Abstract A08: Systematic identification of genetic vulnerabilities synergizing with proteasome and IRE1 inhibition in multiple myeloma cells using ultra-complex shRNA libraries

Multiple myeloma (MM) is characterized by the clonal expansion of malignant plasma cells in the bone marrow. MM remains incurable; thus, effective therapies remain an unmet clinical need. MM cells produce large amounts of paraproteins, which impose a sustained demand on the endoplasmic reticulum (ER) resulting in chronic ER stress. Standard-of-care therapies, like the proteasome inhibitor bortezomib, take advantage of the reliance of MM cells on ER function. By blocking the proteasome, client protein load exceeds the capacity of the proteasome, inducing unmitigatable ER stress and apoptosis. However, most MM cells develop resistance to bortezomib, indicating that genetic escape routes are activated upon selective pressure. Adaptive measures to counteract ER stress are controlled by the unfolded protein response (UPR), which is orchestrated by a network of three intertwined signaling pathways, the most conserved of which is activated by the ER stress sensor IRE1 and its effector, the transcription factor XBP1. IRE1/XBP1 are part of a cell-protective response that, in healthy cells, permits adaptation in response to upsurges in ER function, while in MM, they are thought to promote survival. We show that genetic susceptibilities of MM cells can be identified to guide combination therapies using bortezomib or a chemical inhibitor of IRE1. To this end, we systematically interrogated genetic vulnerabilities involving the ER stress response in MM cells using an unbiased genome-wide RNA interference approach that employs screening of ultra-complex shRNA libraries. Conducting pooled shRNA screens in conventional and microcarrier-based cell culture systems, we show that, unlike bortezomib, IRE1 inhibition alone does not lead to tumor cell death, suggesting that IRE1 is not a single fate determinant. Rather, we show that IRE1 represents an Achilles heel that allows identification of combinatorial genetic vulnerabilities resulting in synthetic lethality. The same rationale was applied to proteasome inhibition, where we identified several genes whose diminished function resulted in synthetic lethality. In this way, we identified novel pathways that when modulated in parallel with proteasome inhibition block escape routes leading to drug resistance. By applying our screening method, we validated known genetic susceptibilities of MM cells, identified new ER-associated MM survival factors, and uncovered vulnerabilities that result in synthetic lethality when combined with pharmacological inhibition of IRE1 or the proteasome. Analyses of published gene expression datasets from MM patient samples suggest that the expression levels of some of the identified genes are predictive of poor survival. Taken together, our data validate the screening method as a powerful tool for the discovery of diagnostic biomarkers and novel therapy targets. Citation Format: Diego Acosta-Alvear, Martin Kampmann, Michael C. Bassik, Crystal P. Lee, Marc A. Shuman, Jonathan S. Weissman, Peter Walter. Systematic identification of genetic vulnerabilities synergizing with proteasome and IRE1 inhibition in multiple myeloma cells using ultra-complex shRNA libraries. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Synthetic Lethal Approaches to Cancer Vulnerabilities; May 17-20, 2013; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(5 Suppl):Abstract nr A08.