Electroporation and LNP-mediated delivery of plasmid DNA-encoded H5N1 influenza virus hemagglutinin support protection against highly pathogenic avian influenza

Highly pathogenic avian influenzas (HPAIs) is a continuing public health threat. Here, we describe the development of plasmid-encoded H5N1 hemagglutinin antigens representing clades that have caused human zoonoses. Electroporation (EP) delivery of clade 2.3.2.1c HA (pCamb) DNA was immunogenic in mice but only partially protective against clade 2.3.4.b challenge. Homologous challenge resulted in complete protection, suggesting that matched clade antigens are important for protection. Contemporary clade 2.3.4.4b HA (pMich) DNA plasmids supported robust cellular and humoral responses when delivered via EP. Co-immunization with plasmids representing both H5 clades supported high titers of binding and neutralizing antibodies against both antigens and complete protection from clade 2.3.4.4b challenge. We formulated the pMich plasmid DNA in lipid nanoparticles (LNPs). A single dose of pMich DNA-LNP supported long-lived immunity in mice that was protective against challenge at both acute and memory timepoints. These data demonstrate that DNA can support robust anti-HPAI immunity and protection.