Frontiers in Immunology (Jan 2023)
CRISPR-mediated rapid arming of poxvirus vectors enables facile generation of the novel immunotherapeutic STINGPOX
- Jack T. Whelan,
- Jack T. Whelan,
- Ragunath Singaravelu,
- Ragunath Singaravelu,
- Ragunath Singaravelu,
- Fuan Wang,
- Fuan Wang,
- Adrian Pelin,
- Adrian Pelin,
- Levi A. Tamming,
- Levi A. Tamming,
- Giuseppe Pugliese,
- Nikolas T. Martin,
- Nikolas T. Martin,
- Mathieu J. F. Crupi,
- Mathieu J. F. Crupi,
- Julia Petryk,
- Bradley Austin,
- Xiaohong He,
- Ricardo Marius,
- Ricardo Marius,
- Jessie Duong,
- Jessie Duong,
- Carter Jones,
- Emily E. F. Fekete,
- Emily E. F. Fekete,
- Nouf Alluqmani,
- Nouf Alluqmani,
- Andrew Chen,
- Stephen Boulton,
- Stephen Boulton,
- Michael S. Huh,
- Matt Y. Tang,
- Zaid Taha,
- Zaid Taha,
- Elena Scut,
- Elena Scut,
- Jean-Simon Diallo,
- Jean-Simon Diallo,
- Taha Azad,
- Taha Azad,
- Brian D. Lichty,
- Brian D. Lichty,
- Carolina S. Ilkow,
- Carolina S. Ilkow,
- John C. Bell,
- John C. Bell
Affiliations
- Jack T. Whelan
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Jack T. Whelan
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Ragunath Singaravelu
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Ragunath Singaravelu
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Ragunath Singaravelu
- Public Health Agency of Canada, Ottawa, ON, Canada
- Fuan Wang
- McMaster Immunology Research Centre, Department of Medicine, McMaster University, Hamilton, ON, Canada
- Fuan Wang
- MG DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, ON, Canada
- Adrian Pelin
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Adrian Pelin
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Levi A. Tamming
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Levi A. Tamming
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Giuseppe Pugliese
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Nikolas T. Martin
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Nikolas T. Martin
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Mathieu J. F. Crupi
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Mathieu J. F. Crupi
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Julia Petryk
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Bradley Austin
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Xiaohong He
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Ricardo Marius
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Ricardo Marius
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Jessie Duong
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Jessie Duong
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Carter Jones
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Emily E. F. Fekete
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Emily E. F. Fekete
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Nouf Alluqmani
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Nouf Alluqmani
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Andrew Chen
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Stephen Boulton
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Stephen Boulton
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Michael S. Huh
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Matt Y. Tang
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Zaid Taha
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Zaid Taha
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Elena Scut
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Elena Scut
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Jean-Simon Diallo
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Jean-Simon Diallo
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Taha Azad
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Taha Azad
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- Brian D. Lichty
- McMaster Immunology Research Centre, Department of Medicine, McMaster University, Hamilton, ON, Canada
- Brian D. Lichty
- MG DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, ON, Canada
- Carolina S. Ilkow
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- Carolina S. Ilkow
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- John C. Bell
- Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON, Canada
- John C. Bell
- Centre for Innovation Cancer Therapeutics, Ottawa Hospital Research Institute, Ottawa, ON, Canada
- DOI
- https://doi.org/10.3389/fimmu.2022.1050250
- Journal volume & issue
-
Vol. 13
Abstract
Poxvirus vectors represent versatile modalities for engineering novel vaccines and cancer immunotherapies. In addition to their oncolytic capacity and immunogenic influence, they can be readily engineered to express multiple large transgenes. However, the integration of multiple payloads into poxvirus genomes by traditional recombination-based approaches can be highly inefficient, time-consuming and cumbersome. Herein, we describe a simple, cost-effective approach to rapidly generate and purify a poxvirus vector with multiple transgenes. By utilizing a simple, modular CRISPR/Cas9 assisted-recombinant vaccinia virus engineering (CARVE) system, we demonstrate generation of a recombinant vaccinia virus expressing three distinct transgenes at three different loci in less than 1 week. We apply CARVE to rapidly generate a novel immunogenic vaccinia virus vector, which expresses a bacterial diadenylate cyclase. This novel vector, STINGPOX, produces cyclic di-AMP, a STING agonist, which drives IFN signaling critical to the anti-tumor immune response. We demonstrate that STINGPOX can drive IFN signaling in primary human cancer tissue explants. Using an immunocompetent murine colon cancer model, we demonstrate that intratumoral administration of STINGPOX in combination with checkpoint inhibitor, anti-PD1, promotes survival post-tumour challenge. These data demonstrate the utility of CRISPR/Cas9 in the rapid arming of poxvirus vectors with therapeutic payloads to create novel immunotherapies.
Keywords
- CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9)
- vaccinia virus (VACV)
- STING agonist
- poxvirus
- oncolytic virus
