accuracy for ED50 estimation was comparably good with both approaches, the precision was up to 90 % higher with concentration-response approach. The difference was most notable when clearance was highly variable between subjects and the top dose was relatively low. The higher precision by the concentration-response analysis lead to better dose selection and up to 20 % higher success rate for the subsequent confirmatory trial. The relatively small difference in success rate translated into a remarkable difference in sample size requirement., Conclusion: By customising these parameters, the approach and the findings can be applied to assessing the value of pharmacokinetic sampling in particular trial situations.

Background: Cyclofem is a combined injectable contraceptive, containing medroxyprogesterone acetate (MPA) and estradiol cypionate. The objective was to characterize the steady-state pharmacokinetics (PK) using tandem liquid chromatography/mass spectrometry and compare these data to a previous PK study of this formulation in US women., Study Design: Fifteen ovulatory, surgically sterile women received three Cyclofem injections, once every 28 days, with serum PK measurements on 23 separate days. Trough levels of estradiol and MPA were obtained on Days 1, 29 and 57, prior to each of the three injections. Steady-state concentrations of MPA and estradiol were assessed during the third treatment month on Days 58, 60, 62, 64, 67, 69, 71, 75, 78 and 85. MPA and serum progesterone levels were measured during the follow-up phase to assess MPA clearance (Days 92, 99, 106, 113, 120, 127, 134 and 141) and return of ovulation (Days 103, 106, 131 and 134)., Results: In the steady state, mean serum MPA concentrations peaked at 1.31 ng/mL at 4.1 days. Mean estradiol levels peaked at 254 pg/mL by 3.3 days. Ovulation was suppressed for at least 77 days post third injection in all but one woman., Conclusions: Once monthly injections of Cyclofem resulted in contraceptive levels of MPA without accumulation of hormones, consistent with a previous US study., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Objective: To examine the electrophysiological, hemodynamic, and antiarrhythmic effects of the novel antiarrhythmic agent AZ13395438., Methods: The ion channel-blocking potency of AZ13395438 was assessed in Chinese hamster ovary cells stably expressing various human cardiac ion channels and in human atrial myocytes. The in vivo electrophysiological, hemodynamic, and antiarrhythmic effects of intravenously administered AZ13395438 were examined in anesthetized rabbits, in anesthetized naive dogs, and in dogs subjected to rapid atrial pacing (RAP) for 8 weeks. Pharmacokinetic/pharmacodynamic (PKPD) modeling was applied to predict the potency of AZ13395438 in increasing atrial and ventricular refractoriness., Results: AZ13395438 potently and predominantly blocked the atrial repolarizing potassium currents I(Kur), I(Ach), and I(to) in vitro. In vivo, AZ13395438 caused a concentration-dependent and selective increase in atrial refractoriness with no or small effects on ventricular refractoriness and repolarization and on hemodynamics in both rabbits and dogs. The PKPD modeling predicted unbound plasma concentrations of AZ13395438 of 0.20 ± 0.039, 0.38 ± 0.084, and 0.34 ± 0.057 µmol/L to increase the right atrial effective refractory period by 20 milliseconds in the rabbit and in the naive and the RAP dogs, respectively. In the RAP dog with atrial fibrillation (AF), AZ13395438 significantly increased AF cycle length and successfully converted AF to sinus rhythm in 12 of the 12 occasions at an unbound plasma concentration of 0.48 ± 0.076 µmol/L. During saline infusion, conversion was seen only in 4 of the 10 occasions (P = .003 vs AZ13395438). Furthermore, AZ13395438 reduced AF inducibility by burst pacing from 100% to 25% (P < .001)., Conclusion: AZ13395438 can be characterized as a mixed potassium ion channel-blocking agent that selectively prolongs atrial versus ventricular refractoriness and shows promising antiarrhythmic efficacy in a clinically relevant animal model of AF.

