Immunocompromised Cas9 transgenic mice for the rapid in vivo assessment of host factors involved in highly pathogenic virus infection

ABSTRACT: Targeting host factors for anti-viral development offers several potential advantages over traditional countermeasures that include broad-spectrum activity and prevention of resistance. Characterization of host factors in animal models provides strong evidence for their involvement in disease pathogenesis, yet the feasibility of performing high-throughput in vivo analysis on lists of genes is problematic. To begin addressing the challenges of screening candidate host factors in vivo, we combined advances in CRISPR-Cas9 genome editing with an immunocompromised mouse model used to study highly pathogenic viruses. Transgenic mice harboring a constitutively-expressed Cas9 allele (Cas9tg/tg), with or without a knockout for type I interferon receptors, served to optimize the in vivo delivery of CRISPR single-guide RNA (sgRNA) using Invivofectamine 3.0, a simple and easy-to-use lipid nanoparticle reagent. Invivofectamine 3.0-mediated liver-specific editing to remove activity of the critical Ebola virus host factor, Niemann-pick disease type C1, in an average of 74% of liver cells protected immunocompromised Cas9tg/tg mice from lethal surrogate Ebola virus infection. We envision immunocompromised Cas9tg/tg mice combined with straightforward sgRNA in vivo delivery will enable efficient host factor loss-of-function screening in liver and other organs to rapidly study their effects on viral pathogenesis and help initiate broad-spectrum host-directed therapeutic development against emerging pathogens.