Development of Spike Receptor-Binding Domain Nanoparticles as a Vaccine Candidate against SARS-CoV-2 Infection in Ferrets

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a causative agent of the CoV disease 2019 (COVID-19) pandemic, enters host cells via the interaction of its receptor-binding domain (RBD) of the spike protein with host angiotensin-converting enzyme 2 (ACE2). Therefore, the RBD is a promising vaccine target to induce protective immunity against SARS-CoV-2 infection. In this study, we report the development of an RBD protein-based vaccine candidate against SARS-CoV-2 using self-assembling Helicobacter pylori-bullfrog ferritin nanoparticles as an antigen delivery system. RBD-ferritin protein purified from mammalian cells efficiently assembled into 24-mer nanoparticles. Sixteen- to 20-month-old ferrets were vaccinated with RBD-ferritin nanoparticles (RBD nanoparticles) by intramuscular or intranasal inoculation. All vaccinated ferrets with RBD nanoparticles produced potent neutralizing antibodies against SARS-CoV-2. Strikingly, vaccinated ferrets demonstrated efficient protection from SARS-CoV-2 challenge, showing no fever, body weight loss, or clinical symptoms. Furthermore, vaccinated ferrets showed rapid clearance of infectious virus in nasal washes and lungs as well as of viral RNA in respiratory organs. This study demonstrates that spike RBD-nanoparticles are an effective protein vaccine candidate against SARS-CoV-2.

Scientific rationale for developing potent RBD-based vaccines targeting COVID-19

“…Preclinical studies have evaluated the protective efficacy of several RBD-specific monoclonals, as well as RBD as a protective immunogen. Investigators evaluating RBD as a vaccine candidate have delivered the antigen either as DNA 81 , mRNA 7,74,[82][83][84][85] , viral vector 86 , soluble monomer or dimer 6,22,[87][88][89][90][91][92][93] , a fusion protein nanoparticle 16,26,[94][95][96][97][98] , or as a virus-like particle (VLP) 91,[99][100][101][102] . Additional RBD-based vaccine candidates in clinical trials (Table 2)…”

“…Vaccine researchers around the world have demonstrated preclinical immunogenicity of multimeric RBD vaccine candidates utilizing a variety of presentation methods including the following: ferritin nanoparticles 94,96,98,108,109 , single-component protein nanoparticles 95 , two-component protein nanoparticles either self-assembling 26 or assembled via SpyTag/Catcher technology 91,99,110 , and VLPs 102,111 . Together, these studies support the assertion that RBD displayed on a particle and co-administered with a suitable adjuvant represents a viable vaccine strategy for SARS-CoV-2, including against VOC.…”

Efficacy and breadth of adjuvanted SARS-CoV-2 receptor-binding domain nanoparticle vaccine in macaques

“…One common approach to enhance the induction of adaptive immune responses is the multimeric presentation of antigen, for example, on the surface of nanoparticles or virus-like particles ( 22 ). Presenting RBD in ordered, multivalent arrays on the surface of self-assembling protein nanoparticles is immunogenic and efficacious in animals ( 23 – 28 ), with improved immunogenicity relative to monomeric soluble RBD and cross-reactive responses to variants ( 17 , 24 , 26 ). However, it is unknown whether RBD nanoparticle vaccines protect against infection in primates, which have become a standard model for benchmarking performance of vaccine candidates by virological and immunologic endpoints.…”

Functionalizing Ferritin Nanoparticles for Vaccine Development

“…Other mutations have also been performed to improve the use of ferritin nanoparticles in nanobiotechnology: (i) the point mutation R64K in Pyrococcus furiosus ferritin eliminated a potential cleavage site [ 9 ], (ii) ligands with affinity for thiol groups were conjugated to engineered ferritin mutants from Archaeoglobus fulgidus and P. furiosus bearing cysteine point mutations in selected topological positions [ 54 ], and (iii) the H. pylori -bullfrog hybrid carrying the N-terminal 2–9 residues from the bullfrog R. catesbeiana L-chain ferritin (N8Q) allowed the formation of radially projecting tails, improving antigen presentation in many ferritin-based vaccine candidates [ 48 , 49 , 50 , 51 , 52 , 53 ].…”