Abstract:

Objective To discuss the relationship between Msh homeobox 1 （MSX1） gene methylation and the clinicopathological characteristics and prognosis of the patients with cervical cancer （CC）， and to analyze the application prospect of MSX1 gene methylation in the diagnosis and treatment of CC. Methods The clinical data， cancer tissues， and adjacent normal tissues were collected from 120 CC patients who underwent surgery between January 2019 and June 2020， and all CC patients were followed up for 3 years； methylation-specific PCR （MSP） was used to detect the methylation status of the MSX1 gene in cancer tissues of the CC patients， who were divided into MSX1 gene hypo-methylation group （n=50） and MSX1 gene hyper-methylation group （n=70）； MSP was used to detect the methylation level of the MSX1 gene in cancer tissues of the CC patients， and the relationship between MSX1 gene methylation status and clinicopathological characteristics and prognosis of the CC patients was analyzed； Western blotting method was used to detect the expression level of MSX1 protein in cancer tissues and adjacent normal tissues of the CC patients； small interfering RNA （siRNA） targeting MSX1 mRNA was designed； the CC cells were divided into control group （transfected with blank vector） and siMSX1 group （transfected with MSX1 siRNA）； real-time fluorescence quantitative PCR （RT-qPCR） method was used to verify the cell transfection efficiency； cell scratch assay was used to detect the migration activity of the cells in two groups； Logistic regression was used to analyze the influencing factors of prognosis in the CC patients. Results The methylation rate in cancer tissues of the CC patients was 58.33%， which was significantly higher than that in adjacent normal tissues （11.67%， χ2=42.725， P<0.01）； the Western blotting results showed that compared with adjacent normal tissue， the expression level of MSX1 protein in cancer tissue of the CC patients was significantly decreased （P<0.01）； high methylation status of the MSX1 gene in cancer tissue of the CC patients was associated with high-risk human papillomavirus （HR-HPV） DNA， lymph node metastasis， and TNM stage （P<0.05）， but not associated with patient age， pathological type， and tumor size （P>0.05）； the cell scratch assay results showed that compared with control group， the expression level of MSX1 mRNA in the cells in siMSX1 group was significantly decreased （P<0.01）， suggesting successful knockout of the MSX1 gene； the cell scratch assay results showed that compared with control group， the migration activity of the cells in siMSX1 group was significantly increased （P<0.01）； the 3-year cumulative survival rate of patients with MSX1 gene methylation was 54.29%， which was significantly lower than that of patients without MSX1 gene methylation （80.00%， χ2=9.717， P=0.002）； the Logistic regression results showed that positive HR-HPV DNA， lymph node metastasis， TNM stage Ⅲ-Ⅳ and MSX1 gene high methylation were independent risk factors for prognosis in CC patients （P<0.05）. Conclusion MSX1 gene methylation is associated with poorer clinicopathological characteristics and adverse prognosis in the CC patients， suggesting its potential as a biomarker for cervical cancer.

Key words: Cervical cancer, MSX1 gene, Methylation, Prognosis, Small interfering RNA