Developing the IVIG biomimetic, Hexa-Fc, for drug and vaccine applications

Abstract

The remarkable clinical success of Fc-fusion proteins has driven intense investigation for even more potent replacements. Using quality-by-design (QbD) approaches, we generated hexameric-Fc (hexa-Fc), a ~20 nm oligomeric Fc-based scaffold that we here show binds low-affinity inhibitory receptors (FcRL5, FcγRIIb, and DC-SIGN) with high avidity and specificity, whilst eliminating significant clinical limitations of monomeric Fc-fusions for vaccine and/or cancer therapies, in particular their poor ability to activate complement. Mass spectroscopy of hexa-Fc reveals high-mannose, low-sialic acid content, suggesting that interactions with these receptors are influenced by the mannose-containing Fc. Molecular dynamics (MD) simulations provides insight into the mechanisms of hexa-Fc interaction with these receptors and reveals an unexpected orientation of high-mannose glycans on the human Fc that provides greater accessibility to potential binding partners. Finally, we show that this biosynthetic nanoparticle can be engineered to enhance interactions with the human neonatal Fc receptor (FcRn) without loss of the oligomeric structure, a crucial modification for these molecules in therapy and/or vaccine strategies where a long plasma half-life is critical.

rIgG1 Fc Hexamer Inhibits Antibody-Mediated Autoimmune Disease via Effects on Complement and FcγRs

“…Hexameric IgG1 Fc molecules were described more recently as a potential carrier for vaccine Ags (36) and subsequently as a therapeutic candidate (21). Initially, in these studies an additional mutation of the Fc sequence at position 310 was proposed (36).…”

“…Initially, in these studies an additional mutation of the Fc sequence at position 310 was proposed (36). This additional mutation appeared to destabilize the hexameric structure (36), and it abolished binding to FcRn (21). Therefore, in their later work the authors focused on a variant with the L309C mutation only (21).…”

“…This additional mutation appeared to destabilize the hexameric structure (36), and it abolished binding to FcRn (21). Therefore, in their later work the authors focused on a variant with the L309C mutation only (21). This molecule was analyzed structurally, for its binding to Fc receptors and other receptors, as well as for complement activation when bound to solid phase.…”

“…This molecule was analyzed structurally, for its binding to Fc receptors and other receptors, as well as for complement activation when bound to solid phase. No analyses of the effect on complement activation under more physiological conditions were performed; likewise, the molecule was not studied in animal models of inflammatory disease (21). More recently, an IgG1/IgG4 hybrid hexameric Fc molecule has also been described (37).…”

“…A trivalent molecule termed Fc3Y showed protection in mouse models of ITP, arthritis, and epidermolysis bullosa acquisita (20). Finally, a hexameric Fc molecule showed increased binding to FcgRs and dendritic cell-specific intercellular adhesion molecule-3 grabbing nonintegrin, appeared to fully activate complement (C1q binding and C5b-9 deposition) (21), and effectively interfered with FcgR function (22). Ab hexamer structures are reported to have superior complement fixing properties; indeed, upon binding to cell-surface Ags, IgG monomers spontaneously arranged into ordered hexamer structures that recruited and activated C1 (23).…”