Simplified procedures and a reduction in sampling errors are important advantages for performing as many chemical analysis steps as possible at the site where a sample or subject is located. Solid-phase microextraction technology addresses this goal, and for our purposes, it also allows for in vivo monitoring of a dynamic living system with minimal disturbance of the system. Here we report the development of a solid-phase microextraction application for in vivo monitoring of circulating blood concentrations of benzodiazepines. A probe based on a polypyrrole extraction phase was developed and used for extraction of drug molecules directly from a peripheral vein with subsequent instrumental quantification. The probe provides good sensitivity and selectivity for the drugs versus the blood matrix, while eliminating the need to draw blood. After sampling, the extracted drugs are quantified by liquid chromatography-tandem mass spectrometry. The limit of detection of the method is ∼3-7 ng/mL for analysis of the benzodiazepines from whole blood, and the method is linear to 1000 ng/mL. The method was used to monitor the pharmacokinetic profiles of diazepam and its metabolites in dogs, and the results compared favorably with profiles determined by conventional methods. Because no blood and only very small amounts of drugs are removed, minimal disruption to the chemical balance in blood occurs. This approach offers the potential for reduced exposure to blood for analytical personnel, simplified, less disruptive sampling, and lower stress levels on animals for pharmacokinetic studies. It also allows for a significant reduction in animal usage for these studies, which is important both ethically and for improving data quality.

An antibody-drug conjugate (ADC) is a unique therapeutic modality composed of a highly potent drug molecule conjugated to a monoclonal antibody. As the number of ADCs in various stages of nonclinical and clinical development has been increasing, pharmaceutical companies have been exploring diverse approaches to understanding the disposition of ADCs. To identify the key absorption, distribution, metabolism, and excretion (ADME) issues worth examining when developing an ADC and to find optimal scientifically based approaches to evaluate ADC ADME, the International Consortium for Innovation and Quality in Pharmaceutical Development launched an ADC ADME working group in early 2014. This white paper contains observations from the working group and provides an initial framework on issues and approaches to consider when evaluating the ADME of ADCs.

Although skin is the largest organ of the human body, cutaneous drug metabolism is often overlooked, and existing experimental models are insufficiently validated. This proof-of-concept study investigated phase II biotransformation of 11 test substrates in fresh full-thickness human skin explants, a model containing all skin cell types. Results show that skin explants have significant capacity for glucuronidation, sulfation, N-acetylation, catechol methylation, and glutathione conjugation. Novel skin metabolites were identified, including acyl glucuronides of indomethacin and diclofenac, glucuronides of 17b-estradiol, N-acetylprocainamide, and methoxy derivatives of 4-nitrocatechol and 2,3-dihydroxynaphthalene. Measured activities for 10 mM substrate incubations spanned a 1000-fold: from the highest 4.758 pmol·mg skin ·h -1 for 17b-estradiol 17-glucuronidation. Interindividual variability was 1.4-to 13.0-fold, the highest being 4-methylumbelliferone and diclofenac glucuronidation. Reaction rates were generally linear up to 4 hours, although 24-hour incubations enabled detection of metabolites in trace amounts. All reactions were unaffected by the inclusion of cosubstrates, and freezing of the fresh skin led to loss of glucuronidation activity. The predicted whole-skin intrinsic metabolic clearances were significantly lower compared with corresponding whole-liver intrinsic clearances, suggesting a relatively limited contribution of the skin to the body's total systemic phase II enzymemediated metabolic clearance. Nevertheless, the fresh full-thickness skin explants represent a suitable model to study cutaneous phase II metabolism not only in drug elimination but also in toxicity, as formation of acyl glucuronides and sulfate conjugates could play a role in skin adverse reactions.

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<P>Background: Although the liver is the primary organ of drug metabolism, the lungs also contain drug-metabolizing enzymes and may, therefore, contribute to the elimination of drugs. In this investigation, the Precision-cut Lung Slice (PCLS) technique was standardized with the aims of characterizing and comparing rat and human pulmonary drug metabolizing activity. </P><P> Method: Due to the limited availability of human lung tissue, standardization of the PCLS method was performed with rat lung tissue. Pulmonary enzymatic activity was found to vary significantly with rat age and rat strain. The Dynamic Organ Culture (DOC) system was superior to well-plates for tissue incubations, while oxygen supply appeared to have a limited impact within the 4h incubation period used here. </P><P> Results: The metabolism of a range of phase I and phase II probe substrates was assessed in rat and human lung preparations. Cytochrome P450 (CYP) activity was relatively low in both species, whereas phase II activity appeared to be more significant. Conclusion: PCLS is a promising tool for the investigation of pulmonary drug metabolism. The data indicates that pulmonary CYP activity is relatively low and that there are significant differences in enzyme activity between rat and human lung.</P>

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