ERBB signal pathway mutation targeted sequencing method for prognosis evaluation of gallbladder carcinoma

A technology for targeted sequencing and signaling pathways, applied in the field of molecular biology

Inactive Publication Date: 2014-09-10
XIN HUA HOSPITAL AFFILIATED TO SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no method for prognostic assessment through targeted sequencing of mutations in the ERBB signaling pathway

Method used

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  • ERBB signal pathway mutation targeted sequencing method for prognosis evaluation of gallbladder carcinoma
  • ERBB signal pathway mutation targeted sequencing method for prognosis evaluation of gallbladder carcinoma
  • ERBB signal pathway mutation targeted sequencing method for prognosis evaluation of gallbladder carcinoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1 Targeted sequencing method for ERBB signaling pathway mutations

[0031] 1. Collect tumor tissue samples from the subject.

[0032] 2. Genomic DNA extraction

[0033] Thaw the tissue block, take an appropriate amount of tissue, wash it in PBS solution, put it into a centrifuge tube (1.5ml) filled with 500μl PBS, cut it with scissors; centrifuge at 12000rpm for 8min, take out and discard the supernatant. Gallbladder cancer cells were first processed into cell suspension, then centrifuged at 10,000 rpm for 1 min, the supernatant was poured out, 200 μl buffer GA was added, and shaken until completely suspended.

[0034] Add 0.45ml TES and mix well, then add 50μl SDS (10%), 20μl proteinase K (20mg / ml), mix thoroughly, incubate at 56°C for 4-6h, shake once every 2h until the system is clear.

[0035] Place it at room temperature, add an equal volume of saturated phenol (500 μl), gently turn it up and down for 5 minutes, centrifuge at 12,000 r / m for 10 minutes, se...

Embodiment 2

[0049] Example 2 ERBB signaling pathway gene mutation can be used as an independent prognostic factor for gallbladder cancer

[0050] 1. Experimental method

[0051] Our research group used the genome-wide exome technology as described in Example 1 to perform targeted sequencing on 57 pairs of gallbladder cancer paired tissues to obtain the frequency of gene mutations, and then conduct pathway analysis for high-frequency mutation genes through the KEGG database, and then according to the patient Gender, age, degree of differentiation of gallbladder cancer, TNM stage, lymphatic metastasis, nerve invasion, lesion location, presence or absence of jaundice, condition of resection margin, ERBB signaling pathway gene mutation, and presence or absence of gallbladder stones were grouped, and the ERBB signal among the groups was compared. Pathway gene mutations.

[0052] SPSS 18.0 was used for data analysis, and count data were analyzed by χ 2 Test (Fisher's exact probability method ...

Embodiment 3

[0069] Example 3 Clinical application of ERBB signaling pathway gene mutation as an independent prognostic factor for gallbladder cancer

[0070] The method described in Example 1 was used to predict the prognosis of a patient with gallbladder cancer, and it was found that the ERBB3 gene of this subject had a mutation, and the mutation site was V104L, suggesting that the prognosis of this patient with gallbladder cancer was poor. According to statistics, the actual survival period of the patient was 7.5 months.

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Abstract

The invention relates to an ERBB signal pathway mutation targeted sequencing method for the prognosis evaluation of gallbladder carcinoma and provides an application of an ERBB signal pathway mutation targeted sequencing reagent / kit for gallbladder carcinoma patients. In the invention, the whole-genome exon technology is adopted for performing mutation target sequencing of the ERBB signal pathway of a gallbladder carcinoma patient; and in combination with the pathological clinical data of the gallbladder carcinoma patient, the Kaplan-Meier survival curve analysis indicates that the lifetime of the patient after operation is closely related to the gene mutation of the ERBB signal pathway, and the multiple-factor analysis of Cox risk regression model indicates that the gene mutation of the ERBB signal pathway can serve as an independent prognosis factor of gallbladder carcinoma. The prompt of screening the ERBB signal pathway related gene mutation of the gallbladder carcinoma cancer is favorable to the gene-level diagnosis and parting of gallbladder carcinoma, the establishment of therapeutic target, the prognosis analysis and the like.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to a targeted sequencing method for ERBB signaling pathway mutations used for gallbladder cancer prognosis assessment. Background technique [0002] Gallbladder cancer is the most common malignant tumor of the biliary system. It is difficult to diagnose early and has a high degree of malignancy. Infiltration and metastasis of the liver and surrounding organs can occur in the early stage of the disease. After diagnosis, the resection rate is only 15% to 47%. The effect of radiotherapy and chemotherapy is inaccurate, the overall 5-year survival rate is only 5%, and only about 16.5% after radical resection, the prognosis is extremely poor. The occurrence of gallbladder cancer is a multi-gene, multi-step pathological process, including a series of changes in genetics. More and more evidence shows that the activation of oncogenes and inactivation of tumor suppressor genes caus...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6874C12Q2531/113C12Q2543/101C12Q2565/501
Inventor 刘颖斌刘赟王慧李茂岚张周李晓光
Owner XIN HUA HOSPITAL AFFILIATED TO SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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