High Programmed Death-1 Expression Research Articles

Periparturient blood T-lymphocyte PD-1 and CTLA-4 expression as potential predictors of new intramammary infections in dairy cows during early lactation (short communication)

BackgroundThe periparturient period in dairy cows is marked by immunosuppression which increases the likelihood of infectious disorders, particularly also mastitis. An in-depth understanding of peripartum leukocyte biology is vital for the implementation of highly successful post-partum disease prevention measures. Immune checkpoint molecules, such as programmed death 1 (PD-1) and cytotoxic T-lymphocyte antigen 4 (CTLA-4), are critical inhibitory receptors expressed on immune cells, particularly T cells, that drive immunosuppressive signaling pathways. However, the potential role of immune checkpoint molecules expression in T-cells on udder health has never been explored. Thus, the association between the occurrence of new postpartum intramammary infections (IMIs) and the expression of programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) on blood T-cells during the peripartum period was investigated.ResultsIn this study, the incidence of IMIs by any pathogen in early lactation was not associated with a higher expression of PD-1 and CTLA-4 in the periparturient period. However, the incidence of IMIs by major pathogens throughout the first month of lactation was significantly associated with higher expression of PD-1 at 14 days before calving (P = 0.03) and CTLA-4 at parturition (P = 0.03) by blood T-cells. Also, the expression of CTLA-4 at D0 (P = 0.012) by T-cells was associated with the occurrence of persistent IMIs during the first month of lactation.ConclusionsTo our knowledge, this is the first report to investigate the expression of PD-1 and CTLA-4 by blood T-lymphocytes during the periparturient period in dairy cows and to explore their relationship with the incidence of new IMIs in the postpartum period. Thus, a comprehensive understanding of leukocyte biology during peripartum would appear to be a prerequisite for the identification of resilient dairy cows or targets innovative (immunological) non-antibiotic approaches in the transition period.

Increased PD-1 expression of bone marrow T-cells in acute myeloid leukaemia patients after stem cell transplantation, and its association with overall survival.

Immune checkpoints are involved in mechanisms by which tumours escape from the host immune system. Our aim was to evaluate acute myeloid leukaemia (AML) patients to determine expression levels of checkpoint molecules according to diagnosis and treatments, and to identify optimal candidates for checkpoint blockade. Bone marrow (BM) samples were obtained from 279 AML patients at different disease status and from 23 controls. Flow cytometric analyses of PD-1 and PD-L1/PD-L2 expression were performed. Programmed death-1 (PD-1) expression levels on CD8+ T-cells at AML diagnosis were increased compared to controls. PD-L1 and PD-L2 expression levels on leukaemic cells at diagnosis were significantly higher in secondary AML than in de novo AML. PD-1 levels on CD8+ and CD4+ T-cells after allo-SCT were significantly higher than those at diagnosis and after CTx. PD-1 expression on CD8+ T-cells increased in the acute GVHD group than in the non-GVHD group. The overall survival of patients with high PD-1 expression on CD8+ T-cells was significantly shorter than that of patients with low PD-1 expression. In conclusion, patients who underwent allo-SCT exhibited high PD-1 expression, suggesting that allo-SCT increases PD-1 expression on T-cells, and the patients with high PD-1 expression on CD8+ T-cells after allo-SCT showed the poor prognosis. For these patients, PD-1 blockade could be an immunotherapeutic strategy.

Galectin-3 interacts with PD-1 and counteracts the PD-1 pathway-driven regulation of T cell and osteoclast activity in Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is an autoimmune disease characterized by joint inflammation and bone erosions. The glycosylated programmed death-1 (PD-1) receptor plays an important role in regulating immune responses and maintaining tolerance. In this study, we focus on two features observed in RA: impaired PD-1 signalling and Galectin-3 (Gal-3) upregulation. We hypothesize that Gal-3 binds PD-1 and PD-1 ligands, potentially contributing to impaired PD-1 signalling. PD-1 and Gal-3 levels in RA synovial fluid (SF) and plasma were evaluated by ELISA. PD-1 and Gal-3 interaction was examined by Surface Plasmon Resonance and ELISA. PD-1, PD-L1 and Gal-3 expression on mononuclear cells from SF and peripheral blood as well as fibroblast-like synoviocytes were examined by flow cytometry. Effects of Gal-3 and PD-L1 on osteoclast formation was evaluated by tartrate-resistant acid phosphatase assay. We show that Gal-3 binds PD-1 and PD-L1. Results demonstrated high expression of PD-1 and Gal-3 on mononuclear cells, especially from SF. Gal-3 inhibited PD-1 signalling when PD-L1 was present. Furthermore, a role of Gal-3 in osteoclast formation was observed in vitro, both directly but also through PD-1:PD-L1 inhibition. Effects of Gal-3 on the PD-1 signalling axis are proposed to be inhibitory, meaning high Gal-3 levels in the complex synovial microenvironment are not desirable in RA. Preventing Gal-3's inhibitory role on PD-1 signalling could, therefore, be a therapeutic target in RA by affecting inflammatory T cell responses and osteoclasts.

