CANCER SCREENING METHOD

• US 20080311570A1
• Filed: 06/15/2007
• Published: 12/18/2008
• Est. Priority Date: 06/15/2007
• Status: Active Grant

• Current Assignee: National Defense Medical Center (Government of The Republic of China)
• Original Assignee: National Defense Medical Center (Government of The Republic of China)
• Inventors: Lai, Hung-Cheng
• Granted Patent: US 7,820,386 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/6886 for cancer immunoassay for cancer G01N33/574
• CPC Class Codes: C12Q 2600/112 Disease subtyping, staging or classification
• CPC Class Codes: C12Q 2600/154 Methylation markers
• CPC Class Codes: C12Q 2600/156 Polymorphic or mutational markers
• CPC Class Codes: C12Q 2600/16 Primer sets for multiplex assays

Abstract

A method for screening cancer comprises the following steps: (1) providing a test specimen; (2) detecting the methylation state of the CpG sequence in at least one target gene within the genomic DNA of the test specimen, wherein the target genes is consisted of SOX1, PAX1, LMX1A, NKX6-1, WT1 and ONECUT1; and (3) determining whether there is cancer or cancerous pathological change in the specimen based on the presence or absence of the methylation state in the target gene; wherein method for detecting methylation state is methylation-specific PCR (MSP), quantitative methylation-specific PCR (QMSP), bisulfite sequencing (BS), microarrays, mass spectrometer, denaturing high-performance liquid chromatography (DHPLC), and pyrosequencing.

Claims

1. A method for screening cancer, characterized in that it comprises of detecting the methylation state of a target gene in the cell of the test specimen as a screening index to determine the presence or absence of the cancer, the method comprising following steps:

step 1;

providing a test specimen;

step 2;

detecting the methylation state of the CpG sequence in at least one target gene within the genomic DNA of the test specimen, wherein the target genes is consisted of SOX1, PAX1, LMX1A, NKX6-1, WT1 and ONECUT1; and

step 3;

determining whether there is a cancer or cancerous pathological change in the specimen based on the presence or absence of the methylation state in the target gene.

2. A method for screening cancer as recited in claim 1, wherein the test specimen is a cervical smear, ascites, blood, urine, feces, sputum, oral mucosa cell, gastric juice, bile, and cervical epithelial cell.

3. A method for screening cancer as recited in claim 1, wherein method for detecting the methylation state of the CpG sequence in the target gene is methylation-specific PCR (MSP), quantitative methylation-specific PCR (QMSP), bisulfite sequencing (BS), microarrays, mass spectrometer, denaturing high-performance liquid chromatography (DHPLC), and pyrosequencing.

4. A method for screening cancer as recited in claim 1, wherein the target gene SOX1 has a nucleotide sequence as depicted in SEQ ID No:

1.

5. A method for screening cancer as recited in claim 1, wherein the target gene PAX1 has a nucleotide sequence as depicted in SEQ ID No:

2.

6. A method for screening cancer as recited in claim 1, wherein the target gene LMX1A has a nucleotide sequence as depicted in SEQ ID No:

3.

7. A method for screening cancer as recited in claim 1, wherein the target gene NKX6-1 has a nucleotide sequence as depicted in SEQ ID No:

4.

8. A method for screening cancer as recited in claim 1, wherein the target gene WT1 has a nucleotide sequence as depicted in SEQ ID No:

5.

9. A method for screening cancer as recited in claim 1, wherein the target gene ONECUT1 has a nucleotide sequence as depicted in SEQ ID No:

6.

10. A method for screening cervical cancer, characterized in that it comprises of detecting the methylation state of the target gene in the cell of the test specimen as a screening index to determine the existence of the cervical cancer, the method comprising following steps:

step 1;

providing a test specimen;

step 2;

detecting the methylation state of the CpG sequence in at least one target gene within the genomic DNA of the test specimen, wherein the target genes is consisted of SOX1, PAX1, LMX1A, NKX6-1, WT1 and ONECUT1; and

step 3;

determining whether there is a cervical cancer or cancerous pathological change in the specimen based on the presence or absence of the methylation state in the target gene.

11. A method for screening cervical cancer as recited in claim 10, wherein the test specimen is a cervical smear, blood, urine, and cervical epithelial cell.

