Cell Line Impact On Antigen Binding Of A Therapeutic Mab

AGUILAR, MF ; ATTALLAH, C; ETCHEVERRIGARAY, M ; OGGERO, M

Copenhage

Congreso; 26th Meeting of the European Society for Animal Cell Technology (ESACT); 2019

European Society for Animal Cell Technology (ESACT)

"Background and novelty: Monoclonal antibodies (mAbs) constitute a large and growing subset of the marketed-biotherapeutics glycoproteins. Although mAbs are bifunctional molecules since variable (V) and constant (C) regions are considered independents domains, and that glycosylation has no effect on antigen binding, there are several studies suggesting the influence of C regions of different mAbs with identical V regions on the antigen binding activity. We developed a novel humanized single-chain variable fragment (scFv) against human interferon-α2b (hIFN-α2b) fused to the human Fcgamma1 region (hz.scFv-Fc). The engineered antibody was subject to a UNL-CONICET patent application to be used as a therapeutic candidate for autoimmune diseases. In this work, we study the impact of the producer cell line on the affinity constant and antigen neutralizing ability of this novel molecule.Experimental approach: The hz.scFv-Fc antibody was produced in CHO-K1, HEK293 and NS0 cells. Affinity constants were measured by competitive ELISA and antigen-neutralizing ability was evaluated by three independent bioassays. N-glycosylation structures were analyzed by hydrophilic chromatography.Results and discussion: Results showed significant differences both in affinity and antigen-neutralization ability between these molecules, where hz.scFv-Fc produced in HEK293 cells presented the lowest affinity constant and antigen-neutralizing ability. After analyzing the N-glycosylation pattern, we found that both, the percentage of occupied N-glycan sites and the structure of the main oligosaccharides differ depending on the host cells. These results evidence that the producer cell line influences the mAb affinity and neutralizing action, leading to the conclusion that the host cells should be carefully taken into account in order to develop a new therapeutic antibody."