CD56dim NK Cell is an Important Factor in T Cell Depletion of cART-Treated AIDS Patients.

PurposeTo investigate factors involved in T-cell depletion in combination antiretroviral therapy (cART)-treated human immunodeficiency virus 1 (HIV-1)-positive patients.Patients and Methods29 HIV-1-positive patients were enrolled. The CD4+, CD8+ T cell subsets and CD56dim NK cells were detected by flow cytometry. The concentrations of cytokines were measured by enzyme-linked immunosorbent assay. Extraction, amplification, and viral load quantification of specimens were performed using the Roche Cobas Ampliprep/Cobas TaqMan HIV-1 test.ResultsCompared with IR group, the total number of red blood cells (RBCs) and lymphocytes (LCs) in INR group was significantly reduced, and there was a significant positive correlation between the number of RBCs and that of LCs. The overall production rates of T cells-related cytokines were lower in INR group. However, the cell-surface expression of programmed death-1 (PD-1) on CD4+ T and CD8+ T cells were markedly elevated in INR group. Moreover, it was found that the proportion and the killing ability of CD56dim NK cells significantly increased in INR patients, and significantly correlated with apoptosis of T lymphocytes.ConclusionA poor immune reconstitution in HIV-positive patients might result from multiple factors, including bone marrow suppression, high PD-1 expression on the surface of CD4+ T cells, and over-activation of T and NK cells. Besides, the activity of NK cells and RBCs count might be important auxiliary indicators for immune reconstitution and provided a reliable guidance for developing strategies to improve immune reconstitution.

Peripheral leukemia burden at time of apheresis negatively affects the clinical efficacy of CART19 in refractory or relapsed B-ALL

Our previous clinical study achieved complete remission (CR) rates of >90% following chimeric antigen receptor T cells targeting CD19 (CART19) treatment of refractory/relapsed B cell acute lymphoblastic leukemia (r/r B-ALL); however, the influence of the leukemia burden in peripheral blood (PB) blasts remains unclear. Here, we retrospectively analyzed 143 patients treated with CART19 (including 36 patients with PB blasts) to evaluate the effect of peripheral leukemia burden at the time of apheresis. One hundred seventeen patients with high disease burdens achieved 91.5% CR or incomplete count recovery CR and 86.3% minimal residual disease-negative CR, and 26 patients with low disease burdens obtained 96.2% MRD− CR. Collectively, 9 of 36 (25%) patients with PB blasts and 2 of 107 (1.87%) patients without PB blasts did not respond to CART19 therapy. The leukemia burden in PB negatively influenced ex vivo cell characteristics, including the transduction efficiency of CD3+ T cells and their fold expansion, and in vivo cell dynamics, including peak CART19 proportion and absolute count, fold expansion, and persistence duration. Further studies showed that these patients had higher programmed death-1 expression in CART19 products. Our data imply that PB blasts negatively affected CART19 production and the clinical efficacy of CART19 therapy in patients with r/r B-ALL.

High PD-1 and Tim-3 Expression Concurrent with Exhausted CD4+ and CD8+ T Cells in Bone Marrow Compared with Peripheral Blood from Patients with Multiple Myeloma

