PD-1／PD-L与慢性病毒性肝炎

PD-1／PD—L信号途径在保持对机体自身成分的免疫耐受中发挥作用,同时在对外来抗原的免疫反应中发挥负向调控作用。病毒慢性持续感染、T细胞耗竭、T细胞或其它组织细胞PD-1／PD—L表达的增加常同时俘在。肝脏多种非实质细胞和肝细胞均可表达PD-L1,具有抑制T细胞活性的作用。目前,PD-1／PD—L与慢性病毒性肝炎的发病机制和治疗对策关系的研究方兴未艾。     

PD-1/PD-L1 blockade in renal cell cancer

Introduction: Immunotherapy using checkpoint inhibitors is providing significant benefit to patients with renal cell carcinoma (RCC), both in overall survival and tolerability of treatment. Given the recent approval of the first checkpoint inhibitor in RCC, this review discusses the background and clinical data for checkpoint inhibition in RCC.

Areas covered: This review introduces and discusses the basic biologic mechanisms of checkpoint inhibitor function and focuses on the current evidence in clinical trials for the use of immunotherapy in RCC.

Expert commentary: Immunotherapy has been a mainstay of therapy in RCC, but the recent approval of nivolumab with ORR of 25% and durable responses has provided a transformative new therapeutic option.     

Immunohistochemical localization of programmed death-1 ligand-1 (PD-L1) in gastric carcinoma and its clinical significance

The present study examined programmed death-1 ligand-1 (PD-L1) detected by immunohistochemical labeling in 102 cases of human gastric carcinoma, 10 adenoma and 10 normal tissues. The relationship between PD-L1 immunolocalization and clinical pathological features, as well as the prognosis of gastric carcinoma, was explored. There was no PD-L1 detectable in normal gastric tissues and very weak immunolabeling in gastric adenomas, but it could be detected in 42.2% of gastric carcinoma tissues. There was no correlation between PD-L1 immunolocalization and patient age, sex, tumor location or the degree of tumor differentiation in the gastric carcinomas. However, PD-L1 immunodetection was significantly correlated to tumor size, invasion, lymph node metastasis and survival time of patients. PD-L1 immunolabeling was significantly enhanced (P<0.01) when the tumor infiltrated into the deep muscular layers, with lymph node metastasis or survival time of less than 2 years, Moreover, multivariate analysis demonstrated that PD-L1 immunodetection could be used as an independent factor to evaluate the prognosis of gastric carcinoma.     

PD-1/PD-L1在非小细胞肺癌中的研究进展及展望

PD-1(programmed cell death-1,程序性死亡受体1)与其配体PD-L1(programmed cell death-ligand 1,程序性死亡配体1)属于CD28/B7家族,是一对共刺激分子,具有负性调控作用。PD-1通过与其配体PD-L1结合调节肿瘤的微环境,使肿瘤细胞免于机体免疫系统的监视和清除。目前已有较多研究显示PD-1/PD-L1在非小细胞肺癌组织中的表达水平与患者的临床病理因素及预后存在显著的相关性。在非小细胞肺癌的治疗领域,以PD-1/PD-L1为代表的免疫治疗成为继手术治疗、化疗、放疗、分子靶向治疗之后的新焦点。PD-1/PD-L1抑制剂在一系列非小细胞肺癌临床试验中也显示出了巨大的临床潜力。本文就PD-1/PD-L1的生物学结构及其在非小细胞肺癌中的作用机制、研究进展及展望作一综述。     

Programmed cell death ligand 1 (PD-L1) expression on gastric cancer and its relationship with clinicopathologic factors

