AB0086 Characterization of a human monoclonal antibody that neutralizes interleukin-20. (23rd January 2014)

• Record Type: Journal Article
• Authors: Pass, J.; Clausen, J.T.; Worsaae, A.; Nørby, P.L.; Andersen, M.D.; Thόrόlfsson, M.; Østergaard, S.; Pedersen, A.A.; Rømer, J.
• Is Part Of: Annals of the rheumatic diseases. Volume 71(2012)Supplement 3
• Page Start: 642
• Page End: 642
• Publication Date: 2014-01-23
• Subjects: Rheumatism -- Periodicals; 616.723005
• DOI: 10.1136/annrheumdis-2012-eular.86
• Languages: English
• ISSNs: 0003-4967

Abstract

Abstract : Background: Interleukin-20 (IL-20) is a member of the IL-10 cytokine family and signals via the IL-20R1/IL-20R2 and IL-20R2/IL-22R1 heterodimer receptor complexes. Elevated amounts of IL-20 have been demonstrated in autoimmune disease tissues such as psoriatic plaques and rheumatoid arthritis (RA) synovium. Neutralization of IL-20 may thus provide a new beneficial approach for treatment of patients with RA. Objectives: Generation and characterization of the neutralizing human anti-IL-20 monoclonal antibody (mAb), NNC0109-0012. Methods: Human mAbs were generated by immunization of transgenic mice bearing the human immunoglobulin G (IgG) locus (BMS/Medarex mice) with recombinant human IL-20. IL-20–specific antibodies were selected based on ELISA. Characterization included stability studies by circular dicroism, dynamic light scattering and size exclusion chromatography, as well as surface plasmon resonance analysis for determination of the kinetic parameters. Epitope mapping was performed by HX-MS and peptide array. A transient luciferase assay was performed using baby hamster kidney (BHK) cells transfected with IL-20R1/IL-20R2 or IL-20R2/IL-22R1 along with a STAT3 reporter construct. Proliferation assays were performed with BaF-3 cells transfected with IL-20R1/IL-20R2 heterodimer. Proliferation and inhibition experiments were conducted using 1 nM IL-20. Results: NNC0109-0012 was selected from a panel of human anti-IL-20 mAbs and was expressed as an IgG4 isotype containing a S241P mutation to reduce formation of half antibodies. A six-month stability study at 25 °C demonstrated insignificant changes in the biophysical parameters for NNC0109-0012. The equilibrium dissociation constant for the interaction between NNC0109-0012 and soluble IL-20, was 0.2 nM as measured by SPR analysis. The epitope for NNC0109-0012 on IL-20 was determined by HX-MS and peptide array. Amino acids V70-F92 constitute the binding interface on human IL-20, with complete protection observed for amino acids R83-F92 as determined by HX-MS experiments. The same epitope was identified using peptide array. An Ala-scan of the binding region showed that residues H79, R83 and N90 are the most critical for binding. Neutralization of the IL-20–mediated activation of IL-20R1/IL-20R2 and IL-20R2/IL-22R1 was demonstrated using a luciferase reporter assay. A concentration of 3 nM NNC0109-0012 completely neutralized the response generated by stimulation with 0.1 nM IL-20. The potency of NNC0109-0012 (IC50 ) was demonstrated to be 0.27 nM when measured as inhibition of proliferation of BaF-3 cells over-expressing the human IL-20 receptor. Conclusions: A novel, neutralizing human anti-IL-20 mAb (NNC0109-0012) has been generated and characterized. NNC0109-0012 binds to human IL-20 with high affinity, and effectively neutralizes the activity of IL-20 in cell-based assays. NNC0109-0012 is being further investigated in patients with RA. Disclosure of Interest: J. Pass Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, J. Clausen Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, A. Worsaae Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, P. Nørby Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, M. Andersen Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, M. Thόrόlfsson Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, S. Østergaard Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, A. Pedersen Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S, J. Rømer Shareholder of: Novo Nordisk A/S, Employee of: Novo Nordisk A/S