Metal ion interactions with mAbs: Part 1.

Area of Science:

• Biopharmaceutical Chemistry
• Protein Degradation Mechanisms
• Antibody Stability

Background:

• Fragmentation in the hinge region of IgG1 monoclonal antibodies (mAbs) compromises product stability, potency, and efficacy.
• Metal ions, particularly Cu(2+), can bind to mAbs, leading to hydrolysis or oxidation and subsequent molecular cleavage.

Purpose of the Study:

• To elucidate the mechanism of Cu(2+)-mediated fragmentation in IgG1 mAbs.
• To investigate the influence of pH on hinge region cleavage products and their formation rates.
• To analyze chemical changes using model peptides derived from the mAb hinge region.

Main Methods:

• Studied Cu(2+)-mediated mAb fragmentation rates as a function of pH, with and without Cu(2+).
• Analyzed fragmentation using model linear and cyclic peptides (SCDKTHTC) from the mAb upper hinge region.
• Investigated chemical changes and degradation products.

Main Results:

• Cu(2+)-mediated fragmentation occurs predominantly via a hydrolytic pathway in solution.
• Cleavage sites and products are dependent on pH and molecular strain.
• Fragmentation rates are significantly slower in acidic environments compared to higher pH.
• Degradation products differ between linear and cyclic peptides, though reaction rates are similar.
• Modifying histidine residues, a potential metal binding site, reduces mAb fragmentation.

Conclusions:

• Cu(2+) induces IgG1 mAb hinge region fragmentation primarily through hydrolysis, influenced by pH and strain.
• Histidine modification offers a strategy to enhance mAb stability by preventing copper-mediated degradation.
• A charged residue may stabilize a structure involved in hydrolysis, forming a copper-binding pocket that increases hinge region susceptibility.