12. A method for screening cervical cancer as recited in claim 10, wherein the test specimen is abnormal cervical smear.

13. A method for screening cervical cancer as recited in claim 10, wherein the test specimen is human papilloma virus testing (HPV testing) positive cervical cell specimen.

14. A method for screening cervical cancer as recited in claim 10, wherein method for detecting the methylation state of the CpG sequence in the target gene is methylation-specific PCR (MSP), quantitative methylation-specific PCR (QMSP), bisulfite sequencing (BS), microarrays, mass spectrometer, denaturing high-performance liquid chromatography (DHPLC), and pyrosequencing.

15. A method for screening cervical cancer as recited in claim 10, wherein the target gene SOX1 has a nucleotide sequence as depicted in SEQ ID No:

1.

16. A method for screening cervical cancer as recited in claim 10, wherein the target gene PAX1 has a nucleotide sequence as depicted in SEQ ID No:

2.

17. A method for screening cervical cancer as recited in claim 10, wherein the target gene LMX1A has a nucleotide sequence as depicted in SEQ ID No:

3.

18. A method for screening cervical cancer as recited in claim 10, wherein the target gene NKX6-1 has a nucleotide sequence as depicted in SEQ ID No:

4.

19. A method for screening cervical cancer as recited in claim 10, wherein the target gene WT1 has a nucleotide sequence as depicted in SEQ ID No:

5.

20. A method for screening cervical cancer as recited in claim 10, wherein the target gene ONECUT1 has a nucleotide sequence as depicted in SEQ ID No:

6.

21. A method for screening ovarian cancer, characterized in that it comprises of detecting the methylation state of the target gene in the cell of the test specimen as a screening index to determine the existence of the ovarian cancer, the method comprising following steps:

step 1;

providing a test specimen;

step 2;

detecting the methylation state of the CpG sequence in at least one target gene within the genomic DNA of the test specimen, wherein the target genes is consisted of SOX1, PAX1, and LMX1A; and

step 3;

determining whether there is an ovarian cancer or cancerous pathological change in the specimen based on the presence or absence of the methylation state in the target gene.

22. A method for screening ovarian cancer as recited in claim 21, wherein the test specimen is ascite, blood, and urine.

23. A method for screening ovarian cancer as recited in claim 21, wherein method for detecting the methylation state of the CpG sequence in the target gene is methylation-specific PCR (MSP), quantitative methylation-specific PCR (QMSP), bisulfite sequencing (BS), microarrays, mass spectrometer, denaturing high-performance liquid chromatography (DHPLC), and pyrosequencing.

24. A method for screening ovarian cancer as recited in claim 21, wherein the target gene SOX1 has a nucleotide sequence as depicted in SEQ ID No:

1.

25. A method for screening ovarian cancer as recited in claim 21, wherein the target gene PAX1 has a nucleotide sequence as depicted in SEQ ID No:

2.

26. A method for screening ovarian cancer as recited in claim 21, wherein the target gene LMX1A has a nucleotide sequence as depicted in SEQ ID No:

3.

27. A method screening liver cancer, characterized in that it comprises of detecting the methylation state of the target gene in the cell of the test specimen as a screening index to determine the existence of the liver cancer, the method comprising following steps:

step 1;

providing a test specimen;

step 2;

detecting the methylation state of the CpG sequence in at least one target gene within the genomic DNA of the test specimen, wherein the target genes is consisted of SOX1, and NKX6-1; and

step 3;

determining whether there is a liver cancer or cancerous pathological change in the specimen based on the presence or absence of the methylation state in the target gene.

The test specimens may be an ascites, blood, urine, feces, gastric juice, bile, and the like.

28. A method for screening liver cancer as recited in claim 27, wherein the test specimens is ascites, blood, urine, feces, gastric juice, and bile.

29. A method for screening liver cancer as recited in claim 27, wherein method for detecting the methylation state of the CpG sequence in the target gene is methylation-specific PCR (MSP), quantitative methylation-specific PCR (QMSP), bisulfite sequencing (BS), microarrays, mass spectrometer, denaturing high-performance liquid chromatography (DHPLC), and pyrosequencing.

30. A method for screening liver cancer as recited in claim 27, wherein the target gene SOX1 has a nucleotide sequence as depicted in SEQ ID No:

1

31. A method for screening liver cancer as recited in claim 27, wherein the target gene NKX6-1 has a nucleotide sequence as depicted in SEQ ID No:

4