Mary E. Keir

PD-1 and Its Ligands in Tolerance and Immunity

Programmed death 1 (PD-1) and its ligands, PD-L1 and PD-L2, deliver inhibitory signals that regulate the balance between T cell activation, tolerance, and immunopathology. Immune responses to foreign and self-antigens require specific and balanced responses to clear pathogens and tumors and yet maintain tolerance. Induction and maintenance of T cell tolerance requires PD-1, and its ligand PD-L1 on nonhematopoietic cells can limit effector T cell responses and protect tissues from immune-mediated tissue damage. The PD-1:PD-L pathway also has been usurped by microorganisms and tumors to attenuate antimicrobial or tumor immunity and facilitate chronic infection and tumor survival. The identification of B7-1 as an additional binding partner for PD-L1, together with the discovery of an inhibitory bidirectional interaction between PD-L1 and B7-1, reveals new ways the B7:CD28 family regulates T cell activation and tolerance. In this review, we discuss current understanding of the immunoregulatory functions of PD-1 and its ligands and their therapeutic potential.

Tissue expression of PD-L1 mediates peripheral T cell tolerance

Programmed death 1 (PD-1), an inhibitory receptor expressed on activated lymphocytes, regulates tolerance and autoimmunity. PD-1 has two ligands: PD-1 ligand 1 (PD-L1), which is expressed broadly on hematopoietic and parenchymal cells, including pancreatic islet cells; and PD-L2, which is restricted to macrophages and dendritic cells. To investigate whether PD-L1 and PD-L2 have synergistic or unique roles in regulating T cell activation and tolerance, we generated mice lacking PD-L1 and PD-L2 (PD-L1/PD-L2−/− mice) and compared them to mice lacking either PD-L. PD-L1 and PD-L2 have overlapping functions in inhibiting interleukin-2 and interferon-γ production during T cell activation. However, PD-L1 has a unique and critical role in controlling self-reactive T cells in the pancreas. Our studies with bone marrow chimeras demonstrate that PD-L1/PD-L2 expression only on antigen-presenting cells is insufficient to prevent the early onset diabetes that develops in PD-L1/PD-L2−/− non-obese diabetic mice. PD-L1 expression in islets protects against immunopathology after transplantation of syngeneic islets into diabetic recipients. PD-L1 inhibits pathogenic self-reactive CD4+ T cell–mediated tissue destruction and effector cytokine production. These data provide evidence that PD-L1 expression on parenchymal cells rather than hematopoietic cells protects against autoimmune diabetes and point to a novel role for PD-1–PD-L1 interactions in mediating tissue tolerance.

PD-1 Regulates Self-Reactive CD8+ T Cell Responses to Antigen in Lymph Nodes and Tissues

PD-1, an inhibitory receptor expressed on activated lymphocytes, regulates tolerance and autoimmunity. We tested the role of PD-1:PD-1 ligand (PD-L) interactions in cross-presentation and the generation and control of CD8+ responses against self-Ag. Ag-naive PD-1−/− OVA-specific OT-I CD8+ T cells exhibited exacerbated responses to cross-presented Ag in mice expressing soluble OVA under the control of the rat insulin promoter (RIP-ovahigh). Following adoptive transfer into RIP-ovahigh recipients, PD-1−/− OT-I T cells expanded in the pancreatic lymph node. In contrast to wild-type OT-I cells, PD-1−/− OT-I T cells secreted IFN-γ and migrated into the pancreas, ultimately causing diabetes. Loss of PD-1 affected CD8+ cells intrinsically, and did not significantly alter the responses of wild-type OT-I T cells adoptively transferred into the same RIP-ovahigh recipient mouse. PD-1:PD-L interactions also limited CD8+ effector cells, and PD-L1 expression on parenchymal tissues protected against effector OT-I T cell attack. Finally, we found that the loss of PD-1 on effector OT-I cells lowers the threshold for Ag recognition in peripheral tissues. These findings indicate two checkpoints where PD-1 attenuates self-reactive T cell responses: presentation of self-Ag to naive self-reactive T cells by dendritic cells in the draining lymph node and reactivation of pathogenic self-reactive T cells in the target organ.

The B7/CD28 costimulatory family in autoimmunity

Summary:  Host defense is dependent on the appropriate induction of immune responses. A central concept in immunology is the ability of the immune system to differentiate foreign from self‐antigens. The failure of the immune response to recognize foreign pathogens can result in infection and disease in the host. The inappropriate response of the immune system to self‐antigens is equally problematic, leading to autoimmune disease. Central and peripheral tolerance mechanisms control self‐reactive T‐cell responses and protect peripheral tissues from autoimmune attack. This review examines the roles of B7/CD28 family members, which can augment or antagonize T‐cell receptor signaling, in the regulation of central and peripheral T‐cell tolerance. We also discuss how B7/CD28 pathways influence both T‐cell‐intrinsic and ‐extrinsic mechanisms of regulation.

