Effect of individual Fc methionine oxidation on FcRn binding: Met252 oxidation impairs FcRn binding more profoundly...

Area of Science:

• Biochemistry
• Immunology
• Pharmacology

Background:

• The neonatal Fc receptor (FcRn) mediates the long serum half-lives of monoclonal antibodies (mAbs) through pH-dependent binding to IgG-Fc.
• The human IgG1 Fc region contains three conserved methionine residues (Met252, Met358, Met428) susceptible to oxidation.
• Oxidation of these methionine residues has been shown to impair FcRn binding and affect the pharmacokinetics of therapeutic antibodies.

Purpose of the Study:

• To quantitatively assess the impact of individual methionine residue oxidation on Fc-FcRn binding.
• To define critical quality attributes related to methionine oxidation in therapeutic antibodies.
• To elucidate the specific roles of Met252, Met358, and Met428 in Fc-FcRn interactions.

Main Methods:

• Generation of two sets of homodimeric site-directed IgG1 mutations: Met to Leu mutants to block oxidation and Met to Gln mutants to mimic oxidation.
• Utilizing mutagenesis approaches to specifically investigate the effects of individual methionine oxidation on Fc-FcRn binding affinity and thermal stability.

Main Results:

• Oxidation of either Met252 or Met428 alone significantly impairs Fc-FcRn binding.
• Met252 oxidation demonstrated a more detrimental effect on FcRn binding compared to Met428 oxidation.
• Met428 oxidation exhibited a greater destabilizing effect on the thermal stability of IgG1-Fc.
• Oxidation of Met358 did not affect Fc-FcRn binding.
• Met to Gln mutations proved useful in mimicking methionine oxidation effects.

Conclusions:

• Individual oxidation of Met252 or Met428 critically impacts Fc-FcRn binding and antibody pharmacokinetics.
• Met252 and Met428 play distinct roles in Fc-FcRn interaction and Fc region stability.
• Met358 is not critical for FcRn binding.
• The Met to Gln mutagenesis strategy is a valuable tool for studying methionine oxidation in Fc-FcRn interactions.