Response of tumours to Hsp90 inhibitors is highly variable and their clinical effects are unpredictable, emphasising the need for a predictive marker. We postulated that sensitivity to Hsp90 inhibitors is connected to basal proteotoxic stress that makes cells dependent on Hsp90. Therefore, we assessed HSF1 as a general sensor of proteotoxic stress and correlated its activity with sensitivity to three separate small molecule Hsp90 inhibitors in seven breast cancer cell lines representing each of the different cancer subtypes. Flow cytometry was used to analyse the viability of breast cancer cell lines after Hsp90 inhibition. HSF1 activity was characterised by Ser326 phosphorylation and the transactivation capacity of HSF1 was determined by qPCR analysis of the ratios of HSF1-dependent (HOP, Hsp70) and HSF1-independent (CHIP) chaperones and cochaperone mRNAs. We show that the sensitivity of breast cancer cell lines to Hsp90 inhibition is highly variable. The basal levels of phosphorylated HSF1 also vary between cell lines and the magnitude of change in HSF1 phosphorylation after Hsp90 inhibition showed a negative correlation with sensitivity to Hsp90 inhibitors. Similarly, the basal transactivation capacity of HSF1, determined by the ratio of Hsp70 or HOP mRNA to CHIP mRNA level, is directly proportional to sensitivity to Hsp90 inhibitors. Increasing basal HSF1 activity by prior heat shock sensitised cells to Hsp90 inhibition. These results demonstrate that endogenous HSF1 activity varies between individual cancer cell lines and inversely reflects their sensitivity to Hsp90 inhibitors, suggesting that basal proteotoxic stress is an important and generalised predictor of response. Mechanistically, the data indicate that high endogenous proteotoxic stress levels sensitise to Hsp90 inhibition due to the inability to respond adequately to further proteotoxic stress. HSF1 activity therefore represents a potential biomarker for therapy with Hsp90 inhibitors, which may be useful for the rational design of future clinical studies.

Regional Centre for Applied Molecular Oncology Masaryk Memorial Cancer Institute Brno Czech Republic

See more in PubMed

Dobbelstein M, Moll U. Targeting tumour-supportive cellular machineries in anticancer drug development. Nat Rev Drug Discov. 2014; 13(3): 179–196. 10.1038/nrd4201 PubMed DOI

Trepel J, Mollapour M, Giaccone G, Neckers L. Targeting the dynamic HSP90 complex in cancer. Nat Rev Cancer. 2010; 10(8): 537–549. 10.1038/nrc2887 PubMed DOI PMC

Neckers L, Workman P. Hsp90 molecular chaperone inhibitors: are we there yet?. Clin Cancer Res. 2012; 18(1): 64–76. 10.1158/1078-0432.CCR-11-1000 PubMed DOI PMC

Cheng Q, Chang JT, Geradts J, Neckers LM, Haystead T, Spector NL, et al. Amplification and high-level expression of heat shock protein 90 marks aggressive phenotypes of human epidermal growth factor receptor 2 negative breast cancer. Breast Cancer Res. 2012; 14(2): R62 10.1186/bcr3168 PubMed DOI PMC

Pick E, Kluger Y, Giltnane JM, Moeder C, Camp RL, Rimm DL, et al. High HSP90 expression is associated with decreased survival in breast cancer. Cancer Res. 2007; 67(7): 2932–2937. 10.1158/0008-5472.CAN-06-4511 PubMed DOI

Wang J, Cui S, Zhang X, Wu Y, Tang H. High expression of heat shock protein 90 is associated with tumor aggressiveness and poor prognosis in patients with advanced gastric cancer. PLoS ONE. 2013; 8(4): e62876 10.1371/journal.pone.0062876 PubMed DOI PMC

Kamal A, Boehm MF, Burrows FJ. Therapeutic and diagnostic implications of Hsp90 activation. Trends Mol Med. 2004; 10(6): 283–290. 10.1016/j.molmed.2004.04.006 PubMed DOI PMC

