Application of miR-130a to diagnosis, treatment and prognosis of ovarian cancer

A technology for ovarian cancer and prognosis evaluation, applied in the field of molecular biology, can solve the problems of lack of clinical practical early diagnosis methods and treatment strategies, unclear understanding of the molecular mechanism of ovarian cancer, etc., achieving strong targeting, promoting proliferation and invasion, inhibiting The effect of autophagy

Active Publication Date: 2017-05-10
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The fundamental reason lies in the unclear understanding of the molecular mechanism of the occurrence and development of ovarian cancer, the lack of clinical and practical early diagnosis methods and effective treatment strategies

Method used

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  • Application of miR-130a to diagnosis, treatment and prognosis of ovarian cancer
  • Application of miR-130a to diagnosis, treatment and prognosis of ovarian cancer
  • Application of miR-130a to diagnosis, treatment and prognosis of ovarian cancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Example 1 Defining the role of miR-130a and TSC1 in ovarian cancer

[0046] Ovarian cancer tissues and control normal fimbria tissues were collected (both tissues were from Qilu Hospital of Shandong University, 2009-2015), and ground on ice with a tissue grinder, added protein lysate, lysed on ice for 20 minutes, and centrifuged at 12000r / min at 4°C for 15 minutes before taking In the supernatant, the protein concentration was measured by BCA method, and western blot and immunohistochemical staining were performed to detect the difference in the content of TSC1 in ovarian cancer tissues and fallopian tube fimbria. The results showed that TSC1 was low expressed in most ovarian cancer tissues ( figure 1 a), grayscale analysis shows that compared with normal fimbria tissue, the content of TSC1 in ovarian cancer tissue is lower, and there is a significant statistical difference between the two (pfigure 1 b) We then performed immunohistochemical staining on tissue chips cont...

Embodiment 2

[0047] Example 2 Determining the Targeted Regulatory Effect of Regulating miR-130a on TSC1

[0048] Expression constructs of miRNA and 3′-UTR:

[0049] miR-130a-3p, miR-27a-3p and miR-204-3p mimics and negative controls were purchased from Gemma Genomics. miR-130a-inhibitor (referring to the sequence 5'-AUGCCCUUUUAACAUUGCACUG-3', shown in SEQ ID NO: 18) and a negative control. Transiently transfected ovarian cancer cell lines, it was found that only after miR-130a was transfected, the TSC1 content decreased ( figure 2 a) At the same time, we found that the phosphorylation level of key proteins in the mTOR signaling pathway was significantly increased after miR-130a was overexpressed, indicating that miR-130a activated the mTOR signaling pathway, whereas the mTOR signaling pathway was inhibited after miR-130a was underexpressed ( figure 2 b) That is, miR-130a is positively correlated with the activation degree of mTOR signaling pathway. In order to further verify that TSC1...

Embodiment 3

[0050] Example 3 Construction of miR-130a overexpressed cell lines.

[0051] The sequence of miR-130a was ligated to the pGIPZ plasmid (commercially available), and after lentivirus packaging, it was transfected into A2780 and SKOV3 cells, and was screened with puromycin. After screening, the contents of miR-130a and TSC1 were detected by western blot and qPCR. After the cell line was successfully established, the proliferation ability of the cells was detected by the plate cloning experiment and the soft agar colony formation experiment, and the invasion ability of the cells was detected by the Transwell experiment.

[0052] 1. Construction of stable cell lines:

[0053] (1) Production of virus particles

[0054] Take Phoenix amphotropic cells in good condition, trypsinize to collect cells and count them according to 3×10 6 The density of each cell / dish is inoculated in a 100mm cell culture dish. The cell culture medium before transfection cannot contain antibiotics, and i...

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Abstract

The invention discloses application of miR-130a to diagnosis, treatment and prognosis of ovarian cancer. The miR-130a is defined, through in-vivo and in-vitro function test, that an mTOR (mammalian Target of Rapamycin) signal path is over-expressed through inhibiting translation of TSC1mRNA (Tuberous Sclerosis 1 Messenger RNA (Ribonucleic Acid)) by the miR-130a, so that proliferation and invasion of ovarian cancer cells are promoted and autophagy of the cells is inhibited. The data testifies that the miR-130a with a reduced endogenous property can be used for inhibiting malignant biological behaviors including growth, invasion and the like of ovarian cancer tumors. Based on the characteristics, a specific miR-130a inhibitor can be designed, the content of TSC1 in a body is recovered and the mTOR signal path is inhibited, so that the aim of treating the ovarian cancer is realized.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to an application of miR-130a in the diagnosis, treatment and prognosis of ovarian cancer. Background technique [0002] Ovarian cancer is a common malignant tumor in gynecology, and its mortality rate has always been the first among gynecological malignant tumors. Although tumor cytoreductive surgery and platinum-taxane combined chemotherapy have improved the 5-year survival rate of patients, there has been no breakthrough in the clinical diagnosis and treatment of ovarian cancer for half a century. The fundamental reason lies in the unclear understanding of the molecular mechanism of the occurrence and development of ovarian cancer, the lack of clinical and practical early diagnosis methods and effective treatment strategies. Therefore, elucidating the molecular mechanism of ovarian cancer development and screening molecular markers related to tumor development and metastasis are...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68A61K45/00A61P35/00
CPCA61K45/00C12Q1/6886C12Q2600/118C12Q2600/156C12Q2600/178
Inventor 刘招舰王玉琼张锡宇
Owner SHANDONG UNIV
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