Within the drug discovery environment, the key process in optimising the chemistry of a structural series toward a potential drug candidate is the design, make and test cycle, in which the primary screens consist of a number of in vitro assays, including metabolic stability, cytochrome P450 inhibition, and time-dependent inhibition assays. These assays are often carried out using multiple drug compounds with chemically diverse structural features, often in a 96 well-plate format for maximum time-efficiency, and are supported using rapid liquid chromatographic (LC) sample introduction with a tandem mass spectrometry (MS/MS) selected reaction monitoring (SRM) endpoint, taking around 6.5 h per plate. To provide a faster time-to-decision at this critical point, there exists a requirement for higher sample throughput and a robust, well-characterized analytical alternative. This paper presents a detailed evaluation of laser diode thermal desorption (LDTD), a relatively new ambient sample ionization technique, for compound screening assays. By systematic modification of typical LDTD instrumentation and workflow, and providing deeper understanding around overcoming a number of key issues, this work establishes LDTD as a practical, rapid alternative to conventional LC-MS/MS in drug discovery, without need for extensive sample preparation or expensive, scope-limiting internal standards. Analysis of both the five and three cytochrome P450 competitive inhibition assay samples by LDTD gave improved sample throughput (0.75 h per plate) and provided comparable data quality as the IC₅₀ values obtained were within 3 fold of those calculated from the LC-MS/MS data. Additionally when applied generically to a chemically diverse library of over 250 proprietary compounds from the AstraZeneca design, make and test cycle, LDTD demonstrated a success rate of 98%., (Copyright © 2011 Elsevier B.V. All rights reserved.)

During nonclinical regulated toxicology studies, blood processing protocol deviations may negatively impact sample integrity and bioanalytical results. Standard practices for resolution of blood processing issues are not well established across the industry. In this article, using an illustrative example, we present a systematic approach to investigate and assess the impact of nonclinical blood processing protocol deviations that involve: assessment of the potential impact of deviations on toxicokinetic evaluation, performance of additional blood processing stability tests to ensure study sample integrity and establishment of study sample results reporting procedures to meet both scientific and regulatory quality requirements. This thorough and systematic approach can serve as a model for other analogous situations.

Accurate prediction of pharmacokinetics (PK) parameters in humans from animal data is difficult for various reasons, including species differences. However, chimeric mice with humanized liver (PXB mice; urokinase-type plasminogen activator/severe combined immunodeficiency mice repopulated with approximately 80% human hepatocytes) have been developed. The expression levels and metabolic activities of cytochrome P450 (P450) and non-P450 enzymes in the livers of PXB mice are similar to those in humans. In this study, we examined the predictability for human PK parameters from data obtained in PXB mice. Elimination of selected drugs involves multiple metabolic pathways mediated not only by P450 but also by non-P450 enzymes, such as UDP-glucuronosyltransferase, sulfotransferase, and aldehyde oxidase in liver. Direct comparison between in vitro intrinsic clearance (CL(int,in vitro)) in PXB mice hepatocytes and in vivo intrinsic clearance (CL(int,in vivo)) in humans, calculated based on a well stirred model, showed a moderate correlation (r² = 0.475, p = 0.009). However, when CL(int,in vivo) values in humans and PXB mice were compared similarly, there was a good correlation (r² = 0.754, p = 1.174 × 10⁻⁴). Elimination half-life (t(1/2)) after intravenous administration also showed a good correlation (r² = 0.886, p = 1.506 × 10⁻⁴) between humans and PXB mice. The rank order of CL and t(1/2) in human could be predicted at least, although it may not be possible to predict absolute values due to rather large prediction errors. Our results indicate that in vitro and in vivo experiments with PXB mice should be useful at least for semiquantitative prediction of the PK characteristics of candidate drugs in humans.

In clinical development, thorough QT interval/corrected QT interval (QT/QTc) studies are performed to demonstrate that new investigational drugs do not change cardiac repolarization. In addition to the analysis recommended by ICH E14, exposure-response modeling has recently gained increased scientific attention. A direct concentration-QTc relationship is usually assumed with this pharmacokinetics (PK) QT analysis. Consequently, unconsidered effect delays might lead to severe bias in estimation. Although literature and the Food and Drug Administration (FDA) recommend checking for hysteresis, little guidance is given for evaluating the presence of lagged effects. Based on simulated delay scenarios, we derive a metric that allows for the detection of relevant effect delays. With this, the necessity for refined modeling of lag times can be easily recognized.

