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Recombinant herpes simplex virus and application thereof

A virus and chimeric virus technology, applied in the field of virology and tumor treatment, can solve the problems of replicating and killing tumor cells that have not yet been found

Active Publication Date: 2018-09-25
XIAMEN UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although scientists have been making various attempts, so far, no oncolytic virus has been found that can replicate and kill tumor cells at a high level in tumor cells without causing serious side effects on normal cells

Method used

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  • Recombinant herpes simplex virus and application thereof
  • Recombinant herpes simplex virus and application thereof
  • Recombinant herpes simplex virus and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0205] Embodiment 1. Construction of recombinant virus OVN and OVH

[0206] (1.1) Culture and titer determination of herpes simplex virus type I (HSV-1)

[0207] The wild-type HSV-1 virus strain KOS was purchased from ATCC Company of the United States (product number VR-1493 TM ), whose genome information has been published in NCBI (GenBank: JQ673480.1). Under the condition of MOI=0.1, the Vero cell (purchased from ATCC Company of the United States, product number CCL-81) was infected with the virus strain KOS TM ). After 48 hours, all cells were collected with a cell scraper and centrifuged to remove cell culture medium. The obtained cell pellet was resuspended in fresh complete medium and stored at -80°C. Subsequently, the cell suspension was repeatedly frozen and thawed (3 times), and then centrifuged to collect the supernatant to obtain the virus liquid. Aliquot the virus solution and store at -80°C.

[0208] Take 1×10 6 The density of the cells, the U-2 OS cells (p...

Embodiment 2

[0229] Example 2. Characterization of recombinant viruses OVN and OVH

[0230] Referring to the method described in Example 1, a recombinant virus dICP0 was constructed, which, compared with the virus strain KOS, lacked two copies of the ICPO gene. Virus strain KOS and recombinant virus dICP0 were used as control viruses to characterize recombinant viruses OVN and OVH. image 3 Compared with the virus strain KOS, the genome modifications contained in the recombinant viruses OVN, OVH and dICP0 are schematically shown; among them, compared with the virus strain KOS, the recombinant virus dICP0 has lost two copies of the ICPO gene; the recombinant virus OVN has lost two copies. Copy of the ICP34.5 gene and ICPO gene; the recombinant virus OVN deleted the double copy of the ICP34.5 gene and the ICPO gene, and the native promoter of the ICP27 gene was replaced by the hTERT core promoter.

[0231] The gene deletion in the recombinant viruses OVN, OVH and dICP0 was verified by PCR m...

Embodiment 3

[0238] Example 3. Evaluation of Replication Ability and Killing Ability of Recombinant Virus OVN / OVH

[0239] Take 5-7.5×10 6Normal cells (L-O2 cells) and tumor cells (U-2 OS cells) in good condition and in the logarithmic growth phase were inoculated in a 6 cm culture plate. Subsequently, cultured cells were infected with viruses KOS, OVN, OVH or dICP0 at a multiplicity of infection of MOI=1. After 48 hours of infection, the state of the cells was observed under a microscope, and photographed and recorded. Subsequently, the virus-infected cells were digested, and the viability of the cells was calculated by trypan blue staining. Cell viability (%)=(number of living cells after virus infection) / (number of control cells not infected with virus)×100. Each group of experiments was set up to repeat in 3 wells, and the experimental results were the average of 3 independent experiments. In addition, referring to the protocol described in Example 1, virus titers at different time...

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Abstract

The invention relates to the field of virology and tumor therapy and particularly provides a recombinant HSV (herpes simplex virus). The recombinant HSV is capable of specifically duplicating and killing tumor cells in tumor cells by a high level but duplicating in normal cells by a low level, so that the recombinant HSV is not only high in tumor cell lethality but also remarkably reduced in sideeffect (especially neurotoxicity). Further, the invention relates to a viral vector constructed on the basis of the recombinant herpes simplex virus, a pharmaceutical composition containing the recombinant herpes simplex virus or the viral vector and application of the recombinant herpes simplex virus or the viral vector. The recombinant herpes simplex virus can be used for infecting and killing the tumor cells and delivering gene medicines into the tumor cells to realize gene therapy.

Description

technical field [0001] The invention relates to the fields of virology and tumor therapy. In particular, the present invention provides a recombinant herpes simplex virus (Herpes Simplex Virus, HSV), which can specifically replicate at a high level in tumor cells and effectively kill tumor cells, but replicate at a low level in normal cells, Therefore, the recombinant herpes simplex virus of the present invention not only has high lethality to tumor cells, but also has significantly reduced side effects (especially neurotoxicity). Further, the present invention relates to a viral vector constructed based on the recombinant herpes simplex virus, a pharmaceutical composition comprising the recombinant herpes simplex virus or the viral vector, and uses of the recombinant herpes simplex virus or the viral vector. The recombinant herpes simplex virus of the present invention can be used to infect and kill tumor cells, and can be used to deliver gene medicine into tumor cells for g...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/01C12N15/869C12N5/10A61P35/00
CPCA61K35/763C12N7/00C12N15/86C07K14/035C12N2710/16643C12N2710/16621A61P35/00C12N5/10C07K14/005C12N2710/16622C12N2710/16632A01K2207/12A01K2227/105A01K2267/0331C12N2710/16651
Inventor 黄承浩罗勇袁权张军夏宁邵
Owner XIAMEN UNIV
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