Nanometer gene vaccine as well as preparation method and application thereof

A gene vaccine and nanotechnology, applied in the field of novel nanogene vaccines with anti-tumor and immune-enhancing activities and their preparation, can solve the problems of high cost, low efficiency of active cellular uptake, fast metabolism, etc.

Inactive Publication Date: 2015-01-21
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the process of induction expression, in vitro purification and renaturation of the recombinant protein has the disadvantages of long time-consuming, cumbersome steps, and high cost, wh

Method used

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  • Nanometer gene vaccine as well as preparation method and application thereof
  • Nanometer gene vaccine as well as preparation method and application thereof
  • Nanometer gene vaccine as well as preparation method and application thereof

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Embodiment Construction

[0045] 1. Construction of pQE31-dsNKG2D-IL-15 prokaryotic expression vector

[0046] 1.1 Primer sequences for PCR amplification: use bioinformatics methods to design corresponding primers, and use the cDNA of human peripheral blood mononuclear cells as a template (see Table 1) to amplify sNKG2D (a) and sNKG2D (b) Gene fragment: The gene fragment of IL-15 was amplified using the pORF hIL-15 vector (from Invivogen) as a template. The reverse complementary sequence of (Gly4Ser)4 (shown in the box in Table 1) was added to the downstream primer sequences of sNKG2D(a) and sNKG2D(b).

[0047] Table 1. Primer sequences for sNKG2D (a), sNKG2D (b) and IL-15 gene amplification

[0048]

[0049] 1.2 The PCR amplification product sequence of sNKG2D(a) is shown as SEQ ID NO: 6, wherein the 1st-6th base is the sequence of the BamH I restriction site; the 7th base is randomized to avoid frameshift mutation of NKG2D Nucleotides added; No. 8-425 is the gene sequence of NKG2D extracellular ...

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Abstract

The invention relates to a nanometer gene vaccine as well as a preparation method and an application thereof, belonging to the technical field of biology. In order to overcome the defects such as high preparation time consumption, complicated steps and high cost existing in recombinant proteins, the invention provides a nanometer gene vaccine. According to the nanometer gene vaccine, based on a genetic engineering technology, a signal peptide sequence, two identical NKG2D extracellular sequences and a sequence of cell factors IL-15 needed by eukaryotic expression are introduced, and an expression vector of fused proteins is constructed and is prepared into the nanometer gene vaccine by using a biocompatible chitosan material. Because the nanometer gene vaccine carries positive charges, the vaccine can be actively devoured by tumor cells or muscle cells, and proteins dsNKG2D-IL-15 linked between the tumor cells and lymphocytes are expressed in the eukaryocyte. Therefore, natural killer cells (NK) or cytotoxic T cells (CTL) in vivo are activated, the specific immune response in vivo can be enhanced, and an antitumor drug is prepared.

Description

technical field [0001] The invention relates to a novel nano-gene vaccine with anti-tumor and immune-enhancing activities and a preparation method thereof, belonging to the field of biotechnology. Background technique [0002] Tumor cells reduce the expression of HLA class I molecules and co-stimulatory molecules, escape the surveillance of T lymphocytes in the body, but activate the activity of NK cells. If the activating signal transmitted by activating receptors on the surface of NK cells is stronger than the inhibitory signal transmitted by inhibitory receptors, NK cells will be activated, secrete cytokines and directly kill tumor cells, known as the body's first anti-tumor line of defense. On the other hand, the fact that tumors escape the monitoring function of NK cells and a large number of experimental evidences prove that the number and function of NK cells in tumor tissues are lower than those in normal tissues. Therefore, how to promote NK cell proliferation and...

Claims

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Application Information

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IPC IPC(8): C12N15/62C12N15/63A61K48/00A61K39/00A61P35/00A61P37/02
Inventor 龚卫娟邵小青陈艳奚菊群肖炜明
Owner YANGZHOU UNIV
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