Reverse vaccinology 2.0: Human immunology instructs vaccine antigen design

Rappuoli, Rino; Bottomley, Matthew J.; D’Oro, Ugo; Finco, Oretta; De Gregorio, Ennio

However, the appropriateness of the HA stem region as a component in a universal flu vaccine was recently confirmed by the finding that bNAbs against this region of HA can be selected from human subjects after infection or vaccination (Kashyap et al., 2008; Throsby et al., 2008; Ekiert et al., 2009 Ekiert et al., , 2011 Sui et al., 2009; Corti et al., 2011b; Dreyfus et al., 2012) . These observations have driven experimental approaches aimed at improving the immune response against the stem (Hai et al., 2012; Schneemann et al., 2012; Kanekiyo et al., 2013) . This aim is complicated because, as for RSV F (see RSV section above), the native HA is a meta-stable membrane glycoprotein that undergoes large conformational changes. To overcome this challenge, Kanekiyo et al. (2013) used a structure-based design to generate a genetic fusion encoding HA from one H1 strain linked to the bacterial protein ferritin. Because of the natural oligomerization of ferritin, the HA-ferritin polypeptide spontaneously selfassembled into 24-mer nanoparticles, each displaying eight trimeric H1 subtype HA "spikes" projecting off the ferritin nanoparticle scaffold, oriented such that the immune system could encounter the ordered arrays of HA in a native-like conformation. When tested in preclinical in vivo models, this elegant multivalent immunogen presenting structurally intact HA trimers was effective in eliciting higher titers of NAbs compared with a similarly adjuvanted trivalent inactivated vaccine (TIV) in both mice and ferrets, providing broader coverage against unmatched H1N1 viruses. Analysis of the specificity of the immune response revealed that the NAbs generated by HA-ferritin nanoparticles were directed against both the sensitive receptor binding site in the HA head and the conserved HA stem region, and the latter was responsible for half of the observed heterosubtype neutralizations. Although it could not be considered a real universal flu vaccine, this HA nanoparticle-based vaccine proved that a potent and broad neutralizing response against the HA stem region can be induced by the use of an appropriate immunogen. However, a potent cross-protective response was observed only in the presence of strong adjuvants such as Ribi or MF59. Such adjuvants may work by improving the development of T follicular helper cells and the germinal center antigen-specific B cell response (Mastelic Gavillet et al., 2015) , thus helping the selection of low-abundance B cell clones recognizing poorly immunogenic epitopes such as those of the HA stem region, as experimentally proved for other model antigens (Victora et al., 2010) .