Tetramerizing tGCN4 domain facilitates production of Influenza A H1N1 M2e higher order soluble oligomers that show enhanced immunogenicity in vivo

Abstract

One strategy for the development of a next generation influenza vaccine centers upon using conserved domains of the virus to induce broader and long-lasting immune responses. The production of artificial proteins by mimicking native -like structure has shown to be a promising approach for vaccine design against diverse enveloped viruses. The amino terminus of Influenza A virus Matrix 2 ectodomain (M2e) is highly conserved among influenza subtypes, and previous studies have shown M2e-based vaccines are strongly immunogenic, making it an attractive target for further exploration. We hypothesized that stabilizing M2e protein in the mammalian system might influence the immunogenicity of M2e with the added advantage to robustly produce the large scale of proteins with native- like fold and hence can act as an efficient vaccine candidate. In this study, we created an engineered construct in which the amino terminus of M2e is linked to the tetramerizing domain tGCN4, expressed the construct in a mammalian system and tested for immunogenicity in BALB/c mice. We have also constructed a stand-alone M2e construct (without tGCN4) and compared the protein expressed in mammalian cells and in E. coli using in vitro and in vivo methods. The mammalian-expressed protein was found to be more stable, antigenic than the E. coli protein, and form higher-order oligomers. In an intramuscular protein priming and boosting regimen in mice, these proteins induced high titers of antibodies and elicited a mixed Th1/Th2 response. These results highlight the mammalian expressed M2e soluble proteins as a promising vaccine development platform.

Identifying Key Drivers of Efficient B Cell Responses: On the Role of T Help, Antigen-Organization, and Toll-like Receptor Stimulation for Generating a Neutralizing Anti-Dengue Virus Response

“…The subclass-specific Ab response may strongly depend on TLR-stimulating factors such as RNA or DNA [ 215 ]. The preferential induction of IgG1 Abs was observed for influenza M2e domain-derived tetramers used for immunizations in BALB/c mice [ 216 ]. Conversely, Wang and co-workers [ 217 ] claimed that oligomers expressed by a DNA-based vaccine derived from the Yersinia pestis V antigen, which included a signal sequence from a human tissue plasminogen activator, elicited a dominant IgG2a response in the BALB/c murine model [ 217 ].…”

Comparative immunogenicity analysis of intradermal versus intramuscular administration of SARS-CoV-2 RBD epitope peptide-based immunogen In vivo

“…Serum humoral antibodies generated against RBD peptide and spike protein were assessed by ELISA as described previously [ 19 ].…”