Application of ythdf2 gene in diagnosis, prevention and treatment of urothelial carcinoma

A urothelial cancer, gene technology, applied in the field of biomedicine, to achieve the effect of promoting proliferation and migration

Active Publication Date: 2022-03-22
SHENZHEN LUOHU PEOPLELS HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It has been reported that YTHDF2 is involved in the occurrence of various malignant diseases such as pancreatic cancer, gastric cancer, and liver cancer, but there is no report on whether it is involved in the occurrence and development of urothelial carcinoma

Method used

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  • Application of ythdf2 gene in diagnosis, prevention and treatment of urothelial carcinoma
  • Application of ythdf2 gene in diagnosis, prevention and treatment of urothelial carcinoma
  • Application of ythdf2 gene in diagnosis, prevention and treatment of urothelial carcinoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1, RT-qPCR detection of differential expression of YTHDF2 gene in urothelial carcinoma tissue

[0024] A total of 39 urothelial carcinoma tissues and corresponding paracancerous tissues were collected. The informed consent of the patients was obtained for the tissue samples and the approval of the organizational ethics committee was obtained. Use Ambion's Trizol reagent to extract RNA samples, and the specific steps are detailed in the instructions. The concentration and purity of the extracted RNA were detected by Nanodrop2000. 1 μg of RNA was used for reverse transcription, and cDNA was obtained by reverse transcription using Vazyme's HiScript Q RT SuperMix. Primers were designed according to the sequences of YTHDF2 gene and RPL13A gene, and the primer sequences were synthesized by Shanghai Sangong. The designed primer sequences are as follows:

[0025] YTHDF2 forward primer: 5'-AGCCCCACTTCCTACCAGATG-3'

[0026] YTHDF2 reverse primer: 5'-TGAGAACTGTTATTTCCC...

Embodiment 2

[0030] Example 2, Differential expression of YTHDF2 gene in human urothelial carcinoma cell lines

[0031] Human urothelial carcinoma cell lines SW780 and UM-UC-3 were cultured in DMEM medium, human urothelial carcinoma cell lines T24 and 5637 were cultured in 1640 medium, and the immortalized bladder tissue cell line Sv-huc-1 was cultured in DMEM / F-12. Culture conditions are 37°C, 5% CO 2 , 90% humidity. The medium was changed in 2 days, the vigorously growing cells were collected, RNA was extracted and reverse-transcribed to obtain cDNA, and RT-qPCR was used to detect the expression of YTHDF2 gene in urothelial carcinoma cell lines. The result is as figure 2 As shown, YTHDF2 gene expression was upregulated in urothelial carcinoma cell lines SW780, UMUC3, T24 and 5637 compared with normal bladder cell lines.

Embodiment 3

[0032] Embodiment 3, the construction of YTHDF2 knockout cell line

[0033] Design sgRNA according to YTHDF2 gene sequence, the designed sgRNA primer sequence is as follows:

[0034] sgRNA1 forward primer: 5'-CACCGTCCATTACTAGTAACATCG-3'

[0035] sgRNA1 reverse primer: 5'-AAACCGATGTTACTAGTAATGGAC-3'

[0036] sgRNA2 forward primer: 5'-CACCGAGTTACTACAGTCCCTCCAT-3'

[0037] sgRNA2 reverse primer: 5'-AAACATGGAGGGACTGTAGTAACTC-3'

[0038] Dissolve the above primers in ddH 2O to a final concentration of 100 μM, add 1 μl of forward and reverse primers to 8 μl ddH 2 In O, annealing was carried out according to the following procedure: 37°C for 30 min, 95°C for 5 min, and cooling down to 25°C at a rate of 0.8°C / s. Take 1 μl of the annealed primer and connect it to the lenticrisprV2 vector. The plasmid carrying YTHDF2 sgRNA was transferred into SW780 and T24 cells by lentivirus, and puromycin was added, and a monoclonal cell line with YTHDF2 knockout was obtained by screening.

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Abstract

The invention discloses the application of YTHDF2 gene in the diagnosis, prevention and treatment of urothelial carcinoma. On the one hand, the present invention discloses the application of a reagent for detecting YTHDF2 gene in preparing products for diagnosing urothelial carcinoma, and on the other hand discloses the application of an inhibitor of YTHDF2 gene expression in preparing a medicament for treating or preventing urothelial carcinoma. The present invention confirms the role of YTHDF2 in urothelial cancer for the first time, YTHDF2 is up-regulated in urothelial cancer tissue, and promotes the proliferation and migration of urothelial cancer cells, YTHDF2 can be used as a clinical diagnosis of urothelial cancer and potential targets for therapy.

Description

technical field [0001] The invention relates to the field of biomedicine, in particular to the application of YTHDF2 gene in the diagnosis, prevention and treatment of urothelial carcinoma. Background technique [0002] Urothelial carcinoma is one of the most common tumors of the urinary system. In my country, urothelial carcinoma ranks first in the incidence of malignant tumors of the male urinary system in China. Higher incidence and postoperative recurrence are the two main characteristics of urothelial carcinoma, but the pathogenesis and recurrence mechanism of urothelial carcinoma is still unclear. Urothelial carcinoma can be divided into non-muscle-invasive urothelial carcinoma (NMIBC) and muscle-invasive urothelial carcinoma (MIBC). 75% of patients were initially diagnosed as NMIBC, and less than 25% of patients were MIBC. At present, the main method for the treatment of urothelial carcinoma is transurethral resection of bladder tumor (TURBT). For low-grade tumors, ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6886
CPCC12Q1/6886C12Q2600/158C12Q2600/118
Inventor 吴松张磊
Owner SHENZHEN LUOHU PEOPLELS HOSPITAL
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