Programmed Death Ligand 1 Interaction Research Articles

Ovarian cancer immunotherapy using PD-L1 siRNA targeted delivery from folic acid-functionalized polyethylenimine: strategies to enhance T cell killing.

Adoptive T cell immunotherapy is a promising treatment strategy for epithelial ovarian cancer (EOC). However, programmed death ligand-1 (PD-L1), highly expressed on EOC cells, interacts with programmed death-1 (PD-1), expressed on T cells, causing immunosuppression. This study aims to block PD-1/PD-L1 interactions by delivering PD-L1 siRNA, using various folic acid (FA)-functionalized polyethylenimine (PEI) polymers, to SKOV-3-Luc EOC cells, and investigate the sensitization of the EOC cells to T cell killing. To enhance siRNA uptake into EOC cells, which over express folate receptors, PEI is modified with FA or PEG-FA so that siRNA is complexed into nanoparticles with folate molecules on the surface. PEI modification with a single functional group lowers the polymer cytotoxicity compared to unmodified PEI. FA-conjugated polymers increase siRNA uptake into SKOV-3-luc cells and decrease unspecific uptake into monocytes. All polymers result in 40% to 50% PD-L1 protein knockdown. Importantly, SKOV-3-Luc cells treated with either PEI-FA or PEI- polyethylene glycol (PEG)-FA/PD-L1 siRNA complexes are up to twofold more sensitive to T cell killing compared to scrambled siRNA treated controls. These findings are the first to demonstrate that PD-L1 knockdown in EOC cells, via siRNA/FA-targeted delivery, are able to sensitize cancer cells to T cell killing.

Subverting the B7-H1/PD-1 pathway in advanced melanoma and kidney cancer.

Ligands for inhibitory immune receptors on T cells may be constitutively expressed on tumor cells or host cells in tumor microenvironment as a consequence of adaptive immunity. Programmed death 1 (PD-1) is 1 such receptor on T cells, which functions as a negative regulator of T cell activity. Tumors that up-regulate programmed death ligand 1 (PD-L1) (B7-H1) may abrogate the host's effector T cell antitumor response. Higher tumoral PD-L1 expression has been linked with inferior clinical outcomes. Multiple cancers including renal cell cancers (RCCs) and melanomas have relatively high levels of PD-L1 on the cell surface. Early evaluations of antibodies that block the interaction of PD-1 and PD-L1 have shown efficacy and a favorable tolerability profile with notable inflammatory toxicities that are generally manageable. Upward of 30% of RCC patients and 50% of melanoma patients achieve objective responses. Durable responses can occur, even in some patients who have discontinued treatment. The developing investigation of PD-1/PD-L1 pathway-blocking agents in RCC and melanoma will likely alter our approaches to the treatment of these 2 deadly diseases.

Abstract 463: Differential cellular localization of programmed death ligand 1 (PD-L1) and enhanced expression of programmed death-1 (PD-1) in melanoma progression.

Abstract Melanoma is an aggressive malignancy with a relatively high metastatic rate, responsible for almost 60% of lethal skin tumors. Although many human cancers such as melanoma express tumor antigens recognized by T cells, host immune responses often fail to control tumor growth for mostly unexplained reasons. Escaping immune destruction is one of the tenets of tumorigenesis; there have been several studies suggesting an important role of PD-1/PD-L1 (B7-H1) interactions in inhibiting the effector functions of cytotoxic T-lymphocyte in tumor. Ghebeh et al., reported that doxorubicin treatment caused decrease in cytosolic B7-H1 and increased nuclear translocalization in breast carcinoma (Breast Cancer Res. 2010;12(4):R48). It is unknown whether differential cellular expression of PD-L1 contributes to the pathogenesis of melanoma. To address the role of PD-L1 and its receptor PD-1 in different stages of malignant melanoma, we have performed semiquantitative immunohistochemical analysis of PD-L1 and PD-1 in benign nevi (n=59), primary melanomas without metastasis (n=22) or with metastasis (n=17), and melanoma metastases (n=32 cases). Benign nevi had significantly higher levels of nuclear PD-L1 immunopositivity (p=0.004) and low levels of PD-1 protein expression (p=0.002). With progression from benign nevi to primary melanoma there was an increase in cytoplasmic positivity for PD-L1 (p=0.03). Also, a significant positive correlation was observed between the percentage of positive cells (p=0.0007) and intensity (p=0.0009) for PD-1 in primary melanomas. Although there was no significant difference in PD-1 levels in between primaries with or without metastasis, a trend was observed for PD-L1 expression: primaries with metastasis having a larger number of tumor cells with cytosolic localization than primaries without metastasis (p=0.07). Interestingly, primary melanomas without metastasis showed significantly larger number of tumor infiltrating lymphocytes (TILs) cells having intense immunoreactivity for PD-L1 protein than did metastatic melanomas (p=0.03). In conclusion, differential cellular localization of PD-L1 protein and high levels of PD-1 protein correlates with disease progression and high-risk histologic features in melanoma and might partially explain the resistance of melanoma cells against the immune system. Citation Format: Nitin Chakravarti, Roland Bassett, Wen-Jen Hwu, Doina Ivan, Jonathan L. Curry, Victor G. Prieto. Differential cellular localization of programmed death ligand 1 (PD-L1) and enhanced expression of programmed death-1 (PD-1) in melanoma progression. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 463. doi:10.1158/1538-7445.AM2013-463

