Programmed Death-ligand 1 mRNA Expression Research Articles

Farnesyltransferase inhibitor FTI-277 inhibits PD-L1 expression on septic spleen lymphocytes and promotes spleen lymphocyte activation.

Farnesyltransferase inhibitors have been tested in clinical trials for the treatment of tumours. In sepsis, the binding of programmed death 1 (PD-1) to programmed death ligand 1 (PD-L1) promotes lymphocyte apoptosis and decreases cytokine expression, thus affecting survival rates. The PD-1/PD-L1 pathway plays an important role in chronic viral infection, bacterial infection and sepsis. However, the precise immunosuppressive and anti-inflammatory functions of this pathway remain poorly understood. In our previous study, the induction of sepsis by caecal ligation and puncture (CLP) resulted in increased farnesyltransferase activity and farnesylated protein levels in the spleen relative to sham treatment. However, the effect of inhibition of farnesyltransferase activity on overall survival rates in patients with sepsis and the specific signalling pathway involved remain to be investigated. In this study, mice with CLP-induced sepsis were treated with farnesyltransferase inhibitor (FTI-277), and PD-L1 expression on septic spleen lymphocytes was examined. Flow cytometric analysis revealed that PD-L1 is expressed constitutively on lymphocytes and that PD-L1 protein expression was up-regulated strongly following CLP. FTI-277 down-regulated PD-L1 mRNA and protein expression on septic spleen lymphocytes in a dose-dependent manner. This effect was associated closely with nuclear factor kappa B (NF-κB). In addition, the significant damping effect of FTI-277 on the PD-L1 signal promoted interferon (IFN)-γ secretion, interleukin (IL)-2 production and splenocyte proliferation in response to anti-CD3+ CD28+ antibodies in mice. Furthermore, FTI-277 reduced spleen lymphocyte apoptosis in septic mice. Therefore, FTI-277 regulates spleen lymphocyte activity via the PD-L1 signalling pathway, with significant anti-inflammatory effects attributable to suppression of the NF-κB pathway. Farnesyltransferase represents a valuable therapeutic target for the treatment of sepsis.

Abstract 458: Choline metabolism in cancer cells modifies PD-L1 expression

Abstract Increased expression of choline kinase-α (Chk-α), the enzyme that converts choline to phosphocholine has been observed in most cancer cells including breast (1). This increase of choline kinase results in increased production of phosphocholine that is used in the synthesis of membrane phosphatidylcholine. We investigated the relationship between Chk-α and the programmed death ligand-1 (PD-L1) expression in triple negative MDA-MB-231 human breast cancer cells. Expression of PD-L1 in response to the secretion of interferon gamma (IFNγ) by cytotoxic T cells and its binding to the immune checkpoint PD-1 on T cells results in cancer cells escaping destruction by the immune system (2). Quantitative real-time PCR analysis (q-RT-PCR) did not reveal any change in PD-L1 mRNA expression between MDA-MB-231 expressing an empty vector control plasmid (231-EV) and MDA-MB-23 stably expressing shRNA against Chk (231-Chk). However, there was a difference in the mean intensity fluorescence (MIF) between 231-EV (325 ±11.5, n=2) and 231-Chk (527±18.5, n=2) in the FACS analysis. Upon induction with IFNγ (10ng/ml) for 24h, there was 1.23 fold increase in PD-L1 mRNA in 231-Chk compared to 231-EV. Interestingly, the MIP for cell surface expression of PD-L1 was 600±21 (n=2) for 231-EV compared to 872±16 (n=2) for 231-Chk cells. Values represent mean + SE. These data suggest that the cell surface increase of PD-L1 in 231-Chk cells could be due to translocation. Increased PD-L1 expression creates an opportunity to deliver PD-L1 targeted nanoparticles carrying therapeutic cargo. Since most conventional treatments result in a decrease of Chk-α and phosphocholine, our data suggest that these treatments may also result in cancer cells escaping immune surveillance through expression of PD-L1. Acknowledgements: This work was supported by NIH R01CA136576 and P50 CA103175. Reference: 1) Glunde K et al, Cancer Res 65, 11034 (2005)., 2) Zou W, Chen L, Nat Rev Immunol 8(6), 467-77 (2008). Citation Format: Balaji Krishnamachary, Marie-France Penet, Yelena Mironchik, Zaver M. Bhujwalla. Choline metabolism in cancer cells modifies PD-L1 expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 458. doi:10.1158/1538-7445.AM2017-458

