The ectodomain of matrix protein 2 (M2e) of human influenza type A virus strains has remained remarkably conserved since 1918. Because M2e-specific immunity has been shown to decrease morbidity and mortality associated with influenza virus infection in several animal models and because natural infection and current vaccines do not appear to induce a good M2e-specific antibody (Ab) response, M2e has been considered as potential vaccine for inducing cross-reactive protection against influenza type A viruses. The high degree of structural conservation of M2e could in part be the consequence of a poor M2e-specific Ab response and thus the absence of pressure for change. To assess this possibility, we studied the course of infection in SCID mice in the presence or absence of passive M2e-specific monoclonal Abs (MAbs). We found that virus mutants with antigenic changes in M2e emerged in 65% of virus-infected mice treated with M2e-specific but not control MAbs. However, the diversity of escape mutants was highly restricted since only two types were isolated from 22 mice, one with a proline-to-leucine and the other with a proline-to-histidine interchange at amino acid position 10 of M2e. The implications of these findings for the use of M2e as a broadly protective vaccine are discussed.Current influenza virus vaccines aim to induce strong antibody (Ab) responses to the ectodomains of hemagglutinin (HA) and neuraminidase (NA) molecules, since these antibodies (Abs) can provide potent protection against infection and/or disease. The main deficiency of this protection is that it targets highly variable viral determinants. This necessitates not only frequent updating of the vaccine to contemporary circulating virus strains but, given that vaccines have to be produced and applied ahead of exposure to epidemic strains, also a correct prediction of these future epidemic strains. Failure to anticipate the emergence of an epidemic strain with significant antigenic changes compared to the vaccine strain will greatly reduce vaccine-induced protection. It would be advantageous, therefore, to expand vaccine-mediated protection to less variable viral targets. One possible way to achieve this may be through induction of 9,10,14,18,21,24,28).M2 is a 97-amino-acid transmembrane protein of influenza type A virus (15, 16). The mature protein forms homotetramers (12, 29) that have pH-inducible ion channel activity (27,29). M2-tetramers are expressed at high density in the plasma membrane of infected cells but are relatively excluded from sites of virus maturation and therefore incorporated only at low frequency into the membrane of mature virus particles (30,33). Most important in the present context are, first, that the sequence of the 24-amino-acid ectodomain of M2 (M2e) has remained remarkably conserved among human epidemic virus strains (Fig. 1A) (20). Indeed, the majority of human epidemic strains isolated since 1918 share the same M2e protein sequence. Second, several studies in mice have shown that M2e-specific Abs restrict influenz...

The current vaccination strategy against influenza A and B viruses is vulnerable to the unanticipated emergence of epidemic strains that are poorly matched by the vaccine. A vaccine that is less sensitive to the antigenic evolution of the virus would be a major improvement. The general feasibility of this goal is supported by studies in animal models that show that immunologic activities directed against relatively invariant viral determinants can reduce illness and death. The most promising approaches are based on antibodies specific for the relatively conserved ectodomain of matrix protein 2 and the intersubunit region of hemagglutinin. However, additional conserved determinants for protective antibodies are likely to exist, and their identification should be encouraged. Most importantly, infection and current vaccines do not appear to effectively induce these antibodies in humans. This finding provides a powerful rationale for testing the protective activity of these relatively conserved viral components in humans.

Background: Matrix protein 2 (M2) is an integral tetrameric membrane protein of influenza A virus (IAV). Its ectodomain (M2e) shows remarkably little diversity amongst human IAV strains. As M2e-specific antibodies (Abs) have been shown to reduce the severity of infection in animals, M2e is being studied for its capability of providing protection against a broad range of IAV strains. Presently, there is little information about the concentration of M2e-specific Abs in humans. Two previous studies made use of ELISA and Western blot against M2e peptides and recombinant M2 protein as immunosorbents, respectively, and reported Ab titers to be low or undetectable. An important caveat is that these assays may not have detected all Abs capable of binding to native tetrameric M2e. Therefore, we developed an assay likely to detect all M2e tetramer-specific Abs.

Background: The M2 ectodomain (M2e) of influenza A virus (IAV) strains that have circulated in humans during the past 90 years shows remarkably little structural diversity. Since M2e-specific antibodies (Abs) are capable of restricting IAV replication in vivo but are present only at minimal concentration in human sera, efforts are being made to develop a M2e-specific vaccine. We are exploring a synthetic multiple antigenic peptide (MAP) vaccine and here report on the role of adjuvants (cholera toxin and immunostimulatory oligodeoxynucleotide) and route of immunization on Ab response and strength of protection.