PrePan

RNA respiratory viruses like Influenza or Coronavirus are the causative agents of pandemics in the 21st century. Preventing these pandemics is one of the greatest challenges for human health. Vaccines are the best cost-effective therapeutical option and usually rely on the viral surface glycoprotein (Spike glycoprotein in Coronavirus and Hemagglutinin in Influenza virus). However, viral glycoproteins accumulate most of the RNA viruses' escape mutations, threatening the vaccines' effectiveness. Therefore, broad-spectrum vaccines that neutralize known and emerging viral strains are urgently required. Here, we propose an alternative approach using conserved structural proteins with slower mutational rates, such as viral nucleoproteins (NPs), as targets for medical intervention. NPs are located inside the virus particles, encapsidating the viral RNA genome by combining two activities: self-polymerization and RNA interaction. In addition to protecting the viral genome from degradation, viral NPs serve as molecular machines for viral replication and transcription. NPs are highly conserved across Influenza and Coronavirus families and are the most abundant protein in infected cells. NPs are ideal targets for antivirals, but they were traditionally not considered antigens for vaccines because of their location inside the viral particle "hidden" from the immune system. However, recent discoveries from our lab and others demonstrated that NPs also regulate the innate and adaptative immune response through distinct mechanisms: (i) strong T cell activation by digested peptides exposed in the cell surface of the infected cells, (ii) direct binding to inflammatory chemokines in the cell surface of infected cells, and (iii) specific monoclonal antibodies (mAbs) for viral NPs that reduce inflammation and pathology in the lung. This project offers the opportunity to generate universal treatments that target conserved internal proteins of the virus that provide a more mutation-resistant therapeutic option for current epidemics and expected future waves of Coronaviruses and Influenza viruses. Here, we describe a stepwise method using purified NPs to generate a panel of specific monoclonal antibodies (mAbs) in mice. We will characterize the affinity and the epitope binging of these mAbs by combining epitope binning assays and electron microscopy analysis (EM). Based on these results, we will test the therapeutic potential of these mAbs in vivo and propose a candidate immunotherapy. To expand our knowledge of the immune response triggered by the viral NPs, we will combine molecular and visualization techniques to unravel the molecular mechanism of transport of the viral NPs to the cell surface. The resulting information will provide insights into the antigenic principles of Influenza and Coronavirus NPs, which can guide the design of universal vaccines.