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      Highly sensitive faecal DNA testing of TWIST1 methylation in combination with faecal immunochemical test for haemoglobin is a promising marker for detection of colorectal neoplasia.

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          Abstract

          Background As TWIST1 methylation is specific to colorectal neoplasia, detection of TWIST1 methylation from faeces samples might be useful for colorectal neoplasia screening. However, because the content of human DNA in faeces is very small, it is very difficult to detect TWIST1 methylation by conventional bisulphite-based methylation assays. Therefore, we developed a new methylation assay without bisulphite treatment, the combined restriction digital PCR assay, and evaluated its sensitivity and specificity in combination with and without the faecal immunochemical test for haemoglobin for colorectal neoplasia detection from faeces samples. Methods For the combined restriction digital PCR assay, DNA was treated with three methylation-sensitive restriction enzymes and an exonuclease, followed by measurement of TWIST1 methylation level by droplet digital PCR. Faecal DNA testing and faecal immunochemical test for haemoglobin were performed on 109 patients with colorectal neoplasia and 10 control individuals. Results Basic performance testing showed that the combined restriction digital PCR assay enabled detection of 0.14% of the TWIST1 methylation level for the lymphocyte DNA. The combined restriction digital PCR assay from faeces samples had a sensitivity of 22.2% (95% confidence interval, 2.8-60.0%) for non-advanced adenoma, 47.1% (95% confidence interval, 23.0-72.2%) for advanced adenoma, and 33.7% (95% confidence interval, 23.7-45.0%) for colorectal cancer, and a specificity of 100.0%. Combination of faecal immunochemical test for haemoglobin and faecal combined restriction digital PCR assay increased sensitivity to 82.4% (95% confidence interval, 56.6-96.2%) for the detection of advanced adenoma. Conclusions We developed the combined restriction digital PCR assay, a possible highly sensitive methylation assay. Combination of faecal combined restriction digital PCR assay with faecal immunochemical test for haemoglobin may provide an alternative screening strategy for colorectal neoplasia, especially for potentially precancerous lesions.

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          Author and article information

          Journal
          Ann. Clin. Biochem.
          Annals of clinical biochemistry
          SAGE Publications
          1758-1001
          0004-5632
          Jan 2018
          : 55
          : 1
          Affiliations
          [1 ] 1 Department of Oncology and Laboratory Medicine, Yamaguchi University Graduate School of Medicine, Ube, Japan.
          [2 ] 2 Department of Gastroenterology and Hepatology, Yamaguchi University Graduate School of Medicine, Ube, Japan.
          [3 ] 3 Department of Gastroenterology, Sentohiru Hospital, Ube, Japan.
          [4 ] 4 Department of Gastroenterological, Breast and Endocrine Surgery, Yamaguchi University Graduate School of Medicine, Ube, Japan.
          [5 ] 5 Department of Translational Research and Developmental Therapeutics against Cancer, Yamaguchi University Graduate School of Medicine, Ube, Japan.
          Article
          10.1177/0004563217691064
          28081635

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