An iPSC-derived human organoid based model of intestinal filovirus infection

Abstract Filoviruses are prime examples of zoonotic viruses that cause fulminant hemorrhagic diseases in humans and nonhuman primates (NHP), resulting in death in up to 90% of human case. The filoviridae family contains six genera in which several species are known to have high fatality rates, all of which are within the Marburgvirus or Ebolavirus genera and will therefore be the focus of this proposal. The largest ever 2014 Ebola virus (EBOV) disease outbreak in West Africa underscored the development and regulatory approval of antiviral countermeasures, including an emergency-use vaccine and therapeutic monoclonal antibodies. While these countermeasures are highly beneficial to block EBOV infection and interfere with disease progression at early stages, they are less suitable for the treatment of late-stage EBOV disease (EVD), which is less studied. Late-stage disease is characterized by gastrointestinal manifestations including diarrhea. At present, no available animal models, including the classical NHP model, can recapitulate the gastrointestinal symptoms of EVD patients. To elucidate the role of the intestinal epithelium in disease initiation and progression, we propose to establish an intestinal filovirus infection model using human induced pluripotent stem cells (hiPSC)-derived human intestinal organoids (HIOs). We hypothesize that infection with EBOV and MARV compromises barrier integrity of the intestinal epithelium and causes abnormal ion flux as the basis for gastrointestinal dysfunction and diarrhea. This project proposes the establishment of a polarized 2-dimentional HIO transwell system that will be used to study the pathophysiological effects of filovirus infection on intestinal epithelial integrity. In Aim 1, we will characterize filovirus-mediated injury by analyzing (1) intestinal barrier and (2) tight junction integrity. We hypothesize that the sepsis-like syndrome that occurs in some patients with severe disease may be due to loss of epithelial barrier integrity in the intestine. In Aim 2, we intend to validate the role of individual genes by genetic ablation and test potential drug candidates as modulators of intestinal epithelial function. We hypothesize that if tight junctions are compromised in infected HIOs, then treatment with small molecule inhibitors targeted at ion exchanger would reverse the leaky junctions by blocking the receptor responsible for the disassemble of the junction. The aims proposed in this study will result in an improved model which more closely recapitulates in vivo intestinal epithelial phenotypes, and thus can be used to inform novel therapeutics targeted at late-stage filovirus infection. The successful, longstanding mentorship of Drs. Mostoslavsky and Mühlberger, the collaborative support of the CReM, the availability of the NEIDL research facilities and the rigorous MTM training program will provide me with the tools and support needed to successfully complete my dissertation work. These departments instill important values about the rigorous approach to academic science by supporting technical training, writing workshops and seminar and lecture series.