Figure 3

Analysis of native Hsp90 complexes in PC3-MM2 and LNCaP cells by Blue Native Western blot and identification of the Hsp90 complex from human tissue. Native Hsp90 complexes were identified by Native Western blot (Figure 3A). 17-AAG, novobiocin and F-4, an early novobiocin analog, had little effect on the Hsp90β complex. Conversely, cytotoxic concentrations of KU174 resulted in potent disruption of the Hsp90β complex (Figure 3A). Dose-dependent effects on Hsp90β, Hsp90α, GRP94 and Hsc70 native complexes were determined following treatment of cells KU174 along with novobiocin and 17-AAG (Figure 3B). Hsp90β complex was also observed in human cancer tissue from two patient samples but to a lesser extent in their normal adjacent tissue (Figure 3C). Hsp60 served as a loading control showing relative equal loading across the patient samples. KU174 also exhibits activity in the androgen dependent cell line LNCaP resulting in Hsp90 complex disruption (Figure 3D).