Immunosuppressive microenvironment plays a crucial role for T cell immunodeficiency in multiple myeloma. Increasing data showed that overexpression of the T cell immunosuppressive receptors, such as programmed death-1 (PD-1), cytotoxic T lymphocyte antigen-4 (CTLA-4) and T cell immunoglobulin mucin-3 (Tim-3) may be related to tumor immunosuppression and poor prognosis, and the malignant bone marrow microenvironment may contribute to such immunosupression. In this study, we compared the difference of PD-1 and Tim-3 expression with exhausted CD3+, CD4+ and CD8+ T cells between bone marrow (BM) and peripheral blood (PB) in patients with multiple myeloma (MM). BM and PB samples from 6 cases (4 males and 2 females, age range: 41-82 years old) with untreated MM were selected to this study, the frequencies of PD-1 and Tim-3 expression in CD3+, CD4+ and CD8+ T cells from BM and PB mononuclear cells were detected by Flow cytometry. We also detected the frequency of CD244 and CD57 expression in both CD4+ and CD8+ T cells, which are the immune markers for T cell exhaustion. The change of the frequency of T cell phenotype was followed up in two cases in different time point after therapy. The results showed that significant higher percentage of PD-1+CD3+T cells was found in BM compared with PB (P= 0.001), particularly in PD1+CD8+T cells was significantly higher in BM (P= 0.017), however, interesting, higher percentage of PD-1+CD4+T cells was showed in PB compared with BM (P= 0.043) (Figure 1). Similarly, the percentage of Tim-3+CD3+T cell in BM seemed high compared with PB, as well as in PD-1+Tim-3+CD3+, CD4+ and CD8+T cells. The percentage of CD57+, CD244+ in CD3+, CD4+ and CD8+ T cells was relative high in BM group in comparison with PB group, although the statistical significance was not significant. However, either PD-1+CD244+ (P= 0.006) or PD-1+CD57+(P = 0.02) in CD3+ T cells was significant higher in BM group than that in PB group. Moreover, PD-1+CD244+(P = 0.000) and PD-1+CD57+ (P = 0.048) in CD8+ T cells were higher distributed in BM than that in PB, while there was not significant different in CD4+ T cells (Figure 2). And the limit data also showed that the numbers of either PD-1+ or PD-1+Tim-3+T cells in different T cell subsets were decreases in both BM and PB when the patient achieved complete remission after therapy (Figure 3). In conclusions, we characterized the different distribution of PD-1 and Tim-3 concurrent with exhausted CD3+, CD4+ and CD8+ T cells between BM and PB in patients with MM. Higher numbers of PD-1+CD244+ or PD-1+CD57+ CD3+ T cells in BM from MM patients may contribute to mediate the BM immunosuppressive microenvironment, higher lever PD-1+CD244+ or PD-1+CD57+CD8+T cells may relate to lower T cell activation and T cell immunodeficiency. [Display omitted] DisclosuresLi:Guangdong Provincial Basic Research Program (No. 2015B020227003: Research Funding; National Natural Science Foundation of China (91642111): Research Funding; Guangzhou Science and Technology Project (No. 2015100010211): Research Funding; Guangdong Provincial Applied Science and Technology Research & Development Program (No. 2016B020237006): Research Funding.

Programmed death-1 mediates venous neointimal hyperplasia in humans and rats

Venous neointimal hyperplasia can be a problem after vein interventions. We hypothesized that inhibiting programmed death-1 (PD-1) can decrease venous neointimal hyperplasia in a rat inferior vena cava (IVC) patch venoplasty model. The rats were divided into four groups: the control group was only decellularized without other special treatment; the PD-1 group was injected with a single dose of humanized PD-1 antibody (4 mg/kg); the PD-1 antibody coated patches group; the BMS-1 (a PD-1 small molecular inhibitor) coated patches group (PD-1 inhibitor-1). Patches were implanted to the rat IVC and harvested on day 14 and analyzed. Immunohistochemical analysis showed PD-1-positive cells in the neointima in the human samples. There was high protein expression of PD-1 in the neointima in the rat IVC venoplasty model. PD-1 antibody injection can significantly decrease neointimal thickness (p < 0.0001). PD-1 antibody or BMS-1 was successfully conjugated to the decellularized rat thoracic artery patch by hyaluronic acid with altered morphology and reduced the water contact angle (WCA). Patches coated with humanized PD-1 antibody or BMS-1 both can also decrease neointimal hyperplasia and inflammatory cells infiltration. PD-1-positive cells are present in venous neointima in both human and rat samples. Inhibition of the PD-1 pathway may be a promising therapeutic strategy to inhibit venous neointimal hyperplasia.

The effect of PD-1 expression on tumor-associated macrophage in T cell lymphoma.

Our study aimed to explore the programmed death 1 (PD-1) expression on tumor-associated macrophage (TAM) in T cell non-Hodgkin lymphoma (T-NHL) and its relationship with lymphoma prognosis. The effect of PD-1 expression on the function of macrophages was also studied. Multispectral image quantitative analysis was applied for detecting PD-1 expression on macrophages in T cell lymphoma tissues. The Kaplan-Meier analysis was performed to evaluate the value of PD-1 expression of TAM in predicting the overall survival of T-NHL. PD-1 overexpression THP-1-derived macrophage was constructed and was cocultured with Jurkat cells to explore the effect of PD-1 on macrophage function. In 17T cell lymphoma cases, the 1-year overall survival rate was significantly lower in patients with higher PD-1 expression on TAMs (0.25 vs 0.86, p < 0.05). After co-cultured with Jurkat cells, classically activated (M1)-related markers on PD-1 overexpressed macrophages were significantly lower than those on controls, while the expressions of alternatively activated (M2) related markers were similar. The PD-1 overexpressed macrophages showed inhibited phagocytosis (4.42% vs 40.7%, p<0.001) and increased IL-10 secretion (144.48pg/ml vs 32.32pg/ml, p<0.001). High PD-1 expression on TAMs in T-NHL may predict poor prognosis. The PD-1 overexpression of macrophages significantly inhibited polarization of M1 macrophages and phagocytosis, and more IL-10 was excreted. These changes may enhance the pro-tumor effects of tumor microenvironment.

