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Human neutralizing antibody to follistatin-like protein and application thereof

A follistatin and protein technology, applied in the direction of antibodies, applications, immunoglobulins, etc., can solve the problems of inability to accurately lock key cytokines, different types of skin fibrosis initiation factors and insufficient understanding of pathological mechanisms, and achieve high application advantage effect

Active Publication Date: 2022-05-10
TIANJIN JIKUN MEDICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The reason why the research on skin fibrosis drugs has failed to make breakthroughs is that the initiating factors and pathological mechanisms of different types of skin fibrosis are not well understood, and the key cytokines involved in the etiology of skin fibrosis cannot be accurately identified

Method used

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  • Human neutralizing antibody to follistatin-like protein and application thereof
  • Human neutralizing antibody to follistatin-like protein and application thereof
  • Human neutralizing antibody to follistatin-like protein and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] Example 1. Activity screening of human FSTL1 antibody

[0067] First, we screened 47 antibodies that bind to human Fstl1 protein from the phage display antibody library, cloned the antibody active sequence (sc-Fv) into the expression vector, transfected 293T cells, and collected the conditioned medium after 2-3 days ( Conditioned medium), which contains a high concentration of scFv, can be directly added to the CAGA-luciferase reporter cell line for luciferase reporter gene detection antibody to Fstl1 neutralizing activity. In the first round of antibody activity screening, we used the conditioned medium obtained by the above method to detect the activity of the anti-Fstl1 human antibody through the TGF-β1 luciferase reporter cell line, and the first round of screening was repeated three times for the luciferase reporter gene detection, screening results ( figure 1 ) shows the increase multiple of luciferase fluorescence intensity in the control group and the experime...

Embodiment 2

[0073] Example 2. Affinity detection of human FSTL1 neutralizing antibodies JK07 and JK04 and affinity comparison with mouse FSTL1 neutralizing antibodies.

[0074] In order to further select more excellent FSTL1 neutralizing antibodies, the two human antibodies JK07 and JK04 with neutralizing activity obtained through activity screening were studied for affinity with the target protein-human FSTL1 protein, using surface plasmon resonance SPR technology The results of the research affinity showed that the affinity of the JK07 antibody to the antigen was 6.17nM, and the affinity of the JK04 antibody to the antigen was 31.17nM; at the same time, the affinity of the human antibody was compared with the mouse FSTL1 neutralizing antibodies 2K6 and 4D22 obtained in previous studies, The affinity of the mouse-derived FSTL1 neutralizing antibody 2K6 to the antigen is 106.8nM, and the affinity of the mouse-derived FSTL1 neutralizing antibody 4D22 to the antigen is 113.1nM. According to ...

Embodiment 3

[0077] Example 3. FSTL1 neutralizing antibody (JK07) can alleviate bleomycin-induced lung fibrosis in mice.

[0078] Male C57BL / 6J (age 8-10 weeks) mice were injected with 2U / kg bleomycin through the trachea, and intraperitoneally injected respectively on the 2nd, 4th, 6th, 8th, 10th, and 12th day of bleomycin treatment 100ug, 200ug, 400ug of FSTL1 neutralizing antibody (JK07) or the corresponding subtype (IgG1), and 200mg / kg of pirfenidone was used as a positive control by intragastric administration 7 days after modeling, and the samples were sacrificed after 14 days of modeling. Detection of pulmonary fibrosis and collagen content.

[0079] Measuring method of dermal thickness: use O8 pathological slide scanner to collect H&E staining image data, use Viewpoint software to measure dermal thickness, the measurement unit is mm, each piece of skin is measured 5 times at different positions, take the average value for statistics, and then calculate the Bole The ratio of the ski...

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PUM

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Abstract

The embodiment of the present invention relates to a human neutralizing antibody of follistatin-like protein and its application, comprising a heavy chain variable region and a light chain variable region, and the heavy chain variable region comprises the following CDRs: amino acid sequence VH CDR1 shown in SEQ ID NO:1, VH CDR2 shown in amino acid sequence as SEQ ID NO:2, and VH CDR3 shown in amino acid sequence as SEQ ID NO:3; The light chain variable region comprises the following CDR: VL CDR1 with amino acid sequence shown in SEQ ID NO:4, VL CDR2 with amino acid sequence shown in SEQ ID NO:5, and VL CDR3 with amino acid sequence shown in SEQ ID NO:6. The human neutralizing antibody of the present invention is an antibody that can block the biological activity of FSTL1, thereby modulating various fibrotic diseases (such as idiopathic pulmonary fibrosis (IPF) and skin fibrosis).

Description

technical field [0001] The invention relates to the field of biomedicine, in particular to a human neutralizing antibody of a follistatin-like protein and an application thereof. Background technique [0002] The information disclosed in this Background section is only for enhancing the understanding of the general background of the present invention and should not be taken as an acknowledgment or any form of suggestion that the information constitutes the prior art that is already known to those skilled in the art. [0003] The essence of visceral fibrosis is the repair response after tissue damage to protect the relative integrity of tissues and organs. The main pathological changes of visceral fibrosis are the increase of fibrous connective tissue and the reduction of parenchymal cells in organ tissues. Continuous progress can lead to organ structural damage, functional decline, and even failure, which seriously threatens human health and life. play an important role in ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/18C12N15/13A61K39/395A61P17/00A61P11/00
CPCC07K16/18A61P17/00A61P11/00C07K2317/24C07K2317/56C07K2317/565C07K2317/567C07K2317/92A61K2039/505
Inventor 杨诚周红刚宁文李霄鹤张亮李建伦东超
Owner TIANJIN JIKUN MEDICAL TECH CO LTD