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Human miR-485-5p antisense nucleic acid and application thereof

An antisense and nucleotide technology, applied in the application field of antisense oligonucleotides, can solve the problems of poor curative effect, low survival rate and poor curative effect in patients with distant metastasis, and achieve growth inhibition and malignant The effect of proliferative capacity

Active Publication Date: 2011-04-27
SUZHOU GENEPHARMA +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] In the past 30 years, although the comprehensive treatment of tumors has been very common clinically, the comprehensive treatment based on surgery and supplemented by radiotherapy and chemotherapy has not significantly improved the survival rate of cancer patients, and the 5-year overall survival rate is still low, hovering At about 30% to 55%, there is no significant improvement, and the 5-year survival rate of middle and advanced patients is even lower, about 20%.
Moreover, these methods have their own limitations, especially for the middle-advanced and relapsed patients, and the curative effect is even worse for those with distant metastasis.

Method used

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  • Human miR-485-5p antisense nucleic acid and application thereof
  • Human miR-485-5p antisense nucleic acid and application thereof
  • Human miR-485-5p antisense nucleic acid and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1, miR-485-5p antisense nucleic acid inhibits the expression of miR485-5p

[0030] The real-time quantitative fluorescence detection test was completed by Shanghai Gemma Pharmaceutical Technology Co., Ltd. The specific experimental steps include:

[0031] Cell culture: U87 cells, cultured in 10% FBS-DMEM medium, 37°C, 5% CO2.

[0032] Cell transfection:

[0033] 1) One day before transfection, inoculate cultured cells in a 24-well plate with an appropriate amount of culture medium without antibiotics, so that the confluence of cells at the time of transfection reaches 30-50%;

[0034] 2) Transfection samples were prepared as follows to prepare oligomers-Lipofecta mine TM 2000 complex: a. Use 50 μl serum-free Opti- Dilute the miR-485-5p inhibitor, negative control, and FAM-labeled negative control in I culture medium to a final concentration of 50 nM, mix gently, and set 3 replicate wells for each transfection;

[0035] b. Gently mix Lipofecta mine before us...

Embodiment 2

[0064] Example 2, Detection of inhibitory activity of MiRNA antisense oligonucleotides on human glioblastoma cell line U87

[0065] Cell culture:

[0066] U87 cells were cultured in 10% FBS-DMEM medium, 37°C, 5% CO2. Collect U87 cells in good growth state, count by centrifugation, and use 2×10 3 Spread each well in a 96-well plate and incubate at 37°C with 5% CO2.

[0067] Transfection:

[0068] 1) One day before transfection, inoculate cultured cells in a 96-well plate with an appropriate amount of culture medium without antibiotics, so that the confluence of cells at the time of transfection reaches 30-50%;

[0069] 2) Transfection samples were prepared as follows to prepare oligomers-Lipofecta mine TM 2000 complex:

[0070] a. Use 25 μl serum-free Opti- Dilute the miR-485-5p antisense oligonucleotide, negative control, and FAM-labeled negative control in I culture medium respectively to a final concentration of 50 nM, mix gently, and set 3 duplicate holes for each tr...

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Abstract

The invention discloses an antisense oligonucleotide used for inhibiting the expression of micro RNA-485-5p, and an application thereof. The antisense oligonucleotide comprises the sequence of 5'-GAAUUCAUCACGGCCAGCCUCU-3' which can be specifically combined with human miR-485-5p. The antisense oligonucleotide can be ribonucleotide, deoxyribonucleotide or chimera of the ribonucleotide and the deoxyribonucleotide, and can be used for modifying any nucleotide in the chain. The miR-485-5p antisense oligonucleotide can be used for effectively inhibiting the expression of miR-485-5p in a human brainglioma cell U87 as well as the growth and propagation of the human brain glioma cell U87, thus effectively treating the brain glioma and other tumours with high expression of miR-485-5p.

Description

technical field [0001] The invention relates to the field of biomedicine. Specifically, the present invention relates to the use of a small molecule microRNA (miRNA), especially the application of antisense oligonucleotides related to the miRNA. The antisense oligonucleotide can be complementary to human miR-485-5p, thereby inhibiting the expression of human miR-485-5p. The present invention also relates to a pharmaceutical composition containing the miRNA antisense oligonucleotide. Background technique [0002] miRNAs are small non-coding RNAs with a length of 20-22bp, usually transcribed by RNA polymerase II (PolII), and generally the initial product is a large one with a cap structure (7MGpppG) and a polyA tail (AAAAA) pri-miRNA. These pri-miRNAs are processed into pre-miRNA precursor products consisting of 70 nucleotides under the action of RNase III Drosha and its cofactor Pasha. RAN-GTP and exportin 5 transport this precursor molecule into the cytoplasm. Subsequen...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11A61K48/00A61P35/00
Inventor 丁侃张佩琢东楠李捷沈孝坤
Owner SUZHOU GENEPHARMA
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