Interleukin-12 over-expression in malignant melanoma B16 cells reduces programmed death-1 expression on T cells in mice with immune reconstitution

To investigate whether interleukin-12 (IL-12) over-expression in malignant melanoma B16 cells affects the expression level of programmed death-1 (PD-1) on T cells in mice during immune microenvironment reconstruction. B16 cells were transfected with an IL-12 expression lentiviral vector, and IL-12 over-expression in the cells was verified qPCR and ELISA. Plate cloning assay was used to compare the cell proliferation activity between B16 cells and B16/IL-12 cells. The expression of IL-12 protein in B16/IL-12 cells-derived tumor tissue were detected by ELISA. C57BL/6 mice were inoculated with B16 cells or B16/IL-12 cells, and 14 days later the proportion of T cells with high expression of PD-1 in the tumor-draining lymph nodes was detected by flow cytometry. Mouse models of immune reconstitution established by 650 cGy X-ray radiation were inoculated with B16 (B16+RT group) or B16/IL-12 (B16/IL-12+RT group) cells, with the mice without X-ray radiation prior to B16 cell inoculation as controls. Tumor growth in the mice was recorded at different time points, and on day 14, flow cytometry was performed to detect the proportion of T cells with high PD-1 expression in the tumor-draining lymph nodes and in the tumor tissue. B16 cells infected with the IL-12-overexpressing lentiviral vector showed significantly increased mRNA and protein levels of IL-12 (P &lt; 0.001) without obvious changes in cell viability (P&gt;0.05). B16/IL-12 cells expressed higher levels of IL-12 than B16 cells in vivo (P &lt; 0.01). In the tumor-bearing mouse models, the proportion of CD4 + PD-1+ T cells was significantly lower in B16/IL-12 group than in B16 group (P &lt; 0.01). In the mice with X-ray radiation-induced immune reconstitution, PD-1 expressions on CD4+ T cells (P &lt; 0.05) and CD8+ T cells (P &lt; 0.01) were significantly higher in B16+ RT group than in the control mice and in B16/IL-12+RT group (P &lt; 0.01 or 0.001); the tumors grew more slowly in B16/IL-12+RT group than in B16 + RT group (P &lt; 0.001). During immune microenvironment reconstruction, overexpression IL-12 in the tumor microenvironment can reduce the percentage of PD-1 + T cells and suppress the growth of malignant melanoma in mice.

Low Expression of Programmed Death 1 (PD-1), PD-1 Ligand 1 (PD-L1), and Low CD8+ T Lymphocyte Infiltration Identify a Subgroup of Patients With Gastric and Esophageal Adenocarcinoma With Severe Prognosis.

Prognosis of gastric and esophageal cancer is poor and treatment improvements are needed. Programmed cell death 1 receptor (PD-1) interaction with its ligand PD-L1 in tumor micro-environment promotes immune tolerance and blocking monoclonal antibodies have entered clinical practice. However, clinical significance of PD-1 and PD-L1 expression in gastric and esophageal adenocarcinomas, particularly in non-Asian patients, is still unclear. Three tissue microarrays including 190 clinically annotated esophageal (n = 31) and gastric (n = 159) adenocarcinomas and 58 paired mucosa specimens, were stained with PD-1, PD-L1, and CD8-specific reagents in indirect immunohistochemistry assays. PD-L1 expression was detectable in 23.2% of cancer specimens. High PD-1 expression was detectable in 37.3% of cases and high CD8+ infiltration in 76%. PD-L1 and high PD1 expression significantly correlated with each other (rs = 0.404, P < 0.0001) and both significantly correlated with CD8+ infiltration (rs = 0.435, P = 0.0003, and rs = 0.444; P = 0.0004, respectively). CD8+ lymphocyte infiltration correlated with improved survival in univariate (P = 0.009), but not multivariate analysis. Most interestingly, multivariate analysis and Kaplan-Meier curves indicate that combined low PD-1/PD-L1 expression and low CD8+ lymphocyte infiltration significantly correlate with poor prognosis. Our data document the clinical significance of a microenvironmental signature including PD-1/PD-L1 expression and CD8+ lymphocyte infiltration in gastric and esophageal adenocarcinomas and contribute to identify a patients' subset requiring more aggressive peri-operative treatments.

B7-Positive and B7-Negative Acute Myeloid Leukemias Display Distinct T Cell Maturation Profiles, Immune Checkpoint Receptor Expression, and European Leukemia Net Risk Profiles.