Abstract 5006: Correlation between PD-L1 expression and tumor clinicopathologic characteristics in early stage breast cancer

Abstract Background: Upregulation of Programmed Death-Ligand 1 (PD-L1) in tumor cells results in the deactivation of cytotoxic T cells in the tumor microenvironment and development of tolerance to the malignant cells. We aimed to investigate the role of PD-L1 expression in early stage breast cancer and its correlation with tumor characteristics as well as assess the relationship between PD-L1 membrane protein and gene expression. Methods: For this single center, retrospective study, women (age >18 years) with early stage breast cancer diagnosed between January 2005 and December 2005 were enrolled. Key inclusion criteria for enrollment were pathologic diagnosis of DCIS or invasive ductal/lobular carcinoma, stage 0-2, with local resection done in 2005. Patients had to have adequate formalin-fixed paraffin-embedded tumor specimens for PD-L1 testing. We hypothesized that higher levels of PD-L1 expression correlates with more aggressive disease and worse prognosis. We used histopathological analysis, medical chart review, and PD-L1 expression via immunohistochemistry (IHC) and reverse transcription polymerase chain reaction (RT-PCR) for data analysis. Chi Square and one way ANOVA analyses were used to compare clinical and pathological characteristics of patients with high and low PD-L1 expression. Results In 2005, 368 women were diagnosed with breast cancer at Providence-Providence Park Hospital of which 31 pathological specimens were obtained for exploratory analysis. PD-L1 protein expression was determined to be positive if more than 1% of the tumor cells showed partial or complete membrane staining for PD-L1 by IHC. All patients with DCIS (n=7) were negative for PDL-1 expression although 3 showed gene expression by PCR. After excluding the DCIS specimens, 17% (4/24) of early stage breast cancer was found to express PD-L1 protein. Seven of 24 specimens (excluding DCIS) were positive for PD-L1 RNA by RT-PCR. There was no relationship between PD-L1 protein and mRNA expression (p=0.552). 75% of the PD-L1 positive specimens by IHC were also ER positive although this was not statistically significant (p=0.312). There was a trend toward larger tumors for those expressing PD-L1 on the cell surface (mean size of 2.7 cm vs 1.7 cm, p= 0.07). All the PD-L1 positive specimens had increased lymphocyte infiltration, although no statistical significance was observed. At 10 years from initial diagnosis, all-cause mortality rate was 29% (7/24). The rate of death in PD-L1 positive patients was 2/4 (50%) compared to 5/20 (25%) in PD-L1 negative group. Conclusion: Expression of PD-L1 in early stage breast cancer could be an important indicator for more aggressive disease and supports investigating checkpoint inhibitors in this population. Further investigation is needed to determine its applicability in clinical decisions and effects on treatment. Citation Format: Hadeel Assad, Juan Cattoni, Subramanyeswara Arekapudi, Nancy Jackson, Hugo Macchi Cattoni, Sherri Motahari, Hema Govil, Anibal Drelichman. Correlation between PD-L1 expression and tumor clinicopathologic characteristics in early stage breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5006. doi:10.1158/1538-7445.AM2017-5006

Effects of programmed death-ligand 1 expression on OK-432 immunotherapy following transurethral resection in non-muscle invasive bladder cancer.

