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Virus vector for prime/boost vaccines, which comprises vaccinia virus vector and sendai virus vector

A technology of vaccinia virus vector and virus vector, which is applied in the direction of virus/bacteriophage, medical preparations containing active ingredients, viruses, etc., and can solve the problem that it is difficult to completely eliminate side effects

Inactive Publication Date: 2013-07-03
HOKKAIDO UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] From the point of view of imparting strong immunity, it is expected that the vaccine can activate both cellular immunity and humoral immunity. Live vaccines can activate both and are effective in obtaining strong immunity and long-lasting immunity. However, due to As the attenuated pathogen, the pathogenic microorganism itself is given to the organism, and it is difficult to completely eliminate the hidden dangers such as side effects caused by the infection of the pathogenic microorganism, strong toxicity due to the reversion of the attenuated pathogenic microorganism (reversion or reversion to the ancestor), etc.

Method used

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  • Virus vector for prime/boost vaccines, which comprises vaccinia virus vector and sendai virus vector
  • Virus vector for prime/boost vaccines, which comprises vaccinia virus vector and sendai virus vector
  • Virus vector for prime/boost vaccines, which comprises vaccinia virus vector and sendai virus vector

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0079] (1) Preparation of vaccinia virus vector carrying the gene encoding human immunodeficiency virus envelope protein

[0080] [1-1] Preparation of m8Δ--env

[0081] Preparation of env gene

[0082] pJW322-env was obtained by inserting the genes encoding the envelope proteins gp160 and gp120 (env) of the human immunodeficiency virus HIV-1 JR-CSF strain (accession number M38429) into the Avr II / Xho I site of pJW322. Then, in order to efficiently express env, the transcription termination sequences of vaccinia virus present at positions 6751 to 6757, 7367 to 7373, and 8305 to 8311 in the env gene were subjected to in vitro mutagenesis (in vitro mutagenesis) method, mutated as described below, and this was designated as pJW322-env2. Here, the amino acid sequence of env was not changed by this mutation.

[0083] 6751st to 6757th

[0084] Before mutation: TTTTTAT (sequence number 1)

[0085] After mutation: TTTCTAT (SEQ ID NO: 2)

[0086] 7367th to 7373rd

[0087] Before...

Embodiment （1

[0124] For the m8Δ--hCD40Lm of the present embodiment (1) [1-2], it is described in the present embodiment (1)[1-1] Western blotting was performed according to the method, and the expression of hCD40Lm was confirmed. Only 10 μg of cells used for electrophoresis was used instead of 1 μg. In the detection of hCD40Lm, anti-CD40L mouse monoclonal antibody was used instead of serum from HIV-1 infected patients. The results are shown in figure 2 .

[0125] Large-scale cultivation of vaccinia virus vector

[0126] By the method described in this example (1) [1-1] , the m8Δ--hCD40Lm of this example (1) was mass-cultured and purified Concentrate and determine the virus titer.

[0127] Preparation of [1-3]m8Δ--env-hCD40Lm

[0128] Preparation of plasmid

[0129] Using the pJW322-env2 of this example (1) [1-1] as a template, PCR was performed with the following primers to amplify and isolate the env gene.

[0130] Forward primer:

[0131] 5'-CTAGAATTCGCCACCATGAGAGTGAAGGGGATCAG...

Embodiment （1）

[0150] For the m8Δ--env-hCD40Lm of this embodiment (1) [1-3], through this embodiment (1)[1-1] ELISA was performed according to the method described in , and the expression of env was confirmed in the plaques. In addition, for m8Δ--env-hCD40Lm of this embodiment (1) [1-3], through this embodiment (1)[1-1] and the method described in this example (1) [1-2] performed western blotting to confirm the expression of env and hCD40Lm. The results are shown in figure 2 .

[0151] Large-scale cultivation of vaccinia virus vector

[0152] By the method described in this example (1) [1-1] , the m8Δ- >-env-hCD40Lm was purified and concentrated, and the virus titer was determined.

[0153] [1-4] Production of m8Δ--hCD40Lm

[0154] Preparation of plasmid

[0155] Using pCA-hCD40Lm3 inserted with the hCD40Lm gene as a template, PCR was performed with the following primers to amplify and isolate the hCD40Lm gene.

[0156] Forward primer: 5'-AAACCCGGGCATGATCGAAACATACAACCAAA-3' (SEQ ID...

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Abstract

To provide a virus vector which can be used for producing a prime / boost vaccine that can activate both cellular immunity and humoral immunity and is effective on infections by pathogenic microorganisms and malignant tumors which are generally believed to be difficult to be treated by vaccine therapy. Provided is a virus vector for prime / boost vaccines, comprising the following components (a) and (b): (a) a vaccinia virus vector which carries a gene encoding an immunogenic polypeptide in such a manner that the gene can be expressed; and (b) a Sendal virus vector which carries the gene encoding the immunogenic polypeptide in such a manner that the gene can be expressed.

Description

technical field [0001] The present invention relates to a viral vector for a prime-boost vaccine, in particular to a viral vector for a prime-boost vaccine that can be used to activate either cellular immunity or humoral immunity . Background technique [0002] Human beings are exposed to the danger of infection by viruses, bacteria, fungi and other various organisms and the infectious diseases caused by them. One way to overcome these infectious diseases is to get vaccinated. Vaccines are roughly divided into live vaccines using pathogenic microorganisms such as attenuated bacteria and viruses themselves, and inactivated vaccines using pathogenic microorganisms killed by chemical treatment or the like or parts thereof that express antigenicity. [0003] From the point of view of imparting strong immunity, it is expected that the vaccine can activate both cellular immunity and humoral immunity. Live vaccines can activate both and are effective in obtaining strong immunity ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/09
CPCA61K39/21A61K2039/545C12N2710/24143C12N2740/16034C12N2760/18843C12N15/86A61K2039/53C12N2740/16134A61K39/12Y02A50/30A61K39/39
Inventor 志田寿利祖父江友好加藤和则井上诚长谷川护
Owner HOKKAIDO UNIVERSITY
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