Abstract
Oncolytic viruses which infect and kill tumour cells can also be genetically modified to express therapeutic genes that augment their anti-cancer activities. Modifying oncolytic viruses to produce effective cancer therapies is challenging as encoding transgenes often attenuates virus activity or prevents systemic delivery in patients due to the risk of off-target expression of transgenes in healthy tissues. To overcome these issues we aimed to generate a readily modifiable virus platform using the oncolytic adenovirus, enadenotucirev. Enadenotucirev replicates in human tumour cells but not cells from healthy tissues and can be delivered intravenously because it is stable in human blood. Here, the enadenotucirev genome was used to generate plasmids into which synthesised transgene cassettes could be directly cloned in a single step reaction. The platform enabled generation of panels of reporter viruses to identify cloning sites and transgene cassette designs where transgene expression could be linked to the virus life cycle. It was demonstrated using these viruses that encoded transgene proteins could be successfully expressed in tumour cells in vitro and tumours in vivo. The expression of transgenes did not impact either the oncolytic activity or selective properties of the virus. The effectiveness of this approach as a drug delivery platform for complex therapeutics was demonstrated by inserting multiple genes in the virus genome to encode full length anti-VEGF antibodies. Functional antibody could be synthesised and secreted from infected tumour cells without impacting the activity of the virus particle in terms of oncolytic potency, manufacturing yields or selectivity for tumour cells. In vivo, viral particles could be efficaciously delivered intravenously to disseminated orthotopic tumours.
Highlights
The first oncolytic virus therapy has been approved in the USA and Europe for the treatment of cancer
These 3’ regions were selected for the insertion of transgene cassettes because they are in non-coding regions of the enadenotucirev genome and located downstream of endogenous promoters responsible for modulating genes controlling the virus life cycle and capsid assembly
An oncolytic virus platform for therapeutic gene delivery to tumours between the linearised pColoAd2.4 shuttle vector and the enadenotucirev linear genome to generate the pColoAd2.4 vector. (G) Schematics showing how transgene cassettes are directly ligated from a sub cloning vector (‘vector 1’) into the pColoAd2.4 vector between AsiSI and SbfI sites
Summary
The first oncolytic virus therapy has been approved in the USA and Europe for the treatment of cancer. Oncolytic viruses were originally developed for their ability to infect, replicate and directly kill human tumour cells while being attenuated in normal cells [6] Their proposed primary mechanism of action was that locally delivered virus particles would rapidly spread and destroy tumours while having limited off-target activity in surrounding healthy tissue. Immune mediated regression has been observed in uninjected distal tumours, which is thought to be mediated by an induced anti-tumour immune response [12, 13] These data have led to oncolytic viruses emerging as a novel class of agent in an expanding group of cancer therapies known collectively as immuno-therapies
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