Abstract
Repurposing approved and abandoned non-oncological drugs is an alternative developmental strategy for the identification of anticancer therapeutics that has recently attracted considerable attention. Due to the essential role of the cellular heat shock response in cytoprotection through the maintenance of proteostasis and suppression of apoptosis, small molecule heat shock response antagonists can be harnessed for targeted induction of cytotoxic effects in cancer cells. Guided by gene expression array analysis and a phenotypic screen interrogating a collection of 3,7-diamino-phenothiazinium derivatives, we have identified the redox-drug methylene blue (MB), used clinically for the infusional treatment of methemoglobinemia, as a negative modulator of heat shock response gene expression in human metastatic melanoma cells. MB-treatment blocked thermal (43 °C) and pharmacological (celastrol, geldanamycin) induction of heat shock response gene expression, suppressing Hsp70 (HSPA1A) and Hsp27 (HSPB1) upregulation at the mRNA and protein level. MB sensitized melanoma cells to the apoptogenic activity of geldanamycin, an Hsp90 antagonist known to induce the counter-regulatory upregulation of Hsp70 expression underlying cancer cell resistance to geldanamycin chemotherapy. Similarly, MB-cotreatment sensitized melanoma cells to other chemotherapeutics (etoposide, doxorubicin). Taken together, these data suggest feasibility of repurposing the non-oncological redox drug MB as a therapeutic heat shock response antagonist for cancer cell chemosensitization.
Highlights
When exposed to cytotoxic stress, mammalian cells activate a number of conserved cytoprotective molecular pathways including the heat shock response, the ER stress/unfolded protein response, and the oxidative stress response [1,2,3]
We demonstrate that methylene blue (MB)-treatment attenuates thermally- and pharmacologically-induced heat shock response gene expression, suppressing Hsp70 (HSPA1A) and Hsp27 (HSPB1) upregulation at the mRNA and protein level, and causing chemosensitization of melanoma cells to geldanamycin cytotoxicity
After demonstrating that MB exposure downregulates protein levels of Hsp70 and Hsp27 in human melanoma cells cultured under standard conditions, we examined the possibility that MB exposure could attenuate cellular heat shock response gene expression elicited upon thermal or pharmacological challenge (Figure 4)
Summary
When exposed to cytotoxic stress, mammalian cells activate a number of conserved cytoprotective molecular pathways including the heat shock response, the ER stress/unfolded protein response, and the oxidative stress response [1,2,3]. The cytosolic chaperone heat shock protein 90 (Hsp90) represents a promising molecular target for chemotherapeutic intervention due to its causative involvement in the stabilization of mutant client proteins and facilitation of oncogenic signaling cascades, and ansamycin-based and non-ansamycin-based small molecule inhibitors of Hsp are currently under clinical evaluation in human cancer patients [11,12]. In the Hsp family, the stress-inducible heat shock protein Hsp ( called Hsp or Hsp, encoded by HSPA1A), an ATP-dependent chaperone overexpressed in many tumor types, has emerged as a causative factor in tumorigenesis and is considered a high quality target for therapeutic intervention aiming at cancer cell chemosensitization [5,10]. We demonstrate that MB-treatment attenuates thermally- and pharmacologically-induced heat shock response gene expression, suppressing Hsp (HSPA1A) and Hsp (HSPB1) upregulation at the mRNA and protein level, and causing chemosensitization of melanoma cells to geldanamycin cytotoxicity. We present prototype data that suggest feasibility of repurposing MB as an investigational drug for the attenuation of heat shock response gene expression causing chemosensitization of human metastatic melanoma cells
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