Unravel the molecular mechanism of XBP1 in regulating the biology of cancer cells.

The Myc family includes proto-oncogenes ( MYC , MYCN , and MYCL ), which regulate several lines of genes coordinating cellular processes, such as proliferation, differentiation, and metabolism 74 . IRE1-XBP1 signaling pathway is involved in both c-Myc- and N-Myc-driven cancers, sustaining the growth and survival of tumor cells through increasing the activity of stearoyl-CoA-desaturase 1 (SCD1). XBP1 inhibition-induced cell apoptosis in a Myc-dependent manner can be partially reversed by exogenous unsaturated fatty acids. Inhibition of SCD1 can suppress IRE1α RNase activity in vivo. In addition, inhibition of IRE1-XBP1 signaling pathway augments the sensitivity of standard chemotherapy drugs to c-Myc-overexpression Burkitt's lymphoma, which is a highly aggressive malignancy 75 . Myc up regulates the transcriptional activity of IRE1-XBP1 signaling pathway through binding to the promoter of IRE1. XBP1 forms a complex with Myc in the nucleus and subsequently increases the transcriptional activity of XBP1. However, XBP1 does not have any effects on Myc transcriptional activity. Inhibition of XBP1 induces cells to be more vulnerable with increased expression of Myc in breast cancer cells 76 , 77 . IRE1α-XBP1s signaling is often up regulated in acute myeloid leukemia (AML) cells. The ratios of XBP1s/XBP1u are associated with poor survival 78 . Inhibitors of IRE1α-XBP1s signaling have been reported to induce caspase-dependent apoptosis and inhibition of proliferation with cell cycle arrest at G1 phage, as indicated by inhibition of CDK4, c-Myc, Bcl-2, and cyclin D1 expression and induction of p21 cip1 and p27 kip1 expression 79 . Consistently, XBP1 expression is also enhanced under hypoxia and essential for osteosarcoma growth and survival through regulating the activity of PI3K/mTOR signaling pathway 80 . In glioma cells, the expression of XBP1 is significantly up regulated in vivo. Knockdown of XBP1 decreases cells viability and ATP production by inhibition of Hexokinase II (HK-II) expression and glycolysis under hypoxia, leading to increased apoptosis and decreased clonogenic survival of glioma cells 81 . Similar scenario has been found in prolactinoma GH3 cells, as indicated that XBP1 significantly promotes cell proliferation and inhibits cell apoptosis 82 , 83 . In BM stromal cells from MM patients, XBP1 deletion compromises the increased expression of VCAM-1, IL-6, and RANKL induced by TNFα and reverses the growth of MM cells and the formation of osteoclast 84 . Ewing's sarcoma (ES) is the frequent pediatric bone tumor with the main pathological mechanism of EWS/FLI1 gene fusion. Recently, the proteomic studies in EWS/FLI1-knock down ES cell lines and human mesenchymal stem cells (hMSCs) show that XBP1 plays a crucial role in cell proliferation. Silence of XBP1 significantly decreases cell viability and induces cell apoptosis in vivo and in vitro 85 . Hepatocyte nuclear factor 4α (HNF4α), phenocopying the effects of XBP1, is a critical developmental mediator of differentiation. Mechanistically, HNF4α binds to the promoter of XBP1 and activates and maintains its expression 86 . In addition, XBP1 activates but not maintains the expression of MIST1 by binding to the promoter of MIST1, which regulates secretory vesicle trafficking in zymogenic chief cells (ZCs) 87 . Thus, HNF4α promotes homeostatic proliferation, which is associated with partial induction and maintenance of XBP1-MIST1 signaling pathway in enzyme-secreting ZCs 86 . Gene expression profile study indicates that XBP1s is involved in regulation of BCL-2 and several other genes related to cell cycles and apoptosis. P53, a tumor suppressor, is often mutated in human cancers. Mutated p53 with defect responses induces tumor development. Deletion of p53 induces the expression of IRE1α by inhibiting the p53-mediated association of IRE1α with synoviolin-1, which promotes the degradation of IRE1α 88 . In irradiated p53-mutated cervical cancer cells, the expression and the stability of IRE1α are significantly increased by RITA (reactivation of p53 and induction of tumor cell apoptosis), which is a small molecule binding to p53 directly and inducing cell apoptosis in wild-type p53 or mutant p53 expressing tumor cells 89 . XBP1s has been found to attenuate the transcriptional expression of Pin1. However, Pin1, in a phosphorylation-dependent manner, binds to XBP1s and forms a complex to stabilize XBP1s through the consensus recognition sequence (Ser/Thr-Pro) and promotes its functions, such as cell proliferation and transformation. Knockout of Pin1 induces decreased expression of XBP1s 19 .

