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Tumour MLH1 promoter region methylation testing is an effective prescreen for Lynch Syndrome (HNPCC)
  1. K Newton1,
  2. N M Jorgensen2,
  3. A J Wallace2,
  4. D D Buchanan3,4,5,
  5. F Lalloo6,
  6. R F T McMahon7,8,
  7. J Hill1,
  8. D G Evans6
  1. 1Department of General Surgery, Manchester Royal Infirmary, Central Manchester University Hospitals NHS Trust, Manchester, UK
  2. 2Genomic Diagnostics Laboratory, Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Saint Mary's Hospital, Manchester, UK
  3. 3Cancer and Population Studies Group, Queensland Institute of Medical Research, Herston, Queensland, Australia
  4. 4Oncogenomics Group, Genetic Epidemiology Laboratory, Department of Pathology, The University of Melbourne, Parkville, Victoria, Australia
  5. 5Centre for Epidemiology and Biostatistics, Melbourne School of Population and Global Health, The University of Melbourne, Parkville, Victoria, Australia
  6. 6Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Saint Mary's Hospital, Manchester, UK
  7. 7Department of Histopathology, Manchester Royal Infirmary, Central Manchester University Hospitals NHS Foundation Trust, Manchester, UK
  8. 8Manchester Medical School, University of Manchester, Manchester, UK
  1. Correspondence to K Newton, Genomic Diagnostics Laboratory, Manchester Centre for Genomic Medicine, Central Manchester University Hospitals NHS Foundation Trust, Saint Mary's Hospital, Oxford Road, Manchester M13 9WL UK; Katynewton2012{at}doctors.org.uk

Abstract

Background and aims Lynch syndrome (LS) patients have DNA mismatch repair deficiency and up to 80% lifetime risk of colorectal cancer (CRC). Screening of mutation carriers reduces CRC incidence and mortality. Selection for constitutional mutation testing relies on family history (Amsterdam and Bethesda Guidelines) and tumour-derived biomarkers. Initial biomarker analysis uses mismatch repair protein immunohistochemistry and microsatellite instability. Abnormalities in either identify mismatch repair deficiency but do not differentiate sporadic epigenetic defects, due to MLH1 promoter region methylation (13% of CRCs) from LS (4% of CRCs). A diagnostic biomarker capable of making this distinction would be valuable. This study compared two biomarkers in tumours with mismatch repair deficiency; quantification of methylation of the MLH1 promoter region using a novel assay and BRAF c.1799T>A, p.(Val600Glu) mutation status in the identification of constitutional mutations.

Methods Tumour DNA was extracted (formalin fixed, paraffin embedded, FFPE tissue) and pyrosequencing used to test for MLH1 promoter methylation and presence of the BRAF c.1799T>A, p.(Val600Glu) mutation 71 CRCs from individuals with pathogenic MLH1 mutations and 73 CRCs with sporadic MLH1 loss. Specificity and sensitivity was compared.

Findingss Unmethylated MLH1 promoter: sensitivity 94.4% (95% CI 86.2% to 98.4%), specificity 87.7% (95% CI 77.9% to 94.2%), Wild-type BRAF (codon 600): sensitivity 65.8% (95% CI 53.7% to 76.5%), specificity 98.6% (95% CI 92.4% to 100.0%) for the identification of those with pathogenic MLH1 mutations.

Conclusions Quantitative MLH1 promoter region methylation using pyrosequencing is superior to BRAF codon 600 mutation status in identifying constitutional mutations in mismatch repair deficient tumours.

  • Genetics
  • Cancer: colon

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