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MUC1-Fc polypeptide vaccine as well as preparation method and application thereof

A polypeptide vaccine, muc1-fc technology, applied in the field of tumor DNA vaccines and viral vector vaccines, can solve problems such as ineffective activation of CTL responses, and achieve the goal of improving antigen bioavailability, good preventive and therapeutic effects, and increased killing rate. Effect

Active Publication Date: 2015-07-29
BENHEALTH BIOPHARMACEUTIC SHENZHEN CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, the currently used MUC1 polypeptide vaccine can only target MHC I or MHC II because the epitope is too small, which has MHC restriction factors and cannot effectively activate the CTL response

Method used

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  • MUC1-Fc polypeptide vaccine as well as preparation method and application thereof
  • MUC1-Fc polypeptide vaccine as well as preparation method and application thereof
  • MUC1-Fc polypeptide vaccine as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] The preparation method of the MUC1 antigen polypeptide with immunoglobulin Fc segment specifically comprises the following steps:

[0047] Fusing the immunoglobulin Fc segment to the MUC1 polypeptide segment to obtain the MUC1 antigen polypeptide sequence with the immunoglobulin Fc segment, its sequence is shown in SEQ ID NO: 1, using the Red / ET homologous recombination method to design the upstream homology arm: (HindIII) CTCTAGCGTTTAAACTT AAGCTT ; downstream homology arm: (BamHI) GGATCC ACTAGTCCAGTGTGGTGGA, the MUC1 antigen polypeptide sequence with immunoglobulin Fc segment is constructed on the pET32a vector, and then expressed and purified in Escherichia coli to obtain the MUC1 antigen polypeptide with immunoglobulin Fc segment.

Embodiment 2

[0049] The preparation method of the MUC1 antigen polypeptide with immunoglobulin Fc segment specifically comprises the following steps:

[0050] Fuse the immunoglobulin Fc segment to the MUC1 polypeptide fragment to obtain the MUC1 antigen polypeptide sequence with the immunoglobulin Fc segment, the sequence of which is shown in SEQ ID NO: 1, using the enzyme-linking method, the restriction site (HindIII) AAGCTT (BamHI) GGATCC, the MUC1 antigen polypeptide sequence with the Fc segment of the immunoglobulin is constructed on the pET29a vector, and then expressed and purified in E. coli to obtain the MUC1 antigen polypeptide with the Fc segment of the immunoglobulin.

Embodiment 3

[0052] Utilize 1,2-dioleoyl-3-trimethylaminoneptane (DOTAP) to coat the MUC1 antigen polypeptide with the immunoglobulin Fc segment of Example 1 to obtain a polypeptide vaccine, and its preparation method comprises the following steps:

[0053] 1) Solvent preparation: prepare a mixture of chloroform and methanol, wherein the volume ratio of chloroform and methanol is 2:1;

[0054] 2) Dissolving 1,2-dioleoyl-3-trimethylaminopropane (DOTAP): Dissolve DOTAP with the solvent in step 1) in a round bottom bottle until the concentration is 1 μM / mL;

[0055] 3) Dry the solvent with a constant flow of nitrogen, so that DOTAP forms a uniform layer of film structure on the container wall;

[0056] 4) After the solvent is blown dry, put it into a vacuum drying oven to dry overnight, so that the solvent can be removed as much as possible;

[0057] 5) Add the MUC1 antigen polypeptide with the immunoglobulin Fc segment of Example 1 diluted with PBS that needs to be coated. The volume should...

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Abstract

The invention relates to an MUC1-Fc polypeptide vaccine as well as a preparation method and an application thereof. The polypeptide vaccine comprises MUC1 antigen peptide containing immune globulin Fc segments, and the sequence of the peptide is represented by SEQ ID NO:1; with the adoption of the genetic engineering technology, the sequence of the MUC1 antigen peptide containing the immune globulin Fc segments is built to a carrier, and the MUC1 antigen peptide is expressed and purified in escherichia coli and coated with carriers such as lipidosome and the like so as to obtain the MUC1-Fc polypeptide vaccine. The MUC1-Fc polypeptide vaccine contains MHC (major histocompatibility complex) I and MHC II binding epitopes simultaneously, thereby being capable of breaking through MHC restrictive factors, effectively activating cytotoxic T lymphocyte (CTL) reaction, having higher killing effect on tumor cells and achieving better preventing and treating effects.

Description

technical field [0001] The invention relates to the field of tumor DNA vaccines and virus vector vaccines, in particular to a MUCl-Fc polypeptide vaccine and its preparation method and application. Background technique [0002] During the occurrence and development of tumor cells, due to gene mutation or expression regulation of cells, some new antigens will be expressed or overexpressed, and these new antigens, as "non-self substances", can be recognized and killed by the body's immune system , the body can resist tumors through natural and acquired immunity. However, tumors can still develop and metastasize under the action of the immune function of the human body, indicating that tumors also have their own protective mechanisms. Tumor cells can evade the body's immune recognition and attack by modifying their own antigens and changing the microenvironment around the tumor, that is, tumor immune escape. The tumor vaccine can break the tolerance of the autoimmune system t...

Claims

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Application Information

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IPC IPC(8): A61K39/00A61K39/395A61P35/00A61P35/04
CPCA61K39/00A61K39/395
Inventor 于浩洋李正成苏静
Owner BENHEALTH BIOPHARMACEUTIC SHENZHEN CO LTD
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