Tubular cell HIV-entry through apoptosed CD4 T cells: A novel pathway

We hypothesized that HIV-1 may enter tubular cells by phagocytosis of apoptotic fragments of HIV-1-infected T cells infiltrating tubular interstitium. The study was designed to evaluate the interaction of programmed death-1 (PD-1) receptors on CD4 T cells and programmed death ligand-1 (PD-L1) on tubular cells (HK2 and HRPTEC, primary tubular cells). Co-cultivation of HIV-1 infected lymphocytes (HIV-LY) with HK2s/HRPTECs resulted in T cell apoptosis, uptake of the apoptosed HIV-LY by HK2s/HRPTECs, tubular cell activation and HIV expression. Cytochalasin-B inhibited tubular cell HIV-LY uptake and anti-PD-L1 antibody inhibited HIV-LY apoptosis and tubular cell HIV-LY uptake, activation and HIV expression. These observations do indicate induction of apoptosis of T cells due to interaction of PD-1 and PD-L1 upon co-cultivation and subsequent phygocytosis of HIV-laden apoptotic bodies by tubular cells and thus the transfer of HIV-1 into tubular cells. These findings identify a novel pathway that facilitates HIV-1 entry into tubular cell.

Blocking of PDL-1 Interaction Enhances Primary and Secondary CD8 T Cell Response to Herpes Simplex Virus-1 Infection

The blocking of programmed death ligand-1 (PDL-1) has been shown to enhance virus-specific CD8 T cell function during chronic viral infections. Though, how PDL-1 blocking at the time of priming affects the quality of CD8 T cell response to acute infections is not well understood and remains controversial. This report demonstrates that the magnitude of the primary and secondary CD8 T cell responses to herpes simplex virus-1 (HSV-1) infection is subject to control by PDL-1. Our results showed that after footpad HSV-1 infection, PD-1 expression increases on immunodominant SSIEFARL peptide specific CD8 T cells. Additionally, post-infection, the level of PDL-1 expression also increases on CD11c+ dendritic cells. Intraperitoneal administration of anti-PDL-1 monoclonal antibody given one day prior to and three days after cutaneous HSV-1 infection, resulted in a marked increase in effector and memory CD8 T cell response to SSIEFARL peptide. This was shown by measuring the quantity and quality of SSIEFARL-specific CD8 T cells by making use of ex-vivo assays that determine antigen specific CD8 T cell function, such as intracellular cytokine assay, degranulation assay to measure cytotoxicity and viral clearance. Our results are discussed in terms of the beneficial effects of blocking PDL-1 interactions, while giving prophylactic vaccines, to generate a more effective CD8 T cell response to viral infection.

