Recombinant virus vector, immune composition containing same, and use

A technology of recombining viral vectors and antigens, applied in the fields of molecular biology and immunology, can solve problems such as not achieving the desired effect, and achieve the effect of prolonging survival time and inhibiting the growth of tumor cells

Inactive Publication Date: 2020-03-27
VACDIAGN BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the separate recombination of tumor antigens and toxins has not yet achieved the desired effect.

Method used

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  • Recombinant virus vector, immune composition containing same, and use
  • Recombinant virus vector, immune composition containing same, and use
  • Recombinant virus vector, immune composition containing same, and use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Shuttle vector pSC65-LMNB construction

[0042] The eukaryotic expression vector pVKD1.0-LMNB expressing the triple fusion tumor antigens LAGE-1, MAGE-A3, NY-ESO-1 and cholera toxin B subunit was provided by Suzhou Industrial Park Weida Biotechnology Co., Ltd. (refer to CN109575142A) , wherein, the amino acid sequence of the LAGE-1 antigen is shown in SEQ ID NO: 1, its encoding nucleic acid sequence is shown in SEQ ID NO: 2; the amino acid sequence of the MAGE-A3 antigen is shown in SEQ ID NO: 3 Its coding nucleic acid sequence is shown in SEQ ID NO: 4; the amino acid sequence of the NY-ESO-1 antigen is shown in SEQ ID NO: 5, and its coding nucleic acid sequence is shown in SEQ ID NO: 6; the described The amino acid sequence of the cholera toxin B subunit polypeptide is shown in SEQ ID NO: 7, and its coding nucleic acid sequence is shown in SEQ ID NO: 8; the amino acid of LMNB fusion protein is shown in SEQ ID NO: 9, and its nucleic acid coding sequence is s...

Embodiment 2

[0045] Example 2 Construction of recombinant vaccinia virus vector rvv-LMNB

[0046] The recombinant vaccinia virus vector was obtained in 143B cells, and the specific method was as follows. On the first day, 143B cells were plated in 6-well cell culture plates (JET, TCP-010-006) ( CRL-8303), 1×10 6 / well and incubated overnight in a 37°C carbon dioxide cell incubator. On the second day, at 0.05 MOI (i.e. 5×10 4 PFU (plaque forming unit) / well) was added to vaccinia virus wild strain 752-1 (provided by Beijing Biological Products), and then placed in a 37°C carbon dioxide cell incubator and incubated for two hours, during which the shuttle vector / transfection reagent was prepared Complex. The shuttle vector is pSC65-LMNB obtained in Example 1, and the transfection reagent is Turbofect (Thermo Fisher Scientific, R0531). For the transfection dose and compounding method, please refer to the instructions of the transfection reagent. After the complex system was completed, t...

Embodiment 3

[0055] Example 3 Amplification preparation and titration of recombinant vaccinia virus vector rvv-LMNB

[0056] The recombinant vaccinia virus vector rvv-LMNB constructed in Example 2, and the wild strain of vaccinia virus were respectively grown in Vero cells ( CCL-81) was amplified, and the amplification method was as follows.

[0057] The day before, prepare a 100% confluent Vero monolayer (1×10 7 cells / dish), a total of 10 dishes.

[0058] Remove the supernatant, replace with the maintenance medium, inoculate the poxvirus to be amplified on the cells (0.01PFU / cell), incubate in a 37°C incubator for 2-3 days, and observe obvious cytopathic changes.

[0059] The cells were scraped and collected, centrifuged at 1800 g for 5 minutes, and the supernatant was removed.

[0060] Resuspend with 5 mL of maintenance medium, and sonicate on ice with an ultrasonic cell pulverizer, the sonication conditions are: 50 watts, 5 seconds of ultrasound / 5 seconds interval, a total of 15 m...

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Abstract

The invention provides a recombinant virus vector, an immune composition containing the same, and a use. The recombinant virus vector comprises a polynucleotide encoding a tumor antigen, and the tumorantigen is one or more selected from a LAGE antigen, a MAGE antigen, or a NY-ESO-1 antigen. A recombinant virus vector vaccine provided by the invention can stimulate tumor-specific immune response,effectively inhibits tumor cell growth, and prolongs the survival time of tumor patients.

Description

technical field [0001] The present invention belongs to the fields of molecular biology and immunology. Specifically, the present invention relates to a recombinant virus vector, an immune composition containing the same and its use, and in particular, the present invention relates to a tumor therapeutic vaccine. Background technique [0002] With the development of tumor biology and immunology, tumor immunotherapy has made great progress and has gradually become an important development direction of current tumor treatment. Especially with the great success of PD-1 / PD-L1 immunotherapy, the development of tumor immunotherapy has become an emerging hotspot in current tumor treatment. [0003] Tumor vaccines stimulate the body's specific immune response against tumors and activate the production of huge immune cells to kill and control the growth of tumor cells, thereby achieving the effect of reducing or controlling tumor growth. The development of tumor vaccines is also a ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/863A61K39/00A61P35/00
CPCA61K2039/5256A61K2039/53A61P35/00A61K39/001184A61K39/001186A61K39/001188A61K2039/812A61K2039/82C12N15/86C12N2710/24043
Inventor 徐建青黄杨张晓燕郜明泉王璐孙甲浩
Owner VACDIAGN BIOTECH
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