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Application of FHL 1 in preparing medicament for treating tumour

A technology of FHL1 and uses, which is applied in the field of use of FHL1 in the preparation of drugs for the treatment of tumors, and can solve the problems of unclear molecular mechanism and the like

Active Publication Date: 2010-03-31
INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although it has been reported that the expression level of FHL1 is decreased in a variety of human tumors, whether FHL1 can inhibit the growth of human tumor cells and the molecular mechanism of how to regulate this growth is not clear.

Method used

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  • Application of FHL 1 in preparing medicament for treating tumour
  • Application of FHL 1 in preparing medicament for treating tumour
  • Application of FHL 1 in preparing medicament for treating tumour

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Embodiment 1, FHL1 mammalian cell expression vector and FHL1 recombinant adenovirus vector

[0048] (1) Method

[0049] 1. Construction of FHL1 mammalian cell expression vector

[0050] The complete coding region sequence of the FHL1 gene was amplified by PCR from a mammary gland cDNA library (purchased from Clontech, USA) according to conventional methods. : After 5 minutes at 94°C, 30 cycles of 94°C for 1 minute, 60°C for 1 minute, 72°C for 1 minute, and finally 72°C for 7 minutes. Kpn I and Xba I enzymes (purchased from Biolabs, UK) were used to digest the PCR products and corresponding mammalian cell expression vectors (pcDNA3, purchased from Invitrogen, USA) at 37°C, and the PCR products and vectors were digested with T4 DNA ligase (purchased from Promega, USA) was ligated at 12-16°C to obtain a recombinant vector (which contains the CMV promoter / enhancer, the complete coding region sequence of the FHL1 gene, the BGH polyA termination signal, the ampicillin...

Embodiment 2

[0082] Example 2, FHL1 inhibits the growth of various tumor cells cultured in vitro

[0083] (1) Method

[0084]The effect of FHL1 on the growth of various tumor cells was determined by crystal violet assay, and the steps were as follows: Digest various tumor cells, including breast cancer cell lines (MCF7, ZR75-1, MDA-MB-468, MDA-MB231), liver cancer cell lines (HepG2, Hep3B, SMMC7721), lung cancer cell lines (A549, Calu3, H460), gastric cancer cell lines (BGC823, MKN-1). After counting the cells, dilute to an appropriate concentration, put 0.5ml of the cell solution into a 24-well plate, so that the number of cells in each well is about 10,000. Transfect these tumor cells with the FHL1 mammalian cell expression vector according to the method described in Example 1 or infect these tumor cells with 20-100 MOI of FHL1 recombinant adenovirus, after cultivating for 1-4 days, remove the medium, and add 0.5 ml to each well 1% glutaraldehyde (fixative solution), stand still fo...

Embodiment 3

[0087] Example 3, FHL1 inhibits anchorage-independent growth of various tumor cells

[0088] (1) Method

[0089] The effect of FHL1 on the anchorage-independent growth of tumor cells was determined by soft agar assay. First, the FHL1 mammalian cell expression vector was transfected into tumor cells according to the method described in Example 1 or the tumor cells were infected with FHL1 recombinant adenovirus at 20-100 MOI, including breast cancer cell lines (MCF7, ZR75-1, MDA-MB -468, MDA-MB231), liver cancer cell lines (HepG2, Hep3B, SMMC7721), lung cancer cell lines (A549, Calu3, H460), gastric cancer cell lines (BGC823, MKN-1), and then use 60mm 2 Cell culture dish, add 3ml DMEM medium containing 0.7% agar (purchased from U.S. Invitrogen Company) in the culture dish, add 3ml DMEM medium containing above-mentioned 20,000 tumor cells and 0.25% agar on it after solidification, each group The cells were set up in 3 parallel samples, and the number of colonies formed by c...

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Abstract

The invention relates to the application of FHL 1 in preparing medicament for treating tumour, in particular to the application of the FHL 1 in preparing the medicament for regulating the expression of cancer genes and / or cancer suppressor genes in cells of mammals and the application of the FHL 1 in preparing the medicament for restraining the growth of tumour cells of the mammals. The inventionalso relates to an expression carrier containing a nucleotide sequence for coding the FHL 1, the application of the expression carrier in preparing the medicament for regulating the expression of thecancer genes and / or the cancer suppressor genes in the cells of the mammals and the application of the expression carrier in preparing the medicament for restraining the growth of the tumour cells ofthe mammals. The expression carrier can effectively suppress the growth of multiple human tumour cells and restrain the growth of the human tumour cells by regulating the expression of the cancer genes and the cancer suppressor genes.

Description

technical field [0001] The present invention relates to a new medical application of four and a half LIM domain proteins, in particular to the application of four and a half LIM domain proteins 1 (FHL1) in the preparation of medicines for treating tumors. Background technique [0002] The four and a half LIM domains protein (Four and a Half LIM domains protein, hereinafter referred to as FHL) family belongs to the LIM superfamily, and its family members are composed of four and a half LIM domains. The LIM domain consists of approximately 50 amino acids, is rich in cysteine, and forms 2 zinc finger structures. The LIM domain mediates many protein-protein interactions and regulates the actions of cytoskeletal proteins, enzymes, and transcription factors. Five members of the FHL family have been found, which are FHL1, FHL2, FHL3, FHL4 and FHL5 / ACT. The expression of FHL members is tissue specific, FHL1, FHL2 and FHL3 are mainly expressed in skeletal muscle and cardiac muscle,...

Claims

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Application Information

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IPC IPC(8): A61K38/17A61K48/00A61P35/00C12N15/12C12N15/86C12N15/861
Inventor 叶棋浓丁丽华杨晓秦玺程龙牛畅黄翠芬
Owner INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE
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