Background: Targeting the immune checkpoints in solid tumors becomes hot recently. Programmed cell death ligand 1 (PD-L1) is ligand for programmed death 1 (PD-1), which is known to negatively regulate T-cell activation. In the present study, we investigated the expression of PD-L1 in tumor specimens of gastric cancer and its relationships with clinicopathological variables and survival. Methods: The expression of PD-L1 in 132 surgically resected specimens of stage II and III gastric cancer was evaluated by immunohistochemistry in microarray tissue. Results: Expression of PD-L1 was observed in 50.8% (67/132) of gastric cancer tumor specimens. Patients whose tumor size over 5cm had a higher positive rate of PD-L1 expression. There was no relationship between the expression of PD-L1 and other clinicopathological variables including age, gender, clinical stage, location as well as histological differentiation. PD-L1 positive patients had significantly poorer survival than negative patients. The 5-year survival rates was 83.1% in those with PD-L1 negative patients and 50.7% for PD-L1 positive patients (P<0.001). The multivariate analysis indicated that both PD-L1 positive and Tumor-node-metastasis stage were independent prognostic factors in gastric cancer patients (P=0.001 and 0.025, respectively). Conclusions: The expression of PD-L1 was found in half of stages II and III gastric cancer patients. Positive of PD-L1 expression indicated poor survival in Chinese stages II and III gastric adenocarcinoma patients. These results may provide the clue for immunotherapy in the adjuvant treatment setting of gastric cancer patients.     

LncRNA XLOC_003810 promotes T cell activation and inhibits PD-1/PD-L1 expression in patients with myasthenia gravis-related thymoma

This study aimed to investigate the effect of long non-coding RNA XLOC_003810 on the activation of CD4⁺ T cells and expression of PD-1/PD-L1 in patients with myasthenia gravis-related thymoma (MG-T). Thymus specimens and thymic mononuclear cells were obtained from MG and MG-T patients or cardiac surgery patients undergoing thoracotomy who were selected as negative controls (NC). XLOC_003810 expression was examined using quantitative real-time PCR (qRT-PCR). Frequency of CD4⁺ T cells and proportion of CD4⁺PD-1⁺ T cells and CD14⁺PD-L1⁺ monocytes were quantified by flow cytometry. The release of inflammatory cytokines was measured by qRT-PCR and enzyme-linked immunosorbent assay. Compared with the NC group, expression of XLOC_003810, frequency of CD4⁺ T cells and the production of inflammatory cytokines were increased in patients with MG and MG-T. XLOC_003810 overexpression significantly increased the frequency of CD4⁺ T cells, facilitated the production of inflammatory cytokines and decreased the proportion of CD4⁺PD-1⁺ T cells and CD14⁺PD-L1⁺ monocytes in the thymic mononuclear cells. In contrast, XLOC_003810 knockdown exerted the opposite effect. Together, XLOC_003810 promotes T cell activation and inhibits PD-1/PD-L1 pathway in patients with MG-T.     

Programmed cell death protein 1 (PD-1) in infection

Exhausted and dysfunctional T cells triggered by infection and cancer render the immune system unable to eliminate these pathogens. Pharmacologic blockade of the surface receptors that inhibit T-cell function has shown remarkable success in patients with various malignancies. In this Review, we discuss the emerging evidence of inhibiting checkpoint pathways as a potential role in controlling or clearing infectious diseases. Though interesting tendencies, much work is still needed in order to develop safe strategies that can be translated into clinically relevant outcomes in patients with infections.     

程序性死亡受体-1及其配体在乙型肝炎慢性化机制中的作用

乙型肝炎病毒（HBV）感染后容易形成慢性化,多数学者认为其主要机制是宿主对HBV各种抗原产生免疫耐受。程序性死亡受体-1（PD-1）是近年新发现的一个负性共刺激信号分子,其配体为PD—L1和PD—12,同属于CD28／B7家族。PD-1／PD—L通路能削弱、限制和／或终止T细胞和抗原递呈细胞（APC）等细胞的活化及效应功能,在慢性HBV感染的免疫耐受机制中发挥了重要作用。     

应用基因芯片筛选慢性乙型肝炎患者PD-1/PD-L通路基因差异表达

目的 应用基因芯片技术筛选慢乙肝病人PD-1/PD-L通路差异性表达的基因.方法 应用基因芯片技术筛选4例慢乙肝病人与2例健康人外周血,获得差异性表达的基因,结合初步构建的PD-1/PD-L信号通路,挑选出一致的转录因子或调控蛋白,并应用Western blot方法检测CD 40L、Bcl-10、GATA-3、NFAT蛋白的水平的表达.结果 与健康人相比,慢乙肝患者有838个异常表达的基因,其中高表达的基因有150个,低表达的基因有688个.在这些异常表达基因中,有13个基因属于所构建的PD-1/PD-L信号通路中的转录因子或调控蛋白,其中表达上调的6个分别是BCL-10、AP、SRY、MYOD1、ELK4、NFAT,表达下调的7个分别是CD 40L、GADS、PDK1、IL-4、GATA3、CREB、RARG.Westem blotting法证实慢乙肝组高表达BCL-10,低表达CD 40L、GATA3,NFAT但与病毒载量不相关.结论 CD40L、BCL-10、GATA3、NFAT在慢性乙型肝炎患者PD-1/PD-L信号传导通路中可能有重要的调控作用,可以做深入的功能分析研究.     

