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Cancer cell/DC fusion tumor vaccine for expressing Endoglin and preparation method for cancer cell/DC fusion tumor vaccine

A technology of tumor cells and fusion cells, which is applied in the field of cancer cell/DC fusion tumor vaccines expressing tumor blood vessel marker molecules and its preparation, and can solve the problems of unsatisfactory therapeutic effects of fusion vaccines

Active Publication Date: 2015-04-29
GUANGXI MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the therapeutic effect of fusion vaccines is still unsatisfactory, and new methods are needed to enhance the effectiveness and targeting of fusion cells to induce CTL

Method used

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  • Cancer cell/DC fusion tumor vaccine for expressing Endoglin and preparation method for cancer cell/DC fusion tumor vaccine
  • Cancer cell/DC fusion tumor vaccine for expressing Endoglin and preparation method for cancer cell/DC fusion tumor vaccine
  • Cancer cell/DC fusion tumor vaccine for expressing Endoglin and preparation method for cancer cell/DC fusion tumor vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0092] Embodiment 1, the preparation of fusion cell

[0093] The method for preparing fusion cells, including S1) and S2):

[0094] S1) introducing Endoglin coding gene into tumor cells to obtain recombinant cells;

[0095] S2) Fusing the recombinant cells with dendritic cells to obtain the fused cells.

[0096] The specific method is as follows:

[0097] 1. Preparation of recombinant cells

[0098] Replace the DNA fragment between the BamHI and XbaI recognition sites on the pLVX-Puro vector with the DNA molecule shown in SEQ ID No.2 in the sequence table, keep the other sequences of pLVX-Puro unchanged, and obtain the recombinant vector pLVX-Puro / Eng , the recombinant vector pLVX-Puro / Eng expresses the Endoglin shown in SEQ ID No.1. The recombinant vector pLVX-Puro / Eng was introduced into HepG2 cells to obtain recombinant cells containing Endoglin coding sequence and EGFP coding sequence, and the recombinant cells were named HepG2(Eng+).

[0099] Wherein, the 1st-1683rd ...

Embodiment 2

[0117] Example 2, DC / HepG2 (Eng+) in vitro induces the generation of T lymphocytes that secrete IFN-γ

[0118] The experiment was repeated three times, and the specific steps of each repeated experiment were as follows:

[0119] The T lymphocytes secreting IFN-γ induced by DC / HepG2 (Eng+) were obtained according to the following method, and the IFN-γ secreted by T lymphocytes was detected by enzyme linked immunospot assay (ELISPOT). ×Washing buffer, biotin-labeled antibody, and enzyme-labeled avidin are all Human IFN-gamma precoated ELISPOT kit (Human IFN-gamma precoated ELISPOT kit is a product of Daktronix Biotechnology Co., Ltd., the catalog number is DKW22-1000-048) The reagents in the specific steps are as follows:

[0120] 1) Take out the well plate in the kit, add 200 μL of incomplete RIPM 1640 medium to each well, let stand at room temperature for 5-10 minutes, and pour off the liquid.

[0121] 2) Add DC / HepG2 (Eng+) 3×10 of Example 1 suspended in incomplete RIPM 164...

Embodiment 3

[0134]Example 3. Therapeutic effect of T lymphocytes secreting IFN-γ induced by fusion cells DC / HepG2 (Eng+) on tumors

[0135] The experiment was repeated three times, and the specific steps of each repeated experiment were as follows:

[0136] The T lymphocytes secreting IFN-γ induced by DC / HepG2 (Eng+) of Example 2, the T lymphocytes secreting IFN-γ induced by DC / HepG2, and the IFN-γ secreted induced by DC / HepG2 (pLVX-Puro) T lymphocytes induced by DC, T lymphocytes secreting IFN-γ induced by DC, T lymphocytes induced by HepG2 (Eng+), T lymphocytes induced by HepG2 and T lymphocytes induced by HepG2 (pLVX-Puro) were suspended in PBS , respectively, the cell content is 10 5 cells / μL of DC / HepG2(Eng+)-induced IFN-γ-secreting T lymphocyte suspension, DC / HepG2-induced IFN-γ-secreting T-lymphocyte suspension, DC / HepG2(pLVX-Puro)-induced secretion IFN-γ T lymphocyte suspension, DC induced IFN-γ secreting T lymphocyte suspension, HepG2(Eng+) induced T lymphocyte suspension, HepG...

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Abstract

The invention discloses a cancer cell / DC (Dendritic Cell) fusion tumor vaccine for expressing Endoglin and a preparation method for the cancer cell / DC fusion tumor vaccine. According to the cancer cell / dendritic cell fusion tumor vaccine for expressing Endoglin and a preparation method for the cancer cell / dendritic cell fusion tumor vaccine, the method for preparing fusion cells comprises the step of fusing tumor cells and dendritic cells to obtain the fusion cells; the tumor cells are recombinant cells for expressing the Endoglin or the recombinant cells which contain the gene for encoding the Endoglin; the Endoglin is the protein of A1) or A2) below: the A1) is the protein of which the amino acid sequence is SEQ ID No.1; the A2) is the protein which has the same function as that of the protein of the A1) and is derived from the protein of the A1) performing substituting and / or deleting and / or adding on one or more amino acid residues of the amino acid sequence which is shown as the SEQ ID No.1.

Description

technical field [0001] The invention relates to a cancer cell / DC fusion tumor vaccine expressing tumor blood vessel marker molecules in the field of biomedicine and a preparation method thereof. Background technique [0002] Immunotherapy for malignant tumors has become a new treatment method besides surgery, radiotherapy and chemotherapy. Among them, tumor-specific cytotoxic T lymphocyte (cytotoxic T lymphocyte, CTL) adoptive therapy has been the research focus of tumor immunotherapy. Fusion of tumor cells and dendritic cells (DC) has been widely accepted as an effective method to induce CTL. However, the therapeutic effect of fusion vaccines is still unsatisfactory, and new methods are needed to enhance the effectiveness and targeting of fusion cells to induce CTL. [0003] Endoglin (also known as CD105, differentiation group 105) is a marker molecule of tumor blood vessels, a homodimeric transmembrane glycoprotein with a molecular weight of 180kDa, and a transforming gr...

Claims

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Application Information

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IPC IPC(8): C12N5/16A61K35/17A61K35/15A61P35/00
Inventor 赵永祥卢小玲陈玥周源
Owner GUANGXI MEDICAL UNIVERSITY
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