The present study aimed to investigate the effect of the negative costimulatory molecule programmed death-ligand 1 (PD-L1) on immunotherapy with OK-432, following transurethral resection of bladder tumors in non-muscle invasive bladder cancer (NMIBC), and to elucidate the underlying mechanism. PD-L1 was detected by immunohistochemical staining in tumor specimens from 55 cases of NMIBC following postoperative immunotherapy with OK-432. The PD-L1 mRNA and protein expression levels were measured in the bladder cancer T24 cell line and the human uroepithelial SV-HUC-1 cell line, following treatment with interleukin (IL)-2, interferon (IFN)-α and IFN-γ, by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis, respectively. PD-L1 was widely expressed in the NMIBC tumors, with 56.4% (31/55) of specimens exhibiting positive staining. When compared with PD-L1-negative patients, PD-L1-positive patients exhibited significantly increased recurrence [48.4% (15/31) vs. 16.7% (4/24)] and progression [16.1% (5/31) vs. 4.2% (1/24)] rates (P<0.05). RT-qPCR and western blotting demonstrated that cytokines IL-2, IFN-α and IFN-γ markedly upregulated PD-L1 mRNA expression rates and protein levels in bladder cancer T24 cells (P<0.05), but had no significant effect in non-tumor SV-HUC-1 cells. In conclusion, PD-L1 expression was negatively-associated with the efficacy of OK-432 intravesical immunotherapy in patients with NMIBC. The results indicated that the involved mechanism occurred via upregulation of PD-L1 by immune cytokines, which in turn suppressed the antitumor effectiveness of the immune system, thereby promoting tumor recurrence and progression.

MP37-01 PROGRAMMED DEATH-LIGAND 1 (PD-L1) EXPRESSION IN PHEOCHROMOCYTOMA

You have accessJournal of UrologyAdrenal1 Apr 2017MP37-01 PROGRAMMED DEATH-LIGAND 1 (PD-L1) EXPRESSION IN PHEOCHROMOCYTOMA Yasuhiro Hashimoto, Atsushi Imai, Shingo Hatakeyama, Takahiro Yoneyama, Takuya Koie, and Chikara Ohyama Yasuhiro HashimotoYasuhiro Hashimoto More articles by this author , Atsushi ImaiAtsushi Imai More articles by this author , Shingo HatakeyamaShingo Hatakeyama More articles by this author , Takahiro YoneyamaTakahiro Yoneyama More articles by this author , Takuya KoieTakuya Koie More articles by this author , and Chikara OhyamaChikara Ohyama More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.1133AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Programmed death ligand-1 (PD-L1), a key target molecule for immunotherapy, is frequently overexpressed in several neoplasms. In the present study, we examined PD-L1 expression in pheochromocytoma because very few reports on this subject are available. METHODS PD-L1 mRNA expression was compared across 184 pheochromocytoma, 492 prostate cancer, and 404 bladder cancer cases based on The Cancer Genome Atlas (TCGA). Furthermore, we enrolled 32 pheochromocytoma patients who were surgically treated at our hospital between June 2005 and February 2016. We conducted PD-L1 immunohistochemistry (IHC) using the SP142 assay. PD-L1 expression was scored at three diagnostic levels (0, 1, and 2). RESULTS A comparison of PD-L1 mRNA expression based on the TCGA revealed that PD-L1 expression was significantly higher in pheochromocytoma than in bladder cancer and prostate cancer (p < 0.001). The SP142 assay of our 32 surgical pheochromocytoma cases revealed that the prevalence of a PD-L1(+) expression (IHC score 1 or 2) in tumor-infiltrating immune cells (TICs) was 25% (eight patients) and that in tumor cells (TCs) was 28.1% (nine patients). The tumor diameter was significantly different between PD-L1(+) TIC patients (3.36 ± 0.35 cm) and PD-L1(-) TC patients (5.37 ± 0.50 cm, non-paired t-test: p = 0.044). In our cohort, there were two cases of malignant pheochromocytomas but none of them were PD-L1(+). CONCLUSIONS PD-L1 expression is relatively higher in pheochromocytoma than that in bladder cancer and prostate cancer based on TCGA. The SP142 assay of our 32 surgical pheochromocytoma cases revealed that the tumor diameter in PD-L1(-) TIC cases was larger than that in PD-L1(+) cases. Further, there were no PD-L1(+) cases of malignant pheochromocytoma. These findings suggest that PD-L1 expression in pheochromocytoma is relatively common and that PD-L1(+) expression in pheochromocytoma may not be associated with tumor aggressiveness. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e474 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Yasuhiro Hashimoto More articles by this author Atsushi Imai More articles by this author Shingo Hatakeyama More articles by this author Takahiro Yoneyama More articles by this author Takuya Koie More articles by this author Chikara Ohyama More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Hypoxia-inducible factor 1α in regulation of programmed death: Ligand 1 in glioma under hypoxia microenvironment.

