Investigation of IRE1/XBP1s pathway and its potential as a therapeutic target in breast cancer

Abstract

X-Box binding protein 1 (XBP1) is an integral component of the unfolded protein response (UPR), a pro-survival mechanism, triggered by the onset of endoplasmic reticulum stress. Activation of the endoplasmic reticulum localized stress sensor IRE1α removes a 26 nucleotide intron from XBP1 mRNA generating the active transcription factor XBP1 spliced (XBP1s). Signalling via this arm of the UPR is generally thought to be pro-survival and represents a mechanism through which cells can endure stressful conditions. High levels of XBP1s are linked to breast cancer, with overexpression reported across a range of subtypes where it correlated with a worse patient outcome. In this study routinely used breast cancer cell lines (MCF-7, T47D, SKBR3, MDA-MB231) were found to have basal protein expression of XBP1s. To determine the relevance of basal XBP1s levels a novel chemical inhibitor of IRE1 was employed which specifically inhibits IRE1 ribonuclease activity and therefore block XBP1 splicing. Addition of the IRE1 inhibitor efficiently reduced basal levels of XBP1s in all cell lines tested. Prolonged treatment with the IRE1 inhibitor significantly reduced cell proliferation in breast cancer cells and induced cell death under reduced serum conditions. Interesting, cell death was found to be mediated through modulation of extracellular factors. Moreover, screening of commonly used anti-cancer drugs revealed that IRE1 inhibition could enhance the cytotoxicity of commonly used anticancer drugs. In parallel to this study a novel downstream target of IRE1/XBP1s was identified; Sestrin 2. Sestrin 2 knockdown highlighted the pro-survival effects of Sestrin 2 during treatment of breast cancer cells with Methotrexate and Bortezomib, presenting a new therapeutic target in breast cancer treatment. In summary, this thesis investigates the role of IRE1/XBP1s signalling in breast cancer and demonstrates the clinical potential of IRE1 inhibition.