Immunophenotypical characterization of immune checkpoint receptor expression in cynomolgus monkeys and human healthy volunteers

China Bioanalysis Forum 2025 -- T cell-based immunotherapies [i.e., the immune checkpoint (IC) T-cell engagers] targeting IC receptors or their ligands, have revolutionized treatment paradigms in in the field of oncology. However, IC receptors are not easily characterized in healthy individuals due to the very low expression in immunologically unchallenged nonclinical species and healthy human donors under homeostatic conditions. Therefore, we have characterized expression of a broad range of IC receptors (CTLA-4, PD-1, PD-L1, 4-1BB, LAG-3, OX-40, TIGIT, GITR and ICOS) in peripheral blood mononuclear cell (PBMC) subpopulations from cynomolgus monkeys and healthy human volunteers, under resting and T-cell stimulatory conditions by multicolor flow cytometry. To our knowledge, this is the first study identifying similarities and differences in the levels of key checkpoint receptors under resting and T-cell stimulation conditions in human and cynomolgus macaque samples. In addition, in order to support clinical trials, we successfully developed an ex vivo fresh whole blood stimulation method to upregulate the expression of multiple IC receptors and validated the immunophenotypical characterization method for these IC receptors.