103 Background: Glioma is a highly lethal tumor that is known for its immune inhibitory capabilities. Despite great progress in treatment modalities, the median survival time of high-grade gliomas patients is only about 11.1 months. Gliomas are aberrantly expressed programmed death–ligand 1 (PD-L1) which results in tumor-induced immune suppression. Hypoxia is well known to induce aggressiveness, promotes tumor progression and resistance to chemotherapy and radiotherapy. Recently, studies have indicated that hypoxia play a prominent role in tumor immune escape. The aim of the study was to explore the relationship between PD-L1 and hypoxia inducible factor 1α (HIF-1α) and the role in the progression of gliomas. Methods: We adopted immunohistochemistry methods to detect expressions of PD-L1 and HIF-1α in 64 surgical specimens (23 cases for low-grade and 41 cases for high-grade gliomas, respectively). Immunoreactivity was analyzed in association with patients’ clinical characteristics or survival time. Expression of the PD-L1 and HIF-1α was analyzed in three GBM cell lines, U343, U373, and U251, under in vitro hypoxia (12, 24 or 72 h at 0.1% O2). Reverse transcriptase-polymerase chain reaction (RT-PCR) was carried out to detect the mRNA expression of PD-L1 and HIF-1α. Results: PD-L1 and HIF-1α protein expression was detected in 81.25% and 84.73% of glioma specimens, respectively. The expression of PD-L1 protein is positively correlated with HIF-1α (r = 0.52; p= 0.013). Kaplan–Meier analysis revealed a significant effect of PD-L1 grade on cumulative overall survival. Patients with negative PD-L1 expression survive longer than those with positive expression ( p= 0.007). PD-L1 and HIF-1α mRNA was most consistently upregulated in relation to duration of in-vitro hypoxia (72h, p= 0.007). There was a significant and positive relationship between the PD-L1 and HIF-1α mRNA expression (r = 0.45; p= 0.01). Conclusions: The present study showed a role for hypoxia/HIF-1α in driving immune escape and provided a potential novel cancer immunotherapy targeting hypoxia to block PD-L1 expression, which may boost the immune system in cancer patients.

Programmed death ligand 1 is expressed in canine B cell lymphoma and downregulated by MEK inhibitors.

Programmed death ligand 1 (PD-L1) expression in antigen-presenting cells and tumors can inhibit T cell-mediated immunity. In this study, PD-L1 mRNA and protein expression was evaluated in canine B cell lymphoma (CLL17-71), large T-cell leukemia (CLGL-90), B cell leukemia (GL-1) and primitive leukocyte round cell neoplasia (CLL-1390). Variable PD-L1 mRNA and protein were observed in these cells with high endogenous expression present in CLL17-71 cells. PD-L1 protein was also observed in canine patient B cell lymphoma tissues using immunostaining. PD-L1 and signal transducer and activator of transcription 1 ( STAT1 ) mRNA expression were reduced in the presence of mitogen-activated protein kinase kinase 1.2 (MEK1/2) inhibitors RDEA119 and AZD6244 in CLL 17-71 cells. RDEA119 had similar effect on PD-L1 and STAT-1 in IFN-γ activated CLL-1390 cells. Overall, these results indicate that PD-L1 is expressed in canine B cell lymphoma. Its inhibition by MEK1/2 inhibitors suggests a possible treatment strategy using targeted drugs which likely could enhance antitumor immune response.

Abstract A18: Investigating the differential expressions of miRNAs and programmed-death-ligand-1 related to the T-bet positive tumor-infiltrating lymphocytes in axillary-node-negative breast cancer

