Inhibition of novobiocin on proliferation of HL-60/Bcr-Abl cells involves disruption of Hsp90 chaperon function

Aim: In an effort to confirm the effects of NB on Bcr-Abl expressing HL-60/Bcr-Abl cells, and the relationship between these effects and the molecular chaperone functions of heat shock protein 90 (Hsp90). Methods: The amounts of Bcr-Abl protein were tested by Western blot. Molecular chaperone functions of Hsp90 were measured by coimmunoprecipitation. Co- immunoprecipitation of Bcr-Abl and its molecular chaperones, the immunoprecipitate was then subjected to Western blot analysis with anti-Abl, anti-Hsp90, or anti-Hsp70 mAb. Proteasome inhibitor ALLnL was used to determine the down-regulation pathway of Bcr-Abl. Results: An exposure of HL-60/Bcr-Abl cells to NB produced down-regulation of intracellular Bcr-Abl protein levels. By binding and inhibiting Hsp90, NB treatment decreased the binding of Bcr-Abl with Hsp90 and Hsp70 and induced the proteasomal degradation of Bcr-Abl, because cotreatment with proteasome inhibitor ALLnL increased Bcr-Abl levels in NP40-insoluble fraction. Conclusions: These studies demonstrate for the first time the activity of NB against HL-60/Bcr-Abl cells and suggest that destruction of Hsp90 chaperon function, resulting in disruption the combination of Bcr-Abl with Hsp90 and cochaperons, may be an important mechanism of NB-mediated effects on HL-60/Bcr-Abl cells.