Detecting esophageal disorders
Provided herein is technology for esophageal disorder screening and particularly, but not exclusively, to methods, compositions, and related uses for detecting the presence of esophageal disorders (e.g., Barrett's esophagus, Barrett's esophageal dysplasia, etc.). In addition, the technology provides methods, compositions and related uses for distinguishing between Barrett's esophagus and Barrett's esophageal dysplasia, and between Barrett's esophageal low-grade dysplasia, Barrett's esophageal high-grade dysplasia, and esophageal adenocarcinoma within samples obtained through endoscopic brushing or nonendoscopic whole esophageal brushing or swabbing using a tethered device (e.g. such as a capsule sponge, balloon, or other device).
1. A method for measuring the methylation level of no more than ten genetic markers at least including ELMO1 and CD1D comprising:
a) extracting genomic DNA from a biological sample of a human individual suspected of having or having Barrett's esophageal dysplasia and/or esophageal adenocarcinoma,
b) treating the extracted genomic DNA with bisulfite,
c) amplifying the bisulfite-treated genomic DNA with primers consisting of a pair of primers specific for ELMO1 and a pair of primers specific for CD1D, and
d) measuring the methylation level of no more than ten genetic markers at least including ELMO1 and CD1D by measuring the methylation of CpG sites in the genetic markers by methylation-specific PCR, quantitative methylation-specific PCR, methylation sensitive DNA restriction enzyme analysis or bisulfite genomic sequencing PCR.
2. The method of claim 1 , wherein the biological sample comprises esophageal tissue obtained through whole esophageal swabbing or brushing.
3. The method of claim 1 , wherein the sample is a stool sample, a tissue sample, a blood sample, or a urine sample.
4. The method of claim 1 wherein the pair of primers specific for ELMO1 consists of SEQ ID Nos: 11 and 12 and the pair of primers specific for CD1D consists of SEQ ID NO: 9 and 10.
5. A method, comprising
a) obtaining a sample comprising DNA from a human individual suspected of having or having Barrett's esophageal dysplasia and/or esophageal adenocarcinoma,
b) treating the obtained DNA with a reagent which selectively modifies unmethylated cytosine residues in the obtained DNA to produce modified residues but which does not modify methylated cytosine residues;
c) measuring the methylation level of no more than ten genetic markers at least including ELMO1 and CD1D in the DNA having the treating of step b).
6. The method of claim 5 wherein the treating of step b) is accomplished through bisulfite modification of the obtained DNA.
7. The method of claim 5 wherein said measuring the methylation level of no more than ten genetic markers at least including ELMO1 and CD1D in the DNA having undergone the treating of step b) is achieved by a technique selected from the group consisting of methylation-specific PCR, quantitative methylation-specific PCR, methylation-sensitive DNA restriction enzyme analysis, quantitative bisulfite pyrosequencing, and bisulfite genomic sequencing PCR.
8. The method of claim 5 wherein the sample comprises esophageal tissue, wherein the esophageal tissue is obtained through whole esophageal swabbing or brushing or use of a sponge capsule device.