Probing the Probes: Fitness Factors For Small Molecule Tools Paul Workman, Ian Collins  Chemistry & Biology  Volume 17, Issue 6, Pages 561-577 (June 2010) DOI: 10.1016/j.chembiol.2010.05.013 Copyright © 2010 Elsevier Ltd Terms and Conditions

Figure 1 Fitness Factors for Chemical Probes Grouped into Four Distinct Areas Chemical properties, biological potency, biological selectivity, and context of use (A), that encompass suggested criteria for evaluating the suitability of chemical probe compounds for exploratory biology (see Baell and Holloway, 2010; Cohen, 2009; Edwards et al., 2009; Frye, 2010; Inglese et al., 2007; Kodadek, 2010; McGovern et al., 2003; Oprea et al., 2007; Rishton, 2003). Threshold values that have been suggested in the literature are tabulated for each of the criteria (B), and a comparison of these properties for drugs, leads, and probes can be found in Table 1. We suggest that although not all probes can, or need to, reach these thresholds in every case, consideration of the criteria will allow a robust assessment of whether the probe is fit-for-purpose, and foster an appreciation of the risk carried forward if significant criteria are not met. Chemistry & Biology 2010 17, 561-577DOI: (10.1016/j.chembiol.2010.05.013) Copyright © 2010 Elsevier Ltd Terms and Conditions

Figure 1 Fitness Factors for Chemical Probes Grouped into Four Distinct Areas Chemical properties, biological potency, biological selectivity, and context of use (A), that encompass suggested criteria for evaluating the suitability of chemical probe compounds for exploratory biology (see Baell and Holloway, 2010; Cohen, 2009; Edwards et al., 2009; Frye, 2010; Inglese et al., 2007; Kodadek, 2010; McGovern et al., 2003; Oprea et al., 2007; Rishton, 2003). Threshold values that have been suggested in the literature are tabulated for each of the criteria (B), and a comparison of these properties for drugs, leads, and probes can be found in Table 1. We suggest that although not all probes can, or need to, reach these thresholds in every case, consideration of the criteria will allow a robust assessment of whether the probe is fit-for-purpose, and foster an appreciation of the risk carried forward if significant criteria are not met. Chemistry & Biology 2010 17, 561-577DOI: (10.1016/j.chembiol.2010.05.013) Copyright © 2010 Elsevier Ltd Terms and Conditions

Figure 2 Fitness Factor Liabilities for Selected Early Chemical Probe Inhibitors of Protein Kinases, the PI3K Family of Lipid Kinases and the HSP90 Molecular Chaperone Important fitness factors that have been improved significantly in subsequent chemical probes and clinical agents acting on these targets have been flagged. The symbols refer to the fitness factors listed in Figure 1, and the evolution of the probes to remove the liabilities shown is discussed in detail in the respective sections of the text. Chemistry & Biology 2010 17, 561-577DOI: (10.1016/j.chembiol.2010.05.013) Copyright © 2010 Elsevier Ltd Terms and Conditions

Figure 3 Structures of Selected Protein Kinase Chemical Probes Discussed in Detail in the Text with Their Main Proposed Targets Indicated Chemistry & Biology 2010 17, 561-577DOI: (10.1016/j.chembiol.2010.05.013) Copyright © 2010 Elsevier Ltd Terms and Conditions

Figure 4 Evolution of Increasingly Selective PKC Inhibitors Based on Staurosporine Percentage inhibitions for UCN-01 and ruboxistaurin were measured at 0.01 μM and 0.1 μM concentrations, respectively, for the same panel of 69 kinases (Bain et al., 2007). IC50 values were determined for sotrastaurin in a panel of 32 kinases (Wagner et al., 2009). The cluster of potent activities for sotrastaurin represents PKC isoform inhibition. All three compounds inhibit several PKC isoforms with IC50 0.001–0.01 μM. Kinase dendrogram (Manning et al., 2002) reproduced courtesy of Cell Signaling Technology, Inc. (www.cellsignal.com). Chemistry & Biology 2010 17, 561-577DOI: (10.1016/j.chembiol.2010.05.013) Copyright © 2010 Elsevier Ltd Terms and Conditions

Figure 5 Structures of Selected PI3 Kinase and HSP90 Chemical Probes Discussed in Detail in the Text with Their Main Proposed Targets Indicated Chemistry & Biology 2010 17, 561-577DOI: (10.1016/j.chembiol.2010.05.013) Copyright © 2010 Elsevier Ltd Terms and Conditions