Abstract Axillary lymph node-negative (ANN) breast cancer (BC) patients generally have a good prognosis; however, 20-30% of ANN patients will experience recurrence. To identify biomarkers that distinguish ANN patients who may benefit from systemic therapy we have been studying characteristics of the breast tumor cells and the tumor microenvironment. We have found that tumor-infiltrating lymphocytes (TILs) that express T-bet (T-box transcription factor) are associated with poor prognosis features such as high grade, hormone receptor negativity and basal subtype. Despite these adverse clinical-pathologic characteristics, T-bet+ TILs cells are associated with a favorable outcome. T-bet is essential for Th1 development and IFN-γ production. To investigate how T-bet+ TILs influence the BC suppression, we are examining molecular differences in T-bet+/high and T-bet-/low tumors including expression of tumor miRNA expression profiles and expression of Programmed-Death-Ligand-1 (PD-L1). Tumors expressing PD-L1 inhibit Programmed-Death-receptor-1-(PD-1)-expressing TILs and evade immune surveillance. Interestingly, PD-L1 is induced by IFNγ, the hallmark cytokine of the T-bet+ TILs, which suggests a potential relationship between the tumor PD-L1 and T-bet+ TILs. Microdissected, formalin-fixed, paraffin-embedded (FFPE) samples of 35 T-bet+/high and 35 T-bet-/low ANN BC tumors are being used to identify the differentially-expressed miRNAs via miRNA sequencing. The sufficiency of the FFPE samples as the miRNA source was confirmed by quantifying miRNA expressions in the 3-5 paired frozen and FFPE samples, and observing the expressions to be highly correlated with both qPCR (r=0.81) and miRNA sequencing (rs=0.73). After identifying the differentially-expressed miRNAs via miRNA sequencing, these miRNAs will be validated via qPCR in both the original and independent BC tumor set. Based on the genes and molecular pathways the validated miRNA(s) may regulate, appropriate in vitro functional assays will be conducted to examine their roles in cellular proliferation, migration and invasion. Furthermore, PD-L1 mRNA expression has been evaluated in 107 ANN tumors and correlations with clinico-pathological characteristics will be determined. Identifying the tumor miRNA(s) associated with T-bet+ TILs and studying their functions may indicate differences between T-bet+/high and T-bet-/low tumors that can be used to improve the prognosis of ANN BC. Characterizing the tumor PD-L1 expression based on T-bet+ TILs and the clinical data may distinguish a subset of ANN BC patients who may benefit from the PD-L1 blockade therapy. Citation Format: Minji Lee, Anna-Marie Mulligan, Shelley Bull, Dushanti Pinnaduwage, Irene Andrulis. Investigating the differential expressions of miRNAs and programmed-death-ligand-1 related to the T-bet positive tumor-infiltrating lymphocytes in axillary-node-negative breast cancer. [abstract]. In: Proceedings of the AACR Special Conference: Function of Tumor Microenvironment in Cancer Progression; 2016 Jan 7–10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2016;76(15 Suppl):Abstract nr A18.

Quantification of PD-L1 mRNA Expression in Lung Adenocarcinoma with EGFR mutation.

e23261Background: Programmed death-ligand 1 (PD-L1) expression on immunohistochemistry is associated with response to PD-1 blockade.However, the molecular mechanism and clinical significance of PD-...

Defective programmed death ligand-1 (PD-L1) expression on monocytes from systemic lupus erythematosus (SLE) patients can be corrected by a soluble T lymphocyte factor (99.15)

Abstract PD-L1 is an inhibitory protein expressed on monocytes (Mo) to maintain peripheral T cell tolerance. We previously reported deficient expression of PD-L1 on Mo from patients with active SLE. To investigate the mechanisms for dysregulated PD-L1 expression in SLE we isolated peripheral blood mononuclear cells (PBMC) from juvenile SLE patients and age-matched controls. PD-L1 expression was measured by flow cytometry and realtime quantitative RT-PCR. Cytokines were analyzed by Luminex. We found that PD-L1 expression on SLE Mo was significantly induced by PBMC or CD4+ T lymphocytes from healthy subjects, and also by supernatant from healthy PBMC. Conversely, PD-L1 expression was decreased in healthy donor Mo by addition of SLE PBMC or supernatant. SLE cells had an increased rate of apoptosis, and inhibition of caspase activity led to decreased apoptosis and upregulation of PD-L1. PD-L1 mRNA expression did not correlate with protein expression. Among 15 cytokines tested, only IL-2 and IL-10 were elevated in plasma of SLE patients with active disease. These results suggest that expression of PD-L1 in juvenile SLE is controlled by caspases at the protein level via soluble factors normally produced by T lymphocytes. Abnormal T lymphocyte activity in SLE may prevent induction of PD-L1, leading to loss of peripheral tolerance. Supported by the Lupus Foundation and the Lupus Research Institute