A Stabilized, Monomeric, Receptor Binding Domain Elicits High-Titer Neutralizing Antibodies Against All SARS-CoV-2 Variants of Concern

• DOI: 10.3389/fimmu.2021.765211
• Publication Date: 2021-12-09T11:06:26Z
• Authors: Shahbaz Ahmed; Mohammad Suhail Khan; Savitha Gayathri; Randhir Singh; Sahil Kumar; Unnatiben Rajeshb...; Sameer Kumar Malladi; Raju S. Rajmani; Petrus Jansen van...; Shane Riddell; Sarah Goldie; Nidhi Girish; Poorvi Reddy; Aditya Upadhyaya; Suman Pandey; Samreen Siddiqui; Akansha Tyagi; Sujeet Jha; Rajesh Pandey; Oyahida Khatun; Rohan Narayan; Shashank Tripathi; Alexander J. McAuley; Nagendrakumar Bal...; Seshadri S. Vasan; Rajesh P. Ringe; Raghavan Varadarajan
• Keywords: Monoclonal antibody; 0301 basic medicine; COVID-19 Vaccines; Molecular biology; Immunology; lyophilization; Coronavirus Disease 2019 Research; FOS: Health sciences; hyperstable mutants; Antibodies, Viral; Diagnostic Methods for COVID-19 Detection; spike protein; Biochemistry; Gene; hamster immunization; Mice; 03 medical and health sciences; Immunogenicity, Vaccine; Neutralization; Cricetinae; Virology; Health Sciences; Animals; neutralizing antibodies; Immunogen; Biology; Antibody; SARS-CoV-2; FOS: Clinical medicine; Mutant; COVID-19; Gastrointestinal Viral Infections and Vaccines Development; RC581-607; Neutralizing antibody; vaccination; Antibodies, Neutralizing; Immunogenicity; Virus; 3. Good health; Chemistry; Infectious Diseases; Mutagenesis; stabilizing mutation; Spike Glycoprotein, Coronavirus; Medicine; Immunologic diseases. Allergy; Titer

Saturation suppressor mutagenesis was used to generate thermostable mutants of the SARS-CoV-2 spike receptor-binding domain (RBD). A triple mutant with an increase in thermal melting temperature of ~7°C with respect to the wild-type B.1 RBD and was expressed in high yield in both mammalian cells and the microbial host, Pichia pastoris, was downselected for immunogenicity studies. An additional derivative with three additional mutations from the B.1.351 (beta) isolate was also introduced into this background. Lyophilized proteins were resistant to high-temperature exposure and could be stored for over a month at 37°C. In mice and hamsters, squalene-in-water emulsion (SWE) adjuvanted formulations of the B.1-stabilized RBD were considerably more immunogenic than RBD lacking the stabilizing mutations and elicited antibodies that neutralized all four current variants of concern with similar neutralization titers. However, sera from mice immunized with the stabilized B.1.351 derivative showed significantly decreased neutralization titers exclusively against the B.1.617.2 (delta) VOC. A cocktail comprising stabilized B.1 and B.1.351 RBDs elicited antibodies with qualitatively improved neutralization titers and breadth relative to those immunized solely with either immunogen. Immunized hamsters were protected from high-dose viral challenge. Such vaccine formulations can be rapidly and cheaply produced, lack extraneous tags or additional components, and can be stored at room temperature. They are a useful modality to combat COVID-19, especially in remote and low-resource settings.