PD-1胞外段cDNA在真核细胞的表达与其功能鉴定

PD-L/PD-1是参与肿瘤免疫逃避的一条抑制性信号途径。为了用可溶性的PD 1受体阻断PD-L/PD-1的相互作用 ,将小鼠PD-1胞外段 (aa1-aa167)作为独立可溶性分子进行了真核表达 ,并对其功能进行鉴定。构建了编码小鼠PD-1胞外段cDNA(sPD 1)的真核质粒表达载体pPD-1A和编码sPD-1-GFP重组蛋白的真核质粒表达载体pPD-1B ;细胞转染实验表明其表达产物主要是分泌到细胞外的可溶性产物 (sPD-1) ,流式细胞仪检测表明sPD-1可有效结合PD-1配体 ;肿瘤细胞杀伤实验表明 ,sPD-1作用于肿瘤细胞或在脾淋巴细胞激活过程中作用于淋巴细胞 ,均可增强Hsp70-H22抗原肽复合物激活的脾淋巴细胞杀伤肿瘤细胞的作用。sPD-1真核表达载体的构建为在肿瘤局部表达抑制性共刺激分子的可溶性受体 ,拮抗肿瘤微环境中负调节因素对T细胞的抑制作用 ,增强机体的抗肿瘤能力 ,提供了一种新的肿瘤基因治疗手段     

Identification and characterization of bovine programmed death‐ligand 2

Previous reports from this group have indicated that the immunoinhibitory programmed death (PD)‐1 receptor and its ligand, PD‐L1, are involved in the mechanism of immune evasion of bovine chronic infection. However, no functional analysis of bovine PD‐L2 in cattle has been reported. Thus, in this study, the molecular function of bovine PD‐L2 was analyzed in vitro. Recombinant PD‐L2 (PD‐L2‐Ig), which comprises an extracellular domain of bovine PD‐L2 fused to the Fc portion of rabbit IgG1, was prepared based on the cloned cDNA sequence for bovine PD‐L2. Bovine PD‐L2‐Ig bound to bovine PD‐1‐expressing cells and addition of soluble bovine PD‐1‐Ig clearly inhibited the binding of PD‐L2‐Ig to membrane PD‐1 in a dose‐dependent manner. Cell proliferation and IFN‐γ production were significantly enhanced in the presence of PD‐L2‐Ig in peripheral blood mononuclear cells (PBMCs) from cattle. Moreover, PD‐L2‐Ig significantly enhanced IFN‐γ production from virus envelope peptides‐stimulated PBMCs derived from bovine leukemia virus‐infected cattle. Interestingly, PD‐L2‐Ig‐induced IFN‐γ production was further enhanced by treatment with anti‐bovine PD‐1 antibody. These data suggest potential applications of bovine PD‐L2‐Ig as a therapy for bovine diseases.     

GSTM1 and mEPHX polymorphisms in Parkinson's disease and age of onset

Both environmental and genetic factors are involved in the development of PD and biotransformation of exogenous and endogenous compounds and may play a role in inter-individual susceptibility. Therefore, we investigated the presence of null genotypes of GSTM1, GSTT1, and two polymorphisms of mEPHX in subjects with Parkinson's disease and in a reference population. The study included 35 male PD patients and a male control group including 283 subjects. Homozygosity of the histidine (H) 113 isoform of mEPHX was significantly increased in PD patients (odds ratio = 3.8 CI 95% 1. 2-11.8) and analysis of allele frequencies displayed an increased frequency of the H-allele among PD patients (odds ratio = 1.9 CI 95% 1.1-3.3). However, a significantly elevated median age for the onset of PD was found among GSTM1 gene carriers (median age = 68 years) compared to PD patients being GSTM1 null genotypes (median age = 57 years). Our observations suggest that (H) 113 isoform of mEPHX, which has been suggested as a low activity isoform, is overrepresented in PD patients and that inherited carriers of the GSTM1 gene postpone the onset of PD. These detoxification pathways may represent important protective mechanisms against reactive intermediates modifying the susceptibility and onset of PD.     

