Vaccinia virus mutants useful for cancer immunotherapy

A vaccinia, virus strain technology, applied in the field of virology, immunotherapy, oncology, can solve the problem of the immune system is not activated

Pending Publication Date: 2020-05-05
MEMORIAL SLOAN KETTERING CANCER CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Despite the presentation of antigens by cancer cells and the presence of immune cells that could potentially respond to tumor cells, in many cases the immune system is either not activated or definitely suppressed

Method used

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  • Vaccinia virus mutants useful for cancer immunotherapy
  • Vaccinia virus mutants useful for cancer immunotherapy
  • Vaccinia virus mutants useful for cancer immunotherapy

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0182] Example 1: Vaccinia C7 inhibits IFN gene induction mediated by STING, TBK1 and IRF3.

[0183] The dual luciferase assay system was used to evaluate the role of vaccinia C7 in the regulation of STING, TBK1 or IRF3-induced IFNB promoter activation in HEK293T cells, which are human embryonic kidneys transformed with SV40 large T antigen Cell line. HEK293T cells were transfected with a plasmid expressing the IFNB firefly luciferase reporter, a control plasmid pRL-TK expressing Renilla luciferase, STING, and vaccinia C7L as indicated. A dual luciferase assay was performed 24h after transfection. By normalizing the firefly luciferase activity with respect to the Renilla luciferase activity, the relative luciferase activity is expressed in arbitrary units. Compared with the IFNB promoter activity in the control sample without STING, the overexpression of STING resulted in 30-fold induction of IFNB promoter activity. Co-transfection of increasing amounts of C7L expression plas...

Embodiment 2

[0184] Example 2: Vaccinia C7 inhibits poly I:C (TLR3) or TRIF-mediated IFN gene induction.

[0185] The TBK1-IRF3 axis is important for signal transduction from multiple sensing pathways, including cGAS-cGAMP-STING, RIG-I or MDA5-MAVS, TLR3-TRIF, and TLR4-TRIF. In order to test whether vaccinia C7 has the inhibitory effect of TRIF signaling, the inventors transfected HEK293T cells with TLR3 expression plasmid, IFN-β-luc reporter and increasing amounts of C7 expression plasmid (10ng, 50ng or 250ng). After 24h, the cells were treated with poly I:C (5 μg / ml). The luciferase activity was measured 24h after poly I:C treatment. Compared with the empty vector control, TLR3 transfection and poly I:C treatment resulted in a 9-fold induction of IFNB promoter activity ( Figure 2A ). The overexpression of C7 resulted in poly(I:C) / TLR3-induced reduction of IFNB promoter activity by up to 90% ( Figure 2A ). To test whether C7 also inhibits TRIF-induced IFNB promoter activity, HEK293T ce...

Embodiment 3

[0186] Example 3: Overexpression of vaccinia C7 in immune cells inhibits IFNB gene induction.

[0187] To evaluate the role of vaccinia C7 in the induction of IFNB gene in immune cells, we generated two cell lines stably expressing vaccinia C7, including murine macrophages RAW264.7 and human THP-1. THP-1 is a human monocytic leukemia cell line, which has been widely used to study the function and immune regulation of human monocytes and macrophages. Briefly, RAW264.7 and THP-1 were transduced with a retrovirus containing a vaccinia C7 expression construct under the CMV promoter and puromycin selection marker. An empty vector with a drug selection marker was also used to generate control cell lines. The drug-resistant cells were obtained and used in the following experiments. Before the THP-1 stable cell lines expressing C7 or with an empty vector were used in experiments, they were differentiated by phorbol-12-myristate-13-acetate (PMA; 20ng / ml) for 3 days. Cells were infecte...

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Abstract

Disclosed herein are methods and compositions related to the treatment, prevention, and/or amelioration of cancer in a subject in need thereof. In particular aspects, the present technology relates tothe use of poxviruses, including a recombinant modified vaccinia Ankara (MVA) virus or vaccinia virus with deletion of vaccinia host-range factor C7 (MVA[delta]C7L and VACV[delta]C7L, respectively),alone or in combination with immune checkpoint blocking agents, as an oncolytic and immunotherapeutic composition. In some embodiments, the technology of the present disclosure relates to a MVA[delta]C7L or VACV[delta]C7L virus further modified to express human Fms-like tyrosine kinase 3 ligand (Flt3L).

Description

[0001] Cross references to related applications [0002] This application claims priority from U.S. Provisional Application No. 62 / 505,713 filed on May 12, 2017, which is hereby incorporated by reference in its entirety. [0003] Federally Funded Research Statement [0004] The present invention was completed with the support of the US government under AI073736, AI095692, AR068118 and CA008748 granted by the National Institutes of Health. The government has certain rights in this invention. [0005] Sequence Listing [0006] This application contains a sequence listing electronically filed in ASCII format and hereby incorporated by reference in its entirety. The ASCII copy created on May 8, 2018 is named 115872-0781_SL.txt and has a size of 492,760 bytes. Technical field [0007] The technology of this disclosure relates generally to the fields of oncology, virology, and immunotherapy. Specifically, the technology of the present invention relates to the use of a poxvirus alone or in c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/285C12N15/863
CPCA61K39/39A61K35/768A61K2039/543A61K2039/572A61K2039/585C12N2710/24121C12N2710/24132C12N2710/24134A61K2039/575A61P35/00A61K39/285A61K2039/525A61K2039/54A61K2039/55588C12N7/00C12N15/86C12N2710/24131C12N2710/24143
Inventor L·邓S·舒曼N·杨T·梅古布J·沃尔乔克
Owner MEMORIAL SLOAN KETTERING CANCER CENT
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