Here, we report on genetically engineered, propagation-incompetent influenza A virus (IAV) particles, so-called defective interfering particles (DIPs) that have been suggested as a promising novel antiviral agent. Typically, IAV DIPs harbor a large internal deletion in one of their eight genomic viral RNA (vRNA) segments. Further, DIPs are capable of hijacking cellular and viral resources upon co-infection with fully infectious standard virus (STV), resulting in an antiviral effect. Besides this replication interference, DIP infection also stimulates innate immunity, adding to the antiviral efficacy.
So far, DIPs were produced in embryonated chicken eggs. To improve scalability and flexibility of processes as well as to increase product quality, we established a cell culture-based DIP production system [1,2]. This includes the development of a genetically engineered virus-cell propagation system that allows production of DIPs without the need to add infectious STV to complement missing gene functions of DIPs. Specifically, the MDCK suspension cell line generated expresses the PB2 protein [2], encoded by segment 1 (S1) of IAV, which is not expressed by “DI244” - a prototypic, well-characterized DIP harboring a deletion in S1. Using this cell culture-based production process in batch [2,3] and perfusion mode [4] at laboratory scale, we show that we can achieve very high DI244 titers of up to 2.6E+11 DIPs/mL. Infections of mice demonstrated that intranasal administration of the produced DI244 material resulted in no apparent toxic effects and in a full rescue of mice co-treated with an otherwise lethal dose of IAV [2].
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