Ab1804

Probing antinuclear antibody specificities by peptide phage display libraries

M.H. Hansen¹, A. Dybwad², Ø. Førre³, M. Sioud¹

¹Institute for Cancer Research, Department of Immunology, The Norwegian Radium Hospital, Montebello; ²The Norwegian Medicines Control Authority; ³The National Hospital, Center for Rheumatic Diseases, Oslo, Norway.

ABSTRACT
Objective
To uncover the specificities of autoantibodies to nuclear proteins (ANA) in patients with juvenile rheumatoid arthritis (JRA).

Methods
Peptide ligands for ANA were selected by panning random peptide phage display libraries on antibodies binding to HEp-2 cells. Positive phage clones were identified by the immunoscreening technique.

Results
Groups of peptides were identified, some of which share the core motifs of KTTTnPY, RVADnL/I or RnNSPL. Perinuclear and nuclear staining of HEp-2 cells were obtained with patient serum antibodies binding to the phage displaying the core peptide motifs. In contrast, no significant reactivity was seen with the antibodies binding to the wild type phage. Antibodies to the phage displaying peptides containing some of the core motifs were detected more frequently in ANA-positive as compared to ANA-negative JRA patients. Homology search with the selected core motifs revealed a significant homology with a number of human nuclear proteins and proteins from potential infectious agents that could serve as trigger in the breakdown of tolerance.

Conclusion
Panning of phage display libraries on antibodies reacting with cellular structures can lead to the identification of their specificities. Thus, the peptide epitopes reported here constitute additional information that may lead to the development of diagnostic tests and the identification of the parental antigens that initiated the B cell responses in patients with JRA.

Key words
Phage display, peptide library, antinuclear antibodies, JRA.

This work was supported by a grant from the Norwegian Foundation for Health and Rehabilitation to M. Sioud.

Please address correspondence and reprint requests to: Mouldy Sioud, DEA Pharm, PhD, Institute for Cancer Research, Department of Immunology, The Norwegian Radium Hospital, Montebello, N-0310 Oslo, Norway.
E-mail: mosioud@embnet.uio.no