Fig 5.

FKBP12-selective ligands do not block the cellular effects of rapamycin. SHSY-5Y cells (A and B) and HeLa cells (C and D) were treated with dimethyl sulfoxide, rapamycin (Rap), or torin-1 (100 nM) in the absence or presence of the FKBP12-selective inhibitor Biricodar (Biri) (40) or compound 44 (cmpd44) (58) or the nonselective FKBP inhibitor FK1706 or FK506 (45) at the indicated concentration. After 60 min, cellular Akt phosphorylation (A and C) and cellular mTOR phosphorylation (B and D) were determined with a homogeneous time-resolved assay. The panselective FKBP inhibitors, but not the FKBP12-selective inhibitors, blocked the effect of rapamycin. For all experiments, mean values from at least three data points are shown. The dimethyl sulfoxide control was set to 100% phosphorylation, while torin-1 was defined as 0% phosphorylation. The values are presented as means ± standard deviations. Two-way analysis of variance and a priori testing were used for statistical analysis. Comparison with dimethyl sulfoxide treatment: ***, P < 0.001; **, P < 0.01; *, P < 0.05. Comparison with treatment with rapamycin alone: +++, P < 0.001; ++, P < 0.01; +, P < 0.05.