Essential role of cooperative NF-κB and Stat3 recruitment to ICAM-1 intronic consensus elements in the regulation of radiation-induced invasion and migration in glioma

Abstract

Although radiotherapy improves survival in patients, GBMs tend to relapse with augmented tumor migration and invasion even after irradiation (IR). Aberrant NF-κB and Stat3 activation and interaction has been suggested in several human tumors. However, possible NF-κB/Stat3 interaction and the role of Stat3 in maintenance of NF-κB nuclear retention in glioblastoma (GBM) still remain unknown. Stat3 and NF-κB (p65) physically interact with one another in the nucleus in glioma tumors. Most importantly, GST pull-down assays identified that Stat3 binds to the p65 transactivation domain (TAD) and is present in the NF-κB DNA-binding complex. Irradiation significantly elevated nuclear phospho-p65/phospho-Stat3 interaction in correlation with increased ICAM-1 and sICAM-1 levels, migration and invasion in human glioma xenograft cell lines 4910 and 5310. ChIP and promoter luciferase activity assays confirmed the critical role of adjacent NF-κB (+399) and Stat3 (+479) binding motifs in the proximal intron-1 in elevating IR-induced ICAM-1 expression. Specific inhibition of Stat3 and NF-κB with Stat3.siRNA or JSH-23 severely inhibited IR-induced p65 recruitment onto ICAM-1 intron-1 and suppressed migratory properties in both cell lines. On the other hand, Stat3C- or IR-induced Stat3 promoter recruitment was significantly decreased in p65-knockdown cells, thereby suggesting the reciprocal regulation between p65 and Stat3. We also observed a significant increase in NF-κB enrichment on ICAM-1 intron-1 and ICAM-1 transactivation in Stat3C overexpressing cells. In in vivo orthotopic experiments, suppression of tumor growth in Stat3.si+IR-treated mice was associated with the inhibition of IR-induced p-p65/p-Stat3 nuclear-colocalization and ICAM-1 levels. To our knowledge, this is the first study showing the crucial role of NF-κB/Stat3 nuclear association in IR-induced ICAM-1 regulation and implies that targeting NF-κB/Stat3 interaction may have future therapeutic significance in glioma treatment.

STAT3 Serine 727 Phosphorylation: A Relevant Target to Radiosensitize Human Glioblastoma

“…Irradiation also induced a dose-dependent increase in pSTAT3-Y705 nuclear accumulation in glioma xenografts (29). The above published data suggest that STAT3 activation might participate in influencing glioma sensitivity to radiotherapy.…”

“…It also supports cell migration by augmenting MMP9 (51) and ICAM-1 (29) expression. It has been also shown that pSTAT3-Y705 activation in tumor cells participates in locally blocking immune cell maturation and activation by modifying tumor secretome (48).…”

“…The canonical Y705 activation can be induced by the binding of the Interleukin 6-family cytokine receptors to the gp130 receptor and the consecutive Janus kinase 2 (JAK2) activation (25). The pSTAT3-Y705 then form homodimers or heterodimers with partner proteins such as STAT1 and NF-κB (p65) (29). Those dimers translocate into the nucleus where they regulate expression of target-genes (5).…”

Role of interleukin-6 in cancer progression and therapeutic resistance

“…As a major effector molecule of NF-kB activation through the STAT-3 pathway, IL-6 appears to be an important component of the NF-kB/IL-6/STAT-3 cascade involved in tumorigenesis [75]. STAT-3 is required for the maintenance of NF-kB activation in tumours [121], whilst IL-6 promotes carcinogenesis through inflammation and cell proliferation [35,44,106]. Since inflammation enhances the growth and progression of gastrointestinal tumours via the activation of IL-6-mediated STAT-3 signalling, it appears that there is a strong link between IL-6, inflammation and tumour promotion [1,117,118].…”

Targeting the STAT3 signaling pathway in cancer: Role of synthetic and natural inhibitors

“…Intercellular adhesion molecule-1 (ICAM-1/CD54) is an inducible cell-surface glycoprotein that mediates adhesion-dependent cellular interactions, and its high expression correlates with increased tumor malignancy and invasiveness. It was observed that irradiation significantly elevated nuclear phospho-p65/phospho-STAT3 interaction in correlation with increased intercellular adhesion molecule-1 (ICAM-1) and soluble-ICAM-1 levels, migration and invasion in human glioma cell lines [169].…”