“…Given that amino acid residues involved in FcRn binding, such as M252 and I253, are located on the 250-helix, the observed displacement of the helix is expected to influence the affinity of Fc for FcRn. 20 Furthermore, the difference in the relative orientation of CH2 with respect to CH3 highlights the dynamic nature of the CH2-CH3 domains at different pHs (Supplementary Figure 1). 22 Recently, deuterium exchange studies of human Fc were performed in the absence or presence of FcRn.…”
Section: Resultsmentioning
confidence: 99%
“…Mutation of either of the two histidine residues significantly reduces the binding affinity of Fc with FcRn. 20,21 In addition to His310 and His435, several other Fc residues are involved in making molecular contacts with FcRn (Figure 1). 10.1080/19420862.2018.1490119-F0001Figure 1.( top ) Structural view of human IgG CH2-CH3 domains ( yellow ) in complex with human FcRn α ( green ) and β2M ( cyan ) domains.…”
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.
“…Given that amino acid residues involved in FcRn binding, such as M252 and I253, are located on the 250-helix, the observed displacement of the helix is expected to influence the affinity of Fc for FcRn. 20 Furthermore, the difference in the relative orientation of CH2 with respect to CH3 highlights the dynamic nature of the CH2-CH3 domains at different pHs (Supplementary Figure 1). 22 Recently, deuterium exchange studies of human Fc were performed in the absence or presence of FcRn.…”
Section: Resultsmentioning
confidence: 99%
“…Mutation of either of the two histidine residues significantly reduces the binding affinity of Fc with FcRn. 20,21 In addition to His310 and His435, several other Fc residues are involved in making molecular contacts with FcRn (Figure 1). 10.1080/19420862.2018.1490119-F0001Figure 1.( top ) Structural view of human IgG CH2-CH3 domains ( yellow ) in complex with human FcRn α ( green ) and β2M ( cyan ) domains.…”
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.
“…The 1519.g57 Fab’ binding site on human FcRn in solution was determined by HDX and was consistent with that determined by X-ray crystallography (data not shown).10.1080/19420862.2018.1505464-F0001Figure 1.Crystal structure of complex of human FcRn (deglycosylated extracellular domain) and 1519.g57 Fab’. (A) The binding epitope of 1519.g57 Fab’ on FcRn α-chain is shown in red; (B) PDB 4N0U – human IgG Fc domain interacting with FcRn, demonstrating the overlapping epitope with that of 1519.g57Fab’, 29 (C) FcRn α-chain sequence, showing residues involved in interaction with 1519.g57 Fab’ (in red ). Residues involved in interaction between human FcRn and IgG (Fc domain) or albumin (as described by Oganesyan et al, 2014, highlighted in yellow or in blue, respectively) are also shown.…”
Section: Resultsmentioning
confidence: 99%
“…Residues involved in interaction between human FcRn and IgG (Fc domain) or albumin (as described by Oganesyan et al, 2014, highlighted in yellow or in blue, respectively) are also shown. The sequence of human FcRn α-chain ECD was taken from Swiss Prot (P55899-1); Residue F44, identified by Oganesyan et al 29 as interacting with albumin, is residue E44 in this sequence and in the sequence of Schmidt et al 30 Dark blue = FcRn α-chain; pale blue = β2 M; magenta = 1519.g57 heavy chain; orange = 1519.g57 light chain; yellow = IgG Fc domain; red = 1519.g57 Fab’ binding epitope on FcRn.…”
Section: Resultsmentioning
confidence: 99%
“…(A) The binding epitope of 1519.g57 Fab’ on FcRn α-chain is shown in red; (B) PDB 4N0U – human IgG Fc domain interacting with FcRn, demonstrating the overlapping epitope with that of 1519.g57Fab’, 29 (C) FcRn α-chain sequence, showing residues involved in interaction with 1519.g57 Fab’ (in red ). Residues involved in interaction between human FcRn and IgG (Fc domain) or albumin (as described by Oganesyan et al, 2014, highlighted in yellow or in blue, respectively) are also shown.…”
Rozanolixizumab (UCB7665), a humanized high-affinity anti-human neonatal Fc receptor (FcRn) monoclonal antibody (IgG4P), has been developed to reduce pathogenic IgG in autoimmune and alloimmune diseases. We document the antibody isolation and compare rozanolixizumab with the same variable region expressed in various mono-, bi- and trivalent formats. We report activity data for rozanolixizumab and the different molecular formats in human cells, FcRn-transgenic mice, and cynomolgus monkeys. Rozanolixizumab, considered the most effective molecular format, dose-dependently and selectively reduced plasma IgG concentrations in an FcRn-transgenic mouse model (no effect on albumin). Intravenous (IV) rozanolixizumab dosing in cynomolgus monkeys demonstrated non-linear pharmacokinetics indicative of target-mediated drug disposition; single IV rozanolixizumab doses (30 mg/kg) in cynomolgus monkeys reduced plasma IgG concentration by 69% by Day 7 post-administration. Daily IV administration of rozanolixizumab (initial 30 mg/kg loading dose; 5 mg/kg daily thereafter) reduced plasma IgG concentrations in all cynomolgus monkeys, with low concentrations maintained throughout the treatment period (42 days). In a 13-week toxicology study in cynomolgus monkeys, supra-pharmacological subcutaneous and IV doses of rozanolixizumab (≤ 150 mg/kg every 3 days) were well tolerated, inducing sustained (but reversible) reductions in IgG concentrations by up to 85%, with no adverse events observed. We have demonstrated accelerated natural catabolism of IgG through inhibition of IgG:FcRn interactions in mice and cynomolgus monkeys. Inhibition of FcRn with rozanolixizumab may provide a novel therapeutic approach to reduce pathogenic IgG in human autoimmune disease. Rozanolixizumab is being investigated in patients with immune thrombocytopenia (NCT02718716) and myasthenia gravis (NCT03052751).
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