INFLUENZA-A SUBTYPE CROSS-PROTECTION AFTER IMMUNIZATION OF OUTBRED MICE WITH A PURIFIED CHIMERIC NS1/HA(2) INFLUENZA-VIRUS PROTEIN

Influenza A/PR/8/34-derived chimeric (D) protein (SK&F 106160) composed of the first 81 amino acids (aa) of NS1 fused to the conserved 157 C-terminal aa of HA(2) (NS11-81-HA(265-222)) was previously, shown to induce H-2(d)-restricted protective cytotoxic T-lymphocyte (CTL) immunity in inbred mice. However, D protein, like other small peptides, exhibited haplotype dependence and was not immunogenic in H-2(b) and H-2(K) mice. A potential use of this antigen in humans and the role of T cells in any protection were evaluated in outbred Swiss and inbred CBF6F(1) (H-2(d/b)) mice. Mice immunized with D protein and challenged by small-particle aerosol with a lethal dose of influenza virus were significantly protected against mortality front influenza A/H1N1 and A/H2N2 (p < 0.05 - < 0.0000001), but not from A/H3N2 and influenza B viruses when compared with control mice. D protein did not induce serum virus-neutralizing antibody but caused virus to be cleared faster in immunized mice. Protection was long-lasting. In vivo depletion of either Lyt2 (CD8+) or L(3)T(4) (CD4+) T cells with monoclonal antibodies led to abrogation of in vitro-generated CTL activity in CF6F(1) mise and significant reduction in the protective efficacy of D protein against virus challenge in both Swiss and CF6F(1) mice. These results suggest that protection was mediated by CD8+ and/or CD4+ cells and not antibody. Thus D protein, via a conserved sequence on the HA(2) polypeptide, has the potential to induce partially cross-reactive CTL that may protect against influenza virus disease in humans.