Methods and compositions for detecting gastrointestinal and other cancers
View Patent ↗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 detecting vimentin methylation in a human subject, comprising:
a) obtaining a sample from a human subject suspected of having or is known to have colon neoplasia; 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 NO: 2 and fragments thereof, and SEQ ID NOS:40-45.
2. The method of claim 1 , wherein the sample is a bodily fluid selected from the group consisting of blood, serum, plasma, a blood-derived fraction, stool, urine, and a colonic effluent.
3. The method of claim 1 , wherein said colon neoplasia is colon cancer.
4. The method of claim 1 , wherein the assay comprises methylation-specific PCR.
5. The method of claim 4 , 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.
6. The method of claim 4 , 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.
7. The method of claim 4 , wherein the compound used to treat DNA is a bisulfite compound.
8. The method of any of claim 1 , wherein the assay comprises using a methylation-specific restriction enzyme.
9. The method of claim 8 , wherein said methylation-specific restriction enzyme is selected from HpaII, SmaI, SacII, EagI, MspI, BstUI, and BssHII.
10. A method for monitoring vimentin methylation in a human subject, comprising:
a) assaying a vimentin nucleic acid in a sample from the human subject suspected of having or is known to have colon neoplasia for the presence or absence of methylation within a nucleotide sequence selected from the group consisting of SEQ ID NO: 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).
11. The method of claim 10 , wherein each sample is a bodily fluid selected from the group consisting of blood, serum, plasma, a blood-derived fraction, stool, urine, and a colonic effluent.
12. The method of claim 10 , wherein said colon neoplasia is colon cancer.
13. The method of claim 10 , wherein the assay comprises methylation-specific PCR.
14. The method of claim 13 , 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.
15. The method of claim 13 , 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.
16. The method of claim 13 , wherein the compound used to treat DNA is a bisulfite compound.
17. The method of claim 10 , wherein the assay comprises using a methylation-specific restriction enzyme.
18. The method of claim 17 , wherein said methylation-specific restriction enzyme is selected from HpaII, SmaI, SacII, EagI, MspI, BstUI, and BssHII.