EPHB4 Regulates the Proliferation and Metastasis of Oral Squamous Cell Carcinoma through the HMGB1/NF-κB Signalling Pathway

Abstract

Background: Malignant proliferation and cervical lymphatic metastasis restrict the prognosis of oral squamous cell carcinoma (OSCC). Erythropoietin-producing human hepatocellular B4 (EPHB4) regulates a series of tumour functions involving tumourigenesis, cancer cell attachment and metastasis. However, the mechanism of EphB4 regulating the malignant progression of OSCC has not been fully elucidated. Methods: EPHB4 expression was analysed in 65 OSCC samples and adjacent noncancerous tissues through immunohistochemistry (IHC). siRNA and overexpression plasmids were transfected into OSCC cells to modify EPHB4 expression, and then, regulatory functions were explored in vitro and in vivo. Co-immunoprecipitation (Co-IP) and mass spectrometry were applied to detect proteins interacting with EPHB4. Subsequently, protein stability assays and NF-κB pathway inhibition assays were used to verify the regulation of EPHB4, high-mobility group box 1 (HMGB1) and nuclear factor-κB (NF-κB) activation. Results: EPHB4 was found to be highly expressed in OSCC tissues, which was related to tumour stage and lymphatic metastasis and resulted in a poor prognosis. Cellular experiments and mouse tongue xenograft models further confirmed that high EPHB4 expression promoted the proliferation and metastasis of OSCC tumours. Mechanistically, co-IP and mass spectrometry studies indicated that EPHB4 could bind to HMGB1 and maintain HMGB1 stability. Downregulation of HMGB1 inhibited the proliferation and metastasis of OSCC cells and inhibited NF-κB phosphorylation activation but did not affect EPHB4 expression. Conclusion:This study revealed the mechanism by which EPHB4 promotes the proliferation and metastasis of OSCC by activating the HMGB1-mediated NF-κB signalling pathway, which can be exploited as a novel marker or therapeutic target to control metastasis and improve the survival rate of OSCC.

A narrative review of the role of Eph receptors in head and neck squamous cell carcinoma

“…This study suggests that EphB4 can act as a tumor suppressor, which is unusual for a tyrosine kinase receptor. This is in discordance with Yi et al's study (Yi et al, 2021) which has demonstrated that EphB4 possesses an oncogenic function. The Bhatia study actually provides clues for explaining this ambiguity, since it was found that EphB4 could be functioning in both ways.…”

“…The expression of EphB4 in 65 samples of oral squamous cell carcinoma (OSCC) was compared to the adjacent noncancerous tissues. A significant association was found between high expression and tumor stage and cervical lymph node metastasis, supporting that expression of EphB4 is a poor predictive marker in OSCC (Yi et al, 2021).…”

“…The expression of EphB4 in 65 samples of oral squamous cell carcinoma (OSCC) was compared to the adjacent noncancerous tissues. A significant association was found between high expression and tumor stage and cervical lymph node metastasis, supporting that expression of EphB4 is a poor predictive marker in OSCC (Yi et al, 2021). On the other hand, Sasabe et al found that ephrin‐B2 immunoreactivity was significantly correlated with lymph node metastasis and clinical stage (Sasabe et al, 2017).…”

“…Yi et al verified that expression of EphB4 in OSCC promoted cancerous cell metastasis in vitro and in vivo. The study revealed the direct effect of EphB4 on metastasis might be related to activating the nuclear factor-κB (NF-κB) signaling pathway via interaction with high-mobility group box 1 (HMGB1) (Yi et al, 2021).This is in line with a recent study by Wu and co-workers who reported suppression of proliferation and metastasis of tumors in lung cancer by targeting an HMGB1-mediated NF-κB signaling pathway by miR-449a (Wu et al, 2019). Together, this indicates the complexity of this group of tyrosine kinase receptors, and further research is needed to elucidate the exact role of each receptor.…”

Contributing Role of High Mobility Group Box 1 Signaling in Oral Cancer Development and Therapy

“…Additionally, targeted knockdown of HMGB1 expression induced a reduction in p-p65, while EPHB4 levels remained unchanged. These data imply that NF-κB pathway functions as a downstream effector pathway of HMGB1 protein in OSCC cells and as a potential regulator of cancer cell proliferation and migration [24].…”