Person:

Sharpe, Arlene

We report that programmed death ligand 2 (PD-L2), a known ligand of PD-1, also binds to repulsive guidance molecule b (RGMb), which was originally identified in the nervous system as a co-receptor for bone morphogenetic proteins (BMPs). PD-L2 and BMP-2/4 bind to distinct sites on RGMb. Normal resting lung interstitial macrophages and alveolar epithelial cells express high levels of RGMb mRNA, whereas lung dendritic cells express PD-L2. Blockade of the RGMb–PD-L2 interaction markedly impaired the development of respiratory tolerance by interfering with the initial T cell expansion required for respiratory tolerance. Experiments with PD-L2–deficient mice showed that PD-L2 expression on non–T cells was critical for respiratory tolerance, but expression on T cells was not required. Because PD-L2 binds to both PD-1, which inhibits antitumor immunity, and to RGMb, which regulates respiratory immunity, targeting the PD-L2 pathway has therapeutic potential for asthma, cancer, and other immune-mediated disorders. Understanding this pathway may provide insights into how to optimally modulate the PD-1 pathway in cancer immunotherapy while minimizing adverse events.

CD80 plays a critical role in stimulation of T cells and subsequent control of infection. To investigate the effect of CD80 on HSV-1 infection, we constructed a recombinant HSV-1 virus that expresses two copies of the CD80 gene in place of the latency associated transcript (LAT). This mutant virus (HSV-CD80) expressed high levels of CD80 and had similar virus replication kinetics as control viruses in rabbit skin cells. In contrast to parental virus, this CD80 expressing recombinant virus replicated efficiently in immature dendritic cells (DCs). Additionally, the susceptibility of immature DCs to HSV-CD80 infection was mediated by CD80 binding to PD-L1 on DCs. This interaction also contributed to a significant increase in T cell activation. Taken together, these results suggest that inclusion of CD80 as a vaccine adjuvant may promote increased vaccine efficacy by enhancing the immune response directly and also indirectly by targeting to DC.

Purpose.

Given that dry eye disease (DED) is associated with T cell–mediated inflammation of the ocular surface and that PD-L1 is an important negative or inhibitory regulator of immune responses constitutively expressed at high levels by corneal epithelial cells, the authors studied the expression and function of PD-L1 in DED.

Methods.

Dry eye was induced in untreated wild-type mice, PD-L1−/− mice, and wild-type mice treated with anti–PD-L1 antibody by exposing these mice to a desiccating environment in the controlled environment chamber modified with subcutaneous administration of scopolamine. Real-time PCR was used to quantify the expression of chemokine gene transcript levels of multiple CC and CXC chemokine ligands and receptors. Epifluorescence microscopy was used to evaluate corneal infiltration of CD3+ T cells after immunohistochemical staining.

Results.

The increased expression of specific chemokine ligands and receptors in PD-L1−/− corneas of normal mice is associated with significant increases in T-cell homing into these corneas. Similar, and more enhanced, increases in T-cell infiltration were observed in PD-L1−/− DED mice or DED mice treated with anti–PD-L1 antibody compared with controls. In addition, the authors found significantly decreased expression of PD-L1 by corneal epithelial cells in DED and significantly increased corneal fluorescein staining score with PD-L1 functional blockade using anti–PD-L1 antibody.

Conclusions.

Downregulation of corneal epithelial PD-L1 amplifies dry eye–associated corneal inflammation and epitheliopathy by increasing the expression of chemokine ligands and receptors that promote T-cell homing to the ocular surface.

