Structure-based design of a broadly protective influenza hemagglutinin stem immunogen

The conserved hemagglutinin (HA) stem region is widely regarded as a highly promising target for the development of a universal influenza vaccine. Through structure-based design and iterative mutagenesis screening, we successfully engineered a soluble trimeric H1 stem-only protein, designated #219, which does not contain heterologous trimerization motifs and thus prevents off-target immune responses. As confirmed by a 2.97 Å-resolution cryo-EM structure, the overall conformation of #219 remains intact. Vaccination of mice with #219 protected against lethal heterologous viral challenges. Specific serum antibodies strongly cross-reacted with HAs of diverse influenza A subtypes, cross-neutralized various H1N1 viruses, and exhibited multiple antibody-effector functions mediating infection-permissive protection. We have developed a promising H1 stem immunogen for broad cross-protection against influenza infection and also proposed an efficacious design strategy for soluble homo-oligomeric proteins.