The insufficient immunogenicity of the original H5N1 vaccine led to the inclusion of MF59 and AS03 adjuvants in influenza vaccines, which enabled the vaccine to achieve U.S. Food and Drug Administration and European Union criteria of seroconversion and seroprotection rates. At the onset of the SOIV-H1N1 pandemic in Europe, these adjuvants were combined with the SOIV-H1N1 vaccines to enhance immunogenicity and allow dose sparing (8
Here, we provide the initial characterization of the effects of oil-in-water adjuvants on the quality of antibodies induced by the SOIV-H1N1 vaccine in different age populations. Our data clearly show that MF59 enhances VN responses by both quantitative and qualitative improvement in HA-specific antibody responses by increasing the diversity of epitope repertoire recognized in the HA1 globular domain and by enhancing the affinity of antibodies specific for native HA. On the basis of the increased magnitude of the effect of MF59 on anti–SOIV-H1N1 responses in younger individuals, and similar findings with the H5N1 vaccine in H5N1-naïve adults, it is likely that the major effect of MF59 is exerted on naïve B cells, increasing the rate of somatic hypermutation. Therefore, MF59 and similar adjuvants have the potential to overcome the tendency in some individuals to generate low-affinity antibodies with pathological potential.
We show that in the face of a novel IAV pandemic, most preexisting serum antibodies are specific for HA2. These antibodies would be induced by previous clinically overt or subclinical infections with influenza viruses or by previous immunizations (mainly in adults) with conventional, nonadjuvanted seasonal influenza vaccines. This is consistent with previous findings that HA2 antibodies are highly cross-reactive, which is expected because of the much higher sequence conservation in HA2 compared to HA1 (13
). The RBD of HA1, where antigenic drift is most extensive, is particularly diverse.
A number of groups have recently described antibodies in mice, ferrets, monkeys, and humans specific for a discontinuous antigenic site in the HA stem region that cross-react between related subtypes (for example, H1-H2-H5) with post-attachment virus-neutralizing activity (31
). Representative monoclonal antibodies (mAbs) have been shown to prevent and treat IAV infections in animal models, raising the prospect of generating vaccines that provide broader immunity within and even across related subtypes (35
). It will be of great interest to examine the effect of MF59 on eliciting such antibodies, and particularly on increasing their affinity, which may be critical in enhancing their in vivo activity.
Here, we revisited the central question of the role of affinity maturation in antiviral immunity. In earlier mouse studies using a panel of mouse mAbs specific for vesicular stomatitis virus (VSV) glycoprotein, protection against VSV infection was independent of immunoglobulin class, affinity, and VN rate constant. Antibody affinity above a minimal threshold (about 2 × 10−7
) provided complete protection from acute infection, and protection depended simply on serum antibody concentration (37
). Furthermore, mAbs generated either early or late after VSV hyperimmunization demonstrated similar high affinity, with no evidence of time-dependent affinity maturation (14
). The authors proposed that for acute viruses, there is little need for affinity maturation because the virus is cleared long before higher-affinity antibodies reach effective concentrations. However, these studies focused on the mouse responses to VSV, for which it appears that a germ-line antibody offers high neutralizing titers against the virus (15
). This is probably an unusual situation whereby very limited affinity maturation is needed, and does not reflect normal antiviral antibody responses. Recent studies demonstrate that acute viral infections (including SOIV) and formalin-inactivated vaccines for measles and respiratory syncytial virus may elicit low-affinity antibodies that form immune complexes in the lungs and activate complement. These non-neutralizing antibodies may contribute to enhanced lung pathology with severe disease and even death after infection (22
Here, we confirm the importance of antibody affinity in anti-influenza immunity by demonstrating strong correlation between 7 M urea–resistant serum antibodies and off-rate constants with VN titers in response to SOIV-H1N1 and H5N1 vaccination. We found that MF59 adjuvant greatly enhanced affinity maturation especially in naïve populations.
How is an oil-in-water adjuvant such as MF59 able to increase antibody affinity? The enhancement observed by MF59 of HA1-specific antibodies could be due to a difference in the requirements for activating naïve HA1-specific as opposed to recall HA2-specific B cells. Indeed, previous findings have shown that vaccination with the MF59-adjuvanted H5N1 vaccine, but not with unadjuvanted vaccine, primes a potent and rapid antigen-specific CD4+
T cell response that is predictive of the high VN antibody levels found after booster immunization (42
Together, our data clearly demonstrate that antibody affinity maturation occurs after influenza vaccination, particularly in naïve individuals. Most importantly, MF59 enhances affinity maturation and offers the promise of improved protection in vivo. These findings provide support for use of adjuvants in influenza vaccines, especially when targeting naïve populations.