Interleukin-6 Production Mediated By The IRE1-XBP1 Pathway Confers Radioresistance In Human Papillomavirus-negative Oropharyngeal Carcinoma

Objective: Endoplasmic reticulum stress(ERS)plays a key role in the pathogenesis and development of tumours and protects tumour cells from radiation damage and drug‐induced stress.We previously demonstrated that EGFR confers radioresistance in human papillomavirus(HPV)-negative human oropharyngeal carcinoma by activating ERS signalling through PERK and IRE1α.In addition,PERK confers radioresistance by activating the inflammatory cytokine NF-κB.However,the effect of IRE1 on radiosensitivity has not yet been fully elucidated.The purpose of this study was to explore the mechanism of radiation resistance induced by IRE1 and to provide a new target for the treatment of radiation resistance patients with HPV(-)oropharyngeal carcinoma.Methods: The expression profiles of IL-6,ERS signalling as well as the clinical information of head and neck cancer samples were obtained from the TCGA database.Pathological tumor sections were obtained from 80 patients with HPV‐negative oropharyngeal squamous cell carcinoma(OSCC)who accepted radical radiotherapy with or without concurrent chemotherapy at the First Hospital of China Medical University between 2005 and 2011.In addition,36 patients with HPV‐negative oropharyngeal squamous cell carcinoma who received radical radiotherapy between 2014 and 2016 were selected.The correlation between IL-6 and the clinical outcomes of the patients was analysed using the Kaplan-Meier method.The expression of IL-6 and ERS signalling activation was analysed by western blotting and RT-PCR.The effects of IL-6 and ERS signalling on cell proliferation,apoptosis,DNA double-strand break and colony formation abilities were assessed by CCK-8 assay,flow cytometry,immunofluorescence and colony formation assay.Results: The present study clarified that IRE1 overexpression was associated with poor outcome in HPV-negative patients treated with radiotherapy(P = 0.0001).Additionally,a significantly higher percentage of radioresistant HPV-negative patients than radiosensitive HPV-negative patients exhibited high IRE expression(66.7% vs 27.8%,respectively;P = 0.001).DNA double strand breakage(DSB)is one of the main ways of radiation therapy to kill tumor cells.Silencing IRE1 and XBP1 increased DNA double-strand break(DSB)and radiation-induced apoptosis,thereby increasing the radiosensitivity of HPV-negative oropharyngeal carcinoma cells.IRE1-XBP1 silencing also inhibited radiation-induced IL-6 expression at both the RNA and protein levels.The regulatory effect of IRE1-XBP1 silencing on DNA DSB-and radiation-induced apoptosis was inhibited by pretreatment with IL-6.Conclusion: These data indicate that IRE1 regulates radioresistance in HPV-negative oropharyngeal carcinoma through IL-6 activation,enhancing X-ray-induced DNA DSBs and cell apoptosis.