title: Methylation analysis and HPV genotyping of self-collected cervical samples from women not responding to screening invitation and review of the literature.
abstract: OBJECTIVE
  To assess the feasibility of partial HPV genotyping and methylation analysis of CADM1, MAL, and miR124-2 genes as triage tests in assaying self-collected cervical samples positive for high-risk HPV on primary screening, and to review the literature regarding host cellular gene methylation analysis of self-collected cervical samples.
  
  
  METHODS
  Women residing in North-East Italy who had failed to respond to the invitation to participate in an organized population-based program were invited to provide a self-sample. Their stored baseline (self-collected) and follow-up (clinician-collected) cervical samples were included in the study. DNA was extracted from HPV-positive (Qiagen's Hybrid Capture 2, HC2) samples. Partial genotyping with separate detection of HPV types 16 and 18 was performed with a hybrid capture-based method and a quantitative PCR assay. Methylation was assayed with a quantitative methylation-specific PCR.
  
  
  RESULTS
  High-risk HPV infection was detected in 48% of baseline and 71% of follow-up HC2-positive samples. Methylation was demonstrated respectively in 15% and 23.5% of baseline and follow-up samples and chiefly involved a single gene (miR124-2). Invalid quantitative PCR results were recorded in 5% of self-collected samples. The specificity of miR124-1, MAL, and CADM1 methylation was 84%, 94%, and 98%, respectively, and the specificity of the three markers combined was 84%. Sensitivity was not estimated due to the lack of CIN2+ samples. The systematic review showed that different methylation assays yield different accuracy values.
  
  
  CONCLUSIONS
  Self-collected samples are suitable for methylation assays included in reflex triage testing. The reproducibility and accuracy of the methylation tests described in the literature should be improved.
authors.affiliation: Immunology and Diagnostic Molecular Oncology Unit, Veneto Institute of Oncology IOV IRCCS, Padua, Italy.
authors.forename: Annarosa
authors.initials: A
authors.lastname: Del Mistro
authors.affiliation: Immunology and Diagnostic Molecular Oncology Unit, Veneto Institute of Oncology IOV IRCCS, Padua, Italy.
authors.forename: Helena
authors.initials: H
authors.lastname: Frayle
authors.affiliation: Immunology and Diagnostic Molecular Oncology Unit, Veneto Institute of Oncology IOV IRCCS, Padua, Italy.
authors.forename: Martina
authors.initials: M
authors.lastname: Rizzi
authors.affiliation: Maternal and Child Department, Local Health Unit 7, Pieve di Soligo-Conegliano, Conegliano, Treviso, Italy.
authors.forename: Gianpiero
authors.initials: G
authors.lastname: Fantin
authors.affiliation: Prevention Department, Local Health Unit 17, Este-Monselice, Este, Padua, Italy.
authors.forename: Antonio
authors.initials: A
authors.lastname: Ferro
authors.affiliation: Department of Life, Health and Environmental Sciences, University of L'Aquila, L'Aquila, Italy.
authors.forename: Paolo Matteo
authors.initials: PM
authors.lastname: Angeletti
authors.affiliation: Interinstitutional Epidemiology Unit, Local Health Authority, Reggio Emilia, Italy; Arcispedale S. Maria Nuova Research Hospital, IRCCS, Reggio Emilia, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Giorgi Rossi
authors.affiliation: Department of Life, Health and Environmental Sciences, University of L'Aquila, L'Aquila, Italy; Epidemiology and Biostatistics Unit, AUSL Teramo, University of L'Aquila, L'Aquila, Italy.
authors.forename: Emma
authors.initials: E
authors.lastname: Altobelli
mesh_terms: D000328:Adult; D002584:Cervix Uteri / Q000821:virology*; D019175:DNA Methylation; D005260:Female; D005838:Genotype; D006801:Humans; D008403:Mass Screening; D008875:Middle Aged; D027383:Papillomaviridae / Q000145:classification / Q000235:genetics*; D030361:Papillomavirus Infections / Q000453:epidemiology / Q000235:genetics* / Q000821:virology*; D002583:Uterine Cervical Neoplasms / Q000453:epidemiology / Q000209:etiology