A novel method to capture methylated human DNA from stool: Implications for colorectal cancer screening

Hongzhi Zou, Jonathan Harrington, Rafaela L. Rego, David A. Ahlquist

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

Background: Assay of methylated DNA markers in stool is a promising approach for colorectal cancer (CRC) screening. A method to capture hypermethylated CpG islands from stool would enrich target analyte and allow optimal assay sensitivity. Methods: Methyl-binding domain (MBD) protein was produced using a pET6HMBD plasmid with MBD DNA sequence cloned from rat MeCP2 gene and bound to a column of nickel-agarose resin. We first established the feasibility of using the MBD column to extract methylated human DNA in a high background of fecal bacterial DNA. To explore the impact of MBD enrichment on detection sensitivity, the tumor-associated methylated vimentin gene was assayed with methylation-specific PCR from stools to which low amounts of cancer cell DNA (0-50 ng) were added and from stools from CRC patients and healthy individuals. Stools from cancer patients were selected with low amounts of human DNA (median 7 ng, range 0.5-832 ng). Results: With MBD enrichment, methylated vimentin was detected in stools enriched with ≥10 ng of cancer cell DNA and in CRC stool with a range of native human DNA amounts from 4 to 832 ng. Without MBD enrichment, methylated vimentin was not detected in the enriched stools and was detected in only 1 cancer stool with high human DNA (832 ng). In stools from healthy individuals methylated vimentin was not detected, with or without MBD enrichment. Conclusions: MBD capture increases assay sensitivity for detecting methylated DNA markers in stool. Applied clinical studies for stool cancer screening are indicated.

Original languageEnglish (US)
Pages (from-to)1646-1651
Number of pages6
JournalClinical Chemistry
Volume53
Issue number9
DOIs
StatePublished - Sep 2007

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Early Detection of Cancer
Colorectal Neoplasms
Screening
Vimentin
DNA
Neoplasms
Genetic Markers
Assays
Bacterial DNA
CpG Islands
Genes
Cells
Nickel
Population Groups
Sepharose
Methylation
DNA sequences
Carrier Proteins
Plasmids
Rats

ASJC Scopus subject areas

  • Clinical Biochemistry

Cite this

A novel method to capture methylated human DNA from stool : Implications for colorectal cancer screening. / Zou, Hongzhi; Harrington, Jonathan; Rego, Rafaela L.; Ahlquist, David A.

In: Clinical Chemistry, Vol. 53, No. 9, 09.2007, p. 1646-1651.

Research output: Contribution to journalArticle

Zou, Hongzhi ; Harrington, Jonathan ; Rego, Rafaela L. ; Ahlquist, David A. / A novel method to capture methylated human DNA from stool : Implications for colorectal cancer screening. In: Clinical Chemistry. 2007 ; Vol. 53, No. 9. pp. 1646-1651.
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