Establishment of in vitro-in vivo correlation (IVIVC) accelerates optimization of desirable drug formulations and/or modification of the manufacturing processes in the scale-up and post-approval periods. This article presents a method of finding the optimal conversion function for establishing Level A point-to-point IVIVC, based on a computer-based evolutionary search technique. Gene expression programming (GEP) is a technique for optimizing a mathematical expression tree with the help of a genetic algorithm. A parameter optimization routine, which minimizes the number of parameters in the mathematical expression trees and estimates the best-fit parameter values, was implemented in the GEP algorithm. Feasibility of the computer program was investigated using the in vitro and in vivo data for sustained release diltiazem formulations. It provided a mathematical equation that, from their in vitro dissolution profiles, successfully predicts the plasma concentration profiles of three different formulations of diltiazem following oral administration. Because the present approach does not use intravenous injection data like conventional IVIVC analyses, it is widely applicable to the evaluation of various oral formulations.

Reliably predicting pharmacokinetic behavior in humans from preclinical data is an important aspect of drug development. The most widely used technique in this regard is allometric scaling. In this review, various approaches developed for predicting pharmacokinetic parameters in humans using interspecies scaling are introduced and discussed. Methods to predict plasma concentration-time profiles in humans after intravenous and oral administration are also reviewed. The reliable prediction of human pharmacokinetics with regard to investigational drugs is aimed, ultimately, at selecting the first in-human dose with which to begin clinical studies. Approaches for the selection of the first in-human dose are also reviewed. Although there have been many trials to compare and optimize interspecies scaling methods, no firm conclusions have been reached. Because interspecies scaling methods are still highly empirical, further effort is needed to improve the reliability of predicting human pharmacokinetics by interspecies scaling.

Innovative attempts have been made to overcome nonproductivity and high expenditure in the clinical stages of new drug development. Microdosing studies using subpharmacological doses provide early insight into the body's disposition toward candidate compounds, and are innovative exploratory trials that can promote productivity in drug development. Highly sensitive analytical technology is crucial in microdosing studies that employ qualitative and quantitative assays of target materials in humans. Accelerator mass spectrometry (AMS) has facilitated the adoption of a human microdosing study in the early phase of clinical drug development. Results derived from AMS microdosing studies using labeled compounds can provide various types of information for candidate selection, including pharmacokinetic characteristics and metabolic profiles of candidate compounds. The applicability of microdosing studies is currently expanding into absolute bioavailability and mass balance studies. Although it remains uncertain whether microdosing adequately predicts the pharmacokinetics of therapeutic doses, further development of microdosing studies using AMS may benefit the field of new drug development and could pose a new challenge to researchers. The use of advanced technology in candidate selection will contribute to improved productivity and competitiveness in pharmaceutical research and development. The introduction of microdosing studies using AMS in Korea will present a newly applicable method for innovative clinical trials and contribute to development potential in global competition.

Direct analysis in real time (DART), a relatively new ionization source for mass spectrometry, ionizes small-molecule components from different kinds of samples without any sample preparation and chromatographic separation. The current paper reviews the published data available on the determination of drugs and drug-like compounds in different matrices with DART-MS, including identification and quantitation issues. Parameters that affect ionization efficiency and mass spectra composition are also discussed., (Copyright © 2011 Wiley Periodicals, Inc.)

Dexpramipexole (KNS-760704; [6R]-4,5,6,7-tetrahydro-N6-propyl-2,6-benzothiazole-diamine) is a novel synthetic amino-benzothiazole in development for the treatment of amyotrophic lateral sclerosis (ALS). Preclinical studies have shown that dexpramipexole is neuroprotective in vitro and in vivo, is highly orally bioavailable and water soluble, and rapidly achieves and maintains high central nervous system concentrations relative to plasma. Two phase 1 clinical studies were conducted to assess the safety, tolerability, and pharmacokinetics (PK) of single and multiple doses of dexpramipexole in 54 healthy male and female adults. The effect of food on the single-dose PK of dexpramipexole was also evaluated. Single doses (50 mg, 150 mg, or 300 mg) and multiple doses (50 mg twice daily, 100 mg twice daily, or 150 mg twice daily) of dexpramipexole over 4.5 days were safe and well tolerated. Dexpramipexole was rapidly absorbed, with time to maximum plasma concentration ranging from 1.8 to 2.6 hours and half-life ranging from 6.4 to 8.1 hours under fasted conditions, and was mostly eliminated in urine as unchanged parent drug (84%-90% of dose). Food had no effect on the single-dose PK of dexpramipexole. These findings support the ongoing development of dexpramipexole for the treatment of ALS and further evaluation of the compound's therapeutic potential in other neurodegenerative diseases.

