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A molecular marker for the diagnosis and treatment of rectal adenocarcinoma

A rectal adenocarcinoma, a pair of technology, applied in the field of biomedicine, can solve the problems of invasiveness, high cost, low sensitivity and specificity, and poor patient compliance

Active Publication Date: 2019-01-18
QINGDAO MEDINTELL BIOMEDICAL CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Clinically, the early diagnosis of rectal adenocarcinoma mainly relies on colonoscopy, imaging examination and serological marker carcinoembryonic antigen (CEA). Colonoscopy is the most reliable method for the diagnosis of rectal adenocarcinoma, but it is invasive and expensive. Poor compliance; CT and ultrasonography are not easy to find small lesions, and the early diagnosis of rectal adenocarcinoma is limited; CEA is a widely used serum marker in clinical practice, with low sensitivity and specificity, and is mainly used in patients with rectal adenocarcinoma treatment monitoring
[0004] At present, the histological diagnosis of rectal adenocarcinoma mainly relies on HE stained sections, and there is no marker that can be used as an independent tissue diagnosis. A new specific marker for the diagnosis, prognosis and treatment of rectal adenocarcinoma has been found becomes very necessary

Method used

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  • A molecular marker for the diagnosis and treatment of rectal adenocarcinoma
  • A molecular marker for the diagnosis and treatment of rectal adenocarcinoma
  • A molecular marker for the diagnosis and treatment of rectal adenocarcinoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Example 1 Screening for gene markers associated with rectal adenocarcinoma

[0060] 1. Sample collection

[0061] Paracancerous tissue and rectal adenocarcinoma tissue samples were collected from 8 cases of rectal adenocarcinoma. The above samples are surgical resection specimens of patients with rectal adenocarcinoma, and all the above samples were obtained with the consent of the organizational ethics committee.

[0062] 2. RNA sample preparation (using E.Z.N.A. kit to operate)

[0063] The tissues obtained above were shredded, put into liquid nitrogen and ground into powder, and RNA was extracted and isolated according to the instructions in the kit. details as follows:

[0064] 1) Isolation of RNA:

[0065] A. Add RNA to tissue homogenate or cells- Reagent II 1ml;

[0066] B. Stand at room temperature for 3 minutes, add 0.2ml chloroform and shake vigorously for 15 seconds;

[0067] C. Place on ice for 10 minutes;

[0068] D. Centrifuge at 12000g for 15min...

Embodiment 2

[0084] Example 2 QPCR sequencing to verify the differential expression of SHROOM4 gene

[0085] 1. Large-sample QPCR verification of differential expression of SHROOM4 gene. According to the sample collection method in Example 1, 70 cases of rectal adenocarcinoma paracancerous tissues and 70 cases of rectal adenocarcinoma tissues were selected.

[0086] 2. The specific operation steps of QPCR are as follows:

[0087] (1) RNA extraction

[0088] The RNA extraction procedure was as described in Example 1.

[0089] (2) reverse transcription

[0090] A. Configure the mixed solution in Microtube: Oligo dT (50μM) 1μl, dNTP mixed solution (10mM) 1μl, RNA template 5μg, add ddH 2 0 to 10 μl, mix well;

[0091] B. Carry out denaturation and annealing reactions on the PCR instrument according to the following reaction conditions: after 5 minutes at 65°C, place it on ice immediately;

[0092] C. Prepare the following reverse transcription reaction solution in the above-mentioned Mic...

Embodiment 3

[0111] Example 3 Overexpression of SHROOM4 gene

[0112] 1. Cell culture

[0113] Human rectal adenocarcinoma cell line HRC-99 was cultured in RPMI1640 medium containing 10% calf serum and 1% P / S in an incubator at 37°C, 5% CO2, and 90% relative humidity. Change the medium once every 2-3 days, and use 0.25% EDTA-containing trypsin for routine digestion and passage.

[0114] 2. Overexpression of SHROOM4 gene

[0115] 2.1 Construction of SHROOM4 gene expression vector

[0116] Amplification primers were designed according to the coding sequence of the SHROOM4 gene (as shown in SEQ ID NO.1), and the primer sequences were as follows:

[0117] Forward primer: 5'-CCGAAGCTTGCCACCATGGAGAACCGGCCTG-3' (SEQ ID NO.7)

[0118] Reverse primer: 5'-CGGGCGGCCGCGAAATTGCTGGGCCCCA-3' (SEQ ID NO.8)

[0119] The coding sequence of the full-length SHROOM4 gene was amplified from the cDNA library of adult fetal brain (clontech company, product number: 638831), and the above cDNA sequence was ins...

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Abstract

The invention discloses a molecular marker for rectal adenocarcinoma diagnosis and treatment. The molecular mark is a SHROOM4 gene which is in a low-expression mode in tissue of a patient with rectal adenocarcinoma, and rectal adenocarcinoma apoptosis can be promoted by improving expression of SHROOM4. According to the molecular marker for the rectal adenocarcinoma diagnosis and treatment, sensibility and specificity of the rectal adenocarcinoma diagnosis are greatly improved, and meanwhile a new target site is provided for a gene therapy of the rectal adenocarcinoma.

Description

technical field [0001] The invention belongs to the field of biomedicine and relates to a molecular marker used for the diagnosis and treatment of rectal adenocarcinoma, and the molecular marker is SHROOM4. Background technique [0002] Rectal adenocarcinoma is a malignant tumor of the digestive system with a high incidence in the world, and its incidence is increasing year by year. In the United States, its morbidity and mortality rate ranks third among malignant tumors. In China, its male and female malignant tumor mortality rate is the fourth, and it has serious harm to human health and life. The occurrence and development of rectal adenocarcinoma is a complex process of multi-gene changes and multi-stage carcinogenesis, including gene activation, inactivation or deletion of tumor suppressor genes, and mutation of mismatch repair genes. Rectal adenoma develops from benign lesions through precancerous lesions to malignant lesions, until the complex evolution process of i...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6886G01N33/68G01N33/574A61K45/00A61P35/00
CPCA61K45/00C12Q1/6886C12Q2600/158G01N33/57407G01N33/57484G01N33/68G01N2333/47
Inventor 宋宏涛王晓云杨承刚
Owner QINGDAO MEDINTELL BIOMEDICAL CO LTD
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