Monoclonal antibodies targeting immune checkpoints have revolutionized oncology. Yet, the effectiveness of these treatments varies significantly among patients, and they are associated with unexpected adverse events, including hyperprogression. The murine research model used in drug development fails to recapitulate both the functional human immune system and the population heterogeneity. Hence, a novel model is urgently needed to study the consequences of immune checkpoint blockade. Dogs appear to be uniquely suited for this role. Approximately 1 in 4 companion dogs dies from cancer, yet no antibodies are commercially available for use in veterinary oncology. Here we characterize two novel antibodies that bind canine PD-1 with sub-nanomolar affinity as measured by SPR. Both antibodies block the clinically crucial PD-1/PD-L1 interaction in a competitive ELISA assay. Additionally, the antibodies were tested with a broad range of assays including Western Blot, ELISA, flow cytometry, immunofluorescence and immunohistochemistry. The antibodies appear to bind two distinct epitopes as predicted by molecular modeling and peptide phage display. Our study provides new tools for canine oncology research and a potential veterinary therapeutic.

Department of Biochemistry and Microbiology University of Victoria Victoria BC Canada

Institute for Adaptive and Neural Computation School of Informatics University of Edinburgh Edinburgh United Kingdom

Institute of Genetic and Molecular Medicine University of Edinburgh Edinburgh United Kingdom

Intercollegiate Faculty of Biotechnology of University of Gdansk and Medical University of Gdansk Gdansk Poland

International Centre for Cancer Vaccine Science University of Gdansk Gdansk Poland

Research Centre for Applied Molecular Oncology Masaryk Memorial Cancer Institute Brno Czechia

The Royal School of Veterinary Studies and The Roslin Institute University of Edinburgh Midlothian United Kingdom

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