Aberrantly methylated genes in pancreatic cancer
The present invention provides a method for detecting pancreatic carcinoma in a subject. The method includes contacting a nucleic acid-containing specimen from the subject with an agent that provides a determination of the methylation state of at least one gene or associated regulatory region of the gene and identifying aberrant methylation of regions of the gene or regulatory region, wherein aberrant methylation is identified as being different when compared to the same regions of the gene or associated regulatory region in a subject not having the pancreatic carcinoma, thereby detecting pancreatic carcinoma in the subject.
1. An amplification primer pair, comprising a forward primer and a reverse primer as set forth in SEQ ID NOS: 5 to 16, 21 to 56, and 61 to 64, wherein the amplification primer pair can amplify a portion of a gene, or associated regulatory region of the gene, selected from the group consisting of reprimo, CLDN5, DR3, LDOC1, LHX1, NEFH, NPTX2, PIG11, SARP2, ST14, SMARCA1, TJP2, and WNT7A, or a combination thereof.
2. The amplification primer pair of claim 1 , which can specifically amplify a methylated 5′ regulatory region of the nucleic acid molecule.
3. The amplification primer pair of claim 2 , comprising SEQ ID NOS: 7 and 8, SEQ ID NOS: 11 and 12, SEQ ID NOS: 15 and 16, SEQ ID NOS: 23 and 24, SEQ ID NOS: 27 and 28, SEQ ID NOS: 31 and 32, SEQ ID NOS: 35 and 36, SEQ ID NOS: 39 and 40, SEQ ID NOS: 43 and 44, SEQ ID NOS: 47 and 48, SEQ ID NOS: 51 and 52, SEQ ID NOS: 55 and 56, or SEQ ID NOS: 63 and 64.
4. An amplification primer pair of claim 1 , which can specifically amplify an unmethylated 5′ regulatory region of the nucleic acid molecule.
5. The amplification primer pair of claim 4 comprising SEQ ID NOS: 5 and 6, SEQ ID NOS: 9 and 10, SEQ ID NOS: 13 and 14, SEQ ID NOS: 21 and 22, SEQ ID NOS: 25 and 26, SEQ ID NOS: 29 and 30, SEQ ID NOS: 33 and 34, SEQ ID NOS: 37 and 38, SEQ ID NOS: 41 and 42, SEQ ID NOS: 45 and 46, SEQ ID NOS: 49 and 50, SEQ ID NOS: 53 and 54, or SEQ ID NOS: 61 and 62.
6. A kit useful for the detection of pancreatic carcinoma in a subject comprising:
a) carrier means compartmentalized to receive a sample therein;
b) one or more containers comprising a first container containing a reagent which modifies unmethylated cytosine and a second container containing primers for amplification of a CpG-containing nucleic acid, wherein the primers are selected from SEQ ID NOS: 5 to 16, 21 to 56, and 61 to 64.
7. The kit of claim 6 , further comprising a third container containing a methylation sensitive restriction endonuclease.
8. The kit of claim 6 , wherein the modifying reagent is bisulfite.
9. The kit of claim 6 , wherein the primers hybridize with a target gene selected from the group consisting of reprimo, CLDN5, DR3, LDOC1, LHX1, NEFH, NPTX2, PIG11, SARP2, ST14, SMARCA1, TJP2, and WNT7A, or a combination thereof.