Kamal A, Thao L, Sensintaffar J, Zhang L, Boehm MF, Fritz LC, et al. A high-affinity conformation of Hsp90 confers tumour selectivity on Hsp90 inhibitors. Nature. 2003; 425(6956): 407–410. 10.1038/nature01913 PubMed DOI

Muller P, Ruckova E, Halada P, Coates PJ, Hrstka R, Lane DP, et al. C-terminal phosphorylation of Hsp70 and Hsp90 regulates alternate binding to co-chaperones CHIP and HOP to determine cellular protein folding/degradation balances. Oncogene. 2013; 32(25): 3101–3110. 10.1038/onc.2012.314 PubMed DOI

Rodrigues LM, Chung YL, Al Saffar NM, Sharp SY, Jackson LE, Banerji U, et al. Effects of HSP90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) on NEU/HER2 overexpressing mammary tumours in MMTV-NEU-NT mice monitored by Magnetic Resonance Spectroscopy. BMC Res Notes. 2012; 5: 250 10.1186/1756-0500-5-250 PubMed DOI PMC

Alexandrova EM, Yallowitz AR, Li D, Xu S, Schulz R, Proia DA, et al. Improving survival by exploiting tumour dependence on stabilized mutant p53 for treatment. Nature. 2015; 523(7560): 352–356. 10.1038/nature14430 PubMed DOI PMC

Miyata Y, Nakamoto H, Neckers L. The therapeutic target Hsp90 and cancer hallmarks. Curr Pharm Des. 2013; 19(3): 347–365. PubMed PMC

Zismanov V, Drucker L, Gottfried M. Combined inhibition of Hsp90 and the proteasome affects NSCLC proteostasis and attenuates cell migration. Anticancer Drugs. 2014; 25(9): 998–1006. 10.1097/CAD.0000000000000140 PubMed DOI

Ruckova E, Muller P, Nenutil R, Vojtesek B. Alterations of the Hsp70/Hsp90 chaperone and the HOP/CHIP co-chaperone system in cancer. Cell Mol Biol Lett. 2012; 17(3): 446–458. 10.2478/s11658-012-0021-8 PubMed DOI PMC

Fogh J, Wright WC, Loveless JD. Absence of HeLa cell contamination in 169 cell lines derived from human tumors. J Natl Cancer Inst. 1977;58(2):209–14. PubMed

Lasfargues EY, Coutinho WG, Redfield ES. Isolation of two human tumor epithelial cell lines from solid breast carcinomas. J Natl Cancer Inst. 1978;61(4):967–78. PubMed

Littlewood-Evans AJ, Bilbe G, Bowler WB, Farley D, Wlodarski B, Kokubo T, et al. The osteoclast-associated protease cathepsin K is expressed in human breast carcinoma. Cancer Res. 1997;57(23):5386–90. PubMed

Soule HD, Vazguez J, Long A, Albert S, Brennan M. A human cell line from a pleural effusion derived from a breast carcinoma. J Natl Cancer Inst. 1973;51(5):1409–16. PubMed

Cailleau R, Olivé M, Cruciger QV. Long-term human breast carcinoma cell lines of metastatic origin: preliminary characterization. In Vitro. 1978;14(11):911–5. PubMed

Keydar I, Chen L, Karby S, Weiss FR, Delarea J, Radu M, et al. Establishment and characterization of a cell line of human breast carcinoma origin. Eur J Cancer. 1979;15(5):659–70. PubMed

Trempe GL. Human breast cancer in culture. Recent Results Cancer Res. 1976;(57):33–41. PubMed

Song CH, Park SY, Eom KY, Kim JH, Kim SW, Kim JS, et al. Potential prognostic value of heat-shock protein 90 in the presence of phosphatidylinositol-3-kinase overexpression or loss of PTEN, in invasive breast cancers. Breast Cancer Res. 2010; 12(2): R20 10.1186/bcr2557 PubMed DOI PMC