Destructive cellular paths underlying familial and sporadic Parkinson disease converge on mitophagy

The knowledge gap separating the molecular and cellular underpinnings of Parkinson disease (PD) and its pathology hinders treatment innovation. Adding to this difficulty is the lack of a reliable biomarker for PD. Our previous studies identify a link of 2 PD proteins, PINK1/PRKN Parkin to a mitochondrial motor adaptor RHOT1/Miro-1, which mediates mitochondrial motility and mitophagy. Here we review our recent paper showing that a third PD protein, LRRK2, also targets RHOT1 and regulates mitophagy, and pathogenic LRRK2 disrupts this function. Notably, we discover impairments in RHOT1 and mitophagy in sporadic PD patients with no known genetic backgrounds, pointing to RHOT1-mediated mitophagy as a convergent pathway in PD. This novelty opens new doors in PD research toward RHOT1-based therapy and biomarker development.     

PD‐1/PD‐L1 expression on CD4+ T cells and myeloid DCs correlates with the immune pathogenesis of atrial fibrillation

Although immuno‐inflammatory response contributes to pathogenesis of AF, molecular and cellular mechanism in this process remains poorly understood. Recently, increasing evidence suggests that Programmed death‐1 (PD‐1)/PD‐1 ligand (PD‐L) pathway may be a potential pathway participating in AF pathogenesis. In this study, we detected the PD‐1 and PD‐L1, 2 expression on peripheral blood function cells by flow cytometry in 91 atrial fibrillation (AF) patients and 35 healthy volunteers. The expression of PD‐1 on CD⁴⁺ T cells and PD‐L1 on myeloid dendritic cells (mDCs) in AF patients is significantly down‐regulated compared with healthy volunteers. In addition, the extent of PD‐1/PD‐L1 down‐regulation is closely related with AF burden. More importantly, Allogeneic mixed leukocyte reactions (MLR) shows that the mDCs PD‐L1 down‐regulation is associated with increased T cell (CD⁴⁺ and CD⁸⁺) proliferation, increased type 1 effector cytokines (IL‐2 and IFN‐γ) secretion, and decreased type 2 effector cytokine (IL‐10) secretion. Then, PD‐L1 up‐regulation by the stimulation of IFN‐α can significantly convert this representation. Collectively, our report suggest that T(CD⁴⁺)/mDCs‐associated PD‐1/PD‐L1 pathway plays a key role in AF immune regulation. PD‐1/PD‐L1 down‐regulation in AF patients promotes T cells function and may contribute to AF pathogenesis.     

Developmental Expression of HNK-1-Reactive Antigens in the Rat Cerebellum and Localization of Sulfoglucuronyl Glycolipids in Molecular Layer and Deep Cerebellar Nuclei

Monoclonal antibody HNK-1-reactive carbohydrate epitope is expressed on proteins, proteoglycans, and sulfoglucuronyl glycolipids (SGGLs). The developmental expression of these HNK-1-reactive antigens was studied in rat cerebellum. The expression of sulfoglucuronyl lacto-N-neotetraosylceramide (SGGL-1) was biphasic with an initial maximum at postnatal day one (PD 1), followed by a second rise in the level at PD 20. The level of sulfoglucuronyl lacto-N-norhexaosyl ceramide (SGGL-2) in cerebellum was low until PD 15 and then increased to a plateau at PD 20. The levels of SGGLs increased during postnatal development of the cerebellum, contrary to their diminishing expression in the cerebral cortex. The expression of HNK-1-reactive glycoproteins decreased with development of the rat cerebellum from PD 1. Several HNK-1-reactive glycoproteins with apparent molecular masses between 150 and 325 kDa were visualized between PD 1 and PD 10. However, beyond PD 10, only two HNK-1-reactive bands at 160 and 180 kDa remained. The latter appeared to be neural cell adhesion molecule, N-CAM-180. A diffuse HNK-1-reactive band seen at the top of polyacrylamide electrophoretic gels was due mostly to proteoglycans. This band increased in its reactivity to HNK-1 between PD 15 and PD 25 and then decreased in the adult cerebellum. The lipid antigens were shown by two complementary methodologies to be localized primarily in the molecular layer and deep cerebellar nuclei as opposed to the granular layer and white matter. A fixation procedure which eliminates HNK-1-reactive epitope on glycoproteins and proteoglycans, but does not affect glycolipids, allowed selective immunoreactivity in the molecular layer and deep cerebellar nuclei.(ABSTRACT TRUNCATED AT 250 WORDS)     

Protectin D1 is generated in asthma and dampens airway inflammation and hyperresponsiveness

