Application of CLEC1B (C-type lectin domain family 1 member B) genes on diagnosis and treatment of bile duct carcinoma

A cholangiocarcinoma and gene technology, applied in the direction of gene therapy, medical preparations containing active ingredients, biochemical equipment and methods, etc., can solve the problems of low radical cure rate, inability to apply early diagnosis of cholangiocarcinoma, lack of diagnostic methods, etc. , to achieve early diagnosis, timely gene diagnosis, and reduce mortality

Active Publication Date: 2015-12-02
BEIJING MEDINTELL BIOMED CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the above method cannot be applied to the early diagnosis of cholangiocarcinoma. When the above method is used for diagnosis, most of the patients have advanced tumors, and the rate of radical surgery is low. At present, there is still a lack of a specific and sensitive diagnostic method for cholangiocarcinoma.

Method used

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  • Application of CLEC1B (C-type lectin domain family 1 member B) genes on diagnosis and treatment of bile duct carcinoma
  • Application of CLEC1B (C-type lectin domain family 1 member B) genes on diagnosis and treatment of bile duct carcinoma
  • Application of CLEC1B (C-type lectin domain family 1 member B) genes on diagnosis and treatment of bile duct carcinoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Example 1 Screening of gene markers associated with cholangiocarcinoma

[0052] 1. Sample collection

[0053] Eight samples of normal bile duct tissue and cholangiocarcinoma tissue were collected. The above samples are surgical resection specimens of patients with cholangiocarcinoma, and all the above samples were obtained with the consent of the organizational ethics committee.

[0054] 2. Preparation of RNA samples (operated using QIAGEN tissue RNA extraction kit)

[0055] 1) Tissue extraction

[0056] In a clean area with less RNase interference, use a mortar containing an appropriate amount of liquid nitrogen to weigh about 20 mg of an isolated lung adenocarcinoma tissue sample, grind it to powder with a pestle, and then transfer the sample to a 2 ml RNase-free in a centrifuge tube. Add 300 μl lysate, place in a homogenizer, grind thoroughly for 1-5min, centrifuge at 12000g, 4°C for 10min, and transfer the supernatant to a new 1.5ml centrifuge tube. Add 600 μl ...

Embodiment 2

[0074] Example 2 QPCR sequencing verification of differential expression of CLEC1B gene

[0075] 1. Large-sample QPCR verification of differential expression of CLEC1B gene. According to the sample collection method in Example 1, 80 cases of cholangiocarcinoma tissue and 80 cases of normal bile duct tissue were selected.

[0076] 2. The specific operation steps of QPCR are as follows:

[0077] (1) RNA extraction

[0078] After collecting the samples, freeze them in liquid nitrogen. After taking them out, put the tissue into a pre-cooled mortar for grinding. After the tissue sample is powdered:

[0079] 1) Add Trizol and place at room temperature for 5 minutes;

[0080] 2) Add 0.2ml of chloroform, vibrate the centrifuge tube vigorously, mix well, and place it at room temperature for 5-10min;

[0081] 3) Centrifuge at 12000rpm for 15min, transfer the upper aqueous phase to another new centrifuge tube (be careful not to absorb the protein material between the two aqueous phas...

Embodiment 3

[0093] Example 3 CLEC1B Gene Overexpression

[0094] 1. Human cholangiocarcinoma cell line QBC939 was cultured at 37°C and 5% CO in DMEM (high glucose) medium containing 10% calf serum. 2 , Cultivated in an incubator with a relative humidity of 90%. The medium was changed once every 2-3 days, and 0.25% trypsin was used for routine digestion and passage.

[0095] 2. Overexpression of CLEC1B gene

[0096] 2.1 Construction of CLEC1B gene expression vector

[0097] Amplification primers were designed according to the coding sequence of CLEC1B gene (as shown in SEQIDNO.1), and the primer sequences were as follows: the forward primer was 5'-CCGGGATCCGCCACCATGCAGGATGAAGAT-3' (SEQIDNO.7), and the reverse primer was 5'-CGGCTCGAGAGGTAGTTGGTCCACCTTGG-3 '(SEQ ID NO. 8). The coding sequence of the full-length CLEC1B gene was amplified from the cDNA library of adult fetal brain (clontech company, catalog number: 638831), and the above cDNA sequence was double-digested with restriction e...

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Abstract

The invention discloses application of CLEC1B (C-type lectin domain family 1 member B) genes on diagnosis and treatment of bile duct carcinoma. RNA (ribonucleic acid)-sep screening is carried out, and a large sample RT-PCR (reverse transcription-polymerase chain reaction) proves that CLEC1B gene expression abnormality is related to initiation and development of the bile duct carcinoma. According to a research result, CLEC1B can also be used for preparing medicines for treating the bile duct carcinoma. By the application of the CLEC1B genes on diagnosis and treatment of the bile duct carcinoma, sensitivity and specificity of diagnosis of the bile duct carcinoma are greatly improved, and a new target spot is provided for gene therapy of the bile duct carcinoma.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to the application of CLEC1B gene in the diagnosis and treatment of cholangiocarcinoma. Background technique [0002] Cholangiocarcinoma refers to malignant tumors that occur in the lining epithelium of the biliary system, and can be divided into two categories: intrahepatic cholangiocarcinoma and extrahepatic cholangiocarcinoma according to the site of occurrence. Intrahepatic cholangiocarcinoma originates from the lining epithelium of any part of the intrahepatic bile duct and its branches to the thin interlobular biliary tree; extrahepatic cholangiocarcinoma is divided into hilar cholangiocarcinoma and distal terminal cholangiocarcinoma. Worldwide, cholangiocarcinoma accounts for 3% of all gastrointestinal tumors and is the second most common hepatobiliary tumor. Due to its characteristics of late discovery, early metastasis, poor prognosis, low surgical resection rate, and insensi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N33/68A61K45/00A61K48/00A61K38/17A61P35/00
CPCA61K38/17A61K45/00A61K48/00C12Q1/6886C12Q2600/158G01N33/57407
Inventor 高倩倩杨承刚
Owner BEIJING MEDINTELL BIOMED CO LTD
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