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Interleukin 17A-targeting antibody, preparation method and applications thereof

An antibody and chimeric antibody technology, applied in the field of medicine, can solve problems such as unclear downstream signals

Active Publication Date: 2018-08-03
HUABO BIOPHARM
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

IL-17B was found to bind IL-17RB, but its downstream signaling remains unclear

Method used

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  • Interleukin 17A-targeting antibody, preparation method and applications thereof
  • Interleukin 17A-targeting antibody, preparation method and applications thereof
  • Interleukin 17A-targeting antibody, preparation method and applications thereof

Examples

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preparation example Construction

[0138] Antibody preparation

[0139] Any method suitable for producing monoclonal antibodies can be used to produce anti-IL-17A antibodies of the invention. For example, animals can be immunized with linked or naturally occurring IL-17A homodimers or fragments thereof. Suitable immunization methods may be used, including adjuvants, immunostimulants, repeated booster immunizations, one or more routes may be used.

[0140]Any suitable form of IL-17 can be used as an immunogen (antigen) for producing non-human antibodies specific to IL-17A and screening the biological activity of the antibodies. The stimulating immunogen can be full-length mature human IL-17A, including natural homodimers, or single / multiple epitope-containing peptides. Immunogens can be used alone or in combination with one or more immunogenicity enhancers known in the art. Immunogens can be purified from natural sources, or produced in genetically modified cells. The DNA encoding the immunogen can be genomi...

Embodiment 1

[0172] Example 1 Anti-human IL-17A mouse monoclonal antibody—preparation method of original antibody

[0173] 1.1 Preparation of hybridoma cells producing mouse monoclonal antibodies

[0174] Firstly, human IL-17A protein was used as antigen, emulsified with adjuvant, and multi-point subcutaneously immunized BALB / c mice to monitor the serum titer of immunized mice. After reaching the requirements, the spleen cells and myeloma (Sp2 / c) mice were collected. 0) The cells are fused and screened by HAT to obtain hybridoma polyclonal cells.

[0175] 1.2 Indirect ELISA—the screening method of hybridoma cells

[0176] Use the ELISA detection method to screen out polyclonals with high specific binding, carry out monoclonal culture, and then use the ELISA method to screen the monoclonal cell lines with high specific binding; through the HT1080 cells to IL-6 release test, screen the cells with functional The effect of monoclonal cell lines, and then use the Biacore method to analyze the...

Embodiment 2

[0185] Example 2 Anti-IL-17A antibody V-gene sequence cloning

[0186] Based on the 5' RACE technique, the DNA sequence encoding the variable region of the mouse antibody expressed by the hybridoma 7D6 / 5H8 was determined. Briefly, heavy and light chain gene-specific cDNAs were prepared using SMART 5'RACE Synthesis Kit (TAKARA, No. 634859) according to the manufacturer's instructions. PCR products were analyzed by agarose gel electrophoresis. The variable regions of both the heavy and light chains are approximately 500 base pairs in size. The amplified PCR product with the appropriate band size obtained from the reaction was cloned into the vector pEASY-Blunt Simple plasmid (Beijing Quanshijin, No.CB111-02), and transformed into Stellar Escherichia coli competent cells (TAKARA, No.636763 )middle. Clones were screened by colony PCR with universal M13 forward or reverse primers, and 2-3 clones from each reaction were selected for DNA sequencing analysis. The results of each s...

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Abstract

The present invention provides an interleukin 17A (IL-17A)-targeting antibody, a preparation method and applications thereof, particularly a new anti-IL-17A monoclonal antibody. According to the present invention, the antibody can bind to IL-17A antigen in a high-specificity manner, has high affinity and low immunogenicity, and is used for preparing drugs for prevention or treatment of IL-17A related diseases such as various inflammations or autoimmune diseases.

Description

technical field [0001] The invention relates to the field of medicine, in particular to an antibody targeting interleukin 17A (also known as IL-17), its preparation method and application. Background technique [0002] So far, six members of the IL-17 family have been discovered: IL-17A (IL-17), IL-17B, IL-17C, IL-17D, IL-17E (IL-25), and IL-17F. These interleukin-17 cytokines can bind to the corresponding receptors to mediate different inflammatory responses. [0003] IL-17A was originally found to be activated by CD4 + secreted by T cells. This type of characteristic T cell subset that secretes IL-17A is called Th17 cells. In addition to Th17 cells, cytotoxic CD8 + T cells (Tc17), γδ T cells, natural killer T cells (NKT-17) and B cells can also express IL-17A under specific conditions. Innate immune cells, including monocytes, neutrophils, natural killer cells, and lymphoid tissue-inducing-like (Lti-like) cells can also produce IL-17A. Recently, in a study of trypano...

Claims

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Application Information

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IPC IPC(8): C07K16/24C12N5/10A61K39/395A61P29/00A61P37/02A61P17/06A61P19/08A61P19/02A61P25/00G01N33/68
CPCG01N33/6869C07K16/244A61K2039/505C07K2317/56C07K2317/33C07K2317/24C07K2317/92G01N2333/54C07K2317/76A61K2039/54A61K2039/545C07K14/7051C07K2317/565C07K2317/622C07K2319/01
Inventor 朱向阳蔡明清于海佳贾慧峰俞玲
Owner HUABO BIOPHARM
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