Mapping and design of SARS-CoV-2 and HIV-1 spike epitopes for inducing neutralizing antibodies

Abstract

The induction of broadly neutralizing antibodies (bNAbs) is essential for the development of effective vaccines against HIV-1 and SARS-CoV-2 infections. To develop an effective HIV-1 vaccine, I constructed nine modified vaccinia Tian-Tan (MVTT) strains to express various Env trimer and determined their immunogenicity in mice and rabbits. Subsequently, to fight newly emerged SARS-CoV-2, I constructed a yeast library displaying random Spike (S) fragments and mapped antigenic epitope landscapes for both monoclonal antibodies and polyclonal sera derived from COVID-19 patients.

Soluble HIV-1 Env proteins, BG505-SOSIP, CNE6-UFO and MG04Bchim, were selected for generating recombinant MVTT (rMVTT) strains based on their high, medium and low stability in forming native-like trimers, respectively. Transmembrane of BG505 (BGTM) and vaccinia hemagglutinin (HATM) were designed for trimer expression on cellular or vaccinia lipid membranes. Nine rMVTT strains were constructed and verified for the correct insertions and expression of the soluble or TM Env proteins. Then, 163 mice were immunized with rMVTTs using various dosages and routes. The HATM design of BG505-SOSIP (MVTTBG505-SOSIP-HATM) enhanced the vaccine immunogenicity for inducing higher levels of antibody responses. Moreover, high dosage of intramuscular MVTTBG505-SOSIP- and MVTTBG505-SOSIP-HATM-primed mice elicited more bNAbs against broad tier-2 HIV-1 pseudoviruses, while MVTTBG505-SOSIP-BGTM primed mice elicited mainly autologous tier-2 NAbs. In contrast, CNE6-UFO- and MG04Bchim-related rMVTTs partially induced antigen specific antibodies but weak NAbs against several pseudoviruses. Furthermore, 12 rabbits were primed with BG505-SOSIP-related rMVTTs intramuscularly. One-time inoculation of rMVTTs induced high amounts of antibody responses. The NAbs, however, appeared at week 7 after the 2nd injection, and robust NAbs developed at week 19 after the 4th dose against multiple autologous and heterologous tier-2 HIV-1 pseudoviruses in several rabbits. MVTTBG505-SOSIP-HATM elicited the lowest levels of antibody responses in rabbits. Rabbit 6 from this group, however, generated the most potent bNAbs, which neutralized 15 strains of HIV-1 pseudoviruses, including 11 from the global panel, with an average IC80 = 1:96. I, therefore, demonstrated that rMVTT-vectored Env trimers likely induced bNAbs in mice and rabbits.

After COVID-19 outbreak, I constructed a yeast surface display (YSD) library, presenting approximately 106 S fragments covering the full-length SARS-CoV-2 S glycoprotein. Using this YSD library, I mapped two conformational epitope regions at the receptor-binding domain (RBD) and the N-terminal domain (NTD), respectively, which were specific to corresponding monoclonal NAbs and were not heat-stable. Moreover, I mapped diverse epitope landscapes using serum samples derived from 12 COVID-19 patients at different disease stages. Among 16 antigenic domains characterized, 2 conformational epitope regions and 6 linear domains were dominant. These dominant epitopes were likely exposed on the surface of trimeric S glycoproteins. Unexpectedly, the domain 7 was likely associated with the disease severity. The average proportion of the domain 7-specific antibodies were 21.77-fold higher in severe patients than that in mild ones. During the natural course of SARS-CoV-2 infection, yeast-pull down experiments revealed that RBD-specific antibodies contained majority of NAbs. Lastly, I demonstrated that cross-reactive antibodies of COVID-19 patients to SARS-CoV and MERS-CoV primarily targeted at the linear epitopes of domains 9, 10 and 14.