Fig. S3.

Inducible and constitutive cytosolic HSP70s are the relevant targets of MAL3-101 in RMS cells. (A and C) Fusion-positive RMS13 (A) or fusion-negative RD (C) cells were transfected with the indicated concentrations of control siRNA or pools of siRNA targeting HSPA1A/B (inducible) and HSPA8 (constitutive) for 72 h and then were treated with MAL3-101. Viability was measured 48 h later by CellTiter-Glo, and IC₅₀ doses were calculated by nonlinear regression. Data shown are from three independent replicates; error bars represent the SEM. (B and D) Immunoblots of cells treated as in A and C demonstrate increasing knockdown with increasing amounts of siRNA. Note that at low concentrations of siRNA, compensatory up-regulation of HSPA1A/B results in more HSP72 (inducible), as in Fig. 3C.