The PD-L1/PD-1 axis expression on tumor-infiltrating immune cells and tumor cells in pediatric rhabdomyosarcoma

BackgroundActivation of immune checkpoints, e.g. PD-1/PD-L1 axis, in cancer microenvironment, enables evasion of host anti-cancer immune response and drives tumor progression. To date, there have been only a few studies analyzing PD-1/PD-L1 expression in pediatric malignancies.AimIn the current study, we aimed to assess PD-L1 and PD-1 expression in pediatric rhabdomyosarcoma (RMS) and to investigate their clinicopathological associations.Materials and methods: The study enrolled 31 children with RMS. Tissue microarrays with representative tumor tissue samples were stained with anti-PD-1 NAT105 clone (Ventana, Roche) and two different antibodies against PD-L1: SP142 (Ventana, Roche) and 22C3 (DAKO). Adequate positive controls were applied. Their expression was assessed in tumor-associated immune cells (TAICs) and in the tumor cells separately.ResultsWe did not detect any positive PD-L1 staining in analyzed tumors using SP142 antibody; however, in 11 cases (35.48%) its expression was revealed by means of 22C3 clone. The staining was restricted to TAICs in all cases, which no reaction in tumor cells. The 5-year relapse free survival (RFS) rate was significantly higher in PD-L1 positive cases (61.5% vs 25.0%, p = 0.024), but it most likely results from more frequent PD-L1 expression in low-stage RMS. PD-1 expression on TAICs was detected in 7 cases and did not influence the prognosis.ConclusionsWe found that PD-L1 expression on TAICs, as detected with the use of 22C3 clone but not SP142 antibody, tends to be associated with low-stage RMS in children. PD-1 expression on TAICs in RMS is neither associated with distinct clinical course nor with clinicopathological features.     

Intrahepatic PD-1/PD-L1 Up-regulation Closely Correlates with Inflammation and Virus Replication in Patients with Chronic HBV Infection

Chronic hepatitis B was characterized by fluctuant immune response to infected hepatocytes resulting in hepatic inflammation and virus persistence. Recently, Programmed Death-1 (PD-1) and its ligand PD-L1 have been demonstrated to play an essential role in balancing antiviral immunity and inflammation in the livers of acute hepatitis B patients, significantly influencing disease outcome. PD-1 up-regulation in peripheral T cells is associated with immune dysfunction in chronic hepatitis B patients. However, the effect of PD-1/PD-L1 on hepatic damage and chronic infective status is still unknown in patients with chronic HBV infection. Here, we report up-regulation of PD-1 and PD-L1 in liver biopsies from 32 chronic HBV patients compared to 4 healthy donors. PD-1/PD-L1 up-regulation was significantly associated with hepatic inflammation and ALT elevation. Moreover, appropriate up-regulation but not overexpression of PD-L1 in the active phase of chronic hepatitis B as well as lower expression of PD-L1 in the inactive phase in liver residential antigen presenting cells (including Kupffer cells and sinusoidal endothelial cells) may contribute to viral inhibition. Our data suggest that the intrahepatic interaction of PD-1 and PD-L1 might play an important role in balancing the immune response to HBV and immune-mediated liver damage in chronic HBV infection.     

Treg suppressive activity involves estrogen-dependent expression of programmed death-1 (PD-1)

