In vivo and in vitro characterization of RVU330 best-inclass dual A2A/A2B adenosine receptor antagonist

The immunosuppressive role of adenosine attracts attention of many researchers as an interesting target for anticancer therapy. High concentrations of adenosine detected in many tumor models affect several subpopulations of infiltrating immune cells e.g. T lymphocytes, dendritic cells and macrophages. In these cells, adenosine signals via either A2A or A2B receptor, therefore the highest efficiency in reversal of adenosine mediated immunosuppression should be achieved by dual antagonism to both receptors. Here, we present data for best-in-class dual A2A/A2B antagonist RVU330 that exerts nanomolar potency in tumor-like adenosine-rich environment. By head-to-head comparison of our in-house developed dual A2A/A2B antagonist to compounds currently pursued in clinical development, we have demonstrated the best-in-class characteristics of our molecule. RVU330 maintains sub- or low nanomolar antagonistic potency in a high adenosine conditions on both A2A and A2B receptor. Multilevel pharmacological characterization spans from receptor binding and cAMP accumulation (assays in cell lines overexpressing target receptors) to functional assays in primary human immune cells which demonstrate the reversibility of immune suppression induced by adenosine. Our compound shows superior activity in antagonizing adenosine-mediated immunosuppression in CD4+ and CD8+ T-cells (IL2 and TNFα production), monocyte derived dendritic cells - moDCs (IL12 and TNFα production, driven mainly by A2B receptors) and macrophages (suppression of VEGF release by M2). High activity in human whole blood assay demonstrates that low nanomolar concentration is sufficient to fully antagonize adenosine. Therapeutic efficacy of RVU330 has been confirmed in syngeneic in vivo mouse model. New best-in-class dual A2A/A2B adenosine receptor antagonists RVU330 combines two crucial activities which addresses distinct, cell type mediated mode of action of adenosine related immunosupression. The dual A2A/A2B antagonistic profile as well as the ability to inhibit both receptors at very high adenosine conditions are key features differentiating RVU330 from other adenosine receptor antagonist. The RVU330 is currently being evaluated in preclinical studies. Citation Format: Michal Galezowski, Katarzyna Dziedzic, Paulina Wegrzyn, Aniela Golas, Magdalena Bonkowska, Karolina Grycuk, Mateusz Ogorek, Aneta Bobowska, Joanna Szeremeta-Spisak, Marcin Nowogrodzki, Grzegorz Satala, Iwona Łozinska-Raj, Przemyslaw Wyrebek, Marcelina Dudek, Anita Janiga, Marek Wronowski, Magdalena Zastawna, Mateusz Świrski, Luigi Stasi, Peter Littlewood, Krzysztof Brzozka, Mateusz O. Nowak. In vivo and in vitro characterization of RVU330 best-in-class dual A2A/A2B adenosine receptor antagonist [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5555.