ECM1 polypeptide epitopes bound to human MHC-class I molecules

A MHC-I, -1ECM1 technology, applied in the direction of medical preparations containing active ingredients, peptides, analytical materials, etc., to achieve the effect of large commercial industrialization, convenient clinical application, and improved accuracy

Active Publication Date: 2016-09-28
魏敏杰
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, using ECM1 as a tumor-associated antigen, using bioinformatics methods to study its CTL epitope, and using it as a new target for tumor immunotherapy, so far, there has been no literature report

Method used

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  • ECM1 polypeptide epitopes bound to human MHC-class I molecules
  • ECM1 polypeptide epitopes bound to human MHC-class I molecules
  • ECM1 polypeptide epitopes bound to human MHC-class I molecules

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1 Screening method for polypeptide epitopes

[0031] The polypeptide sequence obtained in the present invention is directly retrieved from the peptide library through the combination of the supermotif method and the quantitative motif method, and maintains the binding property of MHC-I molecules. Through probabilistic and statistical analysis of these polypeptide sequences, a binding model of the polypeptide and MHC-I class molecules is obtained.

[0032] 1. Obtaining the amino acid sequence of human ECM1

[0033] Find the full-length amino group of natural human ECM1 from the international open shared gene bank NCBI GeneBank

[0034] The acid sequence (540 amino acids in total) is represented as:

[0035] MGTTARAALV LTYLAVASAA SEGGFTATGQ RQLRPEHFQE VGYAAPPSPP

[0036] LSRSLPMDHP DSSQHGPPFE GQSQVQPPPS QEATPLQQEK LLPAQLPAEK

[0037] EVGPPLPQEA VPLQKELPSL QHPNEQKEGM PAPFGDQSHP EPESWNAAQH

[0038] CQQDRSQGGW GHRLDGFPPG RPSPDNLNQI CLPNRQHVVY GPWNLPQSSY

[0039...

Embodiment 2

[0065] Example 2 Computer Simulation of Polypeptide Epitopes Binding to MHC-I Class Molecules

[0066] Use ChemDraw Ultra and ChemDraw3D Ultra in the ChemOffice software package to establish the two-dimensional and three-dimensional structure of the polypeptide epitope obtained above; use SYBYL-X software to analyze the energy and structure of the polypeptide epitope and HLA-A2.1 Modification, simulating the three-dimensional structure of the combination of the two, carrying out molecular dynamics combination simulation and estimation of physiological activity and application value, mainly include the following methods: (1) Construction of polypeptide epitope molecular model. Use ChemDraw Ultra in the ChemOffice software package to construct the two-dimensional structure of the peptide epitope molecule, then import the two-dimensional structure of the peptide epitope into ChemDraw3D Ultra to obtain the three-dimensional model and save it in Mol2 format, and import the three-dim...

Embodiment 3

[0067] Example 3 Synthesis, purification and molecular weight determination of polypeptide epitopes

[0068] The peptide epitope was synthesized by Jill Biochemical (Shanghai) Co., Ltd. using the standard Fmoc protocol, and the purity was detected by high performance liquid chromatography (HPLC), and the molecular weight was determined by mass spectrometry. For its mass spectrogram, see Figure 5 and Figure 7 , it can be seen that the theoretical values ​​of the two polypeptides are close to the measured values, within the allowable error range. HPLC analysis chart see Figure 6 and Figure 8 , it can be seen that the purity is above 95%, indicating that the synthesis effect is good. The above peptides were freeze-dried and stored at -70°C for future use.

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Abstract

The ECM1 polypeptide epitope combined with human MHC-I class molecules is characterized in that: the ECM1 polypeptide epitope combined with human MHC-I class molecules can bind to the binding site of human MHC-I class molecules and can activate specific The polypeptide epitope of cytotoxic T lymphocytes is the following amino acid sequence selected from human extracellular matrix protein-1 (ECM1) and having homology with human ECM1. The ECM1 polypeptide epitope that binds to human MHC-I class molecules is selected from free-type polypeptides, fusion-type polypeptides and chimeric polypeptides in ECM18-16 and ECM1409-417; and one of the five polypeptides A polymer of various forms of monomers. The invention has the advantages of further improving the accuracy of predicting the polypeptide epitope, and simultaneously predicting the polypeptide epitope from the spatial conformation. It is easily synthesized in vitro and easily absorbed by the body after intravenous administration, so it has a very large commercial industrialization value.

Description

technical field [0001] The invention relates to the technical field of biomedical engineering, including screening and obtaining ECM1 polypeptide molecules combined with human major histocompatibility complex (MHC) class I molecules, as well as DNA and RNA encoding these polypeptides, because these polypeptides are compatible with human MHC-I Binding to the binding sites of the class of molecules can activate specific cytotoxic T lymphocytes (CTL cells), thereby effectively killing ECM1-positive tumor tissues and achieving the purpose of treating tumors. Background technique [0002] Extracellular matrix protein-1 (ECM1) is a secreted glycoprotein isolated from the mouse osteogenic stromal cell line MN7 by Mathieu et al. in 1994. Its relative molecular mass is 85KD, and it has a hydrophobic signal peptide and several Glycosylation site. The gene is located on human chromosome 1q21 adjacent to the upper epidermal differentiation complex region, contains 11 exons, and is abou...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K7/06A61K39/00A61P35/00G01N33/68G01N33/574
Inventor 魏敏杰于兆进孙明立陈秋晨
Owner 魏敏杰
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