KU675, a Concomitant Heat-Shock Protein Inhibitor of Hsp90 and Hsc70 that Manifests Isoform Selectivity for Hsp90α in Prostate Cancer Cells

Abstract

The 90-kDa heat-shock protein (Hsp90) assists in the proper folding of numerous mutated or overexpressed signal transduction proteins that are involved in cancer. Inhibiting Hsp90 consequently is an attractive strategy for cancer therapy as the concomitant degradation of multiple oncoproteins may lead to effective antineoplastic agents. Here we report a novel C-terminal Hsp90 inhibitor, designated KU675, that exhibits potent antiproliferative and cytotoxic activity along with client protein degradation without induction of the heat-shock response in both androgen-dependent and -independent prostate cancer cell lines. In addition, KU675 demonstrates direct inhibition of Hsp90 complexes as measured by the inhibition of luciferase refolding in prostate cancer cells. In direct binding studies, the internal fluorescence signal of KU675 was used to determine the binding affinity of KU675 to recombinant Hsp90a, Hsp90b, and Hsc70 proteins. The binding affinity (K d ) for Hsp90a was determined to be 191 mM, whereas the K d for Hsp90b was 726 mM, demonstrating a preference for Hsp90a. Western blot experiments with four different prostate cancer cell lines treated with KU675 supported this selectivity by inducing the degradation of Hsp90a-dependent client proteins. KU675 also displayed binding to Hsc70 with a K d value at 76.3 mM, which was supported in cellular by lower levels of Hsc70-specific client proteins on Western blot analyses. Overall, these findings suggest that KU675 is an Hsp90 C-terminal inhibitor, as well as a dual inhibitor of Hsc70, and may have potential use for the treatment of cancer.

Gambogic acid identifies an isoform-specific druggable pocket in the middle domain of Hsp90β

“…Recently, Blagg and colleagues identified an inhibitor binding to the Hsp90 CTD with moderate specificity for Hsp90α (34). Our data identify GBA as a bioprobe demonstrating Hsp90β-specific inhibition, with a 10-fold binding preference for Hsp90β compared with Hsp90α.…”

“…Although recent drug discovery efforts have moved in this direction (30), to date the only paralog-specific inhibitors identified are for GRP94 (31)(32)(33) as well as an inhibitor with a moderate preference for Hsp90α (34). At present, there are no selective Hsp90β inhibitors, making it difficult to explore the distinct cellular roles for this constitutively expressed Hsp90 protein without resorting to genetic approaches.…”

Targeting the Hsp90 C-terminal domain to induce allosteric inhibition and selective client downregulation

“…Mechanistically, C-terminal ATP site inhibitors are proposed to cause client protein release and degradation through induction of a conformational change that results in separation of the Hsp90 dimer [15]. In general, these types of inhibitors appear less potent than N-terminal inhibitors and none have reached the clinic, although novobiocin derivatives have demonstrated efficacy in prostate cancer cell lines [32,33]. A deguelin derivative was shown to disrupt ATP-binding to the CTD and have anti-proliferative activity in non-small cell lung cancer cell lines [34,35].…”

Hybrid Enzalutamide Derivatives with Histone Deacetylase Inhibitor Activity Decrease Heat Shock Protein 90 and Androgen Receptor Levels and Inhibit Viability in Enzalutamide-Resistant C4-2 Prostate Cancer Cells

“…Association of the AR apoprotein with HSP90 is critical for stabilizing AR in a conformation that allows androgen binding (Veldscholte et al, 1992;Fang et al, 1996). In PCa cells, HSP90 inhibitors induce AR degradation and impair AR nuclear translocation while simultaneously reducing the levels of other oncogenic client proteins, such as p-AKT/AKT, epidermal growth factor receptor, insulinlike growth factor receptor, and survivin (Solit et al, 2002;Saporita et al, 2007;He et al, 2013;Liu et al, 2015). Simultaneous disruption of AR and other aberrant growth/survival networks via HSP90 inhibition is an advantageous treatment strategy for CRPC, because this would silence potentially mutually compensatory oncogenic signaling pathways.…”