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Targeting IRE1α-XBP1 pathway is a novel therapeutic strategy in multiple myeloma
Naoya Mimura, MD
Dana-Farber Cancer Institute
Boston, Massachusetts, USA

Aberrant protein folding results in the accumulation of misfolded/unfolded proteins in the endoplasmic reticulum (ER), which in turn triggers ER stress followed by unfolded protein response (UPR), an adaptive response against ER stress. Since multiple myeloma (MM) cells have high protein synthesis, they are sensitive to ER stress and require strict ER quality control for cell survival.

Upon UPR, IRE1α is activated by auto-phosphorylation, resulting in activation of its endoribonuclease domain to splice XBP1 mRNA from XBP1 unspliced form (XBP1u: inactive) to XBP1 spliced form (XBP1s: active). Since XBP1 is a transcription factor regulating genes which are responsible for protein folding and ER associated degradation (ERAD), IRE1α-XBP1 pathway acts as a pro-survival signaling pathway under the UPR condition. There are some reports on association between MM and XBP1. To begin with, XBP1 is required for the generation of plasma cells. In addition, XBP1 is highly expressed in human MM cells. Moreover, knockdown of XBP1s by siRNA sensitizes MM cells to stress-induced apoptosis. Importantly, it has been reported that the presence of high levels of XBP1s in MM cells is associated with a poor overall survival. My preliminary data also suggest that IRE1α has a crucial role in MM cell survival evidenced by knock-down of IRE1α using shRNA.

In this study I will examine whether IRE1α-XBP1 pathway is a potential novel therapeutic option in MM. I have obtained a small molecule IRE1α endoribonuclease inhibitor MKC-3946 (MannKind, Valencia CA). I confirmed that this compound inhibited the endoribonuclease of IRE1α, leading to inhibition of XBP1 being spliced in MM cells. My preliminary data shows that MKC-3946 induced only modest cytotoxicity in MM cell lines. Conversely, MKC-3946 enhanced growth inhibition by bortezomib in MM cells. I confirmed that bortezomib induced ER stress, evidenced by induction of XBP1s. In contrast, MKC-3946 blocked XBP1s triggered by bortezomib.

These results suggest that inhibition of XBP1 splicing has a therapeutic potential in combination with ER stress inducer such as bortezomib. In this proposal I will further evaluate combination treatments with other agents known to induce ER stress including HSP90 inhibitor and with other agents iucluding dexamethasone, immunomodulatory drugs, HDAC inhibitors, and Akt inhibitors. Moreover, I will examine whether MKC-3946 alone and in combination with ER stress inducers can overcome protective effect by cytokines such as IL6 from bone marrow stromal cells (BMSCs) using our models of MM cells in the bone marrow microenvironment. To elucidate the mechanism of inhibition of XBP1 splicing, I will perform the qRT-PCR in order to evaluate expression of genes responsible for protein folding and ERAD. After having established the efficacy of the MKC-3946 in vitro, I will go on to in vivo study. Using our plasmacytoma model 9 and SCID-hu model, I will determine the efficacy of MKC-3946 therapy on human MM cells in the human BM milieu (human cytokines), as in prior studies 10. These studies will provide the framework for clinical trials of MKC-3946, alone and with bortezomib, to improve patient outcome in MM.

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