Activation of UPR has been related to therapeutic resistance and recurrence. The expression of XBP1 is up regulated in endocrine-resistant breast cancer cells and induces cells resistance to anti-estrogens. Tamoxifen has been widely used for reducing mortality and recurrence in patients with estrogen receptor (EsR)-positive breast tumors. Unfortunately, about half of patients with EsR-positive breast cancer show chemoresistance, which remains a challenge for managing breast cancer. XBP1s expression has been demonstrated to be highly correlated with poor survival in EsR-positive breast cancer among over 170 patients, but not in EsR-negative breast cancer. STF-083010, an inhibitor of XBP1 splicing, can reverse the sensitivity of tamoxifen to resistant MCF-7 cells and delay the progression of breast cancer in a xenograft mammary tumor model 102 . Advanced metastatic breast cancer often expresses EsRα mutations EsRαY537S and EsRαD538G and resists to endocrine therapy, as indicated by robust proliferation when exposing to anti-estrogen. The proliferation in EsRα-mutated breast cancer cells is completely blocked by EsRα biomodulator BHPI, which induces hyper-activation of UPR and low expression of IRE1α-XBP1s signaling pathway estrogen-independently 103 . XBP1 stimulates the transcriptional expression of EsRα in an estrogen-independent manner. Mutually, XBP1 mRNA expression can be up regulated by 17β-estradiol with a positive feedback loop 104 . Activation of XBP1 is closely related to tumor progression and negative chemotherapy in EsRα-positive breast cancers and TNBC 53 . In addition to EsRα, EsRβ decreases the tumor survival in both anti-estrogen-sensitive and -resistant breast cancer cells. Up regulation of EsRβ increases the sensitivity to tamoxifen management, resulting in cell apoptosis in chemoresistant cells. The possible mechanism might be associated with repression of IRE1 and XBP1 expression at the post-transcriptional level 105 . Inhibition of XBP1 re-sensitizes anti-estrogen-resistant breast cancer cells to anti-estrogens. NF-κB signaling also plays a critical role in management of anti-estrogen-resistant breast cancers. Inhibition of NF-κB signaling decreases the anti-estrogen resistance of breast cancer cells mediated by XBP1 overexpression. This might be related to refine-tune the balance between apoptosis and autophagy 106 . FOXC1, a member of Forkhead box transcription factors, is expressed consistently and exclusively in basal-like breast cancers (BLBCs) associated with poor clinical outcome. In estrogen-negative breast cancer, the expression of FOXC1 is up regulated. Overexpression of FOXC1 is associated with reduction of EsRα expression and sensitivity to E2 and tamoxifen by down regulating the expression of XBP1 and insulin receptor substrate 1 (IRS1), leading to increased property of tumorigenesis 107 . Thus, overexpression of XBP1s decreases sensitivity to tamoxifen-induced growth inhibition and increased estrogen-independent cell growth 14 . Both XBP1s and XBP1u show regulatory activity on NF-κB signaling via EsRα. XBP1u is beyond the dominant negative isoform of XBP1s, which exhibits more potent in regulating p65 expression directly. Deletion of p65 might become the promising strategy to re-sensitize XBP1-overexpressing breast cancer cells to anti-estrogens 106 . β-catenin, a critical factor in Wnt signaling pathway, has been implicated in EMT and chemo-resistance. XBP1 binds to the promoter of β-catenin and activates its expression, promoting cell growth. The natural product analogue CYD 6-17 mechanistically targets to the gene expression of β-catenin through attenuating the binding activity of XBP1 in transitional cell carcinoma (TCC), compromising the resistance of TCC to various chemo-therapeutics 108 .