Both the programmed death (PD) 1–PD-ligand (PD-L) pathway and regulatory T (T reg) cells are instrumental to the maintenance of peripheral tolerance. We demonstrate that PD-L1 has a pivotal role in regulating induced T reg (iT reg) cell development and sustaining iT reg cell function. PD-L1(^{−/−}) antigen-presenting cells minimally convert naive CD4 T cells to iT reg cells, showing the essential role of PD-L1 for iT reg cell induction. PD-L1–coated beads induce iT reg cells in vitro, indicating that PD-L1 itself regulates iT reg cell development. Furthermore, PD-L1 enhances and sustains Foxp3 expression and the suppressive function of iT reg cells. The obligatory role for PD-L1 in controlling iT reg cell development and function in vivo is illustrated by a marked reduction in iT reg cell conversion and rapid onset of a fatal inflammatory phenotype in PD-L1(^{−/−}) PD-L2(^{−/−}) Rag(^{−/−}) recipients of naive CD4 T cells. PD-L1 iT reg cell development is mediated through the down-regulation of phospho-Akt, mTOR, S6, and ERK2 and concomitant with the up-regulation of PTEN, all key signaling molecules which are critical for iT reg cell development. Thus, PD-L1 can inhibit T cell responses by promoting both the induction and maintenance of iT reg cells. These studies define a novel mechanism for iT reg cell development and function, as well as a new strategy for controlling T reg cell plasticity.

Regulatory T (T reg) cells are critical for preventing autoimmunity mediated by self-reactive T cells, but their role in modulating immune responses during chronic viral infection is not well defined. To address this question and to investigate a role for T reg cells in exhaustion of virus-specific CD8 T cells, we depleted T reg cells in mice chronically infected with lymphocytic choriomeningitis virus (LCMV). T reg cell ablation resulted in 10–100-fold expansion of functional LCMV-specific CD8 T cells. Rescue of exhausted CD8 T cells was dependent on cognate antigen, B7 costimulation, and conventional CD4 T cells. Despite the striking recovery of LCMV-specific CD8 T cell responses, T reg cell depletion failed to diminish viral load. Interestingly, T reg cell ablation triggered up-regulation of the molecule programmed cell death ligand-1 (PD-L1), which upon binding PD-1 on T cells delivers inhibitory signals. Increased PD-L1 expression was observed especially on LCMV-infected cells, and combining T reg cell depletion with PD-L1 blockade resulted in a significant reduction in viral titers, which was more pronounced than that upon PD-L1 blockade alone. These results suggest that T reg cells effectively maintain CD8 T cell exhaustion, but blockade of the PD-1 inhibitory pathway is critical for elimination of infected cells.

The immune system plays key roles in determining the fate of developing cancers by not only functioning as a tumour promoter facilitating cellular transformation, promoting tumour growth and sculpting tumour cell immunogenicity1–6, but also as an extrinsic tumour suppressor that either destroys developing tumours or restrains their expansion1,2,7. Yet clinically apparent cancers still arise in immunocompetent individuals in part as a consequence of cancer induced immunosuppression. In many individuals, immunosuppression is mediated by Cytotoxic T-Lymphocyte Associated Antigen-4 (CTLA-4) and Programmed Death-1 (PD-1), two immunomodulatory receptors expressed on T cells8,9. Monoclonal antibody (mAb) based therapies targeting CTLA-4 and/or PD-1 (checkpoint blockade) have yielded significant clinical benefits—including durable responses—to patients with different malignancies10–13. However, little is known about the identity of the tumour antigens that function as the targets of T cells activated by checkpoint blockade immunotherapy and whether these antigens can be used to generate vaccines that are highly tumour-specific. Herein, we use genomics and bioinformatics approaches to identify tumour-specific mutant proteins as a major class of T cell rejection antigens following αPD-1 and/or αCTLA-4 therapy of mice bearing progressively growing sarcomas and show that therapeutic synthetic long peptide (SLP) vaccines incorporating these mutant epitopes induce tumour rejection comparably to checkpoint blockade immunotherapy. Whereas, mutant tumour antigen-specific T cells are present in progressively growing tumours, they are reactivated following treatment with αPD-1- and/or αCTLA-4 and display some overlapping but mostly treatment-specific transcriptional profiles rendering them capable of mediating tumour rejection. These results reveal that tumour-specific mutant antigens (TSMA) are not only important targets of checkpoint blockade therapy but also can be used to develop personalized cancer-specific vaccines and to probe the mechanistic underpinnings of different checkpoint blockade treatments.