Irestatin, a potent inhibitor of IRE1α and the unfolded protein response, is a hypoxia-selective cytotoxin and impairs tumor growth
3514 Background: The unfolded protein response (UPR) is an adaptive response to the toxic accumulation of misfolded proteins in the endoplasmic reticulum (ER), and is activated in solid tumors. IRE1a is a core component of the UPR and responds to ER stress through activation of its dual kinase and endonuclease domains. The IRE1a endonuclease splices the mRNA for XBP-1, generateing a potent transcription factor that is required for tumor growth. Methods: We developed a fibrosarcoma cell line expressing a fusion of unprocessed XBP-1 inserted upstream of firefly luciferase. Under ER stress, IRE1a catalyzes the removal of a 26-nt intronic sequence from the XBP-1 mRNA, introducing a shift in reading frame that permits translation of luciferase. We screened a chemical library of 66,000 small molecules for inhibitors of XBP-luciferase activity and identified 12 molecules, termed irestatins, which consistently inhibited the IRE1a signaling module without affecting the activity of a control CMV-luciferase reporter. We pursued several of the most potent irestatins, including irestatin 9389, which inhibited XBP-luciferase activity with an IC50 of ∼25 nM. Results: Irestatin-mediated inhibition of the IRE1a endonuclease impairs the growth of malignant myeloma cells and inhibits the survival of oxygen-starved tumor cells in vitro. Exposure to irestatin 9389 (2.5 μM) had a negligible effect on the survival of HT1080 cells cultured under normal oxygen conditions. However, in cells cultured under hypoxia for 48 hours, irestatin 9389 inhibited colony formation by nearly 80-fold (48% vs. 0.62%). A two-week course of treatment with single-agent irestatin 9389 (50 mg/kg) administered every other day by i.p. bolus injection was well tolerated and strongly inhibited the growth of subcutaneous HT1080 tumor xenografts (1790 ± 380 mm3 vs. 480 ± 210 mm3; P<0.01). Conclusion: Irestatins define a novel class of hypoxia- and ER stress-selective agents targeted to the underlying physiological response to the tumor microenvironment. Intratumoral inhibition of the UPR can potentiate cell death and impair tumor growth. Molecular intervention against central components of the UPR may represent an effective therapeutic strategy in cancer treatment. No significant financial relationships to disclose.