Modelagem e simulação de dinâmica molecular da interação do scFv do anticorpo FMC63 com o receptor de membrana CD19 .

Abstract

A new class of cancer treatment immunotherapy (CAR-T cells) is based on the use of patient-derived T cells that are genetically modified to express a chimeric antigen receptor (CAR). This CAR is responsible for redirect the T cells action against malignant cells that express a specific antigen. The CD19 receptor is a target for immunotherapies because it is a molecule widely expressed in B-cell malignancies. The CAR structure responsible for recognizing an antigen is usually composed of a single-chain variable fragment (scFv). To enable the improvement proposition in the action of CAR-expressing T cells, it is first necessary to understand how the interaction between scFv and the target antigen occurs, considering that it is the central mechanism that activates the cytotoxic response. The scFv of this study is composed of light (VL) and heavy (VH) chain variable domains derived from a murine antibody, FMC63, which are connected by a linker (G4S)3. The crystal structure of the CD19 extracellular domain used in this study has the structure deposited in the PDB, code 6AL5. The Modeller 9.20 program was used to model the scFv structure and the missing loops in the CD19 molecule. Both structures were submitted to molecular dynamics (MD) simulation for 300 ns. Subsequently, the interaction system between scFv and CD19 was constructed by molecular docking through Haddock 2.2 program. The scFv-CD19 system was obtained and simulated for 1800 ns. The molecules structural stability was determined by the root-mean-square deviation (RMSD). The Intermolecular Interaction Potential (IIP) between scFv and CD19 was measured along the simulation. There was a relative VL conformation change observed between 550 ns and 1150 ns. The CD19 structure showed the same RMSD when simulated alone and when simulated together with scFv, indicating that the presence of scFv does not significantly affect its structural equilibrium. The IIP analysis showed a strong attractive interaction between the structures until approximately 1100 ns, where it was possible to observe that the most attractive interaction CDRs are H2 and H3 CDRs. Thus, it was possible to observe that scFv derived from antibody FMC63 lose the interaction with CD19 recognition epitope and that this behavior can be an advantage for its use in the structure of commercially used CARs since they interact with CD19 in the membrane of cancer cells, but only temporarily.