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Use of miR-9 inhibitor in preparation of tumor growth inhibition drugs

A technology of 1.mir-9 and 2.mir-9, applied in the field of application of miR-9 inhibitors in the preparation of drugs for inhibiting tumor growth, can solve problems such as no reports

Inactive Publication Date: 2014-07-30
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the regulation of miR-9 on some non-tumor cells, such as tumor-associated immune cells, has not been reported yet

Method used

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  • Use of miR-9 inhibitor in preparation of tumor growth inhibition drugs
  • Use of miR-9 inhibitor in preparation of tumor growth inhibition drugs
  • Use of miR-9 inhibitor in preparation of tumor growth inhibition drugs

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Regulation of miR-9 on M-MDSC / G-MDSC cell function

[0021] (1) Establishment of tumor model

[0022] A 6-8 week-old C57BL / 6 mouse with a body weight of about 20 g was taken. On the ultra-clean experimental workbench, sterilize the skin of the right armpit of the mouse, shake the cell suspension, and inject the mouse Lewis lung cancer cells subcutaneously, 200 μl each, totaling 5.0×10 6 cells. Three weeks after the inoculation, a lump appeared under the mouse skin, which was hard and poor in mobility, which was the subcutaneous transplanted tumor.

[0023] (2) Separation and purification of MDSCs cells

[0024] Spleen of tumor-bearing mice was aseptically collected, separated and purified using the "Myeloid-Derived Suppressor Cell Isolation Kit" from Miltenyi Biotec, G-MDSC cells were obtained by positive selection with anti-Ly6G antibody, and the negative cells were positively selected by anti-Gr-1 antibody Select to obtain M-MDSC cells. FCM was used to detect cel...

Embodiment 2

[0054] The effect of miR-9 on tumorigenesis and development

[0055] (1) Take C57BL / 6 mice aged 6-8 weeks, and subcutaneously inject mouse Lewis lung cancer cells, 200 μl each, totaling 3.0×10 6 cells. 10 days after implantation of tumor cells, MDSC cells transfected with miR-9 mimics or inhibitors were injected into the local tumor of tumor-bearing mice, 2.0×10 6 cells / only. Cells were injected one week later, and the tumor size was measured every 2 days with the formula volume=0.5×length×width 2 to calculate the tumor volume. At the same time, the survival rate of mice in different treatment groups was observed.

[0056] (2) Take C57BL / 6 mice aged 6-8 weeks and subcutaneously inject mouse Lewis lung cancer cells, 200 μl each, totaling 3.0×10 6 cells. Ten days after implantation of tumor cells, miR-9 mimics or inhibitors were directly injected into the tumor of tumor-bearing mice, and the tumor size and survival rate of the mice were observed.

[0057] like image 3 A...

Embodiment 3

[0059] Expression of miR-9 and MDSC-related factors in patients with lung cancer

[0060] (1) Soybean-sized lung cancer tissue and adjacent normal lung tissue were collected, soaked in 10% formaldehyde for fixation, embedded in paraffin, sectioned, and immunofluorescent stained with FITC-labeled anti-CD11b antibody and APC-labeled anti-CD33 antibody , detect CD11b + CD33 + Number of MDSC cells.

[0061] (2) Grinding tumor tissue and adjacent normal lung tissue, adding Trizol to extract tissue RNA, using specific hsa-miR-9 reverse transcription primers for reverse transcription to obtain cDNA, and then detecting miR in the tissue by real-time fluorescent quantitative PCR -9 expressions.

[0062] hsa-miR-9 specific reverse transcription primer, the sequence is as follows:

[0063] 5'-GTCGTATCCAGTGCGTGTCGTGGAGTCGGCAATTGCACTGGATACGACTCATACA-3' (SEQ ID.5)

[0064] The 10μl reverse transcription system is as follows:

[0065]

[0066] The miRNA reverse transcription procedu...

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Abstract

The present invention discloses a use of a miR-9 inhibitor in preparation of tumor growth inhibition drugs. According to the present invention, the modern biology technology is adopted, and it is found that the miR-9 has the use of regulation of MDSC cells, wherein immunosuppression capability of MDSC cells can be down-regulated by inhibiting miR-9 expression in MDSC cells so as to inhibit tumor growth, such that the miR-9 can be adopted as a target for tumor treatment so as to provide potential application values and practical significance for tumor immunotherapy.

Description

technical field [0001] The invention relates to a new application of miR-9, and the application of miR-9 inhibitors in the preparation of drugs for inhibiting tumor growth belongs to the field of biotechnology. Background technique [0002] MicroRNAs (miRNAs) are non-coding, endogenous small molecule RNAs with a length of about 20-23 nucleotides. miRNA was first discovered in C. elegans in 1993. After years of research, the latest miRNA database shows that more than 20,000 different miRNAs have been found to exist in various organisms. miRNAs can specifically bind to the complementary sequence of the 3' untranslated region (3'UTR) of target gene mRNAs, thereby preventing protein synthesis. Each miRNA can have multiple target genes, and several miRNAs can also regulate the same gene. This complex regulatory network can not only regulate the expression of multiple genes through one miRNA, but also finely regulate the expression of a certain gene through the combination of se...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K45/00A61P35/00
Inventor 王胜军田洁芮棵马洁马斌汤新逸张悦许化溪
Owner JIANGSU UNIV
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