Ebola Virus Life Cycle Modeling In Vivo

ABSTRACT
Sporadic outbreaks of Ebola virus (EBOV) and related filoviruses pose a grave threat for worldwide health
populations, especially those of Western African nations. The largest outbreak of EBOV in 2013 caused nearly
13,000 deaths and had case fatality rates of up to 90%. Despite its near half-century since its emergence in
human populations, there is only one vaccine and only two recently approved therapeutic treatments for Ebola
virus disease (EVD). Detailed molecular study of EBOV biology is necessary to rapidly advance antiviral
development. The largest impediment to detailed molecular study of EBOV is the requirement for high
containment facilities when handling infectious virus. As an alternative, many groups utilize the well-established
life-cycle modeling of both minigenome and transcription and replication virus-like particles (trVLP) systems to
assess molecular viral mechanisms under biosafety level-2 (BSL-2) conditions. To this end, we have generated
a penta-cistronic minigenome (5XMG) construct that contains four of the viral open reading frames and a reporter
gene. The minigenome system remains BSL-2 due to its inability to replicate unless in the presence of the
replication “helper” proteins: NP, VP30, VP35, and L. While this is valuable as a safety mechanism, it inhibits the
use of EBOV modeling in animals. It would be invaluable to have a murine model in which to study the dynamics
of EBOV under BSL-2 conditions in vivo. The proposed study will generate two mouse models for studying EBOV
dynamics using trVLPs by supplementing the necessary helper proteins in trans. We would achieve this through
two specific aims: (Aim 1) In Vivo EBOV Life Cycle Modeling with trVLPs via EBOV Helper Protein Expression
by mRNA-Lipid Nanoparticles (LNPs); (Aim 2) In Vivo EBOV Life Cycle Modeling with trVLPs Supported by
EBOV Helper Protein Expression via Transgenesis. In this model, the proteins missing from our poly-cistronic
minigenome (EBOV VP30, VP35, and L) would be expressed in mice via either mRNA encapsulated LNPs or
as integrated, Cre-recombinase dependent mouse transgenes. Expression of the viral helper proteins will
replicate our 5XMG to amplify and create additional trVLPs that would bud off, infecting additional cells in the
animal’s tissues. Importantly, the proposed system would be a self-contained model for infection with
minigenome-containing trVLPs but will not generate any infectious virus. These tools can have many applications
including the assessment of antiviral host responses as well as being a platform for testing antiviral therapies
like vaccines and small molecule inhibitors.

Grant Number: 1R21AI188373-01
NIH Institute/Center: NIH
Principal Investigator: Michael Barry