A Computationally Designed Hemagglutinin Stem-Binding Protein Provides In Vivo Protection from Influenza Independent of a Host Immune Response

Abstract

Broadly neutralizing antibodies targeting a highly conserved region in the hemagglutinin (HA) stem protect against influenza infection. Here, we investigate the protective efficacy of a protein (HB36.6) computationally designed to bind with high affinity to the same region in the HA stem. We show that intranasal delivery of HB36.6 affords protection in mice lethally challenged with diverse strains of influenza independent of Fc-mediated effector functions or a host antiviral immune response. This designed protein prevents infection when given as a single dose of 6.0 mg/kg up to 48 hours before viral challenge and significantly reduces disease when administered as a daily therapeutic after challenge. A single dose of 10.0 mg/kg HB36.6 administered 1-day post-challenge resulted in substantially better protection than 10 doses of oseltamivir administered twice daily for 5 days. Thus, binding of HB36.6 to the influenza HA stem region alone, independent of a host response, is sufficient to reduce viral infection and replication in vivo. These studies demonstrate the potential of computationally designed binding proteins as a new class of antivirals for influenza.

Massively parallel de novo protein design for targeted therapeutics

“…1a). To design binding interfaces, we superimposed helical segments of the scaffolds on interface helices in previously solved HA and BoNT/B complexes (the previously designed HA binders HB36.6 and HB80.4 14–16 , and the natural target of BoNT/B synaptotagmin-II (Syt-II) 17 ), seeded the newly formed interfaces with hotspot residues from these helices (Extended Data Fig. 1) and designed the remainder of the residues to maximize binding affinity and monomer stability using Rosetta combinatorial sequence optimization (Supplementary Fig.…”

“…HB1.6928.2.3, an affinity-matured, disulfide-containing design, strongly neutralized PR8 and CA09 influenza viruses after 48 hours in culture, with a half-maximal effective concentration (EC 50 ) value for Cal09 (CA09) more than 100-fold lower than the broadly neutralizing antibody FI6v3 18 , or the previously designed HB36.6 14 , on the basis of mass (Fig. 4d; the EC 50 is similar to the antibody on a molar basis).…”

Deep sequencing methods for protein engineering and design

“…The affinity of one designed HA binder, HB36.6, was further improved against seven diverse HA subtypes. HB36.6 showed prophylactic and therapeutic efficacy against lethal challenge of pandemic Influenza in a BALB/c mouse model [15]. …”

Passive immunization with influenza haemagglutinin specific monoclonal antibodies

“…Computationally designed small protein mimics of antibodies that specifically bind to the HA stem of group 1 viruses have been described as an alternative to development of bNAbs for therapy. 62,63 Despite the lack of an Fc region, small protein antibody mimics induced relatively weak cytokine responses and provided significant protection in both prophylactic and therapeutic studies in mice and would potentially be safe and effective even in immunodeficient individuals.…”