Avoidance of Intracellular Hsp90a Inhibition, Including Cell-Impermeable Extracellular Hsp90a (eHsp90a)-Selective Inhibitors and Hsp90ß-Selective Inhibitors as Safer Alternatives

The heat shock protein 90 (Hsp90) family of molecular chaperones mediates the folding and activation of ~400 client proteins, many of which contribute to oncogenesis. As a result, 22 small-molecule Hsp90 pan-inhibitors, which inhibit all four Hsp90 isoforms, have been investigated in the clinic for the treatment of cancer. Unfortunately, detrimental side effects were observed and hindered the clinical development of pan-Hsp90 inhibitors. The two most common on-target toxicities, cardio- and ocular-toxicity, have been attributed to inhibition of the Hsp90a isoform. Consequently, most of these investigational drugs failed in the clinic. Recent work has focused on the development of Hsp90 isoform-selective inhibitors to avoid the complications associated with Hsp90a inhibition. Hsp90a is also secreted extracellularly (eHsp90a) and modulates wound healing, cell motility, and inflammation. Furthermore, eHsp90a levels have been directly correlated with the progression of a variety of cancers, implicating eHsp90a as a cancer biomarker and potential target for cancer. As an alternative strategy, inhibitors of eHsp90a could avoid inhibition of intracellular Hsp90a, and therefore the unintended side-effects, while providing a therapeutic effect. My thesis work herein has focused on the development and validation of cell-impermeable Hsp90a-selective inhibitors, the effects of targeting eHsp90a on cancer migration and invasion, an assessment of whether Hsp90ß-selective inhibitors avoid the toxicities that have plagued Hsp90 pan-inhibitors, and a proteomic analysis to investigate the mechanisms underlying the anti-cancer effects of Hsp90ß-selective inhibitors against triple-negative breast cancer (TNBC). The evidence demonstrate that eHsp90a- and Hsp90ß-selective inhibitors are safer therapeutic alternatives to Hsp90 pan-inhibitors due to their avoidance of intracellular Hsp90a inhibition. Additionally, eHsp90a inhibitors inhibit cancer migration and invasion through unintended binding to eHsp90a’s critical F-5 fragment, which is required for activation of these invasive phenotypes. Furthermore, the proteomic investigation found that Hsp90ß-selective inhibitors inhibit kinase signaling pathways, cell cycle, and DNA repair, to name a few examples, while also identifying RAD9A, CDK1, and RPS9 as potential Hsp90ß-dependent client proteins.