AB0037 Evaluation of the binding kinetics and functional bioassay activity of sarilumab and tocilizumab to the human il-6 receptor (il-6r) alpha
Authors: A. Rafique, J. Martin, M. Blome, T. Huang, A. Ouyang, N. Papadopoulos
Journal: Annals of the Rheumatic Diseases (2013)

<h3>Background</h3> Sarilumab is the first fully human monoclonal antibody (mAb) directed against the interleukin-6 receptor alpha (IL-6Rα). Sarilumab was developed using VelocImmune<sup>®</sup> mice immunized with the human IL-6 (hIL-6) receptor. VelocImmune mice are genetically-engineered to express human antibody variable domain genes in the same robust fashion that the replaced mouse genes are typically expressed. Sarilumab is currently being explored as a new therapeutic modality for the treatment of rheumatoid arthritis. <h3>Objectives</h3> To evaluate the kinetic binding parameters and in vitro functional activity of two monoclonal antibodies directed against IL-6Rα: the fully human mAb sarilumab and the humanized mAb tocilizumab. <h3>Methods</h3> Kinetic binding parameters were measured using Surface Plasmon Resonance (SPR) technology. The ability to block hIL-6 induced activation of the human IL-6Rα was investigated using several bioassays; a human hepatocellular carcinoma cell line HepG2, transfected with a STAT3-luciferase reporter plasmid, as well as a proliferation assay using the human B-lymphoma cell line, DS-1. <h3>Results</h3> Sarilumab bound with high affinity to recombinant monomeric human and monkey IL-6 receptor with a <i>K</i><sub>D</sub> value of 61.9 pM and 71.9 pM, respectively. The binding affinity of sarilumab to the dimeric human IL-6 receptor Fc-fusion was 12.8 pM. Cross-reactivity to mouse IL-6 receptor was not observed using SPR, indicating that sarilumab is specific to human and monkey IL-6 receptor. In contrast, tocilizumab bound to monomeric and dimeric forms of the human IL-6 receptor with a 15-22 fold weaker affinity than that of sarilumab as determined by SPR. In the HepG2 cell luciferase reporter assay, sarilumab effectively blocked luciferase activity induced by 50 pM hIL-6 with an IC<sub>50</sub> of 146 pM and was ∼4 fold more potent than tocilizumab. Similarly, in the DS-1 cell proliferation assay, sarilumab effectively blocked growth induced by 1.0 pM hIL-6 with an IC<sub>50</sub> of 226 pM and was several fold more potent than tocilizumab. <h3>Conclusions</h3> Based on these in vitro assay data, sarilumab has both a higher relative binding affinity for IL-6Rα, blocks IL-6Rα activation, and inhibits IL-6-induced cellular responses such as cell proliferation at lower concentrations than tocilizumab. <h3>Acknowledgements</h3> VelocImmune<sup>®</sup> is a registered trademark of Regeneron Pharmaceuticals, Inc. <h3>Disclosure of Interest</h3> A. Rafique Shareholder of: Regeneron Pharmaceuticals, Inc., Employee of: Regeneron Pharmaceuticals, Inc., J. Martin Shareholder of: Regeneron Pharmaceuticals, Inc., Employee of: Regeneron Pharmaceuticals, Inc., M. Blome Shareholder of: Regeneron Pharmaceuticals, Inc., Employee of: Regeneron Pharmaceuticals, Inc., T. Huang Shareholder of: Regeneron Pharmaceuticals, Inc., Employee of: Regeneron Pharmaceuticals, Inc., A. Ouyang Shareholder of: Regeneron Pharmaceuticals, Inc., Employee of: Regeneron Pharmaceuticals, Inc., N. Papadopoulos Employee of: Regeneron Pharmaceuticals, Inc.