The Novel Costimulatory Programmed Death Ligand 1/B7.1 Pathway Is Functional in Inhibiting Alloimmune Responses In Vivo

The programmed death ligand 1 (PDL1)/programmed death 1 (PD1) costimulatory pathway plays an important role in the inhibition of alloimmune responses as well as in the induction and maintenance of peripheral tolerance. It has been demonstrated recently that PDL1 also can bind B7.1 to inhibit T cell responses in vitro. Using the bm12 into B6 heart transplant model, we investigated the functional significance of this interaction in alloimmune responses in vivo. PD1 blockade unlike PDL1 blockade failed to accelerate bm12 allograft rejection, suggesting a role for an additional binding partner for PDL1 other than PD1 in transplant rejection. PDL1 blockade was able to accelerate allograft rejection in B7.2-deficient recipients but not B7.1-deficient recipients, indicating that PDL1 interaction with B7.1 was important in inhibiting rejection. Administration of the novel 2H11 anti-PDL1 mAb, which only blocks the PDL1-B7.1 interaction, aggravated chronic injury of bm12 allografts in B6 recipients. Aggravated chronic injury was associated with an increased frequency of alloreactive IFN-γ-, IL-4-, and IL-6-producing splenocytes and a decreased percentage of regulatory T cells in the recipients. Using an in vitro cell culture assay, blockade of the interaction of PDL1 on dendritic cells with B7.1 on T cells increased IFN-γ production from alloreactive CD4(+) T cells, whereas blockade of dendritic cell B7.1 interaction with T cell PDL1 did not. These data indicate that PDL1 interaction with B7.1 plays an important role in the inhibition of alloimmune responses in vivo and suggests a dominant direction for PDL1 and B7.1 interaction.

PD-1/PD-L1 interactions inhibit antitumor immune responses in a murine acute myeloid leukemia model

AbstractNegative regulatory mechanisms within the solid tumor microenvironment inhibit antitumor T-cell function, leading to evasion from immune attack. One inhibitory mechanism is up-regulation of programmed death-ligand 1 (PD-L1) expressed on tumor or stromal cells which binds to programmed death-1 (PD-1) on activated T cells. PD-1/PD-L1 engagement results in diminished antitumor T-cell responses and correlates with poor outcome in murine and human solid cancers. In contrast to available data in solid tumors, little is known regarding involvement of the PD-1/PD-L1 pathway in immune escape by hematopoietic cancers, such as acute myeloid leukemia (AML). To investigate this hypothesis, we used the murine leukemia, C1498. When transferred intravenously, C1498 cells grew progressively and apparently evaded immune destruction. Low levels of PD-L1 expression were found on C1498 cells grown in vitro. However, PD-L1 expression was up-regulated on C1498 cells when grown in vivo. PD-1−/− mice challenged with C1498 cells generated augmented antitumor T-cell responses, showed decreased AML burden in the blood and other organs, and survived significantly longer than did wild-type mice. Similar results were obtained with a PD-L1 blocking antibody. These data suggest the importance of the PD-1/PD-L1 pathway in immune evasion by a hematologic malignancy, providing a rationale for clinical trials targeting this pathway in leukemia patients.

Human Corneal Endothelial Cells Expressing Programmed Death-Ligand 1 (PD-L1) Suppress PD-1+T Helper 1 Cells by a Contact-Dependent Mechanism

This study was designed to determine whether human corneal endothelial (HCE) cells could regulate the activation of bystander T cells in vitro. HCE cell lines were established from primary HCE cells. Target-activated T cells were used allogeneic T cells and Jurkat T-cell lines. As an additional target, T-cell clones from uveitis patients were established from aqueous humor via a limiting dilution. T-cell activation was assessed for proliferation by [(3)H]-thymidine incorporation, carboxyfluorescein succinimidyl ester incorporation, or IFNgamma production. Expression of co-stimulatory molecules on IFNgamma-treated corneal endothelial and non-treated cells was evaluated by flow cytometry, RT-PCR, or immunohistochemistry. Expression of co-stimulatory receptors on target T cells was evaluated by flow cytometry. Blocking antibodies was used to abolish the HCE-inhibitory function. HCE cells suppressed both in vitro proliferation and IFNgamma production by CD4(+) T cells via a cell contact-dependent mechanism. HCE constitutively expressed co-stimulatory molecules programmed death-ligand 1 (PD-L1) and PD-L2, and their expression was enhanced by IFNgamma. HCE efficiently inhibited the proliferation of Th1 cells that overexpressed PD-1 among various activated T-cell lines and clones established from patients with uveitis or corneal endotheliitis. A neutralizing mAb for PD-L1, but not PD-L2, blocked the suppressive effect of HCE on Th1 cells. HCE can impair the effector functions and activation of Th1 infiltrating CD4(+) T cells via the PD-1/PD-L1 interaction. The data support the hypothesis that corneal endothelium may contribute to maintenance of the privileged immune status of the anterior chamber of the eye by inducing peripheral immune tolerance.