INTERBIZ 2022: anti-CD39/anti-CD73 Poster submission  

<Abstract>

 Extracellular adenosine triphosphate (eATP) and its metabolized adenosine have emerged as important immune modulators in tumor microenvironment. In response to the tissue damage or cellular stress, ATP is actively released into the extracellular compartment. Contrary to the immunostimulatory function of eATP, its metabolic derivative, adenosine, suppresses the pro-inflammatory activities of the immune cells through A2AR and A2BR receptor signaling. Such conversion is performed by several ectonucleotides, such as CD39 and CD73, which are highly expressed not only in various solid tumors and hematologic malignancies, but also in immune cells, stromal cells, and endothelial cells. Therefore, we speculated that inhibiting their enzymatic activity may enhance the anti-tumor activity by both promoting the immuno-stimulatory eATP and preventing the production of immuno-suppressive adenosine.

 In this purpose, we generated bispecific antibody against CD39 and CD73 through the novel in-house antibody development platform. In brief, four individual human ScFv phage libraries were subjected to three rounds of panning against CD39 and CD73, respectively. Antigen binding polyclonal ScFv phages were regenerated into monoclonal ScFv phages, and were assessed for target binding validation, especially under the cellular context. The CDR region of the candidates with high antigen binding affinity were sequenced, and the seven to ten clones with unique CDR sequences for each protein were subjected to recombinant antibody production. Further selection against the antibodies was performed through the screening of binding epitope and affinity. The selected clones showed suppressive activity against CD39 and CD73, respectively. Importantly, we avoided the clones with similar binding epitope with BY40 (CD39) and Oleclumab (CD73) as they were poorly suppressive. Finally, each of the best clones was utilized in developing a bispecific antibody, which demonstrated the simultaneous binding capability against both CD39 and CD73 in the cellular context.