ColoSeq provides comprehensive lynch and polyposis syndrome mutational analysis using massively parallel sequencing

Colin C. Pritchard, Christina Smith, Stephen J. Salipante, Ming K. Lee, Anne M. Thornton, Alexander Nord, Cassandra Gulden, Sonia S. Kupfer, Elizabeth M. Swisher, Robin L. Bennett, Akiva P. Novetsky, Gail P. Jarvik, Olufunmilayo I. Olopade, Paul J. Goodfellow, Mary Claire King, Jonathan F. Tait, Tom Walsh

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141 Scopus citations

Abstract

Lynch syndrome (hereditary nonpolyposis colon cancer) and adenomatous polyposis syndromes frequently have overlapping clinical features. Current approaches for molecular genetic testing are often stepwise, taking a best-candidate gene approach with testing of additional genes if initial results are negative. We report a comprehensive assay called ColoSeq that detects all classes of mutations in Lynch and polyposis syndrome genes using targeted capture and massively parallel next-generation sequencing on the Illumina HiSeq2000 instrument. In blinded specimens and colon cancer cell lines with defined mutations, ColoSeq correctly identified 28/28 (100%) pathogenic mutations in MLH1, MSH2, MSH6, PMS2, EPCAM, APC, and MUTYH, including single nucleotide variants (SNVs), small insertions and deletions, and large copy number variants. There was 100% reproducibility of detection mutation between independent runs. The assay correctly identified 222 of 224 heterozygous SNVs (99.4%) in HapMap samples, demonstrating high sensitivity of calling all variants across each captured gene. Average coverage was greater than 320 reads per base pair when the maximum of 96 index samples with barcodes were pooled. In a specificity study of 19 control patients without cancer from different ethnic backgrounds, we did not find any pathogenic mutations but detected two variants of uncertain significance. ColoSeq offers a powerful, cost-effective means of genetic testing for Lynch and polyposis syndromes that eliminates the need for stepwise testing and multiple follow-up clinical visits.

Original languageEnglish (US)
Pages (from-to)357-366
Number of pages10
JournalJournal of Molecular Diagnostics
Volume14
Issue number4
DOIs
StatePublished - Jul 1 2012
Externally publishedYes

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ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Molecular Medicine

Cite this

Pritchard, C. C., Smith, C., Salipante, S. J., Lee, M. K., Thornton, A. M., Nord, A., Gulden, C., Kupfer, S. S., Swisher, E. M., Bennett, R. L., Novetsky, A. P., Jarvik, G. P., Olopade, O. I., Goodfellow, P. J., King, M. C., Tait, J. F., & Walsh, T. (2012). ColoSeq provides comprehensive lynch and polyposis syndrome mutational analysis using massively parallel sequencing. Journal of Molecular Diagnostics, 14(4), 357-366. https://doi.org/10.1016/j.jmoldx.2012.03.002