Buffart TE, Carvalho B, van Grieken NC, van Wieringen WN, Tijssen M, Kranenbarg EM, et al. Losses of chromosome 5q and 14q are associated with favorable clinical outcome of patients with gastric cancer. Oncologist. 2012; 17(5): 653–662. 10.1634/theoncologist.2010-0379 PubMed DOI PMC

Jhaveri K, Chandarlapaty S, Iyengar N, Morris PG, Corben AD, Patil S, et al. Biomarkers That Predict Sensitivity to Heat Shock Protein 90 Inhibitors. Clin Breast Cancer. 2016; 16(4): 276–283. 10.1016/j.clbc.2015.11.004 PubMed DOI PMC

Alarcon SV, Mollapour M, Lee MJ, Tsutsumi S, Lee S, Kim YS, et al. Tumor-intrinsic and tumor-extrinsic factors impacting hsp90- targeted therapy. Curr Mol Med. 2012; 12(9): 1125–1141. PubMed PMC

Pearl LH, Prodromou C, Workman P. The Hsp90 molecular chaperone: an open and shut case for treatment. Biochem J. 2008; 410(3): 439–453. 10.1042/BJ20071640 PubMed DOI

Dai C, Sampson SB. HSF1: Guardian of Proteostasis in Cancer. Trends Cell Biol. 2016; 26(1): 17–28. 10.1016/j.tcb.2015.10.011 PubMed DOI PMC

Akerfelt M, Morimoto RI, Sistonen L. Heat shock factors: integrators of cell stress, development and lifespan. Nat Rev Mol Cell Biol. 2010; 11(8): 545–55. 10.1038/nrm2938 PubMed DOI PMC

Chen S, Smith DF. Hop as an adaptor in the heat shock protein 70 (Hsp70) and hsp90 chaperone machinery. J Biol Chem. 1998; 273(52): 35194–35200. PubMed

Ballinger CA, Connell P, Wu Y, Hu Z, Thompson LJ, Yin LY, et al. Identification of CHIP, a novel tetratricopeptide repeat-containing protein that interacts with heat shock proteins and negatively regulates chaperone functions. Mol Cell Biol. 1999; 19(6): 4535–4545. PubMed PMC

Meacham GC, Patterson C, Zhang W, Younger JM, Cyr DM. The Hsc70 co-chaperone CHIP targets immature CFTR for proteasomal degradation. Nat Cell Biol. 2001; 3(1): 100–105. 10.1038/35050509 PubMed DOI

Klijn C, Durinck S, Stawiski EW, Haverty PM, Jiang Z, Liu H, et al. A comprehensive transcriptional portrait of human cancer cell lines. Nat Biotechnol. 2015;33(3):306–12. 10.1038/nbt.3080 PubMed DOI

Caldas-Lopes E, Cerchietti L, Ahn JH, Clement CC, Robles AI, Rodina A, et al. Hsp90 inhibitor PU-H71, a multimodal inhibitor of malignancy, induces complete responses in triple-negative breast cancer models. Proc Natl Acad Sci USA. 2009; 106(20): 8368–8373. 10.1073/pnas.0903392106 PubMed DOI PMC

Bagatell R, Paine-Murrieta GD, Taylor CW, Pulcini EJ, Akinaga S, Benjamin IJ, Whitesell L (2000) Induction of a heat shock factor 1-dependent stress response alters the cytotoxic activity of hsp90-binding agents. Clin Cancer Res. 2000;6(8):3312–8 PubMed

Rodina A, Wang T, Yan P, Gomes ED, Dunphy MP, Pillarsetty N, et al. The epichaperome is an integrated chaperome network that facilitates tumour survival. Nature. 2016; 538(7625): 397–401. 10.1038/nature19807 PubMed DOI PMC