Human Papillomavirus 16 DNA Methylation Patterns and Investigation of Integration Status in Head and Neck Cancer Cases

Abstract

Persistent high-risk human papillomavirus (HPV) infection is a pivotal factor in the progression of cervical cancer. In recent years, an increasing interest has emerged in comprehending the influence of HPV on head and neck squamous cell carcinoma (HNSCC). Notably, it is well established that HPV-associated HNSCC show cases with distinct molecular and clinical attributes compared to HPV-negative cases. The present study delves into the epigenetic landscape of HPV16, specifically its L1 gene and untranslated region (UTR), through pyrosequencing, while the HPV16 DNA physical status was evaluated using E2/E6 ratio analysis in HPV16-positive HNSCC FFPE biopsies. Our findings reveal substantial methylation across six sites within the HPV16 L1 gene and seven sites in the UTR. Specifically, methylation percentages of two L1 CpG sites (7136, 7145) exhibit significant associations with tumor histological grade (p < 0.01), while proving concurrent methylation across multiple sites. The HPV16 DNA physical status was not correlated with the methylation of viral genome or tumor characteristics. This is the first study that examines epigenetic modifications and the HPV16 DNA physical status in Greek HNSCC patients. Our findings suggest an orchestrated epigenetic modulation among specific sites, impacting viral gene expression and intricate virus-host interactions.

Keywords: HNSCC; HPV16; L1; UTR methylation; viral integration.

Figure 1

Box and whisker plot for L1 7136 and L1 7145 expression in relation to tumor grade. Box limits show Q1 and Q3 values. The lines within the boxes correspond to median values, the large circles indicate the median values and smaller circles correspond to measurements. Outliers are the circles outside the whisker limits, (left): L1 7136, (right): L1 7145.

Figure 2

(Left): distribution of different anatomical sites where tumor derived; (Right): histology grading of studied samples.