Programmed death-1 (PD-1):PD-ligand 1 interactions inhibit TCR-mediated positive selection of thymocytes.

Positive selection during thymocyte development is driven by the affinity and avidity of the TCR for MHC-peptide complexes expressed in the thymus. In this study, we show that programmed death-1 (PD-1), a member of the B7/CD28 family of costimulatory receptors, inhibits TCR-mediated positive selection through PD-1 ligand 1 (PD-L1):PD-1 interactions. Transgenic mice that constitutively overexpress PD-1 on CD4+CD8+ thymocytes display defects in positive selection in vivo. Using an in vitro model system, we find that PD-1 is up-regulated following TCR engagement on CD4+CD8+ murine thymocytes. Coligation of TCR and PD-1 on CD4+CD8+ thymocytes with a novel PD-1 agonistic mAb inhibits the activation of ERK and up-regulation of bcl-2, both of which are downstream mediators essential for positive selection. Inhibitory signals through PD-1 can overcome the ability of positive costimulators, such as CD2 and CD28, to facilitate positive selection. Finally, defects in positive selection that result from PD-1 overexpression in thymocytes resolve upon elimination of PD-L1, but not PD-1 ligand 2, expression. PD-L1-deficient mice have increased numbers of CD4+CD8+ and CD4+ thymocytes, indicating that PD-L1 is involved in normal thymic selection. These data demonstrate that PD-1:PD-L1 interactions are critical to positive selection and play a role in shaping the T cell repertoire.

PD-L1 has distinct functions in hematopoietic and nonhematopoietic cells in regulating T cell responses during chronic infection in mice

The inhibitory receptor programmed death 1 (PD-1) is upregulated on antigen-specific CD8+ T cells during persistent viral infections. Interaction with PD-1 ligand 1 (PD-L1) contributes to functional exhaustion of responding T cells and may limit immunopathology during infection. PD-L1 is expressed on both hematopoietic and nonhematopoietic cells in tissues. However, the exact roles of PD-L1 on hematopoietic versus nonhematopoietic cells in modulating immune responses are unclear. Here we used bone marrow chimeric mice to examine the effects of PD-L1 deficiency in hematopoietic or nonhematopoietic cells during lymphocytic choriomeningitis virus clone 13 (LCMV CL-13) infection. We found that PD-L1 expression on hematopoietic cells inhibited CD8+ T cell numbers and function after LCMV CL-13 infection. In contrast, PD-L1 expression on nonhematopoietic cells limited viral clearance and immunopathology in infected tissues. Together, these data demonstrate that there are distinct roles for PD-L1 on hematopoietic and nonhematopoietic cells in regulating CD8+ T cell responses and viral clearance during chronic viral infection.

IFN-α Secretion by Type 2 Predendritic Cells Up-Regulates MHC Class I in the HIV-1-Infected Thymus

The ability of HIV-1 to evade the host immune response leads to the establishment of chronic infection. HIV-1 has been reported to up-regulate MHC I molecules on the surface of thymocytes from HIV-1-infected thymus. We demonstrate in this study that HIV-1 up-regulates MHC I on both HIV-1-infected and uninfected thymocytes in a manner that is independent of Nef, proportional to viral replication, and entirely mediated by IFN-α. IL-3Rα+ type 2 predendritic cells (preDC2) resident in the thymic medulla secrete IFN-α, which acts on IFN-αβR-expressing immature thymocytes to induce MHC I expression. Furthermore, thymic preDC2 are permissive for HIV-1 infection and positive for intracellular p24. These data demonstrate the ability of IFN-α secreted by preDC2 to induce MHC I up-regulation in the HIV-1-infected human thymus.

