Development of Immunotherapeutics for Heartland Virus

Peer Reviewed Medical Research Program (PRMRP) Portfolio: Infectious Diseases Fiscal Year (FY22) PRMRP Topic Area: Viral Diseases FY22 PRMRP Strategic Goal: Prevention: Develop or optimize vaccine strategies, platforms, or compounds, to include active or passive immunoprophylaxis, especially for Dengue, Lassa, and Crimean-Congo Hemorrhagic fever viruses, beta-coronaviruses. Heartland virus (HRTV) is an emerging tick-born disease of significant concern to public health due to the severity of disease and the geographic expansion of its vector. Immunotherapeutics have been shown to be beneficial for treatment of a wide variety of viral infections including SARS-CoV-2, MERS-CoV, Ebola, HIV, Influenza RSV, and Zika. However, to date, none have been developed for HRTV. We believe that HRTV is a good candidate for immunotherapeutic treatment. Here we propose to develop therapeutic monoclonal antibodies to HRTV. HRTV Gn and Gc surface glycoproteins will be expressed in Expi293F cells, and the proteins will be purified using Ni-NTA Agarose. Purified proteins will then be used to inoculate BALB/c mice, and hybridoma cell lines will be generated for production of monoclonal antibodies. Once developed, antibodies will be validated by ELISA; antibodies that display strong binding will then be tested for sensitivity and specificity. Selected antibodies will also be tested for antibody-mediated virus neutralization. Here we will use both the standard PRNT assay as well as previously developed pseudo-typed virus-based assays to quantify neutralization. Neutralizing and binding (but non-neutralizing) antibodies will be mapped in order to identify the optimal targets for virus neutralization. At this stage, we will also evaluate cocktails of neutralizing antibodies in order to maximize their therapeutic effect. Once developed, neutralizing antibodies will be tested using a previously developed AG129 mouse model of HRTV infection. Mice will be inoculated with HRTV, followed by intraperitoneal. administration of mAB or mAB cocktails and monitored for protection against a lethal challenge of HTRV. Future studies will focus on humanizing candidate antibodies for therapeutic use. Less