Acute myeloid leukemia (AML) is generally considered a poorly immunogenic malignancy, displaying a “non-inflamed” leukemia microenvironment (LME), leading to T cell tolerance. However, the immune landscape of AML is much more heterogeneous. Since B7 expression is regarded as a consequence of an interferon-mediated “inflammatory” phenotype, we have investigated by flow cytometry the B7 checkpoint ligands B7.1, B7.2, programmed death ligand 1 (PD-L1), PD-L2, ICOS-L, B7-H3, and B7-H4 on the AML blasts of 30 newly diagnosed patients and their corresponding receptors [cytotoxic T lymphocyte-associated protein 4 (CTLA-4), programmed death 1 (PD-1), and inducible T cell costimulator (ICOS)] on bone marrow (BM) T cell maturation populations. We could thus evidence B7-negative and B7-positive leukemias either with an isolated expression or part of eight different checkpoint ligand “signatures” that always included an inhibitory B7 molecule. B7-positive AMLs encompassed intermediate and adverse European Leukemia Net (ELN) risk cases and displayed mainly central memory CD4+ T cells with high ICOS levels and effector CD8+ T cells with high PD-1 expression. B7-negative cases were rather classified as AML with recurrent genetic anomalies and displayed predominantly naive T cells, with the exception of NPM1 mutated AMLs, which expressed B7-H3. These different B7 immune profiles suggest that specific immunotherapies are required to target the distinct immune evasion strategies of this genetically heterogeneous disease.

CD4+ T cell exhaustion revealed by high PD-1 and LAG-3 expression and the loss of helper T cell function in chronic hepatitis B

BackgroundImmune inhibitory receptors play an important role in chronic infections. However, little is known about their role in hepatitis B virus (HBV) infection. Here, we analyzed the relationship between programmed death-1 (PD-1) and lymphocyte activation gene-3 (LAG-3) expression on CD4+ T cells and HBV disease progression.ResultsPD-1 and LAG-3 expression was significantly higher on CD4+ T cells from HBV patients than on those from the HCs. In addition, a significant positive correlation was found between the PD-1 and LAG-3 expression levels and the ALT(alanine aminotransferase) level. CD4+ T cell function was inhibited by high PD-1 and LAG-3 levels, and CD4+ T cells with high PD-1 and LAG-3 expression lost the ability to secrete IFN-γ, IL-2 and TNF-α. Furthermore, blockade of the PD-1 and LAG-3 pathways reversed the damage to CD4+ T cell proliferation and cytokine secretion.ConclusionsCD4+ T cell exhaustion during chronic HBV had high PD-1 and LAG-3 expression and the absence of helper T cell cytokines, including IFN-γ, IL-2 and TNF-α. After blocking PD-L1 and LAG-3, CD4+ T cell function in chronic hepatitis B patients was partially restored.

Clinical significance of programmed death-1 and programmed death-ligand 1 expression in the tumor microenvironment of clear cell renal cell carcinoma.

Recently, immunotherapy based on blocking immune checkpoints with programmed death‐1 (PD‐1) or PD‐ligand 1 (PD‐L1) Abs has been introduced for the treatment of advanced clear cell renal cell carcinoma (ccRCC), especially tumors resistant to vascular endothelial growth factor‐tyrosine kinase inhibitors (VEGF‐TKIs), but the significance of their expression in the tumor microenvironment is unclear. We investigated these immune checkpoint markers in tumor cells and tumor‐infiltrating immune cells (TIIC) in the tumor microenvironment of 100 untreated and 25 VEGF‐TKI‐treated primary ccRCC tissues. Upregulated expression of PD‐1 and PD‐L1 by TIIC, and PD‐L1 by tumor cells was associated with the histological grade and unfavorable prognosis of RCC patients. High PD‐1 and PD‐L1 expression by TIIC was associated with a poorer response to VEGF‐TKI, whereas PD‐L1 expression by tumor cells did not affect the efficacy of the treatment. Furthermore, increased PD‐1‐positive TIIC and PD‐L1‐positive TIIC were observed in tumors treated with VEGF‐TKIs compared with those in untreated tumors. Our data suggest that PD‐1 and PD‐L1 expression by TIIC in the tumor microenvironment is involved in treatment resistance, and that sequential therapy with immune checkpoint inhibitors could be a promising therapeutic strategy for ccRCC resistant to VEGF‐TKI treatment.

Immune checkpoint markers in gastroenteropancreatic neuroendocrine neoplasia.

