Single-cell RNA Sequencing Reveals Distinct Cellular Factors for Response to Immunotherapy Targeting CD73 and PD-1 in Colorectal Cancer

Recently, the programmed cell death protein 1 (PD-1) blockade has been in spotlight for its clinical success in the immune checkpoint response. This success has led to high demand for the development of novel anti-cancer immunotherapeutic drugs. Adenosine as a well-defined tumorigenic factor is generated extracellular by the ectoenzymes CD73 (also known as NT5E) and CD39 (also known as ENTPD1) from extracellular ATP. Thus development of adenosine pathway blockades has received considerable attention. Here, we aimed to reveal distinct response mechanism of a potent and CD73 ectoenzyme selective inhibitor in comparison with PD-1 blockade in colorectal cancer (CRC) to clarify its value as a novel immunotherapy. METHODS: We used a CD73 inhibitor, and a neutralizing antibody against murine PD-1 as the PD-1 blocker. To understand the distinct mechanism of CD73 inhibitor in comparison with PD-1 blockade, We performed single-cell RNA sequencing (scRNA-seq) of CD45+ tumor-infiltrating lymphocytes from an untreated control (n = 3) and AB680-treated (n = 3) and PD-1-blockade-treated murine CRC in vivo models. We also used flow cytometry analysis to validate our new findinds.