() and () promoter methylation is associated with HPV infection and transcriptional repression in head and neck squamous cell carcinomas.

BACKGROUND

DNA methylation of the immune checkpoint gene has recently been shown to be associated with PD-L1 mRNA expression in various malignancies. This study aimed to investigate the association of and methylation with mRNA expression, immune cell infitration, protein expression and human papilloma virus (HPV) infection in head and neck squamous cell carcinoma (HNSCC) patients.

RESULTS

DNA methylation of and correlates inversely with mRNA expression (: ≤ 0.002; : ≤ 0.014). Methylation of specific CpG-sites of both and were further significantly associated with HPV infection in the TCGA cohort. Immune cell infiltrates correlated significantly with and methylation. In the validation cohort, PD-L1 protein expression was associated with hypomethylation ( = 0.012).

CONCLUSIONS

DNA methylation of and is associated with transcriptional silencing and HPV infection in HNSCCs. Additional studies are warranted to test PD-L1 and PD-L2 methylation as predictive biomarkers for response to immunotherapies (e.g. pembrolizumab and nivolumab) that target the PD-L1/PD-L2/PD-1 immune checkpoint axis.

MATERIALS AND METHODS

and promoter methylation and its mRNA expression were analyzed based on Infinium HumanMethylation450 BeadChip and RNA-Seq (both Illumina, Inc.) data in a representative HNSCC patient cohort ( = 528) enrolled by The Cancer Genome Atlas (TCGA) Research Network. A validation cohort consisting of 168 HNSCC patients treated at the University Hospital Bonn was analyzed regarding and promoter methylation by means of methylation-specific quantitative real-time PCR. PD-L1 protein expression in the validation cohort was quantified via immunohistochemistry (PD-L1 antibody clone 22C3, Dako/Agilent Technologies, Inc.).