Analysis Of PAX 1 Promoter CpG Site Methylation In Cervical Screening And Its Correlation With HPV Infection

[Background and Objective] Cervical cancer is the most common malignant tumor in the female reproductive system.Currently,cervical cancer screening using HPV testing and cervical cytology is the most effective means of prevention,but these methods have limitations.While previous studies have shown that PAX1 methylation has high specificity and sensitivity in cervical cancer screening,few have used a quantitative analysis method to examine the extent of PAX1 methylation and its role in cervical lesion screening.In this study,we aim to explore the diagnostic value of PAX1 methylation testing in cervical intraepithelial neoplasia,identify the optimal methylation site,and develop a prediction model to provide novel insights for cervical cancer screening.Additionally,we will investigate the methylation of PAX1 in cervical exfoliated cells after different types of high-risk HPV infection and analyze their potential correlation.[Methods] This study considered patients who had abnormal results from cervical cancer screening and underwent cervical biopsy during colposcopy at the Department of Gynecology in the Shenzhen Second People’s Hospital between October 2019 and April 2021.A total of 188 subjects were included in the study,and data was collected on their basic information,genotype of the HPV infection,and colpophoscent biopsy results.Quantitative analysis of PAX 1 methylation was performed on 88 sites using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF MS).Based on the pathological results,the patients were divided into five groups: chronic cervicitis(non-malignant,108 cases),CIN I(29 cases),CIN II(21 cases),CIN III(28 cases),and cervical cancer(2 cases).The diagnostic efficacy of different PAX 1 methylation sites for detecting cervical intraepithelial neoplasia(CIN)II or more severe lesions was analyzed and compared with current cervical cancer screening methods.A predictive model was then constructed using logistic regression.Additionally,non-parametric tests were performed to further analyze the correlation between hr-HPV(high-risk human papillomavirus)status and PAX 1 methylation in different patients who were hr-HPV-negative or hr-HPV-positive.[Results] The top five sites with the largest area under the receiver operating characteristic(ROC)curve values were identified as GRCh38/hg38 chr20:X21705687,X21705946,X21706427,X21706285,and X21706637.The diagnostic efficacy of PAX 1 methylation was found to be higher than that of high-risk human papillomavirus(hr-HPV).A predictive model was constructed by combining hr-HPV and PAX 1 methylation sites,with the following formula: prediction index = –5.993 +15.211 × X21705687 + 7.890 × X21706427 + 1.846 × HPV16/18(1,0)+ 1.821 ×other hr-HPV(1,0).The area under the curve(AUC)was 0.868,and the sensitivity and specificity were 0.863 and 0.756,respectively.The Spearman’s correlation coefficients between hr-HPV infection and five different sites(X21705687,X21705946,X21706427,X21706285,and X21706637)were 0.1926,0.20,0.1274,0.1870,and 0.1996,respectively.However,the absolute values of these coefficients were all less than 0.3,indicating weak or no correlation between hr-HPV infection and the respective sites.[Conclusions] The diagnostic efficacy of PAX 1 methylation test applied in cervical cancer screening is higher than that of HPV test.The predictive model constructed by combining hr-HPV testing and PAX 1 methylation can further improve the diagnostic efficacy of high-grade cervical lesions and cervical cancer.PAX 1methylation was significantly associated with cervical lesion grading,but hr-HPV infection of different types did not correlate with PAX 1 methylation in cervical exfoliated cells.