Human Papillomavirus Genotypes and Methylation of CADM1, PAX1, MAL and ADCYAP1 Genes in Epithelial Ovarian Cancer Patients

Abstract

Background: High-risk types of human papillomavirus (HR-HPV) may play a role in the development of epithelial ovarian cancer (EOC). The aim of this study was to determine any HPV genotypes and correlations to CADM1, PAX1, MAL and ADCYAP1 gene methylation in Egyptian EOC patients. Materials and methods: The prevalence of HR-HPV in 100 formalin fixed paraffin embedded EOC tissues was determined using nested polymerase chain reaction (PCR) with MY09/MY11 and GP5+/GP6 + primers to amplify a broad spectrum of HPV genotypes in a single reaction. DNA sequencing was applied to identify HPV genotypes for the positive samples. All samples negative for HPV were re-analyzed for HR-HPV and low-risk HPV subtypes using type specific primers. Results: The prevalence of HPV was 10% in our EOC cases. HPV-16 and HPV-18 were the predominant genotypes followed by HPV−33, all being associated with advanced stages. Other HR-HPV and low risk HPV genotypes were not found. CADM1 was hypermethylated in 100% of patients infected with HPV-16 and HPV-33 and in 75% of patients infected with HPV-18. Hypermethylation of PAX1 was evident in 80% and in 75% of patients infected with HPV-16 and HPV-18 while MAL was hypermethylated in 100% and ADCYAP1 was hypermethylated in 60% and in 75%, respectively. Conclusion: The presence of high risk HPV genotypes among epithelial ovarian carcinoma may reflect an importance of infection in the pathogenesis of EOC. In HR-HPV infected cancers, DNA methylation may be one of the mechanisms triggering the alteration in CADM1, PAX1, MAL and ADCYAP1 gene expression levels.

Keywords: Human papilloma virus genotypes; epithelial ovarian cancer; nested polymerase chain reaction.

Figure 1

PCR Products Images on a 2% Agarose Gel Visualized by UV-Trans-Illumination. Lane M is 100 PCR Marker (Promega), Lanes 1, 2, 3 and 4 were Positive Samples with MY09/MY11 Followed by GP5+/GP6+

Figure 2

Sequencing Data of HPV Genotypes (A) Sequence Alignment of HPV Type 18 Using Basic Local Alignment Search (BLAST). (B) Sequence Alignment of HPV Type 16 Using Basic Local Alignment Search (BLAST). (C) Sequence Alignment of HPV Type 33 Using Basic Local Alignment Search (BLAST)

Figure 3

Methylation Specific PCR for the Studied Genes in EOC Some Samples Infected with HPV. M and UM Indicate the Presence of Methylated and Un-Methylated Target Genes Respectively