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Application of siRNA sequence in preparation of medicine for treating ovarian cancer

A DNA sequence, ovarian cancer technology, applied in the direction of retroRNA virus, DNA / RNA fragments, applications, etc.

Pending Publication Date: 2021-05-25
上海兰天生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, whether LIM domain 2 protein (LIMD2) is related to ovarian cancer has not been reported in the prior art.

Method used

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  • Application of siRNA sequence in preparation of medicine for treating ovarian cancer
  • Application of siRNA sequence in preparation of medicine for treating ovarian cancer
  • Application of siRNA sequence in preparation of medicine for treating ovarian cancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] This embodiment discloses a method for preparing a recombinant vector, comprising:

[0029] Firstly, the DNA sequence group (forward sequence: 5'-GACCCACCCAACTTACCATA-3'(SEQ ID NO: 1); reverse sequence: 5'- AATTCTCCGAACGTGTCAC-3' (SEQ ID NO: 2)) was cloned into the lentiviral vector pLVshRNA-EGFP(2A) Puro vector to obtain pSLenti-U6-shRNA(LIMD2)-CMV-EGFP-2A-Puro vector. Then, use the helper plasmid to package the lentivirus. Next, use the packaged lentivirus to infect ovarian cancer cells A2780, and 24 hours later, use DMEM culture medium containing 5 μg / ml of puromycin to screen for 48 hours, and the obtained ovarian cancer cells after transfection are subjected to cell migration and subcutaneous Tumor formation experiments.

[0030] The schematic diagram of the pLVshRNA-EGFP (2A) Puro vector and the schematic diagram of the pSLenti-U6-shRNA (LIMD2)-CMV-EGFP-2A-Puro vector are respectively as follows figure 1 and figure 2 shown.

Embodiment 2

[0032] Cell migration is an important indicator of tumor metastasis, and Transwell can be used to study the motility of tumor cells.

[0033] There is a permeable polycarbonate membrane on the bottom of the Transwell chamber, the upper chamber is a serum-free cell suspension, and the lower chamber is a serum-containing medium. Certain components in the serum, such as some cytokines, are essential for cell growth , so the cells have tropism for these factors, and the cell tropism is used to promote the transfer of cells to the lower chamber, and different cells will show different motility transfer capabilities here.

[0034] The transwell chamber in the Corning transfer assay kit is composed of a 24-well tissue culture plate and a 12-well cell culture insert. The insert contains a polycarbonate membrane with a pore size of 8 μm. The transferred cells migrate and adhere to the bottom of the polycarbonate membrane.

[0035] This example studies the effect of siRNA on the migrati...

Embodiment 3

[0044] This embodiment relates to nude mouse tumor formation experiments, specifically comprising the following steps:

[0045] 6-week-old female mice were subcutaneously injected with 200 microliters (3×10 6) cell mixture (100 microliters cell PBS+100 microliters Matrigel). Tumor size was measured every 3 days. Tumor volume was calculated according to the following formula: V=a×b 2 / 2. After 3 weeks, the nude mice were euthanized by intraperitoneal injection of 2% pentobarbital sodium (50 mg / kg), and the tumor body was weighed. Part of the tissue was fixed with 10% formaldehyde for subsequent histological testing; the remaining tissue was quickly frozen in liquid nitrogen and stored in a -70°C refrigerator for subsequent molecular biological testing. The injected cell density was 3 x 10 7 / ml.

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Abstract

The invention belongs to the field of biology, and particularly relates to application of a siRNA sequence in preparation of a medicine for treating ovarian cancer. A DNA sequence group for coding the siRNA sequence comprises a forward sequence of which the nucleotide sequence is shown as SEQ ID NO: 1 and a reverse sequence of which the nucleotide sequence is shown as SEQ ID NO: 2. The siRNA can be used for preparing medicines for treating ovarian cancer in the future, and brings good news to ovarian cancer patients.

Description

technical field [0001] The invention belongs to the field of biology, and in particular relates to the application of a siRNA sequence in the preparation of drugs for treating ovarian cancer. [0002] technical background [0003] Due to the lack of early symptoms and signs, most patients with ovarian cancer (OC) are already in the middle and advanced stages when they are discovered, and have the characteristics of high recurrence rate, high mortality rate, and low survival rate, and the 5-year survival rate is only 20% to 25%. %. According to statistics in 2018, there were about 295,414 new cases of ovarian cancer and 184,799 deaths worldwide, posing a huge threat to women's lives and health. The age of menarche, spontaneous abortion, induced abortion, menstrual cycle, dysmenorrhea, and number of pregnancies are risk factors for ovarian cancer, among which induced abortion, spontaneous abortion, and menstrual cycle are the main risk factors. The combination of cytoreductiv...

Claims

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Application Information

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IPC IPC(8): A61K31/713A61P15/08A61P35/00C12N15/867C12N15/113
CPCA61K31/713A61P15/08A61P35/00C12N15/86C12N15/1135C12N2740/15043C12N2310/14C12N2320/30
Inventor 陈里新
Owner 上海兰天生物医药科技有限公司
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