LAG-3 sustains TOX and regulates the CD94/NKG2-Qa-1b axis to govern exhausted CD8 T cell NK receptor expression and cytotoxicity

Exhausted CD8 T cells (Tex) found in chronic viral infection and cancer are defined by sustained co-expression of inhibitory receptors (IRs). Tex can be reinvigorated by blocking IRs, such as PD-1, but synergistic reinvigoration of Tex and enhanced disease control can be achieved by co-targeting multiple IRs including co-blockade of PD-1 and LAG-3, a treatment recently FDA-approved for melanoma. Despite the promise of anti-PD-1/anti-LAG-3 therapy, the cellular and molecular changes intrinsic to CD8 T cells when one or more of these IR pathways are disrupted remain poorly understood. To dissect these IR pathways and their interactions with each other, we used a quadruple co-transfer model to investigate the impact of singular or combined loss of PD-1 and LAG-3 on the differentiation of Tex during chronic viral infection. These analyses revealed distinct roles of PD-1 and LAG-3 in regulating Tex proliferation and cytokine/cytotoxic functions, respectively. Moreover, these studies identified key roles for LAG-3 that complement PD-1 biology including an essential role for LAG-3 in sustaining TOX expression and associated Tex durability. Finally, these studies uncovered a LAG-3-dependent circuit for the generation of an effector-like CD94/NKG2+ subset of Tex capable of enhanced cytotoxicity mediated by recognition of the stress-induced ligand Qa-1b with similar regulation of CD94/NKG2 family members in humans with combination anti-PD-1/anti-LAG-3. These analyses identify a role for LAG-3 in governing CD8 T cell Qa-1b-restricted immune sensing and surveillance. Together, these results disentangled the underlying non-redundant mechanisms of PD-1 and LAG-3 and their synergy in regulating Tex revealing additional immunotherapeutic opportunities based on these pathways.