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AB0037 Evaluation of the binding kinetics and functional bioassay activity of sarilumab and tocilizumab to the human il-6 receptor (il-6r) alpha
  1. A. Rafique1,
  2. J. Martin2,
  3. M. Blome1,
  4. T. Huang1,
  5. A. Ouyang2,
  6. N. Papadopoulos3
  1. 1Therapeutic Antibodies
  2. 2Bioassay, Molecular Biology & Protein Development
  3. 3Therapeutic Proteins, REGENERON PHARMACEUTICALS, INC., Tarrytown, United States

Abstract

Background Sarilumab is the first fully human monoclonal antibody (mAb) directed against the interleukin-6 receptor alpha (IL-6Rα). Sarilumab was developed using VelocImmune® 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.

Objectives 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.

Methods 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.

Results Sarilumab bound with high affinity to recombinant monomeric human and monkey IL-6 receptor with a KD 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 IC50 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 IC50 of 226 pM and was several fold more potent than tocilizumab.

Conclusions 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.

Acknowledgements VelocImmune® is a registered trademark of Regeneron Pharmaceuticals, Inc.

Disclosure of Interest 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.

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