To increase our understanding of important subject characteristics and design variables affecting the performance of oral moxifloxacin in thorough QT studies, population pharmacokinetic and concentration-QTc models were developed by pooling data from 20 studies. A 1-compartment model with first-order elimination described the pharmacokinetics. Absorption delay was modeled using 8 transit compartments. Mean (95% confidence interval) values for oral clearance, apparent volume of distribution, the first-order absorption rate constant, and mean transit time were 11.7 (11.5-11.9) L/h, 147 (144-150) L, 1.9 (1.7-2.1) 1/h, and 0.3 (0.28-0.34) hours, respectively. Overencapsulating the moxifloxacin tablet increased mean transit time by 138% and delayed time to maximum concentration by 0.5 hours but had a minimal effect on overall exposure. Administration with food decreased absorption rate constant by 27%. Women had higher moxifloxacin exposure compared with men, which was explained by lower body weights. A linear model described the concentration-QTc relationship with a mean slope of 3.1 (2.8-3.3) milliseconds per µg/mL moxifloxacin. Mean slopes for individual studies ranged from 1.6 to 4.8 milliseconds per µg/mL. Hysteresis between moxifloxacin plasma concentrations and QTc was modest, and incorporating this delay did not result in a different slope (3.3 milliseconds per µg/mL). There were no differences in slope estimates between men and women or among race categories.

Pharmacologic induction of fetal globin synthesis is an accepted therapeutic strategy for treatment of the beta hemoglobinopathies and thalassemias, as even small increases in hemoglobin F (HbF) levels reduce clinical severity in sickle cell disease (SCD) and reduce anemia in beta thalassemia. Prior generation short chain fatty acid therapeutics, arginine butyrate (AB), and phenylbutyrate, increased fetal and total hemoglobin levels in patients, but were limited by high doses or intravenous (IV) infusion. A fetal globin-inducing therapeutic with convenient oral dosing would be an advance for these classic molecular diseases. Healthy adult human subjects were treated with a novel short chain fatty acids (SCFA) derivative, sodium 2,2 dimethylbutyrate (SDMB), or placebo, with 1 of 4 single dose levels (2, 5, 10, and 20 mg/kg) or daily doses (5, 10, or 15 mg/kg) over 14 days, and monitored for adverse clinical and laboratory events, drug levels, reticulocytes, and HbF assays. SDMB was well-tolerated with no clinically significant adverse events related to study medication. The terminal half-life ranged from 9 to 15 hours. Increases in mean absolute reticulocytes were observed at all dose levels in the 14-day study. The favorable pharmacokinetics (PK) profiles and safety findings indicate that SDMB warrants further investigation for treatment of anemic subjects with beta hemoglobinopathies.

An increasing number of thorough QT (TQT) reports are being submitted to the US Food and Drug Administration's interdisciplinary review team for QT (IRT-QT), requiring time-intensive quantitative analyses by a multidisciplinary review team within 45 days. This calls for systematic learning to guide future trials and policies by standardizing and automating the QT analyses to improve review efficiency, provide consistent advice, and enable pooled data analyses to answer key regulatory questions. The QT interval represents the time from initiation of ventricular depolarization to completion of ventricular repolarization recorded by electrocardiograph (ECG) and is used in the proarrhythmic risk assessment. The developed QT knowledge management system is implemented in the R package "QT." Data from 11 crossover TQT studies including time-matched ECGs and pharmacokinetic measurements following single doses of 400 to 1200 mg moxifloxacin were used for the QT analysis example. The automated workflow was divided into 3 components (data management, analysis, and archival). The generated data sets, scripts, tables, and graphs are automatically stored in a queryable repository and summarized in an analysis report. More than 100 TQT studies have been analyzed using the system since 2007. This has dramatically reduced the time needed to review TQT studies and has made the IRT-QT reviews consistent across reviewers. Furthermore, the system enables leveraging prior knowledge through pooled data analyses to answer policy-related questions and to understand the various effects that influence study results.

