Table 1.

A. HSP90 inhibitory and binding activities of CCT018159 and analogues
CompoundIC50, yeast HSP90 ATPase (μmol/L)IC50, human HSP90β ATPase (μmol/L)IC50, human HSP90β FP (μmol/L)GI50, SRB HCT116 cells (μmol/L)
CCT0181596.6 ± 0.53.2 ± 1.50.60 ± 0.104.1 ± 0.4
CCT0724400.8 ± 0.22.2 ± 0.20.20 ± 0.084.0 ± 0.6
CCT0724530.6 ± 0.11.8 ± 0.20.30 ± 0.026.8 ± 0.7
CCT066961>10053.0 ± 5.07.20 ± 0.3028.0 ± 1.0
B. Relative binding affinity of HSP90 inhibitors to HSP90 in lysates from human cancer (HCT116 and WM266.4) and nontumorigenic cells (HUVEC, MCF10a, and PNT2) as measured by the fluorescence polarization competitive binding assay using a diaryl pyrazole fluorescent probe ( 41)
CompoundIC50 for binding to HSP90β in human cell lysates (μmol/L)
HCT116WM266.4HUVECMCF10aPNT2
CCT0181590.21 ± 0.070.22 ± 0.010.37 ± 0.040.38 ± 0.100.16 ± 0.02
17-AAG0.35 ± 0.130.31 ± 0.070.35 ± 0.050.44 ± 0.090.27 ± 0.04
Geldanamycin0.12 ± 0.030.09 ± 0.010.09 ± 0.020.14 ± 0.020.17 ± 0.05
  • NOTE: (A) Enzyme assays were carried out using full-length yeast Hsp90 or human HSP90β, the latter in the presence of the activating protein AHA1, using the malachite green method ( 34). The fluorescence polarization competitive binding assay was carried out with human HSP90β using a fluorescent pyrazole resorcinol probe ( 41). Cell growth inhibitory activities are also shown; these were carried out in HCT116 human colon cancer cells using the SRB endpoint. Values represent mean ± SE (n = 3).

    (B) The amount of cell lysate that gave an anisotropy reading corresponding to 30 nmol/L human HSP90β was used (4 nmol/L for HCT116 and PNT2 cells and 12 nmol/L for WM266.4, HUVEC, and MCF10a cells). Values represent mean ± SE (n = 3–5).