Cancer immunotherapy has evolved major breakthroughs in the last years. The cell-surface receptor programmed death-1 (PD-1) and its ligand, programmed death ligand-1 (PD-L1), have been detected in various cancer types. However, the analysis on gastroenteropancreatic neoplasia (GEP-NENs) is limited. Therefore, the aim of this study was to characterize GEP-NENs with regard to PD-1/PD-L1 pathway and tumor-infiltrating lymphocytes (TILs). On protein level, we examined TILs, PD-1 and PD-L1 expression in tumor tissue of 244 GEP-NENs using immunohistochemistry. Expression levels were correlated with clinicopathological parameters including long-term survival in an observational study. In total, 244 patients could be included. Most of the patients had a NEN of the small intestine (52.5%) or the pancreas (29.5%). All tumors could be graded by their morphology and Ki67 index, with 57.8% G1, 34% G2 and 8.2% G3 tumors. High TILs (19.6%) and high PD-1 (16.1%) expression showed a significant correlation with shorter patient survival (P < 0.05) and with a higher grading. Furthermore, expression of PD-L1 (8.7%) showed a trend to shorter patient survival. High TILs and PD-1 expression are significantly associated with shorter patient survival and higher grading in GEP-NENs. PD-L1 expression showed a trend to shorter patient survival. Immunotherapy might be a promising therapeutic approach in GEP-NENs especially in tumors with high TILs.

Spleen-Associated Effects on Immunity in Hepatitis B Virus-Related Cirrhosis with Portal Hypertension.

Our study aimed to investigate the histologic and immunological changes of portal hypertension (PH) pre- and postsplenectomy in hepatitis B virus (HBV)-related cirrhosis. Peripheral blood samples were obtained from 30 patients with HBV-related cirrhosis and PH at pre- and postsplenectomy time points and from 15 healthy subjects. Spleen tissue specimens were collected from 15 of the patients with HBV-related cirrhosis and from 8 control patients who had undergone splenectomy due to trauma. Immunohistochemical staining was performed to evaluate the immune effector cells and the expression of negative immune regulators. Flow cytometry was used to investigate the immunophenotypes and percentages. The spleen of cirrhotic patients with PH showed extensive depletion of splenic CD4, CD8, and human leukocyte antigen DR cells along with overexpression of the inhibitory receptors programmed death-1 (PD-1) and T cell immunoglobulin domain and mucin domain-3 and their ligands (PD-L2 and galectin-9). Peripheral blood of patients with PH showed remarkable decrease in proportions of CD8 T cell and natural killer (NK) cells and increase in regulatory T (Treg) cells, as well as high expression of PD-1 in CD4/8 T cells. Compared with presplenectomy patients, cirrhotic patients with PH showed increased proportions of CD8 and NK cells, decreased proportion of Treg cells, and decreased expression of PD-1 in peripheral blood CD4/8 T cells after splenectomy. PH-spleen could lead to peripheral tolerance and immunosuppression in HBV cirrhotic patients, and splenectomy may cause beneficial immunological changes.

The Immune Checkpoint Regulator PDL1 is an Independent Prognostic Biomarker for Biochemical Recurrence in Prostate Cancer Patients Following Adjuvant Hormonal Therapy.

Background: The programmed death 1 (PD1)/programmed death ligand 1 (PDL1) targeted therapies have gained positive outcomes in several tumors, but the evidence of the expression and prognosis value of PD1/PDL1 in high risk prostate cancer was rare.Methods: Immunohistochemical analysis of PDL1/PD1 expression by a validated antibody was performed in a retrospectively collected high risk prostate cancer cohort who received adjuvant hormonal therapy (AHT) after radical prostatectomy (RP). The association between PDL1/PD1 expression and prognosis was determined.Results: In total, 127 patients were enrolled. 49.6% patients were considered PDL1-high expression while the PD1-positive expression proportion was 24.4%. High PDL1 and negative PD1 expression were significantly associated with lower prostate specific antigen (PSA) density (p=0.010 and p=0.033, respectively). Compared with the PDL1-low expression patients, the PDL1-high expression patients had significantly shorter time to PSA nadir (TTN) (P=0.001) and biochemical recurrence (BCR) (P=0.004). In Kaplan-Meier analysis, the PDL1-high expression group (p<0.0001) and the PDL1-high/PD1-negative expression group (p<0.0001) showed markedly lower BCR-free survival in localized disease. Univariate cause-specific Cox proportional hazard regression model concluded total PSA (p=0.047), PDL1-high-expression (p<0.001), PDL1-high/PD1-negative expression (p<0.001) were significant risk factors of shorter progression time to BCR in localized disease. PDL1-high-expression was the independent predictor of time to BCR in multiple Cox regression of all patients (Hazard ratio [HR]: 3.901; 95% Confidence interval [CI]: 1.287-11.824; p=0.016).Conclusions: PDL1 expression is not only highly prevalent in high-risk prostate cancer, but is also an independent biomarker in the prognosis of high-risk prostate cancer received AHT after RP. PDL1/PD1 targeted therapy might be a potentially adjuvant treatment option for high-risk prostate cancer after RP.

