Application of HOXA7 methylation detection reagent to preparation of lung cancer diagnosis reagent

A HOXA7 and detection reagent technology, applied in the field of human HOXA7 gene methylation detection, can solve the problems of limited, difficult to achieve clinical use, and delayed treatment timing.

Active Publication Date: 2020-04-07
CREATIVE BIOSCIENCES (GUANGZHOU) CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

(4) Bone scan: the sensitivity of detection of bone metastases of lung cancer is high, but there is a certain false positive rate
If an early tumor screening product cannot screen out all or most of the patients, those who miss the test will not be able to receive sufficient risk warnings, thus delaying the timing of treatment, which is a huge loss for patients loss
[0018] Although some tumor markers related to lung cancer have been found in the prior art, the sensitivity and specificity of these tumor markers cannot meet the demand due to the limitation of detection reagents or detection methods for these tumor markers. Therefore, the current There is still a need for further research in the field of screening methods that can

Method used

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  • Application of HOXA7 methylation detection reagent to preparation of lung cancer diagnosis reagent
  • Application of HOXA7 methylation detection reagent to preparation of lung cancer diagnosis reagent
  • Application of HOXA7 methylation detection reagent to preparation of lung cancer diagnosis reagent

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0076] Example 1: Selection of Detection Target Genes

[0077] Methylated DNA has obvious advantages as a detection target. Compared with protein markers, DNA can be amplified and easily detected; compared with mutation markers, DNA methylation sites are located in genes The specific site, generally in the promoter region, makes the detection easier and more convenient. In order to complete the present invention, the inventor screened hundreds of genes, and selected better HOXA7, SHOX2, PCDHGA12, HOXD8, GATA3 as candidate detection genes, β-actin gene as an internal reference gene, and studied the methylation of each gene According to the distribution of sites, the primers and probes designed for detection were used for real-time fluorescent quantitative methylation-specific polymerase chain reaction (real-time fluorescent quantitative methylation-specific PCR, qMSP) detection. The detection primers and probes for each gene are as follows:

[0078] The detection primers and ...

Embodiment 2

[0126] Example 2: Detection of HOXA7, SHOX2, PCDHGA12, HOXD8 and GATA3 genes in sputum

[0127] Sample information: A total of 90 cases of sputum samples were tested, including 55 cases of normal control group samples, 35 cases of cancer group control samples, 12 cases of squamous cell carcinoma, 6 cases of small cell carcinoma, 9 cases of adenocarcinoma, and large There were 2 cases of cell carcinoma and 6 cases of unclassified lung cancer.

[0128] Experimental procedure:

[0129] a. Collect sputum samples from patients diagnosed with lung cancer and non-lung cancer patients. After thickening with DTT, centrifuge to separate the pellet and separate the cells, wash them twice with PBS, and then use the DNA extraction kit (HiPure FFPE DNA Kit, D3126-03) to extract DNA.

[0130] b. Use the DNA conversion kit (EZ DNA Methylation Kit, D5002) of ZYMO RESEARCH Biological Company to carry out the bisulfite modification of DNA.

[0131] c. The liquid distribution system is as foll...

Embodiment 3

[0141] Example 3: Detection of HOXA7 and SHOX2 genes in sputum

[0142] A large number of documents show that SHOX2 can be used as a marker for the detection of lung cancer, and there are patents [CN201510203539-method and kit for diagnosing methylation of human SHOX2 gene and human RASSF1A gene-application disclosure], SHOX2 in alveolar lavage fluid, lesion sites It has a high detection rate in samples such as tissue, pleural effusion, and sputum. In order to verify the detection effect of HOXA7, the inventors simultaneously detected the detection efficiency of HOXA7 and SHOX2 gene. In this example, the detection efficiency of SHOX2 gene used the primer and probe sequences disclosed in patent CN201510203539, and the It is expressed as SHOX2_n3 to distinguish it from the SHOX2 gene detected by self-designed primers and probes in Examples 1 and 2 of the present invention.

[0143] The detection primers and probes for each gene are as follows:

[0144] The detection primers an...

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Abstract

The invention belongs to the field of gene diagnosis, and relates to application of a detection reagent using methylation of HOXA7 as a detection object to preparation of a lung cancer diagnosis reagent. The detection reagent uses HOXA7 as a tumor marker for detecting the lung cancer, reaches high sensitivity of 88.6% and high specificity of 95% in sputa, reaches high sensitivity of 76.2% and highspecificity of 95% in lavage fluid, and is higher in detection sensitivity than existing reported tumor markers of the lung cancer; particularly for adenocarcinomas, sensitivity of the detection reagent is greatly promoted; and the detection reagent has great application value for detection and diagnosis on the lung adenocarcinomas.

Description

technical field [0001] The invention belongs to the field of gene diagnosis, and more specifically, the invention relates to the application of a human HOXA7 gene methylation detection reagent in the preparation of a lung cancer detection reagent, and a method for human HOXA7 gene methylation detection. [0002] technical background [0003] Lung cancer is a malignant tumor of the lung originating from the bronchial mucosa, glands or alveolar epithelium. According to the pathological type, it can be divided into: 1. Small cell lung cancer (SCLC): a special pathological type of lung cancer, with obvious tendency of distant metastasis and poor prognosis, but most patients are sensitive to radiotherapy and chemotherapy. 2. Non-small cell lung cancer (NSCLC): other pathological types of lung cancer other than small cell lung cancer, including squamous cell carcinoma, adenocarcinoma, large cell carcinoma, etc. There are certain differences in biological behavior and clinical cour...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886C12Q2600/154C12N15/11C12Q1/68
Inventor 李仕良赵霞牛智通黄龙武赵荣淞吴幽治邹鸿志
Owner CREATIVE BIOSCIENCES (GUANGZHOU) CO LTD
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