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Consistency of the S5 DNA methylation classifier in formalin-fixed biopsies versus corresponding exfoliated cells for the detection of pre-cancerous cervical lesions.

ABSTRACT: Methylation biomarkers are promising tools for diagnosis and disease prevention. The S5 classifier is aimed at the prevention of cervical cancer by the early detection of cervical intraepithelial neoplasia (CIN). S5 is based on pyrosequencing a promoter region of EPB41L3 and five late regions of HPV types 16, 18, 31, and 33 following bisulfite conversion of DNA. Good biomarkers should perform well in a variety of sample types such as exfoliated cells, fresh frozen or formalin-fixed paraffin-embedded (FFPE) materials. Here, we tested the performance of S5 on 315 FFPE biopsies with paired exfoliated cervical samples using four different conversion kits (Epitect Bisulfite, Epitect Fast Bisulfite, EZ DNA Methylation, and EZ DNA Methylation-Lightning). The S5 values from FFPE biopsies for all kits were significantly correlated with those obtained from their paired exfoliated cells. For the EZ DNA Methylation kit, we observed an average increased methylation of 4.4% in FFPE. This was due to incomplete conversion of DNA (73% for FFPE vs. 95% for cells). The other kits had a DNA conversion rate in FFPE similar to the cells (95%-97%). S5 performed well at discriminating

SUBMITTER: Reuter C

PROVIDER: S-EPMC8026949 | biostudies-literature | 2021 Apr

REPOSITORIES: biostudies-literature

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Consistency of the S5 DNA methylation classifier in formalin-fixed biopsies versus corresponding exfoliated cells for the detection of pre-cancerous cervical lesions.

Reuter Caroline C Preece Matthew M Banwait Rawinder R Boer Sabrina S Cuzick Jack J Lorincz Attila A Nedjai Belinda B

Cancer medicine 20210312 8

Methylation biomarkers are promising tools for diagnosis and disease prevention. The S5 classifier is aimed at the prevention of cervical cancer by the early detection of cervical intraepithelial neoplasia (CIN). S5 is based on pyrosequencing a promoter region of EPB41L3 and five late regions of HPV types 16, 18, 31, and 33 following bisulfite conversion of DNA. Good biomarkers should perform well in a variety of sample types such as exfoliated cells, fresh frozen or formalin-fixed paraffin-embe ...[more]

PMID: 33710792

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Project description:BackgroundClinical manifestation of prostate cancer (PCa) is highly variable. Aggressive tumors require radical treatment while clinically non-significant ones may be suitable for active surveillance. We previously developed the prognostic ProstaTrend RNA signature based on transcriptome-wide microarray and RNA-sequencing (RNA-Seq) analyses, primarily of prostatectomy specimens. An RNA-Seq study of formalin-fixed paraffin-embedded (FFPE) tumor biopsies has now allowed us to use this test as a basis for the development of a novel test that is applicable to FFPE biopsies as a tool for early routine PCa diagnostics.MethodsAll patients of the FFPE biopsy cohort were treated by radical prostatectomy and median follow-up for biochemical recurrence (BCR) was 9 years. Based on the transcriptome data of 176 FFPE biopsies, we filtered ProstaTrend for genes susceptible to FFPE-associated degradation via regression analysis. ProstaTrend was additionally restricted to genes with concordant prognostic effects in the RNA-Seq TCGA prostate adenocarcinoma (PRAD) cohort to ensure robust and broad applicability. The prognostic relevance of the refined Transcriptomic Risk Score (TRS) was analyzed by Kaplan-Meier curves and Cox-regression models in our FFPE-biopsy cohort and 9 other public datasets from PCa patients with BCR as primary endpoint. In addition, we developed a prostate single-cell atlas of 41 PCa patients from 5 publicly available studies to analyze gene expression of ProstaTrend genes in different cell compartments.ResultsValidation of the TRS using the original ProstaTrend signature in the cohort of FFPE biopsies revealed a relevant impact of FFPE-associated degradation on gene expression and consequently no significant association with prognosis (Cox-regression, p-value > 0.05) in FFPE tissue. However, the TRS based on the new version of the ProstaTrend-ffpe signature, which included 204 genes (of originally 1396 genes), was significantly associated with BCR in the FFPE biopsy cohort (Cox-regression p-value < 0.001) and retained prognostic relevance when adjusted for Gleason Grade Groups. We confirmed a significant association with BCR in 9 independent cohorts including 1109 patients. Comparison of the prognostic performance of the TRS with 17 other prognostically relevant PCa panels revealed that ProstaTrend-ffpe was among the best-ranked panels. We generated a PCa cell atlas to associate ProstaTrend genes with cell lineages or cell types. Tumor-specific luminal cells have a significantly higher TRS than normal luminal cells in all analyzed datasets. In addition, TRS of epithelial and luminal cells was correlated with increased Gleason score in 3 studies.ConclusionsWe developed a prognostic gene-expression signature for PCa that can be applied to FFPE biopsies and may be suitable to support clinical decision-making.

Dataset Information

Consistency of the S5 DNA methylation classifier in formalin-fixed biopsies versus corresponding exfoliated cells for the detection of pre-cancerous cervical lesions.

Publications

Consistency of the S5 DNA methylation classifier in formalin-fixed biopsies versus corresponding exfoliated cells for the detection of pre-cancerous cervical lesions.

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