Imbalanced proteostasis appears as a hallmark of cancer development and metastasis. Increasing evidence demonstrates that endoplasmic reticulum (ER), an organelle for folding protein and controlling quality, has been showed to play a crucial role during cancer development since the mid-1990s 1 , 2 . Perturbation-induced accumulation of un-folded or mis-folded protein in ER leads to a cyto-protective response called un-folded protein response (UPR). Three critical UPR-associated signaling pathways (IRE1α, PERK, and ATF6) are involved in responding to ER stress by enhancing the ER capacity, promoting ER-associated degradation and chaperone functions, and triggering cell apoptosis if unabated. Cellular adaptation is achieved by activation of UPR. Inositol-requiring enzyme 1α (IRE1α)-X-box-binding protein 1 (XBP1) signaling is the most conserved branch among the three UPR signaling pathways and plays a crucial role in maintaining ER homeostasis. Spliced XBP1 (XBP1s) is the direct downstream factor of IRE1α with activated RNase activity through splicing and removing 26 nucleotide intron from un-spliced XBP1 (XBP1u) ( Figure 1 ), which is firstly identified as a key transcriptional regulator of major histocompatibility complex (MHC) class II in B cells in the early 1990s 3 - 5 . Activation of IRE1α-XBP1s signaling has been implicated in cancer development and metastasis 6 , and targeting XBP1 as an anti-cancer strategy has been reviewed in 2006 by Koong, et al 7 . Cells with XBP1-deficiency significantly decrease their ability of carcinogenesis in nude mice 8 . The expression of XBP1s in nuclei is dramatically related to poor clinical survival. However, no relationships between cytoplasmic XBP1s and survival are found 9 . FK506-binding protein 13 (FKBP13), a chaperone that acts as the downstream factor of XBP1, has been demonstrated to interact with the surplus Ig molecules or mis-folded proteins to the ubiquitin-dependent degradation system in plasma cells, leading to amelioration of ER stress 10 . Thus, XBP1-FKBP13 axis becomes the potential target for managing diseases. In this review, we are focusing on the critical role of XBP1 in mediating the biological activity of cancer cells. We mainly used “XBP1, cancer”, “XBP1, tumor”, “XBP1 carcinoma” as the keywords to primarily search literatures on Pubmed, Europe PMC, and Sciencedirect databases. Next, the functions of XBP1 in cancer as a regulator, not a downstream factor, became the basis for further screening literatures from the titles and the abstracts of articles.

