IKK beta-targeted short peptide and application thereof in inflammatory diseases

An inflammatory response and short peptide technology, applied in the field of biomedicine, can solve problems such as tissue damage, and achieve the effects of weak side effects, small molecular weight, and weak immunogenicity

Active Publication Date: 2020-11-24
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The inflammatory response is a protective host response to infection and tissue damage; normally, the inflammatory response is beneficial for the host to clear

Method used

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  • IKK beta-targeted short peptide and application thereof in inflammatory diseases
  • IKK beta-targeted short peptide and application thereof in inflammatory diseases
  • IKK beta-targeted short peptide and application thereof in inflammatory diseases

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Example 1: Screening of IKIP Action Regions

[0051] The results of the study showed that the IκB kinase binding protein IKIP has a significant binding effect on IKKβ, which inhibits the combination of IKKβ and IKKγ, thereby inhibiting the formation of IKK complex, restricting the entry of p65 into the nucleus, and finally inhibiting the expression of inflammatory cytokines, thereby inhibiting inflammation. The function of the response; cell experiments and animal experiments have shown that the loss of IKIP will lead to a significant increase in inflammatory response. The applicant found that the function of IKIP is mainly dependent on its N-terminal structure, in which the binding protein IKIP of IKKβ competes with IKKγ through 99 amino acids at positions 1-99 of the N-terminus. IKKα / β inhibits the formation of the IKK complex, thereby hindering IKKα / β phosphorylation process and negatively regulate the activation of downstream NF-κB signaling pathway. Therefore, a ...

Embodiment 2

[0063] Example 2: Verifying the effect of short peptides on the expression of inflammatory cytokines

[0064] (1) Short peptides (5 μM) F0, F1, and F2 were added to mouse peritoneal macrophages in advance for pretreatment, and then stimulated by adding LPS, PGN, TNFα, and IL-1β 2 hours later, at different time points by RT- The mRNA levels of inflammatory cytokines TNF-α and IL-6 were detected by PCR.

[0065](2) Short peptides (5μM) F0, F1, and F2 were added to mouse peritoneal macrophages in advance for pretreatment, and then stimulated by adding LPS and PGN respectively after 2 hours. After 24 hours of culture, the inflammatory cytokine TNF-α was detected by ELISA , IL-6 secretion level.

[0066] (3) Short peptides (5μM) F0, F1, and F2 were added to mouse peritoneal macrophages in advance for pretreatment. After 2 hours, LPS, PGN, TNFα, IL-1β and other stimuli were added, and Western Blot was used at different time points. Detect the phosphorylation level of adapter prote...

Embodiment 3

[0070] Example 3: Verifying the effect of short peptides on the IKK complex

[0071] (1) 293T cells, overexpressing IKKβ-HA plasmid, adding short peptides F0, F1, F2 (5μM) 24 hours after transfection, adding IP Buffer 2 hours later to lyse the cells. Take 50 μL of cell lysate as Input, and add 50 μL of StreptavidinBeads to the remaining samples. After shaking for 4 hours in a silent mixer, wash the beads with 1000 g of IP Buffer for 4 minutes*4 times. The combination of short peptide and IKKβ was detected by Western Blot.

[0072] (2) 293T cells overexpressed IKKα-Myc, IKKβ-Myc, and IKKγ-Myc, and 24 hours after transfection, short peptide F1 (5 μM) was added. After continuing to culture for 2 hours, IP Buffer lysed the cells. Take 50 μL of cell lysate as Input, and add 50 μL of Streptavidin Beads to the remaining samples. After shaking for 4 hours in a silent mixer, wash the beads with 1000 g of IP Buffer for 4 min*4 times. Western Blot was used to detect the combination of...

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Abstract

The invention belongs to the field of biological medicines, and particularly relates to an IKK beta-targeted short peptide and application thereof in inflammatory diseases. In order to provide a moreeffective treatment means for inhibiting inflammatory reaction, the invention provides the IKK beta-targeted short peptide. The short peptide comprises an N-terminal amino acid 46-60 region derived from I kappa B kinase binding protein IKIP, and the N terminal of the short peptide is fused with a cell-penetrating peptide and has a polypeptide marker. The short peptide can target IKK beta and inhibit combination of IKK beta and IKK gamma, thus, formation of an IKK complex is inhibited, expression of inflammatory cytokines TNF-alpha and IL-6 is reduced, and inflammatory reaction is inhibited. Ananimal model shows that the short peptide can effectively treat arthritis and inflammatory enteritis. Compared with full-length proteins, the short peptide has the advantages of small molecular weight, easiness in synthesis, weak immunogenicity and relatively weak side effects; and the cell-penetrating peptide has better absorption, and the polypeptide marker is easy to track and observe, and thus, a reference is provided for clinically treating the inflammatory diseases.

Description

technical field [0001] The invention belongs to the field of biomedicine, and specifically relates to a short peptide targeting IKKβ or a pharmaceutical composition and / or auxiliary pharmaceutical composition for inhibiting inflammatory response or preventing / treating inflammatory response diseases or a short peptide targeting IKKβ Use of the peptide in the preparation of products for inhibiting inflammatory responses or preventing or treating inflammatory response diseases. Background technique [0002] The information disclosed in this background section is only intended to increase the understanding of the general background of the present invention, and is not necessarily taken as an acknowledgment or any form of suggestion that the information constitutes the prior art already known to those skilled in the art. [0003] The inflammatory response is a protective host response to infection and tissue damage; normally, the inflammatory response is beneficial for the host t...

Claims

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Application Information

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IPC IPC(8): C07K19/00A61K38/17A61K47/64A61P29/00
CPCC07K14/47A61P29/00C07K2319/10A61K38/00
Inventor 高成江张磊柳翰森吴海峰赵学英郑义刘冰玉
Owner SHANDONG UNIV
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