Biol. Chem., Vol. 390, pp. 175–179, February 2009 • Copyright  by Walter de Gruyter • Berlin • New York. DOI 10.1515/BC.2009.021 2009/250 Article in press - uncorrected proof Short Communication Murine and human cathepsin B exhibit similar properties: possible implications for drug discovery Dejan Caglic ˇ 1 , Gregor Kosec 1 , Lea Bojic ˇ 1 , Thomas Reinheckel 2 , Vito Turk 1 and Boris Turk 1, * 1 Department of Biochemistry and Molecular and Structural Biology, Joz ˇ ef Stefan Institute, Jamova 39, SI-1000 Ljubljana, Slovenia 2 Department of Molecular Medicine and Cell Research, Albert Ludwigs University of Freiburg, Stefan-Meier-Str. 17, D-79104 Freiburg, Germany * Corresponding author e-mail: boris.turk@ijs.si Abstract Validation of drug targets and subsequent preclinical studies are usually carried out on animal disease models, with mouse being the most commonly used. However, results from mouse models cannot always be directly related to human disease. Major discrepancies between the properties of murine and human variants were observed during the evaluation of compounds targeting cathepsins S and K. It is important, therefore, to know whether similar differences exist between murine and human cathepsin B. Thus, both enzymes were expressed and biochemically characterized. The enzymes exhibited similar biochemical properties, indicating that cathepsin B transgenic mouse models could be useful for studying its role in human pathologies. Keywords: animal disease models; enzyme kinetics; human cathepsin B; murine cathepsin B. Cysteine cathepsins form a group of proteases that were long believed to be involved primarily in intracellular pro- tein turnover. However, there is growing evidence for their specific intra- and extracellular functions, linking them to bone remodeling, precursor protein activation, MHC class II antigen processing and presentation, and other functions. Disturbances in the regulation of the intracel- lular activities and uncontrolled release of cathepsins from lysosomes have been implicated in many disease states, such as osteoporosis, inflammatory diseases, and tumor progression (Turk et al., 2001; Vasiljeva et al., 2007; Zavas ˇnik-Bergant and Turk, 2007; Brix et al., 2008). Cathepsins S and K have been validated as drug targets for inflammatory and immune disorders, and for osteo- porosis, respectively (Grabowska et al., 2005; Turk, 2006; Vasiljeva et al., 2007; Weidauer et al., 2007). Several compounds targeting cathepsin K are now in advanced clinical trials for treating osteoporosis (Turk, 2006), and an inhibitor of cathepsin S entered phase I clinical trials for psoriasis treatment in 2006 (Vasiljeva et al., 2007). Validation of drug targets and all preclinical studies are normally carried out using appropriate animal disease models, with mouse being the most commonly used (Zambrowicz and Sands, 2003). However, results from mouse models cannot always be directly related to human disease. Namely, a problem was observed during evaluation of compound(s) targeting cathepsins S and K, arising from the major differences observed between the human and mouse enzymes. The compounds that were developed against the human enzymes were found to have approximately 2–3 orders of magnitude weaker affinities for their mouse orthologs (Marquis et al., 2001; Gustin et al., 2005). This was very surprising as the amino acid sequences of the corresponding human and mouse enzymes are very similar. However, structural analysis of the enzymes revealed that several important residues in the active site cleft are different, which affects substrate and inhibitor binding (Marquis et al., 2001). Therefore, compounds developed against the human enzyme could fail to exhibit a therapeutic effect in a mouse disease model. As a consequence, alternative models had to be developed to enable preclinical evaluation of the com- pounds. For example, in the case of cathepsin K, two alternative models have been developed, one based on human-derived cells (Kung Sutherland et al., 2003) and the other on ovariectomized cynomolgus monkeys, since monkey cathepsin K is sequentially identical to human cathepsin K (Stroup et al., 2001). There is increasing evidence that cathepsin B could be a drug target for several pathologies, based mainly on its role in cancer progression and osteoarthritis (Mohamed and Sloane, 2006; Vasiljeva et al., 2007). Cathepsin B was thus observed to be overexpressed in various human tumors, with its membrane association and/or secretion additionally increasing its pathological charac- ter (reviewed in Mohamed and Sloane, 2006). Gene knock-out studies of cathepsin B revealed its role in angiogenesis, cell proliferation, tumor growth and remod- eling of the extracellular matrix (Gocheva et al., 2006; Vasiljeva et al., 2006, 2008). However, a partial compen- satory effect of cathepsin X was observed in a mammary tumor mouse model, suggesting at least a partial func- tional redundancy (Vasiljeva et al., 2006). In addition, in osteoarthritis and rheumatoid arthritis cathepsin B was suggested to be involved by degrading collagen and aggrecan in affected joints (reviewed in Yan and Sloane, 2003; Vasiljeva et al., 2007), in addition to activating the fibrinolytic cascade by converting pro-urokinase to uro- kinase (Ikeda et al., 2000). Taking advantage of the unique biochemical properties of cathepsin B, several Brought to you by | Purdue University Libraries Authenticated Download Date | 6/1/15 4:16 PM