SUMMARY As part of the Pan-Coronavirus Vaccine (PanCoVac) Consortium, the goal of Research Project 1 (RP1) is the ‘Design and evaluation of pan-CoV vaccines’. A comprehensive characterization of the B and T cell responses to vaccination will be carried out by Research Project 2 (RP2, ‘Immunological responses to pan-CoV vaccines. In Aim 1 of RP1 (‘Focusing immune responses towards the stem of the spike protein’), several strategies will be tested to increase the levels of antibodies directed at the conserved, immune-subdominant epitopes in the stem of the SARS-CoV-2 spike protein (to major viral antigen), while avoiding strong antibody responses directed at immunodominant epitopes in the head of the spike protein. These strategies include chimeric spikes composed of SARS-CoV-2 immune-subdominant stem epitopes and immunodominant head epitopes of unencountered coronaviruses, ‘outdiluting’ antibody responses to the immunodominant epitopes by using cocktails of spike proteins with multiple mutations in key amino acid positions, glycan-shielding of immunodominant head epitopes, ‘inverted antigens’ in which the spike protein will be presented to the immune system in an inverted orientation (and thus become more accessible), and ‘headless’ spike proteins lacking portions of the immunodominant head epitopes. Some of these approaches may be tested in combination, and may be tested with sequence-optimized stem epitopes based on ancestral reconstruction (a computational approach to deduce the most likely common progenitor sequence). In Aim 2 (‘Eliciting broadly reactive immune responses to the head of the spike protein’), strategies will be tested to direct the immune responses away from the most sequence-diverse epitopes and towards more conserved epitopes in the head. In addition, conserved heard epitopes will be sequenced-optimized to be recognized by cross-protective antibodies. These modifications will be introduced into diverse spike proteins, and vaccine cocktails of diverse, mutant spike proteins will then be tested for their immunogenicity and protective efficacy. All antigens in Aims 1 and 2 will be designed in collaboration with a structural biologist, and for selected antigens and/or antigen/antibody complexes, X-ray crystallography and Cryo-EM will be carried out. In Aim 3 (‘Immunogenicity and protective efficacy of broadly reactive antigens’), the novel vaccinates will be tested in mice for their immunogenicity; samples from vaccinated mice will be provided to RP2 for B and T cell analysis. Selected vaccine candidates (those with broader immune responses) will next be tested in mice and Syrian hamsters for their ability to provide protection against different coronaviruses. For candidate vaccines that provide broad protection (compared to controls), we will also assess the durability of immune responses, and the effect of vaccination on virus transmissibility. Moreover, these vaccinate candidates will be tested in an mRNA lipid nanoparticle vaccine platform provided by Daiichi Sankyo, a pharmaceutical company.

PROJECT NARRATIVE Per Funding Opportunity Announcement (FOA) Number PAR-20-072: Project Narrative: Do not complete. Note: ASSIST screens will show an asterisk for this attachment indicating it is required. However, eRA systems only enforce this requirement in the Overall component and applications will not receive an error if omitted in other components.