Non-invasive detection of endometrial cancer through exfoliated cervical cell gene methylation signatures: Evaluation of CELF4/GALR1/ZNF486 methylation biomarkers.

Abstract

Objective: The current non-invasive screening methods for endometrial cancer (EC), such as transvaginal ultrasound (TVUS), lack sufficient specificity, leading to unnecessary invasive diagnostic procedures. The detection of cancer-specific deoxyribonucleic acid (DNA) methylation alterations in cervical cytology samples presents a promising, minimally invasive alternative. This study aimed to evaluate the diagnostic potential of novel DNA methylation biomarkers (CUGBP Elav-like family member 4 methylation [CELF4^m ], Galanin receptor 1 methylation [GALR1^m ], Zinc finger protein 486 methylation [ZNF486^m ]) in cervical exfoliated cells for EC screening.

Material and methods: This case-control study enrolled patients scheduled for diagnostic or surgical curettage at the hospital from September 2024 to June 2025. A total of 104 women underwent endometrial evaluation (32 type I EC, 3 type II EC, 61 benign lesions [BL], 8 atypical hyperplasia). Quantitative methylation-specific polymerase chain reaction assessed biomarker performance against histopathological diagnosis.

Results: All three markers (CELF4^m, GALR1^m, ZNF486^m ) showed significant differential ΔCp values between BL and EC groups (all P < 0.01), with moderate diagnostic accuracy (area under the ROC curve [AUCs]: 0.654-0.704). The single-marker GALR1^m outperformed endometrial thickness (ET) measured by TVUS in specificity (60.9% vs. 27.5%, P < 0.001) while maintaining comparable sensitivity (80.0% vs. 74.3%, P = 0.527). A two-marker panel (GALR1^m /ZNF486^m , Model 1) achieved sensitivity (97.1% [95% confidence interval [CI]: 91.6-100%]) with modest specificity (49.3% [95% CI: 37.5-61.1%]). Among three-gene models, Model 3 demonstrated the highest overall performance (AUC: 0.733 [95% CI: 0.657-0.808]; sensitivity: 91.4% [95% CI: 82.1-100%]), whereas Model 4 balanced sensitivity (80.0% [95% CI: 66.7-93.3%]) and improved specificity (63.8% [95% CI: 52.4-75.1%]).

Conclusion: DNA methylation signatures in cervical cells show superior diagnostic accuracy to conventional ET measured by TVUS, particularly through multi-marker panels. This non-invasive approach represents a promising EC screening strategy.