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Sialyltransferase recombinant plasmid and application of sialyltransferase recombinant plasmid and pemetrexed the preparation of medicament for inhibiting proliferation and invasion of bladder cancer

A technology of sialyl and pemetrexed, applied in medicine, in the field of plasmids overexpressing sialyltransferase ST3GAL4 gene, can solve the problem that the effect of pemetrexed is less researched and other problems, and achieve inhibition of proliferation and invasion , the effect of good application prospects

Inactive Publication Date: 2019-03-29
DALIAN MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, the role of pemetrexed in bladder cancer is still little research

Method used

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  • Sialyltransferase recombinant plasmid and application of sialyltransferase recombinant plasmid and pemetrexed the preparation of medicament for inhibiting proliferation and invasion of bladder cancer
  • Sialyltransferase recombinant plasmid and application of sialyltransferase recombinant plasmid and pemetrexed the preparation of medicament for inhibiting proliferation and invasion of bladder cancer
  • Sialyltransferase recombinant plasmid and application of sialyltransferase recombinant plasmid and pemetrexed the preparation of medicament for inhibiting proliferation and invasion of bladder cancer

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Experimental program
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Effect test

Embodiment 1

[0032] The present invention entrusts a company to construct an overexpression vector of ST3Gal4. The Gene ID of ST3Gal4 is 6484 (BC010645), the species is human, the insertion size is 1002bp, the pcDNA3.1(-) vector is used, the size is 5428bp, and the tag used is HA , the endonuclease used is EcoR I / Hind III. Such as figure 1 a.

[0033] The nucleotide sequence of the ST3Gal4 insertion sequence is shown in SEQ ID No.1; the nucleotide sequence of pcDNA3.1(-) is shown in SEQ ID No.2.

[0034] Bladder cancer cells were overexpressed ST3Gal4 plasmid by lipofection technology. The positive clone of the ST3Gal4 recombinant expression vector was identified by EcoRI / Hind III double digestion and DNA sequencing. The sequencing reaction was provided and completed by Nanjing Biotech Co., Ltd. The sequencing results were in full agreement with the expected results.

[0035] ST4Gal4 sequence:

[0036] Sense strand: 5'-TGAATCTGCCCACTTCGACC-3'

[0037] Antisense strand: 5'-TTCGCACCCGCT...

Embodiment 2

[0040] 1. Cell culture

[0041] (1) Cell Recovery

[0042] The frozen cells were taken out from the liquid nitrogen, placed in PE gloves, shaken in a 37°C water bath until they melted, then quickly transferred to a 5mL centrifuge tube, added 3mL of medium, and centrifuged at 1000rpm for 4min. After resuspending the cell pellet with pre-warmed medium, transfer it to a cell culture flask and culture it in an incubator.

[0043] (2) Cell replacement and passage

[0044] The human bladder urothelial cell line (SV-HUC-1) was cultured in F12 / DMEM medium containing 10% fetal bovine serum and double antibodies (penicillin 100 U / mL, streptomycin 100 μg / mL). Human bladder cancer cell line (RT4) was cultured in MCCOY'S5A medium containing 10% fetal bovine serum and double antibodies (penicillin 100 U / mL, streptomycin 100 μg / mL). Human bladder cancer cell line (T24, 5637) was cultured in 1640 medium containing 10% fetal bovine serum and double antibodies (penicillin 100 U / mL, streptomy...

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Abstract

The invention discloses a sialyltransferase recombinant plasmid and application of the sialyltransferase recombinant plasmid and pemetrexed in the preparation of a medicament for inhibiting proliferation and invasion of bladder cancer. The growth and proliferation of bladder cancer cells can be effectively inhibited by constructing a plasmid overexpressing a sialyltransferase ST3Gal4 gene, and theplasmid plays a role in the growth and proliferation of bladder cancer cells by mediating a PI3K-AKT signaling pathway. Further, after cell transfection and with the combined effect with pemetrexed,the recombinant plasmid can further inhibit the ability of bladder cells to proliferate and migrate. The sialyltransferase ST3Gal4 can be used as an indicator of bladder cancer disease. Therefore, thetechnology of the present invention provides a good application prospect in preparing a medicament for inhibiting proliferation and invasion of bladder cancer, and provides a new target for early diagnosis and treatment of bladder cancer.

Description

technical field [0001] The invention belongs to the technical field of biopharmaceuticals, and in particular relates to a plasmid overexpressing the gene of sialic acid acyltransferase ST3GAL4 and its application in combination with pemetrexed to inhibit the proliferation and invasion of bladder cancer. Background technique [0002] At present, malignant tumor is one of the only diseases that threaten human health and life. In my country, the morbidity and mortality of bladder cancer are the highest among urinary system tumors, and the incidence has been increasing in recent years. [0003] Patients with bladder cancer have a high recurrence rate after surgical resection, and the use of BCG infusion therapy will have great adverse reactions, which is extremely harmful to the body. Due to the poor therapeutic effects of traditional surgery, radiotherapy, chemotherapy and other means, and side effects, surgery combined with drug treatment emerged as the times require, and ear...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K38/45A61K31/519A61P35/00A61P13/10
CPCA61K31/519A61K38/45A61K48/005A61P13/10A61P35/00C12Y203/01A61K2300/00
Inventor 汪淑晶张嘉宁杨德勇王施丹张晗
Owner DALIAN MEDICAL UNIVERSITY
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