Properties of Human IgG1s Engineered for Enhanced Binding to the Neonatal Fc Receptor (FcRn)

Abstract

We describe here the functional implications of an increase in IgG binding to the neonatal Fc receptor. We have defined in a systematic fashion the relationship between enhanced FcRn binding of a humanized anti-respiratory sincytial virus (RSV) monoclonal antibody (MEDI-524) and the corresponding biological consequences in cynomolgus monkeys. The triple mutation M252Y/S254T/T256E (YTE) was introduced into the Fc portion of MEDI-524. Whereas these substitutions did not affect the ability of MEDI-524 to bind to its cognate antigen and inhibit RSV replication, they resulted in a 10-fold increase in its binding to both cynomolgus monkey and human FcRn at pH 6.0. MEDI-524-YTE was efficiently released from FcRn at pH 7.4 in both cases. We show that MEDI-524-YTE consistently exhibited a nearly 4-fold increase in serum half-life in cynomolgus monkeys when compared with MEDI-524. This constituted the largest half-life improvement described to date for an IgG in a primate. For the first time, we demonstrate that these sustained serum levels resulted in an up to 4-fold increase in lung bioavailability. Importantly, we also establish that our non-human primate model is relevant to human. Finally, we report that the YTE triple substitution provided a means to modulate the antibodydependent cell-mediated cytotoxicity (ADCC) activity of a humanized IgG1 directed against the human integrin ␣ v ␤ 3 . Therefore, the YTE substitutions allow the simultaneous modulation of serum half-life, tissue distribution and activity of a given human IgG1.

Extending human IgG half-life using structure-guided design

“…These include residues that make direct contact with FcRn, as well as peripheral and non-surface exposed residues that have the potential to modify the interaction surface and/or influence the dynamics of the 250-helix. Indeed, some of these identified positions have been previously investigated, such as within the half-life enhancing mutants YTE, 10 LS (M428L/N434S), 13 AAA (T307A/E380A/N434A), 27 QL (T250Q/M428L) 11 and V308P. 28 In our analysis, we first investigated the role of each residue in silico , and identified the network of interactions mediated by that residue (Figure 2).…”

“…10–13 The FcRn affinity-enhancing mutants in these studies were identified from phage display coupled with directed mutagenesis, 14–16 alanine scanning, 17 or, in select cases, from molecular modeling and rational design. 11,13 The prototypical example of an FcRn affinity-enhancing Fc mutant is the M252Y/S254T/T256E (YTE) mutation which, when incorporated into motavizumab IgG1, is able to extend serum half-life in humans by more than four-fold.…”

Generation and characterization of a high affinity anti-human FcRn antibody, rozanolixizumab, and the effects of different molecular formats on the reduction of plasma IgG concentration

“…We also present in vivo data in human FcRn-transgenic mice and in cynomolgus monkeys to investigate mechanism of action, and to allow characterization of pharmacokinetics (PK), pharmacodynamics (PD) and safety. The cynomolgus monkey was selected as a suitable species for investigation of the PK, PD and safety of human IgG, and mutants thereof, because human and cynomolgus monkey IgG4 bind comparably to cynomolgus monkey FcRn 26,27 and have similar Fc effector functions. 28 …”

Non-neutralizing antibodies increase endogenous circulating Ang1 levels

“…These mutations are reported to increase the half-life of antibodies in cynomolgus monkey up to 4 fold. 24 We therefore compared mAB-NN in wild-type IgG1 format with mAB-NN in IgG1 with YTE mutations in a multiple-dose cynomolgus monkey study. The molecules were administered once a week for three weeks, followed by a post-treatment observation period.…”