Preparation method of breast cancer-specific epitope polypeptide-loaded dendritic cell vaccine and kit thereof

A technology of dendritic cells and antigenic epitopes, applied in animal cells, vertebrate cells, blood/immune system cells, etc., can solve the problems of cancer recurrence, poor prognosis, and inability to obtain patients, and achieve a radical cure for recurrence and metastasis , the effect of overcoming tolerance

Inactive Publication Date: 2014-05-14
BEIJING HONGRUNYUAN BIOLOGICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The existing technology, that is, the conventional treatment methods such as surgery, chemotherapy and radiotherapy can remove or kill most of the breast cancer in the body, but there are still some re...

Method used

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  • Preparation method of breast cancer-specific epitope polypeptide-loaded dendritic cell vaccine and kit thereof
  • Preparation method of breast cancer-specific epitope polypeptide-loaded dendritic cell vaccine and kit thereof
  • Preparation method of breast cancer-specific epitope polypeptide-loaded dendritic cell vaccine and kit thereof

Examples

Experimental program
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Effect test

preparation example 1

[0048] Preparation example 1 Human Her2 / neu, BCRP and GEP100 protein HLA-A2010 positive epitope peptide prediction

[0049] The bioinformatics and molecular analysis system (BIMAS) was used to predict the binding of HLA polypeptides, and the HLA epitope polypeptides that specifically bind to human Her2 / neu, BCRP and GEP100 proteins were screened, and 6 kinds of HLA-A0201 binding affinity over 100 were selected. The amino acid sequence of is shown in Table 1:

[0050]

[0051]

[0052] Preparation Example 2 Dendritic cell acquisition

[0053] Take 100 mL of peripheral blood by Ficoll-Hypaque density gradient centrifugation to obtain mononuclear cells. Peripheral blood mononuclear cells were resuspended in RPMI1640 medium and added to a 6-well plate to adhere. At 37℃, 5% CO 2 After 90 minutes of incubation in the incubator, the non-adherent cells were washed and collected for induction of CIK cells. Adherent cells were induced by adding complete medium RPMI1640, containing 5% au...

Embodiment 3

[0054] Example 3 DCs loaded with Her2 / neu, BCRP and GEP100 protein epitope polypeptide

[0055] After 5 days of cultivation, 3×10 6 Immature DCs use the HER2 / NEU, BCRP and GEP100 protein SEQ ID NO. 1-6 of the HER2 / NEU, BCRP and GEP100 protein SEQ ID NO. 1-6 polypeptide composition with more than one epitope or three epitope polypeptide compositions of the three antigens At 37℃, 5% CO 2 Load in the incubator for 2 hours. The antigen-loaded DCs were harvested by centrifugation (10min, 1000rpm). The DCs obtained were washed 3 times with normal saline, and then resuspended in normal saline to a concentration of 3×10 6 Mature DC / mL, and adding human albumin with a final concentration of 2% by mass to volume, thereby preparing a DC vaccine loaded with a composition of Her2 / neu, BCRP and GEP100 protein epitope polypeptide.

Embodiment 1

[0056] The Her2 / neu, BCRP and GEP100 protein SEQ ID NO. 1-6 of the polypeptide composition of more than one epitope in SEQ ID NO. 1-6 described in Example 1 load dendritic cells through CD86-PE, CD80-PE, CD40-FITC, CD83 -PE, CD11c-FITC and HLA-DR-PerCP (BD company) staining, flow cytometry to detect DC phenotype changes after loading. The test results showed that the phenotype of the prepared DC was CD11c+ / HLA-DR+ 95.0%, CD11c+ / CD83+ 79.8%, CD86+ / HLA-DR+ 89.9%, CD80+ / HLA-DR+ 90.5% and CD40+ / HLA-DR+ 85.8%, in line with the specific phenotype of DC cells, and high expression of costimulatory molecules.

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Abstract

The invention provides a preparation method of a breast cancer-specific epitope polypeptide-loaded dendritic cell vaccine, and the method comprises the following steps: loading dendritic cells with identified human leukocyte antigen A201 positive epitope polypeptide of human epidermal growth factor receptor 2, breast cancer drug resistance protein and IQ motif and Sec7 structural domain 1 protein which specifically express in breast cancer (namely polypeptide comprising the amino acid sequence shown in any of SEQ ID No.1-6) to prepare into the dendritic cell vaccine which can induce strong breast cancer-targeting cytotoxic T lymphocyte response. The invention also provides a kit comprising the breast cancer-specific epitope polypeptide-loaded dendritic cell vaccine.

Description

Technical field [0001] The present invention relates to a method for preparing a dendritic cell vaccine loaded with a breast cancer specific epitope polypeptide, a kit using the method, and a dendritic cell and vaccine obtained by the method. In particular, the present invention relates to the use of human epidermal growth factor receptor 2, breast cancer resistance protein, IQ motif and Sec7 domain 1 protein expressing human epidermal growth factor receptor 2, breast cancer resistance protein and Sec7 domain 1 protein. A method for preparing morphocytes, a kit using the method, and a dendritic cell vaccine obtained by the method. Background technique [0002] At present, breast cancer is one of the most important diseases threatening women’s health. China is one of the countries with the fastest increase in the incidence of breast cancer, with about 470,000 patients. The incidence of breast cancer is now increasing at a rate of 3% per year. Has become the number one killer of u...

Claims

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Application Information

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IPC IPC(8): A61K39/00A61P35/00C12N5/0784
Inventor 沈丽黄启明郝丽敏
Owner BEIJING HONGRUNYUAN BIOLOGICAL TECH
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