Potentially, a set of XBP1 monoclonal antibodies (MAbs) has been developed for investigation on expression profile and functions of XBP-1 109 . XBP1 expression is related to poor clinical outcomes in patients with several cancers, including breast cancer 9 and pulmonary adenocarcinoma (pADC) 110 . The poor clinicopathological effects of XBP1 in breast cancer have been showed no correlation with the parameters, such as age, T stage, N stage, TNM stage, pathology grade, estrogen receptor, progesterone receptor, HER2 status, or luminal subtype 9 . In pADC, increased expression of XBP1s is associated with the poor prognosis of patients with ALK translocation. In contrast, increased expression of GRP78 is related to the poor prognosis of patients with EGFR -mutated pADC 110 . However, deactivation of IRE1-XBP1s signaling pathway could be involved in the strategy of different cancer management. Kaposi's sarcoma-associated herpesvirus (KSHV) undergoes latent and lytic phases in life. Viral IL-6 (vIL-6), an analog of human IL-6, stimulates the proliferation and differentiation of B cells and angiogenesis and is induced by replication and transcription activator (RTA) in the lytic phase. XBP-1s has been demonstrated to bind to the promoter of vIL-6 and activate its expression, increasing the survival of KSHV-MCD lymph node plasmablasts 111 . KSHV also potently infects cells of primary effusion lymphoma (PEL), which is co-infected with Epstein-Barr virus (EBV). Similar to transactivation of KSHV-RTA by XBP1, the EBV immediate-early promoter Zp is also up regulated and subsequently increases the generation of the lytic-cycle trans-activator BZLF1 by XBP1 112 , 113 . In contrast, Lai, et al (2011) found that XBP1s did not activate the expression of EBV BZLF1 and BRLF1 in PELs, suggesting that XBP1 specifically activates the KSHV lytic cycle in dually infected PELs 114 . Binding of XBP1 in the promoter of IL-6 is also identified in melanocytes and melanoma cells, promoting cells proliferation and progression. Furthermore, IL-6 subsequently activates the intracellular JAK-STAT3 signaling pathway in an autocrine/paracrine manner 61 . Activation of IRE1α-XBP1 signaling increases the proliferation of HCC cells through up regulation of IL-6 promoter activity and activation of STAT3 signaling 115 . Human pancreatic tumors are extremely hypoxic due to insufficient angiogenesis. Vascular endothelial growth factor (VEGF) is a key pro-angiogenic factor produced by HIF-1α under hypoxia. XBP1 and HIF-1α are essential for tumor angiogenesis. Unfortunately, VEGF does not involve in XBP1-mediated angiogenesis directly. Overexpression of a dominant-negative form of IRE1α decreases blood vessel formation significantly. However, these can be effectively reversed by overexpression of XBP1s 116 .

The progression of diseases is usually verified as a collective consequence involving various tissues and the governing signaling pathways. In the context of cancer cells, stressful elements, such as hypoxia, genetic mutations, and metabolic dysfunction, induce accumulation of faulty proteins in the ER. UPR ameliorates these hostile conditions and promotes cancer cells to adaptation and survival. However, hyper-activation of ER stress may leads to cell apoptosis. Although various investigations have documented many possible roles of ER stress and IRE1α-XBP1s signaling pathway in the progression of cancer, there still needs an accurate understanding of what the exact status of ER stress and the contribution of UPR in the pathogenesis are in cancer. The biological processes in various cancer cells and in tumor-infiltrating immune cells are regulated by the molecular signaling pathways, which ultimately induce the progression of cancers. Activation of IRE1α-XBP1s signaling has been implicated in cancer development and progression. Mechanistically, active XBP1 with transactivation domain functions as a transcription factor to regulate the downstream target genes expression by binding to specific sites, such as CRE or CRE-like sites. XBP1 forms a complex with HIF1α and signatures of hypoxia-driven gene expression. XBP1 is involved in tumor metastasis and drug resistance. In addition, XBP1 is necessary for terminal differentiation of B cells into immunoglobulin-secreting plasma cells and coordination of cellular structural and functional changes. Epigenetic studies indicate that IRE1-XBP1 signaling pathway is of physiological requirements. This suggests that dysfunctions of XBP1 may lead to imbalanced homeostasis and produce poor prognosis in clinic. The immunosuppressive activity of XBP1 also promotes cancer cells survival against T cell cytotoxic functions and immunotherapeutics. The potential XBP1 peptides-based vaccination and/or combination with immune-modulatory drug administration have been developed for effective management for several cancers. XBP1 acts as the biomarker of cancer development and progression and its inhibition becomes the strategy for clinical cancer management. However, there are some questions to be answered. Under physiological or pathological environment, what are the stimuli for activating IRE1α-XBP1s signaling? What are the different functions of XBP1s and XBP1u in the development and progression of cancers? Consequently, the molecular network regulated by XBP1 is still obscure.