Use of Mass Cytometry to Profile Human T Cell Exhaustion

Mass cytometry has become an important technique for the deep analysis of single cell protein expression required for precision systems immunology. The ability to profile more than 40 markers per cell is particularly relevant for the differentiation of cell types for which low parametric characterization has proven difficult, such as exhausted CD8⁺ T cells (T_EX). T_EX with limited effector function accumulate in many chronic infections and cancers and are subject to inhibitory signaling mediated by several immune checkpoints (e.g., PD-1). Of note, T_EX represent considerable targets for immune-stimulatory therapies and are beginning to be recognized as a major correlate of successful checkpoint blockade approaches targeting the PD-1 pathway. T_EX exhibit substantial functional, transcriptomic and epigenomic differences compared to canonical functional T cell subsets [such as naïve (T_N), effector (T_EFF) and memory T cells (T_MEM)]. However, phenotypic distinction of T_EX from T_EFF and T_MEM can often be challenging since many molecules expressed by T_EX can also be expressed by effector and memory T cell populations. Moreover, significant heterogeneity of T_EX has been described, such as subpopulations of exhausted T cells with progenitor-progeny relationships or populations with different degrees of exhaustion or homeostatic potential that may directly inform about disease progression. In addition, T_EX subsets have essential clinical implications as they differentially respond to antiviral and checkpoint therapies. The precise assessment of T_EX thus requires a high-parametric analysis that accounts for differences to canonical T cell populations as well as for T_EX subset heterogeneity. In this review, we discuss how mass cytometry can be used to reveal the role of T_EX subsets in humans by combining exhaustion-directed phenotyping with functional profiling. Mass cytometry analysis of human T_EX populations is instrumental to gain a better understanding of T_EX in chronic infections and cancer. It has important implications for immune monitoring in therapeutic settings aiming to boost T cell immunity, such as during cancer immunotherapy.