Yeast virus-derived stimulator of the innate immune system augments the effiacacy of a viral-vector based immunotherapeutics
M.-C. Claudepierrea, et al.
Journal of Virology, February 2014
To identify novel stimulators of the innate immune system, we constructed a panel of eight HEK293-cells lines, double-positive for human Toll-like receptors (TLR) and a NF-κB-inducible reporter gene. Screening a large variety of compounds and cellular extracts detected a TLR3 activating compound in a microsomal yeast extract. Fractionation of this extract identified a RNA molecule of 4.6 kb, named Nucleic Acid Band 2 (NAB2) that was sufficient to confer the activation of TLR3. Digests with single- and double-strand-specific RNases showed the double-strand nature of this RNA, and its sequence was found to be identical to the genome of the dsRNA L-BC virus of Saccharomyces cerevisiae. A large scale production and purification process of this RNA was established based on chemical cell lysis and dsRNA-specific chromatography. NAB2 complexed with the cationic lipid Lipofectin, but neither NAB2 nor Lipofectin alone, induced the secretion of IL-12(p70), IFNα, IP-10, Mip-1β and IL-6 in human monocyte-derived dendritic cells. While NAB2 activated TLR3, Lipofectin-stabilized NAB2 signaled also via the cytoplasmic sensor for RNA recognition MDA-5. Significant increase of RMA-MUC1 tumor rejection and survival was observed in C57BL/6 mice after prophylactic vaccination with MUC1-encoding MVA and NAB2+Lipofectin. This combination of immunotherapeutics strongly increased the percentage of infiltrating Natural Killer (NK) cells and plasmacytoid dendritic cells (pDCs) at the injection sites, cell types which can modulate innate and adaptive immune responses.