Design and evaluation of an HSP70-targeting PROTAC in synergy with an HSF1 inhibitor for enhanced antitumor activity

Heat shock protein (HSP) 70 represents a clinically promising anti-tumor target, yet the development of effective inhibitors faces numerous technical challenges. To address this, we developed novel non-ATP site Proteolysis-targeting Chimeras (PROTACs) that selectively degrade HSP70 by engaging the E3 ubiquitin Ligase CRBN. However, the PROTACs exhibited limited degradation activity, potentially due to heat shock response-mediated HSP70 upregulation. To circumvent this resistance mechanism, we explored combination therapy with the heat shock factor 1 (HSF1) inhibitor DTHIB to disrupt the heat shock feedback loop, markedly enhancing HSP70 degradation. The combination strategy showed synergistic and selective anti-tumor activity across a panel of Cancer cell lines. This success relied on the distinct profile of C4, which preferentially targets cytosolic HSP70 and, unlike conventional inhibitors, effectively circumvents compensatory HSP70 upregulation.