Elevated cleavage of human immunoglobulin gamma molecules containing a lambda light chain mediated by iron and histidine

Monoclonal antibodies in liquid formulation undergo nonenzymatic hydrolysis when stored at 5 °C for extended periods. This hydrolysis is enhanced at extreme pH and high temperature. In this study we discover that iron in the presence of histidine also enhanced cleavage of human immunoglobulin gamma (IgG) molecules containing a lambda light chain when incubated at 40 °C. The level of cleavage was concentration dependent on both iron and histidine levels. Enhanced cleavage with iron and histidine was not observed on IgG molecules containing a kappa light chain. Using CE-SDS to quantify levels of Fab + Fc, the Fab arm, and free light chain (LC) and heavy chain (HC) fragments, we show that cleavage resulted in elevated levels of free light and heavy chain fragments. Using MS we find elevated scission between residues E/C on the LC resulting in LC fragment 1–215. We also observed enhanced cleavage between S/C residues of the HC resulting in HC fragment 1–217. The corresponding Fab + Fc fragment beginning with cys-218 was not found. Instead, we find elevation of a Fab + Fc fragment that began with aspartic acid (cleavage between C/D). Further studies to understand how iron and histidine enhance cleavage of lambda light chain IgG molecules are ongoing.