Study On The Role And Mechanism Of SQLE In Oral Squamous Cell Carcinoma

Objective : Head and neck squamous cell carcinoma(HNSCC)is one of the most common malignant tumors in the world.More than half of the cases occur in the oral cavity.Oral squamous cell carcinoma(OSCC)is the most common pathological type of oral cancer,accounting for about 90% of the new cases annually.OSCC can occur in any part of the oral cavity,including tongue,gingiva,floor of mouth,palate and buccal mucosa.OSCC often leads to chewing,swallowing and speech dysfunction,which seriously affects the quality of life of patients.Advanced OSCC shows high recurrence rate and metastasis rate,the 5-year survival rate is always low.The molecular mechanism of OSCC is complex.It is the result of a variety of interdependent molecular mechanisms,involving not only the expression changes of specific genes and proteins,but also the changes of metabolic process.Squalene epoxidase(SQLE)is a monooxygenase that catalyzes the conversion of squalene to 2,3(S)-oxidosqualene.SQLE is the second rate-limiting enzyme in the cholesterol synthesis pathway,and its inhibition effectively reduces cholesterol synthesis.For a long time,it has been known that SQLE is highly amplified in a variety of cancers and drives abnormal metabolism;thus,it is considered a metabolic oncogene.The overexpression of SQLE has been shown to promote the proliferation and metastasis of lung squamous cell carcinoma,hepatocellular carcinoma,and esophageal squamous cell carcinoma and is associated with poor prognosis of prostate cancer and breast cancer.Another study identified a subset of neuroendocrine tumors,especially small cell lung cancers,showing significant sensitivity to the inhibition of SQLE.At present,the catalytic domain of human SQLE has been identified,which provides the possibility of the rational development of new targeted drugs.However,the biological mechanism of SQLE in OSCC has not been systematically elucidated.Therefore,this study aimed to evaluate the role of SQLE in the occurrence and development of OSCC through bioinformatic analysis,and validation experiments.Methods:1.In this study,transcriptome data and clinical data from TCGA and GEO databases were used to compare SQLE m RNA expression in HNSCC tumor tissues and normal tissues.Univariate Cox regression analysis and Kaplan-Meier survival curve were used to evaluate the prognostic significance of SQLE in HNSCC patients.Genomic data of TCGA patients were used to evaluate the mutation,copy number variation and methylation level of SQLE gene in HNSCC.Hu RI and Linked Omics were used to identify the SQLE related protein-protein interaction network and coexpressed genes,and these genes were enriched by Metascape.TIMER 2.0 was used to explore the correlation between SQLE and immune infiltration of HNSCC.2.Through immunohistochemical experiments,we selected 40 cases of OSCC tumor samples and 15 cases of adjacent normal samples to further verify SQLE protein expression difference in tumor tissues and normal tissues,and to explore its correlation with the clinicopathological characteristics of OSCC patients.3.We constructed the si RNA of SQLE and successfully transfected it into oral squamous cell carcinoma cell line CAL-27.The knockdown efficiency of SQLE was verified by western blot.Next,CCK-8 assay,Ed U assay and wound healing tests were used to evaluate the effect of SQLE on the proliferation and migration of CAL-27 cells.Results: 1.SQLE m RNA expression in HNSCC was significantly higher than that in normal tissues(P <0.01).With the increase of copy number and demethylation of SQLE gene,the m RNA expression level of HNSCC patients increased(P < 0.01).Univariate Cox regression analysis and Kaplan-Meier survival analysis showed that HNSCC patients with high expression of SQLE had worse overall survival(OS)and progression-free survival(PFS)(P<0.01).The results of immune infiltration analysis showed that the copy number and m RNA expression of SQLE were negatively correlated with the infiltration levels of CD8 + T cells,regulatory T cells,follicular helper T cells and activated mast cells,and positively correlated with the infiltration levels of M0 macrophages and resting mast cells in HNSCC(P<0.01).2.SQLE protein expression in OSCC was significantly higher than that in normal tissues(P<0.05),and SQLE protein expression in patients with T3 / T4 OSCC was significantly higher than that in T1 / T2 OSCC(P<0.05).3.Western blot analysis showed that the expression of SQLE protein in CAL-27 cells transfected with SQLE si RNA was significantly lower than that in the control groups(P <0.05).The results of CCK-8 and Ed U assays showed that compared with the control groups,the proliferation ability of CAL-27 cells was significantly decreased after SQLE knockdown(P<0.05).The results of wound healing tests showed that the migration ability of CAL-27 cells after SQLE knockdown was significantly lower than that of the control groups(P<0.05).Conclusion: 1.The m RNA expression level of SQLE in HNSCC is higher than that in normal tissues.High expression of SQLE is associated with poor prognosis of HNSCC.Copy number amplification and DNA demethylation promote the overexpression of SQLE in HNSCC,which may be related to the negative immune regulation of HNSCC.2.The protein expression level of SQLE in OSCC was higher than that in normal tissues,and correlated with T stage of OSCC patients.3.Knockdown of SQLE inhibited the proliferation and migration of OSCC cells.