Abstract LB163: Blocking adenosine production with ORIC-533, a CD73 inhibitor with best-in-class properties, reverses immunosuppression in high-AMP environments

Adenosine is recognized for its anti-inflammatory and immunosuppressive properties. Signaling through adenosine receptors expressed on immune cells initiates a cascade that diminishes activation and proliferation of cytotoxic T-cells, impairs activity of natural killer cells and CD4+ effector T-cells, and promotes the expansion of immunosuppressive cell types. CD73, a cell surface ecto-5'-nucleotidase, converts AMP to adenosine and is a major catalyst of adenosine generation in the tumor microenvironment. Overexpression of CD73 is observed in many solid tumors and may correlate with unfavorable clinical outcomes; therefore, reducing the level of adenosine via inhibition of CD73 is a strategy for enhancing antitumor immune responses and overcoming therapeutic resistance. We have developed ORIC-533, a potent, orally bioavailable, AMP-competitive, small molecule inhibitor of CD73. ORIC-533 is highly selective for CD73 and exhibits picomolar potency in biochemical and cellular assays, blocking adenosine production from AMP. Expression of CD73 on immune cells is sufficient to drive immunosuppression in vitro when AMP is present. We have previously shown that ORIC-533 can rescue activation of CD8+ T- cells exposed to AMP and restore proliferation, cytokine secretion and CD25 expression at low nanomolar concentrations. To understand the role of CD73 in dendritic cell (DC) priming of helper CD4+ T-cells, we utilized co-culture of allogeneic immature DC and CD4+ T-cells, where AMP to adenosine conversion severely suppresses activation of helper CD4+ T-cells. Treatment with ORIC-533 completely restored activation of CD4+ T-cells in this setting. To better define the biologically relevant AMP concentrations likely to be encountered in tumors, we determined the concentration of AMP in a panel of primary patient tumors. We discovered that intratumoral AMP levels can reach up to 500 micromoles, emphasizing the requirement for CD73 inhibitors to be efficacious in a high AMP environment. We previously demonstrated that nanomolar concentrations of ORIC-533, unlike other CD73 inhibitors tested, can efficiently rescue cytotoxic T-cell function in the presence of millimolar AMP concentrations. Surprisingly, in these studies we found that inhibitors of adenosine receptors, A2A or A2A/B, can only rescue CD8+ T-cell function in the context of low micromolar AMP or adenosine and thus are likely to be ineffective in patient tumors with high AMP. Taken together, these preclinical r