Axitinib (AG-013736) is a potent investigational drug that has antitumor activity in patients with metastatic renal cell carcinoma and other types of cancers. In this study, ion mobility spectrometry and "direct analysis in real time" (DART™) mass spectrometry were used to rapidly identify AG-013736 in drug substance samples and 1mg Axitinib tablets. The plasmagrams of the sample solutions exhibited a major peak with a reduced ion mobility that was within ±0.0002cm(2)V(-1)s(-1) of that for AG-013736 in an external reference standard solution. The DART ionization source was coupled with both a time-of-flight mass spectrometer and a lower-resolution ion trap mass spectrometer. Samples were analyzed by this technique in as little as 5s with minimal to no sample preparation required. The isotopic masses of the protonated dimer ions of AG-013736 were used to identify AG-013736 in the active tablet. Both techniques were also used to develop low-level limit tests for rapidly verifying the presence or absence of AG-013736 in blinded clinical supplies of active and matching placebo tablets of Axitinib., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The option, for practical and ethical reasons, to replace animal plasma with human plasma for calibration standards was successfully applied to 73 analytical methods developed in our laboratory during the last years. The animals used for obtaining blank plasma could then be reduced with a number corresponding to about 25% of mice or 5% of rats in ordinary one-month toxicology studies. This is of important public concern and also in accordance with the 3R-strategy. The methods were successfully validated for determination of drug concentrations in plasma from rat, dog, mouse, rabbit and cynomolgus monkey. Reproducibility of study samples from dosed animals was established, showing a mean accuracy of 100.8% with a CV of 7.2% (n=1339). The purpose of this paper is to present a scientific basis for the alternative approach to adopt human plasma matrix for calibration standards, which will reduce animal use, without compromising the quality of appropriately validated assays. Additional advantages are cheaper and simplified plasma maintenance and the possibility to validate methods for several species in the same analytical batch., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Understanding the metabolism of a novel drug candidate in drug discovery and drug development is as important today as it was 30 years ago. What has changed in this period is the technology available for proficient metabolite characterization from complex biological sources. High-efficiency chromatography, sensitive MS and information-rich NMR spectroscopy are approaches that are now commonplace in the modern laboratory. These advancements in analytical technology have led to unequivocal metabolite identification often being performed at the earliest opportunity, following the first dose to man. For this reason an alternative approach is to shift from predicting and extrapolating possible human metabolism from in silico and nonclinical sources, to actual characterization at steady state within early clinical trials. This review provides an overview of modern approaches for characterizing drug metabolites in these early clinical studies. Since much of this progress has come from technology development over the years, the review is concluded with a forward-looking perspective on how this progression may continue into the next decade.

Evaluating biliary excretion, a major elimination pathway for many compounds, is important in drug discovery. The bile duct-cannulated (BDC) rat model is commonly used to determine the percentage of dose excreted as intact parent into bile. However, a study using BDC rats is time-consuming and cost-ineffective. The present report describes a computational model that has been established to predict biliary excretion of intact parent in rats as a percentage of dose. The model was based on biliary excretion data of 50 Bristol-Myers Squibb Co. compounds with diverse chemical structures. The compounds were given intravenously at <10 mg/kg to BDC rats, and bile was collected for at least 8 h after dosing. Recoveries of intact parents in bile were determined by liquid chromatography with tandem mass spectrometry. Biliary excretion was found to have a fairly good correlation with polar surface area (r = 0.76) and with free energy of aqueous solvation (DeltaG(solv aq)) (r = -0.67). In addition, biliary excretion was also highly corrected with the presence of a carboxylic acid moiety in the test compounds (r = 0.87). An equation to calculate biliary excretion in rats was then established based on physiochemical properties via a multiple linear regression. This model successfully predicted rat biliary excretion for 50 BMS compounds (r = 0.94) and for 25 previously reported compounds (r = 0.86) whose structures are markedly different from those of the 50 BMS compounds. Additional calculations were conducted to verify the reliability of this computation model.

