Modelagem computacional e expressão heteróloga de nanocorpos com potencial inibidor de VLA-4

Abstract

The α4β1 or VLA 4 integrin is a transmembrane protein widely expressed on the surface of immune cells, mainly on monoc ytes and T lymphoc ytes, being involv ed in the mechanism of leukocyte extravasation, an essential step f or the development of inflammator y processes. As a r esult, VLA 4 is associated with chronic inflammatory diseases of great public health impact, such as A sth ma, R heumatoid A rthritis and M ul tiple S clerosis, thus being an important target for therapies for h uman pathologies A commercialize d anti VLA 4 compoun d is Natalizumab, a monoclonal antibody used in the treatment of M ultiple S clerosis, associated, howe ver, with high pr oduction cost and increased risk for the opportunistic infection o f Progressive Multi focal Leukoencephalopathy, in add ition to not being s pecific for this integrin . In this context, the objective of this work is to propose a three dimensional structure for three nanobodies with VLA 4 inhibitory potential and carry out thei r productio n in a heterologous expression system, as initial strategies for the development of VLA 4 antagonists in the VHH format. Thus, the three dimensional structures of these antibodies, obtained by comparative modeling using the MODELLER program, are presented, as we ll as the soluble expression in E. coli SHuffle pLysS, at 15 º C, for 16 h, and efficient purification by affinity chromatography . In summary, the results are promising in the development of anti VLA 4 nanobodies and guide the n ext work steps, w hich i nclude the r efinement of the modeling of the CDRs of the pre dicted structures, refinement of the purification me thodology, quantific ation of the yield and functional analysis of these nanobodies