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Mixed polypeptide vaccine, its preparation and application

A technology of polypeptide vaccines and hybrid peptides, which is applied in the direction of drug combinations, peptide/protein components, medical preparations containing active ingredients, etc., and can solve problems such as inability to be used repeatedly for a long time, allergic immune reactions of patients’ antibodies, and unsuitability for autoimmune diseases. , to achieve the effect of simple process

Inactive Publication Date: 2004-10-13
龚小迪
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the use of foreign antibodies for treatment will induce patients to produce allergic immune responses to the antibodies, and cannot be used repeatedly for a long time, so it is not suitable for autoimmune diseases that require long-term treatment

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0154] Example 1: Chemical synthesis of mouse MCP1 amino-terminal hybrid peptide (named N-MCP1-TT) Since the mouse is selected as a model to test the efficacy of the vaccine, the vaccine must be prepared with a mouse chemokine amino-terminal hybrid peptide. produced as an autoantigen. Now take mouse N-MCP1-TT as an example to illustrate its chemical synthesis method. The specific sequence is as follows:

[0155] QPDAVNAPLTCCYSFTQYIKANSKFIGITELKK

[0156] It is formed by linking the amino terminal fragment of mouse MCP1 with a special fragment of tetanus toxin through peptide bonds, and named it N-MCP1-TT.

[0157] Standard methods of chemical synthesis have been described above in the Preparation Methods section. For example, the specific method we adopt is synthesized by using an automatic peptide synthesizer. According to the sequence of the selected hybrid peptide, the amino acid lysine at the carboxyl end of the sequence that has been coupled to the resin is used as th...

Embodiment 2

[0165] Example 2: Gene recombination and expression in Escherichia coli of N-terminal fragment hybridization peptide (N-IL8-DNA) of IL-8

[0166] The gene for the amino-terminal fragment of the chemokine IL-8 (Appendix 1, SEQ ID NO: 8) was obtained from a human monocyte mRNA library by RT-PCR. The 5'-primer used for this was 5'-ttgctagctccaccatgacttccaagctgg-3'; the 3'-primer was: 5'-tatagcggccgcggagtatgtctttat-3'. The cDNA of this gene was first cloned into the PCR2.1 plasmid vector. Then the IL-8 amino terminal fragment gene (named N-IL8) was excised with restriction endonucleases NheI and HindIII, and transcloned into the pCDNA3.1 expression plasmid vector (this vector contains the DNA fragment of CpG, which can be found in expression in mammalian cells). The plasmid pcDNA3.1 carrying the gene was transfected into Escherichia coli DHα5, and the bacteria were mass-cultured in the culture medium containing Ampicillin antibiotic. The plasmid carrying the hybrid peptide gene...

Embodiment 3

[0174] Example 3: Mouse MCP1 amino-terminal hybrid peptide (N-MCP1-TT) and mouse MIP2 amino-terminal hybrid peptide (N-MIP2-TT) mixed polypeptide vaccine inhibits arthritis in mice. MRL-lpr mice spontaneously developed arthritis at around 24 weeks of mouse age. By intradermally injecting complete Freund's adjuvant into mice, arthritis can be induced early (at about 14 weeks), and the symptoms are obvious, which is conducive to the observation of curative effect. Figure 1a shows that during the onset of arthritis, a large number of monocytes and neutrophils infiltrated into the joint cavity (identified by immunostaining, that is, staining the infiltrating leukocytes with antibodies against various leukocyte surface antigens, and then Each type of leukocytes was counted separately, and the number accounted for the percentage of the total invading cells). The chemokine gene expression was studied by limited PCR method (that is, only 20-30 cycles of amplification during PCR). Th...

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Abstract

The present invention provides a mixed polypeptide vaccine, it can induce body interior to produce antibody directed against self-body several different inflammatory chemotatic factors, and it is made up by mixing hybrid peptides of different species, in which the hybrid peptide of every species is made up by covalently connecting amino tail end of fragment of specific chemotatic factor and immunological recognition fragment. The invention also provides preparation method of these products, specially adopts chemical synthetic production method and provides test method of the product. Said invented mixed vaccine is easily prepared, and is suitable for curing auto immune diseases long-term dependent on medicine.

Description

technical field [0001] The invention relates to medical preparations, in particular to a mixed polypeptide vaccine capable of inhibiting autoimmune diseases and its preparation, production and application. technical background [0002] Proteins are generally composed of more than 50-80 amino acids and are compounds with a certain three-dimensional spatial conformation. Polypeptide: generally composed of less than 50 amino acids, a compound with a certain three-dimensional spatial conformation. Small peptide: composed of less than 30 amino acids. Generally do not have a three-dimensional spatial conformation. Chemokines: Proteins secreted by animals that induce the movement of various leukocytes. Antigen: A substance that elicits an immune response in an animal, including the production of antibodies. Antigens are generally non-self substances. Autoantigen: A substance that elicits an immune response in an animal, including the body's own substances that produce antibodi...

Claims

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Application Information

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IPC IPC(8): A61K38/16A61K39/00A61K39/395A61K45/06A61P29/00A61P37/02
CPCY02A50/30
Inventor 龚疆红
Owner 龚小迪
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