Extending human IgG half-life using structure-guided design

Abstract

Engineering of antibodies for improved pharmacokinetics through enhanced binding to the neonatal Fc receptor (FcRn) has been demonstrated in transgenic mice, non-human primates and humans. Traditionally, such approaches have largely relied on random mutagenesis and display formats, which fail to address related critical attributes of the antibody, such as effector functions or biophysical stability. We have developed a structure- and network-based framework to interrogate the engagement of IgG with multiple Fc receptors (FcRn, C1q, TRIM21, FcγRI, FcγRIIa/b, FcγRIIIa) simultaneously. Using this framework, we identified features that govern Fc-FcRn interactions and identified multiple distinct pathways for enhancing FcRn binding in a pH-specific manner. Network analysis provided a novel lens to study the allosteric impact of half-life-enhancing Fc mutations on FcγR engagement, which occurs distal to the FcRn binding site. Applying these principles, we engineered a panel of unique Fc variants that enhance FcRn binding while maintaining robust biophysical properties and wild type-like binding to activating receptors. An antibody harboring representative Fc designs demonstrates a half-life improvement of > 9 fold in transgenic mice and > 3.5 fold in cynomolgus monkeys, and maintains robust effector functions such as antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity.

Antigen physiochemical properties allosterically effect the IgG Fc-region and Fc neonatal receptor affinity

“…To further assess the putative Fab and FcRn interactions, we analyzed the IgG-FcRn complex structure by negative stain electron microscopy, and observed several distinct classifications of IgG-FcRn complex ( Figure 2 ). The Fab arms of IgG appear to be highly dynamic as previously observed, 31 with multiple conformations previously predicted by Booth et al 32 These include the canonical Y-shaped conformation ( Figure 2(a and d )), a T-shaped conformation ( Figure 2(b and e )) or a mixed Y/T conformation ( Figure 2(c and f )). In addition, we observed that the soluble form of FcRn can bind to the IgG at either 1:1 ( Figure 2(b -d)) or 2:1 ( Figure 2(e -f)) stoichiometry.…”

Fc Sialylation Prolongs Serum Half-Life of Therapeutic Antibodies

“…Structurally, the Del mutation is well tolerated, as demonstrated by the conserved thermal stability. In this aspect, the Del mutation is an improvement over the well-known FcRn + YTE variant, which increases mAb serum t 1/2 in humans at the cost of highly decreasing the thermal stability of the mAb (83). In addition, as assessed by two in silico algorithms that predict epitope binding to human HLA alleles (Antitope [iTope and TCED] and Lonza [Epibase]), the Del mutation failed to generate neo-epitopes with consensus HLA-binding residues.…”

Characterization of two in vivo challenge models to measure functional activity of monoclonal antibodies to Plasmodium falciparum circumsporozoite protein

“…Monoclonal antibodies (mAbs) to CSP have been proposed as new infection-blocking interventions [9][10][11][12], and protection by mAbs can be expected to wane with the half-life of the antibody in serum. While altering the sequence of mAb Fc regions has been used to extend serum half-life of mAbs [13], an important additional approach for improving the durability of protection is to identify mAbs effective at lower concentrations. Identification and development of such potent antibodies will require assays that reliably measure their functional activity.…”