In vitro characterization of M2e-based mRNA vaccine against avian influenza virus

Jana, Shraman (2025) In vitro characterization of M2e-based mRNA vaccine against avian influenza virus. Masters thesis, Indian Institute of Science Education and Research Kolkata.

Text (MS Dissertation of Shraman Jana (20MS018)) 20MS018_Thesis_file.pdf - Submitted Version Restricted to Repository staff only Download (1MB)

Abstract

The rapid spread of avian influenza virus (AIV) across domestic poultry and cattle continuously poses a major global pandemic threat with periodic transmission events among humans. This reinforces the quick development of effective vaccination platforms. Having started from a sceptic idea, messenger RNA (mRNA) vaccines have been successfully translated to clinical settings over the last few decades. The reported success of this vaccine technology in the COVID-19 pandemic has marked its potential to address emerging infectious diseases such as AIV due to its fast development and huge scalability. But to overcome the prevailing seasonal problems of antigenic drift and shift associated with this virus, highly conserved and broadly cross-protective viral antigens should instead be explored as major vaccine targets. Therefore, the main aim of our study was in vitro characterization of an mRNA vaccine construct encoding a four-tandem repeat sequence of Type A influenza virus Matrix protein 2 ectodomain (4xM2e) being reported as a conserved and cross-protective viral antigen in several studies. To further increase the immunogenicity of this vaccine construct, a second construct had been designed in which Pan HLA-DR epitope (PADRE) and a segment of chicken invariant chain (Ii) have been incorporated along with 4xM2e. The gene constructs were successfully cloned into a Linearized Template Vector and functional mRNAs were synthesized by in vitro transcription and capping reactions. As a delivery platform, lipid nanoparticles had been formulated and subjected to biophysical analyses. Finally, in vitro protein expression from mRNAs had been validated using both lipofectamine and lipid nanoparticles as transfection reagents. Future in vivo studies are required to investigate the immunogenicity and protective efficacy of the in vitro synthesized mRNAs against the target pathogens.

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