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Tumor necrosis factor (TNF) binding peptide and TNFR1 blocking peptide, and application thereof in treatment of ulcerative colitis

A peptide-binding technology for ulcerative colitis, applied in DNA/RNA fragments, medical preparations containing active ingredients, peptides, etc., can solve problems such as poor biological stability, high price, and limited sources

Inactive Publication Date: 2012-01-18
HUAZHONG UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Although the above-mentioned commercialized anti-TNF-α reagents (anti-TNF antibody, soluble TNF receptor) have achieved good clinical therapeutic effects, there are still a series of problems, such as potential side effects, limited sources, poor biological stability, expensive etc.
At present, clinical trials have found that the above-mentioned anti-TNF-α reagents (anti-TNF antibodies, soluble TNF receptors) have side effects such as red blood cell and thrombocytopenia, tuberculosis infection, upper respiratory tract, urinary tract infection, rash, fever, hypo / hypertension, etc. Severe cases may cause tumors

Method used

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  • Tumor necrosis factor (TNF) binding peptide and TNFR1 blocking peptide, and application thereof in treatment of ulcerative colitis
  • Tumor necrosis factor (TNF) binding peptide and TNFR1 blocking peptide, and application thereof in treatment of ulcerative colitis
  • Tumor necrosis factor (TNF) binding peptide and TNFR1 blocking peptide, and application thereof in treatment of ulcerative colitis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0082] Example 1 Screening and Specific Identification of TNFR1 Blocking Peptides

[0083] With the help of phage display technology, a 12-amino-acid linear peptide (TNFR1 blocking peptide) that can bind to TNFR1 was screened from the phage 12-line peptide library using hrTNFR1 protein as bait. The specific screening process was the same as in 7.1 of Example 7 below.

Embodiment 2

[0084] Example 2 Effect of TNF receptor blocking peptide (12-line peptide) on rat peritoneal macrophage function

[0085] The following TNFR1 blocking peptide (12-line peptide) powders were all synthesized by the Third Military Medical University.

[0086] 2.1 The effect of TNF receptor blocking peptide on TNF-α-induced rat peritoneal MΦ respiratory burst was detected by NBT method: rat peritoneal macrophages were routinely isolated, and TNF-α was added to 3×105 / ml rat peritoneal macrophages (5ng / ml) 100μl and / or TNFR1 blocking peptide (10μg / ml) 100μl, set 3 duplicate wells for each group; then add 100μl / well NBT (2mg / ml), put in 37 ℃ CO2 incubator for 45 minutes; Add 100 μl / well of 70% methanol solution, fix for 5 min; discard the supernatant, wash 3 times with 70% methanol solution; air dry, finally add 120 μl / well 2M KOH solution and 140 μl / well DMSO, mix immediately Measure its OD value at 630nm. The result is as figure 1 : Exogenous TNF2α can significantly enhance the r...

Embodiment 3

[0089] Example 3 Effect of TNFR1 blocking peptide on adjuvant arthritis in rats

[0090] 3.1 Establishment of adjuvant arthritis in rats Wistar rats, ♂, weighing 150-200 g, were provided by the Experimental Animal Center of Tongji Medical College, Huazhong University of Science and Technology. Complete Freund's adjuvant. Experimental grouping: control group: injection of normal saline 0.1ml / rat; model group: intradermal injection of Freund's complete adjuvant (F5881, purchased from Sigma, USA) 0.1ml / rat in the right foot pad; treatment group 1: injection Immediately after the adjuvant, inject TNFR1 blocking peptide 0.1ml / body, the doses are 0.5 and 1.0mg / kg body weight, qd, each dose 4, continuous treatment 10d, treatment group 2: the method and dosage are the same as treatment group 1, continuous After 20 days of treatment, the ankle joint swelling (cm) of the rats was measured with a vernier caliper, and the joint tissues were routinely sliced ​​for HE staining. After the ...

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Abstract

The invention relates to tumor necrosis factor (TNF) binding peptide and TNFR1 blocking peptide. The amino acid sequences of the TNF binding peptide and the TNFR1 blocking peptide are Cys-Lys-His-Gln-Trp-His-lys-His-Cys and Cys-lys-His-Ala-Leu-His-Arg-His-Cys; both the TNF binding peptide and the TNFR1 blocking peptide can competitively inhibit the TNF-alpha from bonding with the TNFR1 in vivo and in vitro to antagonize biological functions of the TNF-alpha, and have better treatment effect in trinitro-benzene-sulfonic acid (TNBS)-induced experimental ulcerative colitis rat model.

Description

[0001] This application is a divisional application of a Chinese patent application with application number 200810236895.3. The application number of the original application is: 200810236895.3; the application date is: December 19, 2008; the invention name is: TNF-α binding peptide and TNFR1 blocking peptide and Its application in the treatment of TNF-related diseases. technical field [0002] The invention relates to biotechnology, in particular to small molecular polypeptides such as TNF-binding peptides, TNFR1 blocking peptides and TNFR1 blocking peptide-Fc fusion proteins capable of antagonizing the biological activity of tumor necrosis factor-α (TNF-α). Background technique [0003] TNF-α is an important class of pro-inflammatory cytokines. freed. Clinical studies have confirmed that: in the occurrence and development of various infectious or non-infectious inflammations and their complications (such as autoimmune diseases such as rheumatoid arthritis, endotoxic shock...

Claims

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Application Information

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IPC IPC(8): C07K7/06C12N15/11A61K38/08A61P1/04
Inventor 李卓娅尹丙姣王晶梁慧芳黄丽霞何亚萍
Owner HUAZHONG UNIV OF SCI & TECH
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