The Plasma Cell Transcription Factor XBP1 is Required To Mitigate The Unfolded Protein Response In Ph⁺ ALL

The transcription factors X-box binding protein 1 (XBP1) and PR domain zinc finger protein 1 (PRDM1) are critical for terminal differentiation of B cells into plasma cells but have no known role at earlier stages of B-cell development. Additionally, XBP1 is a critical effector molecule during the unfolded protein response (UPR), which is required to salvage ER stress and to promote cell survival. Previous work identified XBP1 and PRDM1 as central requirements for plasma cell development and as critical mediators of cell survival in plasma cell-derived multiple myeloma.

We report the unexpected finding that XBP1 and PRDM1 genes are hypermethylated at their promoters and not expressed in normal pre-B cells, whereas pre-B cell-derived Ph⁺ ALL cells express both molecules at surprisingly high levels. Interestingly, XBP1 expression levels are linked to BCR-ABL1 kinase activity and transcriptionally activated by STAT5 and repressed by the tumor suppressor BACH2.

Furthermore, high expression levels of XBP1 at diagnosis predict poor overall survival (OS) and relapse-free survival (RFS) of leukemia patients in two clinical trials for patients with high risk acute lymphoblastic leukemia (n=207; COG P9906 trial; p=8.95e-06 and ECOG E2993; n=215; p=0.017). A multivariate analysis showed that high expression of XBP1 is an independent predictor of poor outcome regardless of other established risk factors such as white blood cell count (WBC) and minimal residual disease (MRD) (n=207; COG P9906 trial; WBC; p=8.95e-06 and MRD; p=5.85e-05). Patients with a high expression of XBP1 and low expression of BACH2 profile had the worst RFS. In addition, high levels of XBP1also correlated with positive MRD status at day 29 after onset of chemotherapy.

To study the function of Xbp1 in Ph⁺ ALL, we developed a genetic system for inducible deletion of Xbp1 in a mouse model of Ph⁺ ALL based on bone marrow B cells from mice carrying loxP-flanked allele of Xbp1 (Xbp1^fl/fl). On the basis of this model, bone marrow B cell precursors were transformed by BCR-ABL1 in the presence of IL7. Inducible Cre-mediated deletion of Xbp1 was achieved by transduction of leukemia cells with tamoxifen (4-OHT)-inducible Cre. Interestingly, 4-OHT-induced deletion of Xbp1 in Ph⁺ ALL-like leukemia cells caused rapid cell death within two days of induction. Xbp1-deletion resulted in extensive apoptosis, cellular senescence and cell cycle arrest associated with increased levels of p53, p21 and Arf. Similarly, in an in vivo setting, deletion of Xbp1 substantially prolonged survival of NOD-SCID transplant recipient mice (n=7; p=0.007). Mechanistically, deletion of Xbp1leads to increased expression of the pro-apoptotic molecule CHOP and phosphorylation of the stress MAP kinases p38 and JNK.

To investigate the potential clinical relevance of these findings, we used recently identified small-molecule inhibitors for XBP1 activation (Papandreou et al., 2011; Volkmann et al., 2011; Kriss et al., 2012). Small-molecule inhibition of XBP1 activation overall mimicked the findings in genetic experiments. Importantly, pharmacological targeting of XBP1 also induced cell death in multiple patient-derived cases of Ph⁺ ALL carrying the T315I mutations, which confers far-reaching TKI-resistance and caused reduction of leukemia-initiating cells (LICs) in patient-derived TKI-resistant Ph⁺ ALL cells in vivo and significantly prolonged survival of leukemia-bearing NOD-SCI mice (n=5 per group).

Collectively, these studies identify XBP1 as an important survival factor and as a potential therapeutic target to overcome drug-resistance in Ph⁺ ALL. Like transformed plasma cells in multiple myeloma, Ph⁺ ALL cells are selectively sensitive to ER stress and critically dependent on XBP1 and likely other factors of the UPR pathway. Clinical validation of this concept could lead to improved treatment options for patients with TKI-resistant Ph⁺ ALL.

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