Novel oncolytic adenovirus EM/VSV-G Ad5-P and application thereof in preparation of antitumor drugs

A VSV-G, oncolytic virus technology, applied in the field of tumor immunotherapy and tumor treatment, to achieve high expression and ensure the effect of anti-tumor treatment

Pending Publication Date: 2021-12-24
NANJING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Exosome-mimetic technology is currently only used to wrap drugs for targeted delivery 42-44 , so far there has been no report on the use of its packaged virus
Moreover, it is widely believed that the technique is unlikely to be useful for encapsulating therapeutic entities as large as viruses

Method used

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  • Novel oncolytic adenovirus EM/VSV-G Ad5-P and application thereof in preparation of antitumor drugs
  • Novel oncolytic adenovirus EM/VSV-G Ad5-P and application thereof in preparation of antitumor drugs
  • Novel oncolytic adenovirus EM/VSV-G Ad5-P and application thereof in preparation of antitumor drugs

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] Example 1 Infection efficiency of adenovirus Ad5 to cells with low CAR expression is low

[0071] CAR (the coxsackie and adenovirus receptor) is the main receptor of adenovirus Ad5 infected cells. Early studies have demonstrated that adenovirus Ad5 has low infection efficiency for cells with low CAR expression. 23,29 We found that the fluorescence intensity of 293T, A549, HCC-LM3 and Hepa1-6 cells stained with anti-CAR-PE was significantly increased compared with the isotype treatment group by flow cytometric analysis, indicating that 293T, A549, HCC-LM3 and Hepa1-6 cells High expression of CAR on the cell surface ( figure 1 a); while the surface of B16-F10, CT26.WT and H22 cells were stained with CAR antibody, there was no significant change in fluorescence intensity compared with the isotype treatment group; indicating that these cell lines do not express CAR; while K562 and Jurkat cells were stained with CAR antibody After that, compared with the isotype treatment ...

Embodiment 3

[0072] Example 3 EM / VSV-G Ad5-GFP can resist the effect of anti-Ad5 neutralizing antibody and the carried VSV-G mediates its entry into cells

[0073] To further prove that EM / VSV-G Ad5-GFP is encapsulated in vesicles, and that such encapsulation brings new properties, we used anti-Ad5 neutralizing antibody and VSV-G neutralizing serum for EM / VSV- G Ad5-GFP was processed for subsequent analysis. After treating EM / VSV-G Ad5-GFP and Ad5-GFP viruses with anti-Ad5 neutralizing antibodies at dilutions of 1 / 32, 1 / 64, and 1 / 128, the infection efficiency of Ad5-GFP virus was only that of the PBS-treated control group 5.66±2.1%, 9.3±2.02% and 12.6±3.84%, while the infection efficiency of EM / VSV-G Ad5-GFP was 61.0±3.10%, 71.3±3.7% and 81.6±1.2% ( figure 2 f) It can be seen that after treatment with anti-Ad5 neutralizing antibody, the Ad5-GFP virus almost lost its ability to infect, while the EM / VSV-G Ad5-GFP was infected by the exosome-like outer membrane and the presence of VSV-G. C...

Embodiment 4

[0074] Example 4 EM / VSV-G Ad5-GFP has high infection efficiency to cell lines with low CAR expression

[0075]Previous studies have found that adenovirus Ad5 has a low infection efficiency for cells with low CAR expression. Here we want to observe through experiments whether EM / VSV-G Ad5-GFP virus is different from Ad5-GFP virus and has the same effect on cell lines with low CAR expression. High infection efficiency. Here we tested the infection efficiency of EM / VSV-G Ad5-GFP and Ad5-GFP viruses on cells with low CAR expression. Surprisingly, compared with Ad5-GFP, the infection efficiency of EM / VSV-G Ad5-GFP was extremely significantly improved in CAR low-expressing cell lines B16 / F10, CT26.WT, H22, K562 and Jurkat increased to 8.00±0.54 times, 2.83±0.04 times, 14.99±1.09 times, 11.35±0.37 times and 6.25±0.58 times ( image 3 a,b). In addition to detecting the infection efficiency of EM / VSV-G Ad5-GFP and Ad5-GFP viruses on CAR low-expressing cells, we also detected the inf...

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Abstract

The invention relates to the field of tumor treatment, in particular to application of oncolytic adenovirus EM / VSV-G Ad5-P in preparation of antitumor drugs. The invention discloses an antitumor effect of the oncolytic adenovirus EM / VSV-G Ad5-P in vivo and in vitro. The exosome-like coated oncolytic adenovirus EM / VSV-G Ad5-P, hereinafter referred to as a novel virus, is prepared by using cells expressing VSV-G and utilizing an exosome-like technology. The novel virus realizes heavy targeting through VSV-G, infects a CAR low-expression cell line, increases the gene transduction efficiency of the virus, can resist the neutralization effect of a virus antibody, and also can increase the virus yield. In-vitro research finds that the infection efficiency, the oncolytic ability and the PD-1 expression quantity of the novel virus are remarkably improved in a CAR low-expression cell line. In-vivo experiments show that the novel virus can resist neutralization of an Ad5 antibody, prolong a virus treatment window and continuously express PD-1, so that the novel virus can remarkably activate anti-tumor immunity and finally remarkably prolong survival of mice.

Description

technical field [0001] The invention relates to the field of tumor treatment, especially the field of tumor immunotherapy, and in particular to the application of an oncolytic adenovirus EM / VSV-G Ad5-P in the preparation of antitumor drugs. Background technique [0002] Malignant tumors are still an important disease that threatens human health worldwide 1 . In recent years, tumor immunotherapy has made remarkable achievements, targeting immune checkpoint receptors, such as cytotoxic T-lymphocyte-associated protein 4 (CTLA4), programmed cell-death receptor 1 (programmed cell-death protein 1PD -1) and programmed cell-death ligand 1PD-L1 tumor immunotherapy is a revolutionary advance in tumor treatment 2 . In particular, anti-PD1 / PDL1 antibody therapy has been shown to be effective in at least 15 types of tumors 3-6 , several PD-1 / PD-L1 inhibitors have been approved for clinical treatment worldwide 6 . With the in-depth research and numerous clinical trials, it has been ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/01A61K35/761A61P35/00A61P35/02
CPCC12N7/00C07K14/70521A61K35/761A61P35/00A61P35/02C12N2710/10021
Inventor 魏继武吴俊华张永辉吴俊艺张海林刘淑雯马丁
Owner NANJING UNIV
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