Cancer screening method

A screening and detection technology, applied in the field of cancer screening using methylated DNA as a biomarker

Active Publication Date: 2013-12-18
NAT DEFENSE MEDICAL CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] It can be seen that there are still many deficiencies in the above-mentioned existing cervical cancer screening methods, which are not a good design and need to be improved urgently

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Experimental program
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Effect test

Embodiment 1

[0039] Example 1 Materials and Methods

[0040] 1. Material

[0041] The test materials include a series of complete cervical lesion samples, including: squamouscell carcinoma (SCC, n=20), adenocarcinoma (AC, n=20), and normal cervical samples (n=10) ). All cervical samples, ovarian samples, and colorectal cancer samples were taken from Taipei Military General Hospital. The genomic DNA (genomic DNA) of each sample was extracted with QIAamp DNA set (QIAGEN) and used to compare and analyze DNA methylation in the whole genome Situation. In addition, Bioanalyzer (Agilent) was used to test the quality of genomic DNA before analysis. In this example, 10 μg fragmented DNA was used for MeDIP (Methyl DNA IP).

[0042] 2. Perform DNA methylation analysis using MeDIP and CpG island-Plus-Promoter arrays (CpGisland-Plus-Promoter arrays)

[0043] First, take Bioruptor TM UCS-200 (Diagenode) fragments genomic DNA to 300~1,000bp. Take 30μl polyclonal Anti-5'-methyl cytosine antibody (Abcam) in...

Embodiment 2

[0055] Example 2 Screening of methylation target genes in cervical cancer

[0056] After screening by the CpG island-Plus-Promoter arrays (CpG island-Plus-Promoter arrays), four target genes were screened for possible high methylation in cervical cancer cells. They are DBC1 (SEQ ID No:1). ), PDE8B (SEQ ID No: 2), PTPRR (SEQ ID No: 3), and ZNF582 (SEQ ID No: 4). The detailed information is shown in Table 3; from Table 3, we can see that these four genes except DBC1 have Besides being known to be related to bladder cancer, few studies have shown the connection between these genes and cervical cancer.

[0057] Table 3 Details of the methylated genes in cervical cancer cells screened by the CpG island-Plus-Promoter array

[0058]

[0059]

Embodiment 3

[0060] Example Trisulfite sequencing (BS) analysis of target gene methylation status in cervical lesion samples

[0061] Target gene: PTPRR

[0062] Test sample group:

[0063] 1. HeLa cervical cancer cell line (HeLa_0);

[0064] 2. HeLa cervical cancer cell line (HeLa_10D) treated with 10μM DNA methyltransferase inhibitor 5'-aza-2'-deoxycytidine (AZC);

[0065] 3. Simultaneous treatment of 10μM DNA methyltransferase inhibitor 5'-aza-2'-deoxycytidine (AZC) and 0.33μM TSA (Sigma Chemical Co., St.Louis, MO) HeLa cervical cancer cell line ( HeLa_DT);

[0066] 4. Cervical adenocarcinoma sample (AC);

[0067] 5. Sample of cervical squamous cell carcinoma (SCC);

[0068] 6. Control group (normal): normal cervical blood DNA is used as a control group without methylation.

[0069] After the aforementioned test samples were modified with sulfite, the sulfite sequencing (BS) was used to analyze whether the target gene (PTPRR) has hypermethylation in each test sample. The results are shown in Figure...

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Abstract

The invention relates to a cancer screening method which comprises the following steps that: (1) providing a tested body; (2) testing the CpG sequence methylation state of at least one target gene in genome DNA of the tested body, and the target gene comprises PTPRR, ZNF582, PDE8B and DBC1; (3) judging whether the tested body has cancer or lesions before cancer according to that whether the methylation state of the at least one target gene exists; and the methylation state detection method is methylation-specific PCR, MSP, quantitative methylation-specific PCR, QMSP, bisulfite sequencing (BS), microarrays, massspectrometer, denaturing high-performance liquidchromatography (DHPLC) and the like.

Description

Technical field [0001] The present invention relates to a method of cancer screening, in particular to a method of cancer screening using methylated DNA as a biomarker. Background technique [0002] Cervical cancer is one of the leading causes of death for women in the world and Taiwan. According to the World Health Organization (WHO) statistics in 2002, cervical cancer is the second leading cause of cancer deaths in women worldwide, second only to breast cancer; regular cervical cancer screenings Screening is the best way to prevent cervical cancer. There are two main ways to screen for cervical cancer. One is the most common Pap smear test (Pap smear), and the other is the human papilloma virus test (HPV). testing); Pap smear is to remove the secretions from the neck of the uterus and observe whether there are cancerous lesions in the epithelial cells that fall off with a microscope to detect cervical cancer at an early stage; while the HPV test is polymerase-linked Reaction (...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 赖鸿政
Owner NAT DEFENSE MEDICAL CENT
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