Abstract LB-115: An orally bioavailable inhibitor of CD73 reverts intratumoral immunosuppression and promotes anti-tumor responses
Authors: Metzger Todd , Blank Brian , Chan Brenda , Chen Chelsea , Chen Yuping , Du Xiaohui , Duong Frank , Friedman Lori , Junttila Melissa , Kawai Hiro , Kong Wayne , Moore Jared , Pham Johnny , Rew Yosup , Sun Daqing , Sun Jessica , Sutimantanapi Dena , Wu Kejia , Yeh Chien-Hung , Yuen Natalie , Zavorotinskaya Tatiana 
Published: September 2020

Abstract
The 5' ecto-nucleotidase CD73 is dynamically expressed in distinct biological contexts as a means to increase the adenosine metabolite, which functions to limit immune activation and prevent excessive inflammation. CD73 activity is required for the final step of adenosine production. In the context of tumors, excessive CD73-mediated adenosine generation impairs anti-tumor immune responses. Here, we show that an orally bioavailable small molecule inhibitor of CD73 can prevent adenosine generation, relieve adenosine-driven immunosuppression within the tumor in vivo and generate anti-tumor efficacy.
In vitro analysis confirmed the substantial suppressive effects of adenosine on murine CD8+ T cells, cross-presenting dendritic cells, and macrophages. NECA, a stable analog of adenosine, impaired immature bone-marrow precursor differentiation into CD103+ dendritic cells, a subset which is critical for tumor-derived antigen presentation to naïve T cells. Interestingly, NECA also inhibited the ability of macrophages to upregulate the M1 marker, CD80, in both the presence and absence of additional polarizing stimuli. Furthermore, OR-558, a CD73 small molecule inhibitor, could fully inhibit CD73-mediated adenosine production and completely restore T cell activation at sub-nanomolar concentrations.
Prior work in CD73 germline knock-out animals has shown that EG7 syngeneic tumor growth is highly dependent on CD73. Building on this knowledge, we next we sought to evaluate CD73 inhibition on AMP to adenosine conversion ex vivo in EG7 syngeneic tumor extracts and indeed observed that OR-558 treatment could significantly impair adenosine production within the tumor milieu. This finding translated to the in vivo tumor setting, where single agent anti-tumor efficacy was observed with continuous OR-558 treatment. Anti-tumor efficacy coincided with significantly decreased intra-tumoral adenosine levels and immune modulation, such as increased intratumoral T cell activation, dendritic cell maturation, and M1 macrophage polarization. Lastly, we determined the bioavailability of orally dosed OR-558 in EG7 tumor-bearing mice and found that significant single agent anti-tumor efficacy was also achievable with once-daily oral dosing of OR-558. Taken together, these data demonstrate that adenosine has pleiotropic immunosuppressive effects and inhibition of CD73-mediated adenosine production in vivo is sufficient to reverse these immune effects and enhance anti-tumor immunity. These results support clinical development of orally bioavailable CD73 small molecule inhibitors.
Citation Format: Todd Metzger, Brian Blank, Brenda Chan, Chelsea Chen, Yuping Chen, Xiaohui Du, Frank Duong, Lori Friedman, Melissa Junttila, Hiro Kawai, Wayne Kong, Jared Moore, Johnny Pham, Yosup Rew, Daqing Sun, Jessica Sun, Dena Sutimantanapi, Kejia Wu, Chien-Hung Yeh, Natalie Yuen, Tatiana Zavorotinskaya. An orally bioavailable inhibitor of CD73 reverts intratumoral immunosuppression and promotes anti-tumor responses [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 LB-115.