Usage of HOXA7 and HOXA9 methylation detection reagents in preparation of lung cancer diagnostic reagents

A technology of HOXA7 and HOXA9, which is applied in the direction of DNA/RNA fragments, recombinant DNA technology, microbial measurement/inspection, etc., can solve the loss of patients, the sensitivity and specificity of tumor markers cannot meet the needs, and there is no non-invasive screening method for lung cancer and other problems to achieve the effect of improving the detection effect

Active Publication Date: 2020-04-07
CREATIVE BIOSCIENCES (GUANGZHOU) CO LTD
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  • Summary
  • 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 c...

Method used

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  • Usage of HOXA7 and HOXA9 methylation detection reagents in preparation of lung cancer diagnostic reagents
  • Usage of HOXA7 and HOXA9 methylation detection reagents in preparation of lung cancer diagnostic reagents
  • Usage of HOXA7 and HOXA9 methylation detection reagents in preparation of lung cancer diagnostic reagents

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0089] Example 1: Selection of Detection Target Genes

[0090] 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 the gene Specific sites, usually in the promoter region, make detection easier and more convenient. In order to complete the present invention, the inventor screened a large number of genes, selected representative HOXA7, HOXA9, SHOX2, PCDHGA12, HOXD8, and GATA3 as candidate detection genes, and β-actin gene as an internal reference gene to study the methylation sites of each gene According to the distribution, 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:

[0091] The detection p...

Embodiment 2

[0148] Example 2: Detection of HOXA7, HOXA9, SHOX2, PCDHGA12, HOXD8 and GATA3, and combinations of HOXA7 and HOXA9 in sputum

[0149] Sample information: A total of 90 sputum samples were tested, including 55 samples from the normal control group, 35 samples from the cancer group, and 12 cases of squamous cell carcinoma, 6 cases of small cell carcinoma, 9 cases of adenocarcinoma, and large cell carcinoma in the 35 samples of the cancer group. There were 2 cases of carcinoma and 6 cases of lung cancer not clearly classified.

[0150] Experimental procedure:

[0151] a. Collect sputum specimens 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 Magen’s DNA extraction kit (HiPure FFPE DNA Kit, D3126-03) to extract DNA.

[0152] b. Use the DNA conversion kit (EZ DNAMethylation Kit, D5002) of ZYMO RESEARCH Biological Company to carry out ...

Embodiment 3

[0169] Example 3: Detection of HOXA7, HOXA9 and SHOX2 genes in sputum

[0170] A large number of documents show that SHOX2 can be used as a marker for the detection of lung cancer, and a patent shows [CN201510203539 Method and kit for diagnosing methylation of human SHOX2 gene and human RASSF1A gene - application disclosure], SHOX2 can be used in alveolar lavage fluid, lesion tissue , pleural effusion, sputum and other samples have a high detection rate. In order to verify the detection effect of HOXA7 and HOXA9, the inventors simultaneously detected HOXA7 and HOXA9 SHOX2_n3 genes. In this example, the detection efficiency of the SHOX2 gene uses the primer and probe sequences disclosed in the patent CN201510203539, and the SHOX2 gene is expressed as SHOX2_n3, to distinguish it from the self-designed primer and probe sequences used in Examples 1 and 2 of the present invention. Needle detection of the SHOX2 gene.

[0171] detection results.

[0172] The detection primers and ...

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Abstract

The invention belongs to the field of genetic diagnosis, and relates to application of detection reagents, which use sputum as test samples as well as methylation of HOXA7 and HOXA9 as detect targets,in preparation of lung cancer diagnostic reagents. For the first time, the invention utilizes the HOXA7 and the HOXA9 in the sputum together as a tumor marker for lung cancer detection; and the tumormarker in the sputum has a sensitivity up to 97.1% and a specificity up to 90.9%, so that the tumor marker has higher detection sensitivity compared with existing reported tumor markers of the lung cancer. Especially for adenocarcinoma, the detection sensitivity of the tumor marker is even as high as 100%, so that the tumor marker is of very great application values to adenocarcinoma detection and diagnosis.

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 and human HOXA9 gene methylation detection reagent in the preparation of lung cancer diagnostic reagents, and a method for human HOXA7 gene and HOXA9 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 biolog...

Claims

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Application Information

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