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Characterization of a replicating expanded tropism oncolytic reovirus carrying the adenovirus E4orf4 gene

Gene Therapy volume 25, pages 331–344 (2018)Cite this article

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Abstract

While the mammalian orthoreovirus type 3 dearing (reovirus T3D) infects many different tumour cells, various cell lines resist the induction of reovirus-mediated cell death. In an effort to increase the oncolytic potency, we introduced transgenes into the S1 segment of reovirus T3D. The adenovirus E4orf4 gene was selected as transgene since the encoded E4orf4 protein induces cell death in transformed cells. The induction of cell death by E4orf4 depends in part on its binding to phosphatase 2A (PP2A). In addition to the S1-E4orf4 reovirus, two other reoviruses were employed in our studies. The reovirus rS1-RFA encodes an E4orf4 double-mutant protein that cannot interact with PP2A and the rS1-iLOV virus encoding the fluorescent marker iLOV as a reporter. The replacement of the codons for the junction adhesion molecule-A (JAM-A) binding head domain of the truncated spike protein blocks the entry of these recombinant viruses via the reovirus receptor JAM-A. Instead these viruses rely on internalization via binding to sialic acids on the cell surface. This expands their tropism and allows infection of JAM-A-deficient tumour cells. Here we not only demonstrate the feasibility of this approach but also established that the cytolytic activity of these recombinant viruses is largely transgene independent.

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Acknowledgements

We would like to thank Dr. Philip Branton for providing us with the plasmid pcDNA3-FLAG-B55a that we used for generating the 911-Flag.PP2A cell line. We are grateful to Dr. Geertje van der Horst (LUMC, Department of Urology) for providing the PC-3M-Pro4/Luc and UMUC-3 cell lines and Dr. Ruud Verrijk (Dr. Reddy’s Research & Development, Leiden, The Netherlands) for discussions on statistical methods. Also the Forensic Laboratory for DNA Research, of the LUMC department of Human Genetics is gratefully acknowledged for STR analyses of the human cell lines.

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  1. Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, The Netherlands

    Vera Kemp, Iris J. C. Dautzenberg, Steve J. Cramer, Rob C. Hoeben & Diana J. M. van den Wollenberg

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  1. Vera Kemp
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Correspondence to Diana J. M. van den Wollenberg.

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Kemp, V., Dautzenberg, I.J.C., Cramer, S.J. et al. Characterization of a replicating expanded tropism oncolytic reovirus carrying the adenovirus E4orf4 gene. Gene Ther 25, 331–344 (2018). https://doi.org/10.1038/s41434-018-0032-9

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