Clinically relevant aberrant Filip1l DNA methylation detected in a murine model of cutaneous squamous cell carcinoma.

Kevin Roth,Sandra Steyaert,Maureen Higgins,Charlotte M Proby,Elena V Knatko,Tim De Meyer,Albena T Dinkova-Kostova,Louis Coussement

doi:10.1016/j.ebiom.2021.103383

Abstract

BackgroundCutaneous squamous cell carcinomas (cSCC) are among the most common and highly mutated human malignancies. Understanding the impact of DNA methylation in cSCC may provide avenues for new therapeutic strategies.MethodsWe used reduced-representation bisulfite sequencing for DNA methylation analysis of murine cSCC. Differential methylation was assessed at the CpG level using limma. Next, we compared with human cSCC Infinium HumanMethylation BeadArray data. Genes were considered to be of major relevance when they featured at least one significantly differentially methylated CpGs (RRBS) / probes (Infinium) with at least a 30% difference between tumour vs. control in both a murine gene and its human orthologue. The human EPIC Infinium data were used to distinguish two cSCC subtypes, stem-cell-like and keratinocyte-like tumours.FindingsWe found increased average methylation in mouse cSCC (by 12.8%, p = 0.0011) as well as in stem-cell like (by 3.1%, p=0.002), but not keratinocyte-like (0.2%, p = 0.98), human cSCC. Comparison of differentially methylated genes revealed striking similarities between human and mouse cSCC. Locus specific methylation changes in mouse cSCC often occurred in regions of potential regulatory function, including enhancers and promoters. A key differentially methylated region was located in a potential enhancer of the tumour suppressor gene Filip1l and its expression was reduced in mouse tumours. Moreover, the FILIP1L locus showed hypermethylation in human cSCC and lower expression in human cSCC cell lines.InterpretationDeregulation of DNA methylation is an important feature of murine and human cSCC that likely contributes to silencing of tumour suppressor genes, as shown for Filip1l.FundingBritish Skin Foundation, Cancer Research UK

Highlights

IntroductionDespite the beneficial effects of ultraviolet radiation (UVR) for e.g. vitamin D production and skin nitric oxide mediated blood pressureResearch in contextEvidence before this studyCutaneous squamous cell carcinoma (cSCC) is the most common human malignancy
Despite the beneficial effects of ultraviolet radiation (UVR) for e.g. vitamin D production and skin nitric oxide mediated blood pressureAbbreviations: UVR, Ultraviolet radiation; NMSC, non-melanoma skin cancer; Cutaneous squamous cell carcinomas (cSCC), cutaneous squamous cell carcinoma; ssUV mcSCC, solar simulated mouse cSCC; Tet, Ten-eleven-translocase;RRBS, reduced representation bisulfite sequencing; VS, ventral skin; DS, dorsal skin; iHS, independent healthy skin; CpG, 50Cytosinephosphate-Guanine-30; CGI, CpG island; differentially methylated regions (DMRs), differentially methylated region; Filip1l, Filament A interacting protein 1-like; iso, isoform; normal human keratinocytes (NHK), normal human keratinocyte; AK, actinic keratosis; 5mC, 5-methylcytosine; 5hmC, 5hydroxymethylcytosineResearch in contextEvidence before this studyCutaneous squamous cell carcinoma is the most common human malignancy
As DMRs cannot be readily compared between mouse and human, as well as between different platforms (RRBS versus Infinium array), we focused on genes containing differentially methylated CpGs in both species

Summary

Introduction

Despite the beneficial effects of ultraviolet radiation (UVR) for e.g. vitamin D production and skin nitric oxide mediated blood pressureResearch in contextEvidence before this studyCutaneous squamous cell carcinoma (cSCC) is the most common human malignancy. We have previously developed a preclinical mouse model that is histopathologically and genetically remarkably similar to human cSCC. This model presents a unique opportunity to study cSCC biology and, more importantly, test treatment and prevention strategies to improve cSCC management. Genes were considered to be of major relevance when they featured at least one significantly differentially methylated CpGs (RRBS) / probes (Infinium) with at least a 30% difference between tumour vs control in both a murine gene and its human orthologue. A key differentially methylated region was located in a potential enhancer of the tumour suppressor gene Filip1l and its expression was reduced in mouse tumours. Interpretation: Deregulation of DNA methylation is an important feature of murine and human cSCC that likely contributes to silencing of tumour suppressor genes, as shown for Filip1l.

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