〔Abstract〕 Objective To investigate the regulatory role of ABCG5 in head and neck squamous cell carcinoma.Methods Immunohistochemical staining was used to detect the expression of ABCG5 in 105 patients with hypopharyngeal carcinoma,and the relationship between the expression and the long-term survival time of the patients was analyzed.Immunowestern blotting and flow cytometry were used to detect the expression of ABCG5 in head and neck squamous cell carcinoma cell lines.Flow sorting was employed to investigate the differences in proliferation and metastasis,spheroidization ability,and expression of stem cell molecules between ABCG5 positive and negative cells.si RNA interference was used to further demonstrate whether ABCG5 affects the malignant phenotype.Results ABCG5 was associated with poor prognosis in patients with hypopharyngeal carcinoma.ABCG5 was enriched in stem cells of head and neck squamous cell carcinoma.ABCG5-positive cells had stronger proliferation(P<0.001),metastasis(P<0.01),and spheroid formation abilities(P<0.001) than negative cells.The expression levels of some stem cell molecules(SOX2、NANOG、SOX9、OCT4、CD44) in ABCG5-positive cells were higher than those in ABCG5-negative cells.The expression of epithelial cell protein E-cadherin was lower in ABCG5-positive cells than that in ABCG5-negative cells,and the expression of interstitial cell proteins N-cadherin,Slug,Snail 1,and Vimentin were higher(P<0.01).Moreover,interfering with ABCG5 expression significantly inhibited tumor cell spheroid formation(P<0.001),as well as the expression of proteins related to cancer stem cells and epithelial-mesenchymal transition.Conclusion ABCG5 has a potential biological role in maintaining the function of head and neck squamous cell carcinoma stem cells.