Estrogen [17-beta-estradiol (E2)] is a potent driver of the FoxP3+ regulatory T cell (Treg) compartment. Recently, Tregs were further characterized by intracellular expression of the negative co-stimulatory molecule, programmed death-1 (PD-1). To clarify the role of PD-1 versus FoxP3 in E2-enhanced Treg suppression, we evaluated both markers and functional suppression in wild-type, estrogen receptor knockout (ERKO) mice and PD-1 KO mice. We demonstrate that intracellular PD-1 expression is also E2 sensitive, since E2 treatment increased intracellular PD-1 levels in CD4+FoxP3+ cells, and PD-1 expression and Treg suppression were reduced in ERKO mice. Surprisingly, PD-1 KO mice retained normal levels of FoxP3 expression, but Tregs from these mice lacked functional suppression. However, E2 pre-treatment of PD-1 KO mice partially restored functional Treg suppression without enhancing FoxP3 expression. Thus, functional Treg suppression in immunized mice without E2 pre-treatment was more closely linked to PD-1 expression than to FoxP3 expression. However, although enhanced PD-1 expression was E2 dependent, functional suppression was still enhanced by E2 pre-treatment in the absence of PD-1. These data clearly demonstrate that E2 can affect multiple regulatory elements that influence Treg suppression, including both PD-1-dependent and PD-1-independent pathways.     

EGFR和PD-L1双靶向嵌合抗原受体构建及表达

目的:构建肿瘤相关抗原表皮生长因子受体特异性嵌合抗原受体(EGFR-CAR)和程序性死亡受体-配体1(PD-L1)抗体双修饰慢病毒载体表达系统。方法:人PD-L1-Fc蛋白免疫BALB/c小鼠,经细胞融合、亚克隆筛选高分泌PD-L1特异性抗体的稳定杂交瘤,酶联免疫吸附试验(ELISA)和Western blot检测抗体特异性,流式细胞术(FACS)鉴定对PD-1配受体封阻性能,Fortebio测定抗体亲和力,抗体全长测序,经保留鼠源CRD1、CRD2和CRD3人源化改造后构建单链抗体(single-chain variable fragment,scFv);人EGFR单克隆抗体杂交瘤系,经5′RACE技术扩增其轻链和重链可变区(VL和VH)基因,构建scFv,克隆至真核载体pcDNA3.1表达鉴定。基因合成EGFR-CAR(引入CD137协同信号胞内功能域)与PD-L1-scFv借助2A序列连接,克隆入慢病毒pLVX-EF1a-IRES-ZsGreen1表达载体,使用Lenti-X Packaging Single Shots (VSV-G)共同转染293T细胞,获得包装病毒,感染293V细胞,FACS测定CAR膜表达,ELISA检测CAR感染293V细胞培养上清中PD-L1-scFv表达情况,转染激活人外周血T细胞,验证CAR膜表达。结果:获得PD-L1抗体11E3,具备高度配受体封阻性能,经人源化改造后,亲和力稳定(2.67×10 -10 mol/L),EGFR-scFv获得有效表达。进一步构建了EGFR-CAR和PD-L1双修饰慢病毒分泌型CAR(CTC0537-1)及膜表达型CAR(CTC0537-2),其病毒感染293V细胞阳性率为10%。CTZ0431-1感染293V细胞后,细胞膜表面表达EGFR-scFv,检测培养上清存在PD-L1-ScFv;CTZ0431-2感染293V细胞后,细胞膜表面EGFR-scFv和PD-L1-scFv有效表达,双表达病毒感染活化T细胞的CAR表达率为39.3%。 结论:成功构建了EGFR-CAR和PD-L1-scFv双表达慢病毒载体,EGFR-CAR中度结合亲和力,此为EGFR靶向和PD-L1抗体双修饰CAR-T细胞的实体瘤治疗研究提供了关键工具。     

Clinical significance of programmed death ligand-1 (PD-L1) in colorectal serrated adenocarcinoma

Preliminary research results with antibody of the negative costimulatory molecule programmed cell death ligand-1 (PD-L1) suggested its expression on tumor cells associated with various tumor grade and postoperative prognosis. However, to date, there is no information of PD-L1 expression in colorectal serrated adenocarcinoma (SAC) and its clinical relevance. Therefore, the purpose of this study is to investigate the clinical significance of PD-L1 expression in a large cohort of patients with SAC. Here, we first retrospectively identified all SAC collected at our institution between August 2008 and May 2013. The expression levels of PD-L1 were examined by immunohistochemistry in 120 patients with SAC. We further evaluated the correlation between expression data and clinical parameters, including patient age, sex, tumor size, location, grade, primary tumor classification (pT), lymph node metastasis (pN), distant metastases (pM), and vascular invasion. Strong PD-L1 expression was detected in 25% of SAC. Higher expression of PD-L1 was significantly associated with pN (P=0.003) and pM (P=0.014). Survival analysis showed that patients with higher expression of PD-L1 had a poorer prognosis (P=0.045). However, multivariate regression analysis did not support PD-L1 as an independent prognostic factor (P=0.430). Our data suggest that PD-L1 may represent a new biomarker of metastasis and prognosis for patients with SAC, but as a target in the treatment of SAC is less certain.     

