BEYOND BINDING: THE CDR EFFECT ON INTER-CHAIN INTERACTIONS OF MONOCLONAL ANTIBODIES

Monoclonal antibodies (mAbs)  have stood out among high-precision therapies, which led to the design of these molecules to improve their therapeutic properties. The main focus of this optimization is the complementarity-determining regions (CDRs) that directly affect their binding property. However, the impact of these changes on the protein dynamics is underexplored. Here, we investigated the impact of CDRs composition on interactions of the light and heavy chains of antibodies. We used Pertuzumab and Trastuzumab, two clinically approved mAbs, that differ only in their paratope composition. All-atom Molecular Dynamics (MD) simulations (five replicas of 200 ns each) were performed for each antibody in antigen-bound and unbound conditions. Inter-chain interaction energy was calculated with MM/PBSA analysis and decomposed by amino acids. The residues contributing ≤ –2.5 kcal/mol were selected for further investigation. Inter-chain contacts were calculated with a custom Python script. Two residues were considered in contact if they are within 3 Å for ≥ 75 % of the frames. Interestingly, we found that CDR residues TYR 91 (CDRL3) and PHE 99A / TYR 99B (CDRH3) directly contributed to Pertuzumab’s inter-chain energy of interaction, but not for Trastuzumab. Moreover, the TRP 110, a non-CDR residue of the variable domain (named as framework region– FW), is a strong contributor only in Trastuzumab systems (unbound = -4.03 ± 0.03 kcal/mol; antigen-bound = -3.28 ± 0.04 kcal/mol). Despite both antibodies having this residue, its strong contribution only in Trastuzumab indicates that CDR changes impact the energetic function of other residues. Regarding the contacts, CDR-CDR interactions occur only in Pertuzumab, but CDR-FW and FW-FW contacts were observed in all systems. Also, the FW-FW contact partners differ between these mAbs, indicating paratope also indirectly influences the inter-chain network. Additionally, the differences were not limited to the variable domains, as the PHE118 residue in the constant domain of light-chain display different contacts profile between Pertuzumab and Trastuzumab, showing that CDR changes propagate to other domains. These results demonstrate that CDR composition impacts inter-chain interactions both directly and indirectly, highlighting the inter-chain analysis as a new topic to be considered in antibody design.