Heat shock factor 1 (HSF1) was originally identified as a master regulator of the classical ‘cytoprotective’ heat shock response. However, a large body of evidence has now verified the importance of HSF1 to tumorigenesis and cancer progression. HSF1 is activated by various elements of the cancer state, reprogramming the transcriptome in a way that is overlapping with, but distinct from, the canonical heat-shock response. Also, there is a strong correlation between the expression of activated HSF1 in tumors and adverse clinical outcomes. This evidence indicates that the inhibition of HSF1-mediated transcription could be a viable strategy in cancer treatment. Inhibiting the HSF1 stress pathway represents an attempt at targeting non-oncogene addiction and proteotoxic stress, which has been proposed to be advantageous. However, HSF1 is a ligandless transcription factor and is unlikely to be amenable to standard drug discovery strategies and direct inhibition with small molecules. Therefore, we proposed that inhibitors of HSF1-mediated transcription, which antagonize the HSF1 pathway but without necessarily binding directly to HSF1, could be discovered and developed via a cell-based phenotypic screen. We carried out a high throughput Arrayscan assay of 200,000 compounds to measure the inhibition of HSF1-mediated HSP72 expression stimulated by pre-treatment with an HSP90 inhibitor. We identified a singleton hit with a bisamide core, CCT245232. This compound showed potent growth inhibition in a range of human cancer cell lines but had poor physicochemical properties leading to an unacceptable pharmacokinetic profile. Improvement of the physicochemical properties of CCT245232 whilst maintaining potency versus our cell-based assays led to the orally bioavailable tool compound CCT251236. This compound shows potent growth inhibition (GI50 values in low nanomolar range) of human ovarian cancer cell lines in vitro and good efficacy against human ovarian cancer xenografts in nude mice in vivo. We applied chemo-proteomic strategies to identify the molecular target using a probe based on CCT251236 and discovered pirin as a high affinity molecular target. Binding of CCT251236 to recombinant pirin was confirmed in biophysical assays. CCT251236 recapitulates the reported anti-migratory phenotype for a pirin ligand although binding to pirin alone does not explain the cellular phenotype observed with our chemical tool. We are currently using CCT251236 as a chemical probe while further optimizing its properties to identify a clinical candidate.
Citation Format: Matthew D. Cheeseman, Nicola E. Chessum, Carl S. Rye, Elisa A. Pasqua, Michael J. Tucker, Birgit Wilding, Lindsay E. Evans, Susan Lepri, Meirion Richards, Swee Y. Sharp, Salyha Ali, Martin Rowlands, Lisa O'Fee, Asadh Miah, Angela Hayes, Alan T. Henley, Marissa Powers, Robert te Poele, Emmanuel De Billy, Loredana Pellegrino, Florence Raynaud, Rosemary Burke, Robert L. van Montfort, Suzanne A. Eccles, Keith Jones, Paul Workman. Discovery of chemical probe CCT251236: An orally bioavailable efficacious pirin ligand from an HSF1 phenotypic screen [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-304. doi:10.1158/1538-7445.AM2017-LB-304