Tumor microenvironment classification based on T-cell infiltration and PD-L1 in patients with mismatch repair-proficient and -deficient colorectal cancer.

The classification of tumor microenvironments according to the presence or absence of tumor infiltrating lymphocytes (TILs) and programmed death ligand-1 (PD-L1) expression has been used to predict the efficacy of immune checkpoint inhibitor antibodies in several cancer types, not including colorectal cancer (CRC). The current study investigated the TIL/PD-L1 status of patients with CRC, particularly patients who presented as mismatch repair-proficient (pMMR) and mismatch repair-deficient (dMMR). A total of 243 patients with CRC were enrolled and defined as pMMR (121 patients) or dMMR (122 patients). Using Pearson's χ2 test and multivariable multinomial logistic regression analysis, the associations between MMR status, TIL presence and PD-L1 expression were investigated, in addition to the association between TIL/PD-L1 status and clinicopathological features. The results demonstrated that the dMMR group more frequently exhibited TIL+ (85/122 vs. 61/121) and PD-L1+ (49/122 vs. 32/121) phenotypes compared with the pMMR group. PD-L1+ expression was identified in 42.4% of TIL+ cases in the dMMR group, while only 18.0% of TIL+ cases were PD-L1+ in the pMMR group. High programmed death-1 expression and dMMR status were revealed as two independent risk factors for TIL+ PD-L1+ status. In conclusion, compared with the pMMR group, the dMMR group was more likely to present with a TIL+ PD-L1+ status, which suggests that a TIL+ PD-L1+ tumor microenvironment may partly contribute to the improved response of dMMR patients to anti-PD-1/L1 therapy.

Tumor Mutational Burden and Other Immunotherapy Markers in Histiocytic Neoplasms Using Next Generation Sequencing

IntroductionHistiocytic neoplasms are rare hematological malignancies that have protean clinical manifestations and can pose significant management challenges. Recently, vemurafenib was approved by the US-FDA for treatment of BRAF-V600-mutant Erdheim-Chester disease (ECD). However, there is a lack of FDA-approved therapies for other histiocytic neoplasms such as Langerhans cell histiocytosis (LCH) and Rosai-Dorfman disease (RDD). Over the last 5 years, immune checkpoint inhibitors such as programmed death-1 (PD-1) and programmed death-ligand 1 (PD-L1) inhibitors have shown significant improvement in outcomes among patients with several hematological and solid organ malignancies. In order to identify appropriate treatment candidates for these therapies, predictive biomarkers have been developed in various cancers. Evidence from solid tumors has suggested a favorable response to checkpoint inhibitor therapy with higher tumor mutational burden (TMB), defined as the number of mutations within a tumor genome. Next generation sequencing (NGS) of various tumors has shown an association between TMB of < 5 mutations/megabase (mut/Mb) to be associated with an absence of benefit from checkpoint inhibitors. In addition, high levels of PD-1/PD-L1 expression and microsatellite instability (MSI) are also correlated with response to therapy. The latter may be a result of somatic or germline alterations in DNA mismatch repair genes. In this study, we report the results for these biomarkers using NGS in patients with histiocytic neoplasms.MethodsWe utilized TempusTM NGS platform to analyze the tissue specimen of patients with histiocytic neoplasm. The Tempus xO Assay (Tempus Labs; Chicago, IL) combines a 1,711 gene targeted somatic and germline DNA sequencing panel with RNA-sequencing to detect both germline and somatic single nucleotide polymorphisms, indels, copy number variants, and gene rearrangements causing chimeric mRNA transcript expression in a wide array of solid tumor types. The assay utilizes formalin-fixed paraffin-embedded tumor samples and matched blood samples. TMB was calculated and reported as somatic mutations in tumor tissue per million base-pairs or mut/Mb. RNA sequencing was utilized to assess for PD-L1 and PD-1 gene expressions as compared to matched tumor and normal reference sets. Both PD-L1 and PD-1 gene expressions were reported as percentiles. DNA mismatch repair status was predicted by analysis of alterations in five common mismatch repair genes in somatic and germline DNA (MSH2, MSH6, MLH1, PMS2, and EPCAM). If there were no alterations identified in these genes, the mismatch repair status was predicted as microsatellite stable (MSS).ResultsA total of 13 patients with histiocytic neoplasms were included in the study. The distribution of individual histiocytic neoplasms was as follows: RDD (n=9), ECD (n=3), and LCH (n=1). The median TMB for RDD and ECD patients was 0.17 mut/Mb. For the one patient with LCH, the TMB level was 0.51 mut/Mb. Individual TMB levels are shown in figure 1. PD-L1 and PD-1 expression levels are depicted in table 1. Compared to normal reference sets, the PD-L1 expression was elevated in one patient each with RDD and ECD, and PD-1 expression was elevated in two patients each with RDD and ECD. For both ECD patients with higher PD-1 expression, NGS also showed presence of BRAF-V600E in the tumor tissue. The LCH patient had a low level of PD-L1 and PD-1 expression. For patients where evaluation of DNA mismatch repair was feasible on the tissue specimen (n=4), none showed related somatic or germline alterations.ConclusionsIn our series, the histiocytic neoplasms RDD, ECD, and LCH demonstrated low levels of TMB. None of the RDD and ECD patients were found to have alterations in DNA mismatch repair genes. Other markers of immunotherapy such as PD-L1/PD-1 expression appeared to be higher in ECD patients with BRAF-V600E, but in only a small subset of RDD patients. The low TMB seen in our study suggests that these histiocytic neoplasms may be less likely to respond to immune checkpoint inhibitors such as anti-PD-1 and anti-PD-L1 agents as compared to the tumors with high TMB. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Clinical and histopathological significance of PD-1 expression in cutaneous lesions of adult T-cell leukemia–lymphoma

