Yeast-Expressed SARS-CoV Recombinant Receptor-Binding Domain (RBD219-N1) Formulated with Aluminum Hydroxide Induces Protective Immunity and Reduces Immune Enhancement

Abstract

We developed a severe acute respiratory syndrome (SARS) subunit recombinant protein vaccine candidate based on a highyielding, yeast-engineered, receptor-binding domain (RBD219-N1) of the SARS beta-coronavirus (SARS-CoV) spike (S) protein. When formulated with Alhydrogel ® , RBD219-N1 induced high-level neutralizing antibodies against both pseudotyped virus and a clinical (mouse-adapted) isolate of SARS-CoV. Here, we report that mice immunized with RBD219-N1/Alhydrogel ® were fully protected from lethal SARS-CoV challenge (0% mortality), compared to ~ 30% mortality in mice when immunized with the SARS S protein formulated with Alhydrogel ® , and 100% mortality in negative controls. An RBD219-N1 formulation Alhydrogel ® was also superior to the S protein, unadjuvanted RBD, and AddaVax (MF59-like adjuvant)formulated RBD in inducing specific antibodies and preventing cellular infiltrates in the lungs upon SARS-CoV challenge.Specifically, a formulation with a 1:25 ratio of RBD219-N1 to Alhydrogel ® provided high neutralizing antibody titers, 100% protection with non-detectable viral loads with minimal or no eosinophilic pulmonary infiltrates. As a result, this vaccine formulation is under consideration for further development against SARS-CoV and other emerging and re-emerging beta-CoVs such as SARS-CoV-2.

Prospects for a safe COVID-19 vaccine

“…SARS-CoV-1 vaccines comprising inactivated whole virus (with virus inactivation by formalin or UV irradiation), recombinant spike protein (expressed in baculovirus), or chimeric viral-like particles have elicited cellular immunopathology when administered to mice despite the presence of high titers of neutralizing antibodies (50). In these studies, an alum adjuvant was shown to reduce immunopathology compared to non-adjuvanted vaccines, a finding confirmed in mouse immunization experiments with the SARS-CoV-1 spike protein receptor binding domain formulated with alum (51). Other studies have highlighted the importance of inducing Th1 responses as well as CD8+ T cells after vaccination of mice as a means to enhance protective immunity and prevent cellular immunopathology (45,(52)(53)(54).…”

Design of a highly thermotolerant, immunogenic SARS-CoV-2 spike fragment immunogen

“…Recently, various RBD derived subunit vaccine candidates have been tested for immunogenicity employing varying fragment lengths, fusion adaptors (Fc, dimers), and adjuvants. No antibody dependent enhancement of infection, immunopathologies, or Th2 bias have been observed with the SARS-CoV-2 RBD subunit derivatives examined so far [40][41][42][43] . Three independent studies used RBD-Fc fusions with one study using RBD residues 331-527, another used RBD-Fc from Sino Biologicals (residues not mentioned) and a third used a full-length S1-Fc fusion (residues 14-685) reporting viral neutralizing antibody titers of ~100-400, 1280 and NT 50 derived from pseudoviral neutralizations of 378 respectively 41,42,44 .…”

“…Multiple studies employing a variety of vaccine formulations and modalities have now demonstrated that SARS-CoV-2 viral neutralization titers in small animals including mice and guinea pigs are predictive of immunogenicity in macaques and humans ( Figure 5E) [17][18][19][20]22,23,[33][34][35][36][37][40][41][42][43]45 . Despite promising immunogenicity in several cases, all of the above liquid formulations were either refrigerated or frozen prior to use.…”

“…The RBD fragment could also be expressed at high yield in the microbial host Pichia pastoris, and was properly folded, stable and immunogenic. Interestingly, an alhydrogel adjuvanted formulation of a related SARS-CoV-1 RBD construct was recently shown to be immunogenic and protect mice from SARS-CoV-1 challenge 40 . Unfortunately, in the present study when pRBD was used as an immunogen, the elicited antibodies were poorly reactive with either the mammalian cell expressed RBD or the corresponding Spike ectodomain.…”

Capsid-like particles decorated with the SARS2-CoV-2 receptor-binding domain elicit strong virus neutralization activity

“…The urgent need of an effective SARS-CoV-2 vaccine, to contain the worldwide pandemic and prevent new viral outbreaks, has led to a global effort involving a wide range of vaccine technologies. These include genetic-based (mRNA and DNA) principles 16,17 , replicating/non-replicating viral vectors (measles 18 , adenovirus 19,20 , baculovirus) recombinant proteins or peptides 21 , virus-like particles (VLPs)/nanoparticles or inactivated and live-attenuated viral vaccines [22][23][24] . In fact, more than 120 SARS-CoV-2 candidate vaccines are currently registered by WHO, of which 21 are currently undergoing clinical testing 25 .…”