We explored the mechanism of action of CD39 antibodies that inhibit ectoenzyme CD39 conversion of extracellular ATP (eATP) to AMP and thus potentially augment eATP–P2-mediated proinflammatory responses. Using syngeneic and humanized tumor models, we contrast the potency and mechanism of anti-CD39 mAbs with other agents targeting the adenosinergic pathway. We demonstrate the critical importance of an eATP–P2X7–ASC–NALP3-inflammasome–IL18 pathway in the antitumor activity mediated by CD39 enzyme blockade, rather than simply reducing adenosine as mechanism of action. Efficacy of anti-CD39 activity was underpinned by CD39 and P2X7 coexpression on intratumor myeloid subsets, an early signature of macrophage depletion, and active IL18 release that facilitated the significant expansion of intratumor effector T cells. More importantly, anti-CD39 facilitated infiltration into T cell–poor tumors and rescued anti–PD-1 resistance. Anti-human CD39 enhanced human T-cell proliferation and Th1 cytokine production and suppressed human B-cell lymphoma in the context of autologous Epstein–Barr virus–specific T-cell transfer. Significance: Overall, these data describe a potent and novel mechanism of action of antibodies that block mouse or human CD39, triggering an eATP–P2X7–inflammasome–IL18 axis that reduces intratumor macrophage number, enhances intratumor T-cell effector function, overcomes anti–PD-1 resistance, and potentially enhances the efficacy of adoptive T-cell transfer. This article is highlighted in the In This Issue feature, p. 1631

CD96 is a novel target for cancer immunotherapy shown to regulate NK cell effector function and metastasis. Here, we demonstrated that blocking CD96 suppressed primary tumor growth in a number of experimental mouse tumor models in a CD8⁺ T cell–dependent manner. DNAM-1/CD226, Batf3, IL12p35, and IFNγ were also critical, and CD96-deficient CD8⁺ T cells promoted greater tumor control than CD96-sufficient CD8⁺ T cells. The antitumor activity of anti-CD96 therapy was independent of Fc-mediated effector function and was more effective in dual combination with blockade of a number of immune checkpoints, including PD-1, PD-L1, TIGIT, and CTLA-4. We consistently observed coexpression of PD-1 with CD96 on CD8⁺ T lymphocytes in tumor-infiltrating leukocytes both in mouse and human cancers using mRNA analysis, flow cytometry, and multiplex IHF. The combination of anti-CD96 with anti–PD-1 increased the percentage of IFNγ-expressing CD8⁺ T lymphocytes. Addition of anti-CD96 to anti–PD-1 and anti-TIGIT resulted in superior antitumor responses, regardless of the ability of the anti-TIGIT isotype to engage FcR. The optimal triple combination was also dependent upon CD8⁺ T cells and IFNγ. Overall, these data demonstrate that CD96 is an immune checkpoint on CD8⁺ T cells and that blocking CD96 in combination with other immune-checkpoint inhibitors is a strategy to enhance T-cell activity and suppress tumor growth.

Critical immune-suppressive pathways beyond programmed death 1 (PD-1) and programmed death ligand 1 (PD-L1) require greater attention. Nectins and nectin-like molecules might be promising targets for immunotherapy, since they play critical roles in cell proliferation and migration and exert immunomodulatory functions in pathophysiological conditions. Here, we show CD155 expression in both malignant cells and tumor-infiltrating myeloid cells in humans and mice. Cd155^–/– mice displayed reduced tumor growth and metastasis via DNAM-1 upregulation and enhanced effector function of CD8⁺ T and NK cells, respectively. CD155-deleted tumor cells also displayed slower tumor growth and reduced metastases, demonstrating the importance of a tumor-intrinsic role of CD155. CD155 absence on host and tumor cells exerted an even greater inhibition of tumor growth and metastasis. Blockade of PD-1 or both PD-1 and CTLA4 was more effective in settings in which CD155 was limiting, suggesting the clinical potential of cotargeting PD-L1 and CD155 function.

A key feature of immune functional impairment with age is the progressive involution of thymic tissue responsible for naive T cell production. In this study, we identify two major phases of thymic epithelial cell (TEC) loss during aging: a block in mature TEC differentiation from the pool of immature precursors, occurring at the onset of puberty, followed by impaired bipotent TEC progenitor differentiation and depletion of Sca-1lo cTEC and mTEC lineage-specific precursors. We reveal that an increase in follistatin production by aging TECs contributes to their own demise. TEC loss occurs primarily through the antagonism of activin A signaling, which we show is required for TEC maturation and acts in dissonance to BMP4, which promotes the maintenance of TEC progenitors. These results support a model in which an imbalance of activin A and BMP4 signaling underpins the degeneration of postnatal TEC maintenance during aging, and its reversal enables the transient replenishment of mature TECs.