Manipulation of immunodominant variable epitopes of norovirus capsid protein elicited cross-blocking antibodies to different GII.4 variants despite the low potency of the polyclonal sera

Norovirus is a major cause of viral acute gastroenteritis. The predominant genotype, GII.4, presents a continuous emergence of new variants that could escape immunity developed in previous infections. The emergence of GII.4 variants is associated with multiple mutations on five antigenic sites (A, C, D, E, and G) of the major viral capsid protein. Antigenic site A includes most immunodominant epitopes, while antigenic site G shifted its immunodominance during the evolution of GII.4 noroviruses. This interrelation of immunodominance and high variability challenges the development of universal vaccines for GII.4 noroviruses. In this study, we designed an immunogen that aims to elicit antibodies toward conserved regions by mutating epitopes from antigenic sites A and G. Immunization of mice with mutant virus-like particles (VLPs) in which antigenic sites A and G were resurfaced with alanine mutations (12∆AG) elicited multiple cross-blockade monoclonal antibodies against different GII.4 variants. However, the carbohydrate-blockade titers at the serum level were low as compared with wild-type VLPs as the result of low frequency and/or low potency of these cross-blockade antibodies. This study confirmed that cross-reactive antibodies can be elicited by mutating highly variable epitopes, but further studies are required to overcome the low immunogenicity of these conserved epitopes for the development of vaccine candidates that could elicit broadly protective responses.IMPORTANCEThe fast-evolving nature of RNA viruses is a major obstacle for vaccine design and development. Protective antibodies are often directed to highly variable sites, so viruses could rapidly escape from immunity acquired from previous infections or vaccination. Here, we designed mutant norovirus virus-like particles (VLPs) in which the immunodominant and highly variable epitopes were resurfaced into alanine for refocusing immune responses toward conserved epitopes. The mutant VLPs could elicit multiple broadly cross-reactive antibodies against different GII.4 noroviruses that emerged in the 1970s-2010s; however, the frequency of these cross-reactive antibodies was low at the serum level. While further investigations are required to enhance the potency of these cross-reactive responses, this study opens a new avenue for the rational development of efficacious norovirus vaccines.