Identification of an Allosteric Small-Molecule Inhibitor Selective for the Inducible Form of Heat Shock Protein 70

Abstract

Summary Inducible Hsp70 (Hsp70i) is overexpressed in a wide spectrum of human tumors and its expression correlates with metastasis, poor outcomes, and resistance to chemotherapy in patients. Identification of small molecule inhibitors selective for Hsp70i could provide new therapeutic tools for cancer treatment. In this work, we used fluorescence-linked enzyme chemoproteomic strategy (FLECS) to identify HS-72, an allosteric inhibitor selective for Hsp70i. HS-72 displays the hallmarks of Hsp70 inhibition in cells, promoting substrate protein degradation and growth inhibition. Importantly, HS-72 is selective for Hsp70i over the closely related constitutively active Hsc70. Studies with purified protein show HS-72 acts as an allosteric inhibitor, reducing ATP affinity. In vivo HS-72 is well-tolerated, showing bioavailability and efficacy, inhibiting tumor growth and promoting survival in a HER2+ model of breast cancer. The HS-72 scaffold is amenable to resynthesis and iteration, suggesting an ideal starting point for a new generation of anticancer therapeutics targeting Hsp70i.

Mutations in the Yeast Hsp70, Ssa1, at P417 Alter ATP Cycling, Interdomain Coupling, and Specific Chaperone Functions

“…The thermofluor binding assay did not reveal multiple transition peaks, as has been seen using differential scanning calorimetry [38,54,55]. Instead, only one transition is evident, as previously demonstrated using this method [56]. The slope of the transitions was qualitatively similar for all three proteins and no significant shift in the curve was seen for the mutant proteins compared to wild type Ssa1.…”

Combined chemical–genetic approach identifies cytosolic HSP70 dependence in rhabdomyosarcoma

“…This specialization in function suggests that pharmacologic inhibition of HSP70 will provide a therapeutic window in certain cancer subsets. Recently discovered small molecule HSP70 inhibitors have shown some activity in select preclinical cancer models (10)(11)(12)(13)(14). The availability of these HSP70 inhibitors creates,…”

Isoform-selective Genetic Inhibition of Constitutive Cytosolic Hsp70 Activity Promotes Client Tau Degradation Using an Altered Co-chaperone Complement

“…Therefore, the design and development of Hsp70 isoform-selective inhibitors will likely be essential for rationally targeting these proteins for therapeutic purposes in the future, and although challenging due to structural similarities between Hsp70 isoforms, it is possible. An Hsp72-selective inhibitor was recently developed and showed good efficacy for treating cancer with fewer side effects (72). Development of Hsc70 selective inhibitors could hold similar promise for tauopathies.…”