Flowchart: Preparation: Hsp90









Text Box: Novobiocin


Text Box: Hsp90



Text Box:   nNos 

Text Box:                            


Text Box: Estrogen 





Hsp90 Inhibitors Identified from a Library of Novobiocin Analogues.

Yu XM, Shen G, Neckers L, Blake H, Holzbeierlein J, Cronk B, Blagg BS.

Department of Medicinal Chemistry and The Center for Protein Structure and Function, The University of Kansas, 1251 Wescoe Hall Drive, Malott 4070, Lawrence, Kansas 66045-7582, Urologic Oncology Branch, National Cancer Institute, NIH, Rockville, Maryland 20850, and Department of Urology, The University of Kansas Medical Center, Kansas City, Kansas 66103.

Novobiocin is a C-terminal inhibitor of the Hsp90 protein folding machinery, which is responsible for the conformational maturation of numerous proteins involved in cancer growth and survival. Due to novobiocin's poor inhibitory activity ( approximately 700 muM), very little attention has been paid toward the development of novobiocin analogues for Hsp90 inhibition. In this study, a parallel library of 20 novobiocin derivatives was prepared and the biological activity of each evaluated by Western blot analysis of Hsp90 client proteins. A4 was found to be a potent inhibitor of Hsp90 as determined by its ability to cause the degradation of several Hsp90 client proteins in both breast and prostate cancer cell lines. In the presence of 1 muM A4, several Hsp90 client proteins were degraded, including AKT, Her2, Hif-1alpha, and the androgen receptor.

PMID: 16159253 [PubMed - as supplied by publisher]














Bradykinin mimics ischemic preconditioning by generating reactive oxygen species (ROS). To identify intermediate steps leading to ROS generation, rabbit cardiomyocytes were incubated in reduced MitoTracker Red that becomes fluorescent after exposure to ROS. Fluorescence intensity in treated cells was expressed as % of that in paired untreated cells. Bradykinin (500nM) caused 51+/-16% increase in ROS generation (p<0.001). Co-incubation with either bradykinin B2 receptor blocker HOE140 (5 micro M) or free radical scavenger N-(2-mercaptopropionyl) glycine (1mM) prevented this increase confirming the response was receptor-mediated and ROS were actually being measured. Closing mitochondrial KATP (mKATP)channels with 5-hydroxydecanoate (5HD, 1mM) prevented increased ROS generation. Bradykinin-induced ROS generation was blocked by L-NAME (200 micro M) implicating nitric oxide as an intermediate. Blockade of guanylyl cyclase with ODQ (10 micro M) aborted bradykinin-induced ROS generation, but not that from diazoxide, a direct opener of mKATP channels. The PKG blocker 8-Br-cGMPS (25 micro M) eliminated bradykinin's effect. Conversely, direct activation of PKG with 8-pCPT-cGMP (100 micro M) increased ROS generation (39+/-15%, p<0.004) similar to bradykinin. This increase was blocked by 5HD. Finally, the nitric oxide donor SNAP (1 micro M)increased ROS by 34+/-6%. This increase was also blocked by 5HD. In intact rabbit hearts bradykinin (400nM) decreased infarction from 30.5+/-3.0% of the risk zone in control hearts to 11.9+/-1.4% (p<0.01). This protection was aborted by either L-NAME (200 micro M) or ODQ (2 micro M)(35.4+/-5.7% and 30.4+/-3.0% infarction, resp., pNS vs control). Hence, bradykinin preconditions through receptor-mediated production of nitric oxide which activates guanylyl cyclase. The resulting cGMP activates PKG that opens mKATP. Subsequent release of ROS triggers cardioprotection.