BackgroundAdult T-cell leukemia–lymphoma (ATL) is a mature T-cell malignancy caused by human T-cell leukemia virus type I infection and is known to exhibit cutaneous involvement in 50% or more patients. Few studies have evaluated the clinicopathological significance of programmed death-1 (PD-1) expression in the cutaneous lesions of ATL. MethodsSkin biopsy specimens from 29 ATL patients with cutaneous lesions were evaluated regarding the clinicopathological feature, survival outcome, and PD-1 expression level on infilitrated CD3+CD4 + CD25+ cells. The optimal cut-off point of PD-1 expression for clinicopathological feature and outcome was determined as the value of the maximum Youden index by receiver operating characteristic (ROC) analysis. ResultsPD-1 was expressed broadly from zero to 90% on the skin biopsy specimens of the 29 patints, with the median value of 50%. The PD-1-expression level was significantly higher in the poorer-prognosis eruption group (nodulotumoral, erythrodermic and purpuric types) (P = 0.003), in the poorer histopathological infiltration patterns (diffuse and nodular) (P = 0.007), and in the poorer infiltrating cell-size group (large-sized cells) (P = 0.017) than in the corresponding group. ROC curve analyses showed that the optimal cut-off value for PD-1-expression level to predict the poorer-prognosis eruption, the poorer- histopathological infiltration pattern, the poorer infiltration cell size, and the poorer outcome (death) was 60%, 50%, 50%, and 80%, respectively. Patients with high PD-1 expression had a shorter median survival time than those with low PD-1 expression (18.2 months vs. 26.0 months), but the difference was not statistically significant. ConclusionsATL patients with cutaneous lesions in which PD-1 were highly expressed have more advanced dermatological and histopathological patterns and possibly worse survival than those with low PD-1 expression on cutaneous lesions. Further large-scale studies are warranted to verify these findings.

Expression Patterns, Prognostic Value, and Intratumoral Heterogeneity of PD-L1 and PD-1 in Thymoma and Thymic Carcinoma.

Thymic epithelial tumors (TETs) including thymoma and thymic carcinoma are rare tumors with little data available to guide treatment. Immunotherapy with checkpoint blockade has shown promising activity, but data regarding the expression patterns and prognostic implications of programmed death 1 (PD-1) and its ligand (PD-L1) in TETs have yielded conflicting results. Intratumoral heterogeneity of PD-1/L1 expression has been shown in other cancers, but has not been described in the TET literature. We performed a retrospective single-center review of 35 patients with resected TET. PD-1/L1 expression was assessed by immunohistochemistry using PD-1 clone: NAT105 and PD-L1 clone: 22C3. Tumor samples from 35 patients were evaluated including 32 patients with thymoma and 3 patients with thymic carcinoma. PD-L1 expression was detected in 83% (29 of 35) tumor samples, including 100% (3 of 3) of thymic carcinoma patients and 81% (26 of 32) of thymoma patients. PD-1 expression was detected in 77% (27 of 35), including 33% (1 of 3) of thymic carcinoma patients and 81% (26 of 32) thymoma patients. High PD-1 expression was associated with lower grade tumors. Unlike prior studies, PD-L1 expression was not associated with higher grade tumors or higher stage. Neither PD-L1 nor PD-1 expression was significantly associated with survival. Three patients with thymoma had multiple tumor sections evaluated for expression of PD-1/L1, with differing expression patterns of both PD-L1 and PD-1 observed in two patients. This study confirms high expression of PD-L1 and PD-1 in TET and shows for the first time intratumoral heterogeneity of PD-L1 and PD-1 in thymoma patients.