Endothelial Programmed Death-1 Ligand 1 (PD-L1) Regulates CD8+ T-Cell–Mediated Injury in the Heart

Background— PD-L1 and PD-L2 are ligands for the inhibitory receptor programmed death-1 (PD-1), which is an important regulator of immune responses. PD-L1 is induced on cardiac endothelial cells under inflammatory conditions, but little is known about its role in regulating immune injury in the heart. Methods and Results— Cytotoxic T-lymphocyte–mediated myocarditis was induced in mice, and the influence of PD-L1 signaling was studied with PD-L1/L2–deficient mice and blocking antibodies. During cytotoxic T-lymphocyte–induced myocarditis, the upregulation of PD-L1 on cardiac endothelia was dependent on T-cell–derived interferon-γ, and blocking of interferon-γ signaling worsened disease. Genetic deletion of both PD-1 ligands [PD-L1/2(−/−)], as well as treatment with PD-L1 blocking antibody, transformed transient myocarditis to lethal disease, in association with widespread polymorphonuclear leukocyte–rich microabscesses but without change in cytotoxic T-lymphocyte recruitment. PD-L1/2(−/−) mice reconstituted with bone marrow from wild-type mice remained susceptible to severe disease, which demonstrates that PD-L1 on non–bone marrow–derived cells confers the protective effect. Finally, depletion of polymorphonuclear leukocytes reversed the enhanced susceptibility to lethal myocarditis attributable to PD-L1 deficiency. Conclusions— Myocardial PD-L1, mainly localized on endothelium, is critical for control of immune-mediated cardiac injury and polymorphonuclear leukocyte inflammation.

The Programmed Death-1 Ligand 1:B7-1 Pathway Restrains Diabetogenic Effector T Cells In Vivo

Programmed death-1 ligand 1 (PD-L1) is a coinhibitory molecule that negatively regulates multiple tolerance checkpoints. In the NOD mouse model, PD-L1 regulates the development of diabetes. PD-L1 has two binding partners, programmed death-1 and B7-1, but the significance of the PD-L1:B7-1 interaction in regulating self-reactive T cell responses is not yet clear. To investigate this issue in NOD mice, we have compared the effects of two anti–PD-L1 Abs that have different blocking activities. Anti–PD-L1 mAb 10F.2H11 sterically and functionally blocks only PD-L1:B7-1 interactions, whereas anti–PD-L1 mAb 10F.9G2 blocks both PD-L1:B7-1 and PD-L1:programmed death-1 interactions. Both Abs had potent, yet distinct effects in accelerating diabetes in NOD mice: the single-blocker 10F.2H11 mAb was more effective at precipitating diabetes in older (13-wk-old) than in younger (6- to 7-wk-old) mice, whereas the dual-blocker 10F.9G2 mAb rapidly induced diabetes in NOD mice of both ages. Similarly, 10F.2H11 accelerated diabetes in recipients of T cells from diabetic, but not prediabetic mice, whereas 10F.9G2 was effective in both settings. Both anti–PD-L1 mAbs precipitated diabetes in adoptive transfer models of CD4+ and CD8+ T cell-driven diabetes. Taken together, these data demonstrate that the PD-L1:B7-1 pathway inhibits potentially pathogenic self-reactive effector CD4+ and CD8+ T cell responses in vivo, and suggest that the immunoinhibitory functions of this pathway may be particularly important during the later phases of diabetogenesis.

Programmed Death 1 Ligand (PD-L) 1 and PD-L2 Limit Autoimmune Kidney Disease: Distinct Roles

The programmed death 1/programmed death 1 ligand (PD-L) pathway is instrumental in peripheral tolerance. Blocking this pathway exacerbates experimental autoimmune diseases, but its role in autoimmune kidney disease has not been explored. Therefore, we tested the hypothesis that the programmed death 1 ligands (PD-L1 and PD-L2), provide a protective barrier during T cell- and macrophage (Mφ)-dependent autoimmune kidney disease. For this purpose, we compared nephrotoxic serum nephritis (NSN) in mice lacking PD-L1 (PD-L1−/−), PD-L2 (PD-L2−/−), or both (PD-L1/L2−/−) to wild-type (WT) C57BL/6 mice. Kidney pathology, loss of renal function, and intrarenal leukocyte infiltrates were increased in each PD-L−/− strain as compared with WT mice. Although the magnitude of renal pathology was similar in PD-L1−/− and PD-L2−/− mice, our findings suggest that kidney disease in each strain is regulated by distinct mechanisms. Specifically, we detected increased CD68+ cells along with elevated circulating IgG and IgG deposits in glomeruli in PD-L2−/− mice, but not PD-L1−/− mice. In contrast, we detected a rise in activated CD8+ T cells in PD-L1−/− mice, but not PD-L2−/− mice. Furthermore, since PD-L1 is expressed by parenchymal and hemopoietic cells in WT kidneys, we explored the differential impact of PD-L1 expression on these cell types by inducing NSN in bone marrow chimeric mice. Our results indicate that PD-L1 expression on hemopoietic cells, and not parenchymal cells, is primarily responsible for limiting leukocyte infiltration during NSN. Taken together, our findings indicate that PD-L1 and PD-L2 provide distinct negative regulatory checkpoints poised to suppress autoimmune renal disease.