Current Drug Metabolism - Volume 11, Issue 7, 2010

Glucuronidation is one of the main phase II metabolic reactions in humans and animals. A variety of analytical techniques and methods have been used for the detection and quantification of glucuronides of both endogenous and xenobiotic compounds from different biological samples of humans and animals. Drug metabolism has been extensively studied with both in vitro and in vivo experiments under various conditions. The purpose of this review is to explore in detail the benefits and drawbacks of different liquid chromatography- mass spectrometric (LC/MS) methods and techniques in detection and identification of all forms of glucuronide conjugates from in vitro, biological, and environmental samples. The entire analytical procedure is covered, from sample treatment, separation, and ionization to qualitative and quantitative analyses. The aim of this review is not to cover every published paper where glucuronides are identified and/or quantified, but rather to focus on special cases where a new analytical approach or technical development has led to a better, more specific, or more comprehensive detection, identification, or quantitation of glucuronide conjugates.

The in vitro metabolic stability assays are indispensable for screening the metabolic liability of new chemical entities (NCEs) in drug discovery. Intrinsic clearance (CLint) values from liver microsomes and/or hepatocytes are frequently used to assess metabolic stability as well as to quantitatively predict in vivo hepatic plasma clearance (CLH). An often used approximation is the so called wellstirred model which has gained widespread use. The applications of the well-stirred model are typically dependent on several measured parameters and hence with potential for error-propagation. Despite widespread use, it was recently suggested that the well-stirred model in some circumstances has been misused for in vitro in vivo extrapolation (IVIVE). In this work, we follow up that discussion and present a retrospective analysis of IVIVE for hepatic clearance prediction from in vitro metabolic stability data. We focus on the impact of input parameters on the well stirred model; in particular comparing “reference model” (with all experimentally determined values as input parameters) versus simplified models (with incomplete input parameters in the models). Based on a systematic comparative analysis and model comparison using datasets of diverse drug-like compounds and NCEs from rat and human, we conclude that simplified models, disregarding binding data, may be sufficiently good for IVIVE evaluation and compound ranking at early stage for cost-effective screening. Factors that can influence prediction accuracy are discussed, including in vitro intrinsic clearance (CLint) and in vivo CLint scaling factor used, non-specific binding to microsomes (fum), blood to plasma ratio (CB/CP) and in particular fraction unbound in plasma (fu). In particular, the fu discrepancies between literature data and in-house values and between two different compound concentrations 1 and 10 μM are exemplified and its potential impact on prediction performance is demonstrated using a simulation example.

Bleomycin is a potent chemotherapeutic agent that can mediate cell killing by attacking the DNA. It is used in combination with other antineoplastic agents to effectively treat lymphomas, testicular carcinomas and squamous cell carcinomas of the cervix, head and neck. However, resistance to bleomycin remains a persistent limitation in exploiting the full therapeutic benefit of the drug for other types of cancers. Herein, we review recent findings from both yeast and human cells showing that uptake of bleomycin-A5 is a key mechanism that limits toxicity of the drug. We also discuss how the mammalian transporter hCT2 (SLC22A16) could be used to predict the outcome of tumor responses towards bleomycin therapy, and highlight the importance of further exploring this permease with respect to its regulation and pharmacological substrates for treating a wide range of cancers.

The human breast cancer resistance protein (BCRP/ABCG2) is the second member of the G subfamily of the large ATPbinding cassette (ABC) transporter superfamily. BCRP was initially discovered in multidrug resistant breast cancer cell lines where it confers resistance to chemotherapeutic agents such as mitoxantrone, topotecan and methotrexate by extruding these compounds out of the cell. BCRP is capable of transporting non-chemotherapy drugs and xenobiotiocs as well, including nitrofurantoin, prazosin, glyburide, and 2-amino-1-methyl-6-phenylimidazo [4,5-b]pyridine. BCRP is frequently detected at high levels in stem cells, likely providing xenobiotic protection. BCRP is also highly expressed in normal human tissues including the small intestine, liver, brain endothelium, and placenta. Therefore, BCRP has been increasingly recognized for its important role in the absorption, elimination, and tissue distribution of drugs and xenobiotics. At present, little is known about the transport mechanism of BCRP, particularly how it recognizes and transports a large number of structurally and chemically unrelated drugs and xenobiotics. Here, we review current knowledge of structure and function of this medically important ABC efflux drug transporter.

Tyrosine kinase inhibitors (TKIs) are a new class of highly-selective and molecularly targeted anticancer agents. Most of these newly developed TKIs are hydrophobic, thus allowing them to rapidly penetrate the cell membrane to reach their specific intracellular targets. However, their therapeutic potential could be significantly hindered by the overexpression of certain ATP binding cassette (ABC) membrane transporters, which extrude hydrophobic drugs and result in cellular resistance to TKIs by tumor cells. Moreover, it has been recently demonstrated that some TKIs could upregulate ABC transporters in tumor cells, thereby effectively reducing their intracellular accumulation and antitumor efficacy. On the other hand, other TKIs were found to interact with ABC transporters and reverse multi-drug resistance (MDR) of tumor cells. In this review, the interaction of several TKIs, currently in clinical use or being developed in clinical trials, with the MDR-related ABC transporters, in particular ABCB1, ABCC1 and ABCG2, will be discussed.