Methods and compositions for detecting gastrointestinal and other cancers
This application describes methods and compositions for detecting and treating vimentin-associated neoplasia. Differential methylation of the vimentin nucleotide sequences has been observed in vimentin-associated neoplasia such as neoplasia of the upper or lower gastrointestinal tract, pancreas, and/or bladder.
1. A method for determining vimentin methylation in a human subject, comprising:
a) obtaining a sample from a human subject; and b) assaying a vimentin nucleic acid in the sample for the presence or absence of methylation within a nucleotide sequence selected from the group consisting of SEQ ID NOs: 2 and fragments thereof, and SEQ ID NOS:40-45, wherein the sample is obtained from a subject suspected of having or is known to have an esophageal neoplasia.
2. The method of claim 1 , wherein the sample is obtained from an esophageal brushing.
3. The method of claim 1 , wherein the esophageal neoplasia is Barrett's esophagus or Barrett's esophagus with high grade dysplasia.
4. The method of claim 1 , wherein said esophageal neoplasia is adenocarcinoma of the esophagus.
5. The method of claim 1 , wherein the assay comprises methylation-specific PCR.
6. The method of claim 5 , comprising: a) treating DNA from the sample with a compound that converts non-methylated cytosine bases in the DNA to a different base; b) amplifying a region of the compound-converted vimentin nucleotide sequence with a forward primer and a reverse primer; and c) analyzing the methylation patterns of said vimentin nucleotide sequences.
7. The method of claim 5 , comprising: a) treating DNA from the sample with a compound that converts non-methylated cytosine bases in the DNA to a different base; b) amplifying a region of the compound-converted vimentin nucleotide sequence with a forward primer and a reverse primer; and c) detecting the presence and/or amount of the amplified product.
8. The method of claim 5 , wherein the compound used to treat DNA is a bisulfite compound.
9. The method of any of claim 1 , wherein the assay comprises using a methylation-specific restriction enzyme.
10. The method of claim 9 , wherein said methylation-specific restriction enzyme is selected from HpaII, SmaI, SacII, EagI, MspI, BstUI, and BssHII.
11. A method for monitoring vimentin methylation in a human subject, comprising:
a) assaying a vimentin nucleic acid in a sample from the human subject for the presence or absence of methylation within a nucleotide sequence selected from the group consisting of SEQ ID NOs: 2 and fragments thereof, and SEQ ID NOS:40-45 for a first time; and
b) at a later time, assaying a vimentin nucleic acid in another sample from the same human subject for the presence or absence of methylation within a nucleotide sequence assayed in step a), wherein the sample is obtained from a subject suspected of having or is known to have esophageal neoplasia.
12. The method of claim 11 , wherein the sample is obtained from an esophageal brushing.
13. The method of claim 11 , wherein the esophageal neoplasia is Barrett's esophagus or Barrett's esophagus with high grade dysplasia.
14. The method of claim 11 , wherein said esophageal neoplasia is adenocarcinoma of the esophagus.
15. The method of claim 11 , wherein the assay comprises methylation-specific PCR.
16. The method of claim 15 , comprising: a) treating DNA from a sample with a compound that converts non-methylated cytosine bases in the DNA to a different base; b) amplifying a region of the compound-converted vimentin nucleotide sequence with a forward primer and a reverse primer; and c) analyzing the methylation patterns of said vimentin nucleotide sequences.
17. The method of claim 15 , comprising: a) treating DNA from the sample with a compound that converts non-methylated cytosine bases in the DNA to a different base; b) amplifying a region of the compound-converted vimentin nucleotide sequence with a forward primer and a reverse primer; and c) detecting the presence and/or amount of the amplified product.
18. The method of claim 15 , wherein the compound used to treat DNA is a bisulfite compound.
19. The method of claim 11 , wherein the assay comprises using a methylation-specific restriction enzyme.
20. The method of claim 19 , wherein said methylation-specific restriction enzyme is selected from HpaII, SmaI, SacII, EagI, MspI, BstUI, and BssHII.