Worldwide up to 650,000 people die from influenza each year, including an average of 42,000 people within the United States, approximately 80% of whom are above the age of 65 [1,2,3]. High rates of hospitalization and death occur despite seasonal vaccines and available therapies [1, 4]. The influenza vaccine must be updated annually because influenza virus lacks proofreading mechanisms during replication resulting in a high mutation rate. These mutations are especially prevalent in the most immunogenic proteins of influenza, and their accumulation is referred to as antigenic drift. [5, 6] The seasonal influenza vaccine has been utilized in the United States since 1945, but the necessity of reformulating the seasonal vaccine annually is an expensive and time consuming collaborative global effort [3, 7].
Despite extensive screening and development each year, the seasonal vaccine’s efficacy can be limited. Dependent on the degree of mutation after the selection of the vaccine virus sequence each year, efficacy ranges between 10 and 60% [7, 8]. For example, during the 2013–2014 influenza season, the vaccine was 52% effective and vaccinated adults were 52–79% less likely to die as a result of influenza; however, the next year (2014–2015), effectiveness of the vaccine dropped to 19% [8,9,10].
Influenza A is capable of an additional mechanism of change, antigenic shift. Antigenic shift is the rapid change and development of a new influenza virus and occurs when two different serotypes of influenza A co-infect the same cell and exchange RNA segments . This genetic recombination usually involves a newly human-adapted hemagglutinin (HA) (e.g. from birds or swine) or a highly mutated HA to which the human population is naïve [5, 11]. If antigenic shift occurs after strain selection, there is little to no protection against a potentially pandemic influenza strain . This was the case in 2009, when the A/California/04/2009 pandemic virus dramatically shifted, becoming more antigenically similar to the 1918 “Spanish flu” than to the seasonal H1N1 strains between 1977 and 2008 .
Despite the clear need for a vaccine that is universally applicable to seasonal and pandemic strains of influenza A, no universal influenza vaccine has been FDA approved. The AuNP-M2e + sCpG vaccination, first published in 2014, utilizes M2e as a potentially universal target for influenza A, because of the high level of conservation in the M2e peptide sequence between serotypes and isolates and its expression on both the surface of virions and infected cells [12,13,14,15,16]. M2e is the extracellular N-terminal portion of M2 and has been considered an excellent candidate for influenza A vaccination or treatment since its discovery by Lamb, et al. in 1981 [11, 12, 17]. However, there has been limited success in actualizing that potential . Well over 30 M2e vaccines have been developed and published using a variety of adjuvants, including four which have entered clinical trials [12, 19].
AuNP-M2e + sCpG vaccination seems particularly promising because it is easy and inexpensive to produce, has a short manufacturing time, is egg-free, and can be lyophilized so that it is stable long term at room temperature , making it not only easy to stockpile but also feasible for large scale production. This vaccine is demonstrated to be very efficacious 21 days post vaccination in protecting against lethal challenge with H1N1 A/PR/8/1934, pH1N1 A/CA/04/2009, H3N2 A/Victoria/3/75, and H5N1 A/Vietnam/1203/2004, indicative of a highly cross reactive memory immune response [15, 20]. The vaccine has also been demonstrated to maintain elevated but gradually decreasing antibody titers and to be protective against H1N1 A/PR/8/1934 lethal challenge up to eight months post vaccination in mice . These publications successfully demonstrate that intranasal vaccination with AuNP-M2e + sCpG in healthy, young 12–14 week old BALB/c mice (vaccinated at 6–8 weeks and infected 42 days post vaccination) and healthy, adult 54–56 week old BALB/c mice (vaccinated at 6–8 weeks and infected 8 months post vaccination) induces an M2e-specific memory response, which is protective against lethal challenge.
However, adults over 65 years old are at the highest risk during most influenza seasons, constituting 71–85% of deaths and 54–70% of hospitalizations related to seasonal influenza, estimated by the CDC . Many studies have identified being over 65 years of age as one of the most significant risk factors for fatality from influenza infection A from current circulating strains of H3N2 and pH1N1 [22, 23]. This seems to be predominantly caused by immunosenescence, or the decreased efficiency of the immune system as a result of aging . In response to influenza this is characterized as decreased antibody-mediated and cellular immunity and decreased responsiveness to vaccines, as decreased thymus function limits the induction of new and memory responses to antigens [24,25,26]. Adults over 65 typically experience altered clinical presentation of influenza A with reduced fever symptoms but increased respiratory symptoms, including coughing and wheezing . These patients also have increased rates of deadly complications, namely pneumonia to which people over 65 are already at increased risk [26, 27].
Further, many vaccines lose efficacy during aging resulting in a partial loss of protection and potentially requiring booster vaccinations to remain effective . Regular boosters are required for tetanus and diphtheria vaccines, and boosters for pertussis and polio are often recommended [24, 29, 30]. To determine if the AuNP-M2e + sCpG vaccine is a viable option as a universal vaccination and to begin to test it if might require re-administrations during a lifetime, we tested if the AuNP-M2e + sCpG vaccination loses efficacy after aging mice to geriatric age (defined as 18 months or 72 weeks old) .
We vaccinated BALB/c mice at 3–6 weeks of age and challenged them 15 months after vaccination with a lethal challenge of H1N1 A/PR/8/1934. At the time of challenge, these mice were approximately 18–24 months old and retain significant M2e-specific antibody titers in total IgG, IgG1, IgG2a, and IgG2b. Further, the antibody titers increase upon infection with influenza A (H1N1 PR8) and remain elevated for at least 3 months, suggesting the elderly mice retain effective M2e-specific memory B cells. These mice are significantly protected from lethal influenza challenge (H1N1, 8.3 PFU). These results suggest that AuNP-M2e-CpG is an excellent candidate as a universal influenza vaccine as it maintains lifelong protection in mice despite aging to geriatric age.