In this study we investigated the correlations between the single crystal structure, the crystal habit and morphology, and surface energetics of an investigational pharmaceutical compound. Crystal structures of both the anhydrous form (Form A) and monohydrate form (Form B) have been solved from single crystal X-ray analysis. The external morphology of Form A solid was predicted by molecular modeling using attachment energies to be thick plate-like with two dominant faces (100) and (002). The external morphology of Form B was predicted to be needle-like with a dominant face (101 ). The predicted morphologies were confirmed by optical micrographs and the Miller indices of the dominant faces were complemented by X-ray powder diffraction (XRPD) method. Contact angle measurements showed that the anhydrous form has better wettability as predicted from crystal structure and morphology.

The aim of the study was to evaluate the use of imaging in the development of neuropharmacological drugs. All New Drug Applications (NDAs) approved from 1995 through 2004 in the Division of Neuropharmacological Drug Products at the Food and Drug Administration were surveyed for imaging studies. Imaging literature was also reviewed with respect to antipsychotics and antidepressants. One hundred and six NDAs (35 new molecular entities (NMEs)) were approved; 15 of these NDAs (10 NMEs) had imaging studies. The primary imaging modality was positron emission tomography. Imaging was primarily conducted for drugs used in schizophrenia, depression, multiple sclerosis, and migraine. The majority evaluated receptor occupancy or proof of concept. Examples (including literature) are discussed as pertinent to dosage, efficacy, safety, or further development of a drug or class of drugs. Imaging contributes to optimal clinical development of central nervous system (CNS)-active drugs. Opportunities are available for its broader use, contributing to improved understanding of the clinical pharmacology of CNS-active drugs.

A major challenge in selecting an appropriate stationary phase for diastereomeric separation is that it is difficult to predict which of the commercially available stationary phases could achieve the required liquid chromatographic (LC) separation. This work describes the selection and evaluation of a porous graphitic carbon (PGC) column coupled with tandem mass spectrometry (MS/MS) for the simultaneous quantitation of an experimental drug candidate (I), its two diastereomeric metabolites (II and III), and its demethylated metabolite (IV) in rat plasma. In addition, we investigated the PGC column for the separation of another drug candidate (VI), its two diastereomeric metabolites (VII and VIII) and its ketone metabolite (IX). The PGC column showed excellent chromatographic resolution for the two diastereomers II and III, as well as for VII and VIII. In contrast, the required resolution for the diastereomers II and III could not be achieved using silica-bonded C(18), C(30), phenyl, perfluorinated, polar embedded and polar end-capped phases. The PGC column showed ruggedness with excellent reproducibility of retention times, peak symmetry and response over a period of more than 400 injections of a plasma acetonitrile-precipitation extract. Excellent accuracy and precision were achieved, with accuracy of 94-108% and intra- and inter-run precision within 9%. This work indicates that PGC is a valuable addition to the repertoire of LC columns used for quantitative LC/MS/MS bioanalysis, especially where the separation and quantitation of diastereomeric analytes is involved., (Copyright (c) 2006 John Wiley & Sons, Ltd.)

Introduction: There is a continuing need for increased throughput in the examination of new chemical entities (NCEs) in terms of the pharmacokinetic (PK) parameters. The aim was to validate a new study method designed to improve throughput and reduce inter-animal variability and animal number requirement in routine bioavailability and plasma PK studies of NCEs in awake rats., Methods: The design uses a new method for intravenous (iv) administration via the saphenous vein in combination with serial blood sampling via the tail vein. The multiple sampling method was compared with single sampling (decapitation) and the effect on haematocrit (Hct) levels was studied. Direct injection in the saphenous vein was compared to iv administration using an indwelling jugular catheter., Results: Using structurally different NCEs, it was shown that a combination of direct injection via the saphenous vein and multiple sampling from the tail vein produces comparable plasma concentrations and subsequent PK results to the comparator methods. Furthermore, Hct levels remained within recommended levels using a total blood sampling volume of up to 2.1 ml/day for rats with a body weight of around 250 g., Discussion: The new model increases throughput by avoiding the time required for preparative surgery, increases quality by allowing inter-animal comparison of major PK parameters as concentration time curves can be obtained from each animal, and reduces the number of animals required.