Immunoglobulin G (IgG) is the primary antibody in blood and human serum, with IgG1 being its predominant subclass. These antibodies possess two fragments: the fragment antigen binding domain (Fab), responsible for pathogen recognition, and the fragment crystallizable (Fc) domain, which interacts with the immune cells to induce potent effector functions. These effector functions, such as Antibody-dependent cellular cytotoxicity (ADCC), are triggered when receptors on Natural Killer (NK) cells, like CD16a (FcγRIIIa), interact with the Fc portion of IgG1 antibodies. The strength of this response is determined by the binding affinity, which is affected by glycosylation patterns on both the Fc region and the CD16a receptor. Therefore, controlling Fc glycosylation is a key strategy for developing high-affinity antibodies. Research has demonstrated that when glycans are absent from the Fc region, complex formation does not occur, highlighting the importance of studying glycan effects.
The goal of this study, in collaboration with Amgen, is to understand how specific glycan architecture on the IgG1 Fc impacts its binding affinity with the CD16a receptor. The Fc has two potential glycosylation sites, one on each arm, and it has been observed in experiments that the presence of biantennary high mannose and biantennary fucose greatly reduces the strength of binding. However, it is not yet clear how combinations of the aforementioned glycan types along with other complex glycans impact the binding affinity. To elucidate this, we have performed large-scale all-atom Molecular Dynamics (MD) simulations of the IgG1 Fc-CD16a complex with varying degrees of glycosylation. We performed binding energy calculations using MMPBSA, and our results corroborate the findings that the addition of fucose to a complex glycan negatively impacts binding affinity. The underlying mechanism was identified to be the disruption of protein-protein interactions between the Fc region and CD16a receptor, thereby orienting the complex to a less energetically favorable conformation. To obtain improved binding energy measurements, Free Energy Perturbation calculations using the BFEE2 tool are being adopted. We expect to be able to accurately predict the affinities for a combination of different complex glycans with high mannose. The findings would help us to design antibodies that will enable modulation of improved receptor binding, leading to improved affinities and improved antibody efficacy.