Modelagem e simulação de dinâmica molecular de fragmentos de scFv derivados do anticorpo m971

Abstract

Acute Lymphoid Leukemia is considered one of the most aggressive types of cancer. Thus, Adoptive Cell Therapy based on CAR (Chimeric Antigen Receptor) is an example of one of the immunotherapies developed that seeks to achieve remission of this malignancy and is based on the expression of a CAR, which has the function of rescaling the action of T cells, enabling them to be able to recognize specific antigens on the surface of tumor cells. CAR is responsible for antigen recognition in tumor cells and, in the structure of CAR, the single-chain variable fragment (scFv), which is formed by the variable light (VL) and variable heavy (VH) domains of a monoclonal antibody (mAb ) and the linker, which connects the two portions, is responsible for the interaction with the tumor antigen. Recently, studies of CAR obtained from the m971 antibody have experienced a great growth, because the said antibody recognizes the protein, CD22, present exclusively in B cells and has an inhibitory function, making it an attractive target for therapies based on CARs . In view of this, the present work aimed initially to carry out a comparative study, via Molecular Dynamics (DM) simulation, of structural determinants of scFv models in aqueous solution, in order to verify the influence of linker size on the structure of scFvs derived of the m971 antibody. The crystallographic coordinates of the Fab portion of the m971 antibody are deposited in the Protein Data Bank, code 7052. Therefore, to obtain m971-derived scFv models, the MODELLER program was used with the missing residues modeling protocol only to add the linker between the VH and VL domains. Then, the scFvs were submitted to DM simulation for 500 ns. Subsequently, analyzes of structural determinants of the scFvs submitted to DM simulation were carried out. The results of the RMSD analyzes suggest that the VH and VH+VL domains of the long scFv have greater structural movement than the VH and VH+VL domains of the short scFv, with mean RMSD values of the VH+VL domain of the short scFv linker and VH+VL domain of scFv with long linker, 0.19 ± 0.01 and 0.22 ± 0.02nm, respectively. Finally, the results obtained in this work make it possible to carry out a more complete study later, considering the scFv's interacting with the CD22 antigen, in order to verify whether the change in the size of the scFv linker (scFv with long linker to scFv with short linker) will benefit the scFv-CD22 interaction and, consequently, may increase the effectiveness of CAR against the CD22 antigen, a marker of leukemic cells.