Programmed cell death 1 (PD-1) and its ligand PD-L1 are required for allograft tolerance

Programmed cell death-1 (PD-1, CD279) and its widely expressed, inducible ligand, PD-L1 (CD274), together dampen T cell activation, but whether they are essential for allograft tolerance is unknown. We show, using gene-deficient mice and blocking mAbs in wild-type mice, that costimulation blockade is ineffectual in PD-1(-/-) or PD-L1(-/-) allograft recipients, or in wild-type allograft recipients treated with anti-PD-1 or anti-PD-L1 mAb. Alloreactive PD-1(-/-) CD4 and CD8 T cells had enhanced proliferation and cytokine production compared to wild-type controls, and anergy could not be induced in PD-1-deficient CD4 T cells. We conclude that without inhibitory signals from PD-1 ligation, alloantigen-induced T cell proliferation and expansion cannot be regulated by costimulation blockade, and peripheral tolerance induction cannot occur.     

Chemopreventive agents induce programmed death-1-ligand 1 (PD-L1) surface expression in breast cancer cells and promote PD-L1-mediated T cell apoptosis

Chemotherapy has been widely used in cancer treatment. However, the prognosis of the cancer patients following chemotherapy has not been substantially improved. Alternative strategies such as immunotherapy and their combinations with chemotherapy are now being considered. Yet, the effects of chemotherapy on the immune responses of cancer cells are not clear. Cancer immunoresistance and immune escape are major obstacles in immunotherapy. In the present studies, we examined the effects of chemopreventive agents, paclitaxel, etoposide and 5-fluorouracil, on the surface expression of programmed death-1-ligand 1 (PD-L1), a negative regulator of T cell anti-tumor immunity. Interaction of PD-L1 on cancer cells with programmed death receptor 1 (PD-1) on T cells has been reported to inhibit the proliferation of tumor-reactive cytotoxic T cells and induce T cell apoptosis, which could be an important mechanism in the development of cancer immunoresistance. We demonstrated that those chemopreventive agents were able to induce PD-L1 surface expression in human breast cancer cells, which then promoted PD-L1-mediated T cell apoptosis. Our studies reveal a potential link between chemotherapy and cancer immunoresistance.     

Clinicopathological value of programmed cell death 1 (PD-1) and programmed cell death ligand 1 (PD-L1) expression in synovium of patients with rheumatoid arthritis

The etiology of rheumatoid arthritis (RA) is thought to involve dysfunction of the programmed cell death 1/programmed cell death ligand 1 (PD-1/PD-L1) pathway; PD-1 negatively regulates autoimmunity by interacting with its ligand, PD-L1. We therefore investigated PD-1/PD-L1 expression in synovial tissue of patients with RA. We immunohistochemically stained synovial specimens from 51 patients with RA and assessed the association between PD-1/PD-L1 expression and rheumatoid factor (RF), the total count of infiltrating T cells, C-reactive protein (CRP), and Krenn’s synovitis score. PD-1 expression on infiltrating lymphocytes was detected in 34/51 RA cases (66.7%), while PD-1 expression was very mildly correlated only with the number of total infiltrating T cells (R²?=?0.1011, P?=?0.0230). On the other hand, PD-L1 expression on synovial lining cells was observed in 37/51 RA cases (72.5%). Furthermore, a higher PD-L1 expression was significantly associated with RF positive state (P?=?0.0454), and the correlations between PD-L1 expression and the number of infiltrating T cells (R²?=?0.5571, P?<?0.0001), CRP (R²?=?0.4060, P?<?0.0001), and Krenn’s synovitis score (R²?=?0.7785, P?<?0.0001) were confirmed. PD-1 was expressed on infiltrating lymphocytes, while PD-L1 was expressed on synovial lining cells; the expression of PD-L1 on synovial lining cells was significantly correlated with the active state of the disease. These data suggest that PD-1/PD-L1 pathway may have an important role in the pathogenesis of RA.