Non-invasive diagnosis of vulvar dysplasia using cervical methylation markers—a case control study

Abstract

Background

Diagnostic screenings for vulvar squamous intraepithelial lesions (VSIL) are limited and without information on disease trends. A panel of six methylation markers (ASTN1, DLX1, ITGA4, RXFP3, SOX17, ZNF671; GynTect® assay) has shown promise in diagnosing cervical intraepithelial neoplasia (CIN). Given the similarities between the carcinogenesis of cervix and vulva, this study aimed to investigate the suitability of these markers for diagnosing vulvar lesions.

Methods

One hundred twenty-one vulvar FFPE samples and 237 vulvar cell smears with different VSIL grades, HPV status, and with or without lichen sclerosus and planus were tested. Additionally, dysplasia-free vulvar cell smears from patients with cervical dysplasia were analyzed. The expression of DNA methyltransferases (DNMTs) in the FFPE samples was measured.

Results

The markers demonstrated high specificity in vulvar smears, with sole 5.45% of dysplasia-free smears testing positive. Yet, 75.00% of vulvar carcinoma smears appear positive in the methylation kit, similar to VHSIL (VIN III) smears with 77.78%. In FFPE samples, dysplasia-free samples from the tumor microenvironment of high-grade vulvar neoplasia showed 43.75% positivity. The positivity rates for VSIL and carcinoma samples were 76.92%, 64.71%, 64.71%, and 80.49%, respectively. DNMT3a expression was the highest in VLSIL (VIN I) samples, while DNMT1 was only expressed in VHSIL (VIN III) and carcinoma samples. Lichen sclerosis and planus showed a high false positive rate of 45.45% for dysplasia-free and 54.54% for smears with dVIN. Cervical HSIL was associated with a significantly higher number of positive results in the kit than in patients without cervical dysplasia.

Conclusions

The findings suggest that the methylation markers comprising GynTect® may be suitable for detecting vulvar neoplasia, as they exhibit high sensitivity. Nonetheless, adjustments are needed for comparable specificity. Lichen should be considered in result interpretation, and the kit should be used with caution for patients with lichen. Moreover, we observed methylation changes as an early event with the highest positivity of VLSIL. Surprisingly, changes in methylation pattern are not as local as presumed. Cervical SIL led to changed methylation in the vulva. Patients with positive kit results should be monitored regularly for all genital dysplasia. This sheds new light on the epigenetics in cancer.