Intrinsic proteotoxic stress levels vary and act as a predictive marker for sensitivity of cancer cells to Hsp90 inhibition

Abstract

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.

Prodigiosin/PU-H71 as a novel potential combined therapy for triple negative breast cancer (TNBC): preclinical insights

“…All treatment groups decreased the levels of EGFR protein expression. Our finding is in harmony with two recent studies that explained that targeting HSP90α provides the likelihood of simultaneously disrupts EGFR is one of the HSP90α client proteins and a hallmark of cancer 26 , 98 .…”

“…All treatment groups decreased the levels of EGFR protein expression. Our finding is in harmony with two recent studies that explained that targeting www.nature.com/scientificreports/ HSP90α provides the likelihood of simultaneously disrupts EGFR is one of the HSP90α client proteins and a hallmark of cancer 26,98 . Amongst other types of breast cancers, TNBC has the most extensive vascularisation with significantly high expression levels of VEGF.…”

The sensitivity to Hsp90 inhibitors of both normal and oncogenically transformed cells is determined by the equilibrium between cellular quiescence and activity

“…The differential sensitivity of cancer over normal cells to Hsp90 inhibitors and thus their differential dependence on Hsp90 has been widely accepted in the community, and yet, the evidence has remained somewhat anecdotal and circumstantial and the mechanisms very poorly understood [13–24]. Nevertheless, this has formed the basis for numerous clinical trials; currently, a search with the keyword "Hsp90" in clinicaltrials.gov yields 124 hits, of which 18 are active and/or recruiting.…”

“…Due to unique features of the N-terminal ATP binding pocket of Hsp90, specific competitive inhibitors of Hsp90 have been developed [11,12]. Intriguingly, cancer cells were found to be more sensitive to Hsp90 inhibitors than normal cells, conceivably reflecting their increased dependency on Hsp90 chaperone activity; however, although a number of molecular mechanisms have been proposed to explain this difference, the precise reasons have remained controversial and incompletely characterized [13–24]. The difference in sensitivity suggested a therapeutic window and prompted high hopes for the treatment of cancer and led to a large number of clinical trials.…”

Proteotoxic Stress and Cell Death in Cancer Cells

“…Clearly, impacting these adaptive mechanisms has important consequences to the survival of cancer cells [209,210]. This dependence has attracted interest in developing therapeutic approaches aimed at switching-off these adaptations and thus unleashing all the dramatic consequences of the unresolved proteotoxic stress [210][211][212][213][214][215][216][217][218]. In some circumstances, adaptations to proteotoxic stress can favor the resistance to other therapeutic regiments, as observed for HSF1 and the resistance to the receptor tyrosine kinase (RTK) inhibitor lapatinib in breast cancer [219].…”