Protectins are newly identified natural chemical mediators that counter leukocyte activation to promote resolution of inflammation. In this study, we provide the first evidence for protectin D1 (PD1, 10R,17S-dihydroxy-docosa-4Z,7Z,11E,13E,15Z,19Z-hexaenoic acid) formation from docosahexaenoic acid in human asthma in vivo and PD1 counterregulatory actions in allergic airway inflammation. PD1 and 17S-hydroxy-docosahexaenoic acid were present in exhaled breath condensates from healthy subjects. Of interest, levels of PD1 were significantly lower in exhaled breath condensates from subjects with asthma exacerbations. PD1 was also present in extracts of murine lungs from both control animals and those sensitized and aerosol challenged with allergen. When PD1 was administered before aeroallergen challenge, airway eosinophil and T lymphocyte recruitment were decreased, as were airway mucus, levels of specific proinflammatory mediators, including IL-13, cysteinyl leukotrienes, and PGD(2), and airway hyperresponsiveness to inhaled methacholine. Of interest, PD1 treatment after aeroallergen challenge markedly accelerated the resolution of airway inflammation. Together, these findings provide evidence for endogenous PD1 as a pivotal counterregulatory signal in allergic airway inflammation and point to new therapeutic strategies for modulating inflammation in asthmatic lung.     

An optimized protocol for the in vitro generation and functional analysis of human PD1/PD-L1 signal

Programmed cell death-1 (PD1) is an inhibitory receptor expressed on the activated T and B cells. Binding of PD1 to its ligands, PD-L1 and PD-L2 has led to deliver an inhibitory signal into the activated T cells. Recently, blocking PD1/PD-L1 pathway has emerged as a new treatment paradigm across a broad spectrum of malignancies. Remarkable clinical responses of monoclonal antibodies specific for PD-1 or its ligands in patients with many different types of cancer, attracted several pharmaceutical companies and researchers to investigate the agents that block PD1/PD-L1 signal. The safety and efficacy of the agents are needed to examine in the preclinical studies. In this study, we optimized a facile and cost-effective protocol for in vitro generation and functional analysis of human PD1/PD-L1 pathway. Activation of CD8?+?CD279?+?T cell was performed by anti-CD3 and D28 antibodies and the recombinant PD-L1 was used for inactivation of T cells through PD1/PD-L1 pathway. In this protocol, T-cell cytokine production (IL-2 and IFN-γ) and proliferation assay confirmed that a measurable PD1/PD-L1 signal was generated. We expected that in vitro PD1/PD-L1 signal that has been optimized in this study will serve as a valuable protocol for preclinical studies involving PD1/PD-L1 pathway.     

Oxidative modifications and down-regulation of ubiquitin carboxyl-terminal hydrolase L1 associated with idiopathic Parkinson's and Alzheimer's diseases

Alzheimer's disease (AD) and Parkinson's disease (PD) are the two most common neurodegenerative diseases that occur either in relatively rare, familial forms or in common, sporadic forms. The genetic defects underlying several monogenic familial forms of AD and PD have recently been identified, however, the causes of other AD and PD cases, particularly sporadic cases, remain unclear. To gain insights into the pathogenic mechanisms involved in AD and PD, we used a proteomic approach to identify proteins with altered expression levels and/or oxidative modifications in idiopathic AD and PD brains. Here, we report that the protein level of ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1), a neuronal de-ubiquitinating enzyme whose mutation has been linked to an early-onset familial PD, is down-regulated in idiopathic PD as well as AD brains. By using a combination of two-dimensional gel electrophoresis and mass spectrometry, we have identified three human brain UCH-L1 isoforms, a full-length form and two amino-terminally truncated forms. Our proteomic analyses reveal that the full-length UCH-L1 is a major target of oxidative damage in AD and PD brains, which is extensively modified by carbonyl formation, methionine oxidation, and cysteine oxidation. Furthermore, immunohistochemical studies show that prominent UCH-L1 immunostaining is associated with neurofibrillary tangles and that the level of soluble UCH-L1 protein is inversely proportional to the number of tangles in AD brains. Together, these results provide evidence supporting a direct link between oxidative damage to the neuronal ubiquitination/de-ubiquitination machinery and the pathogenesis of sporadic AD and PD.     

A plasmodesmata-localized protein mediates crosstalk between cell-to-cell communication and innate immunity in Arabidopsis

Plasmodesmata (PD) are thought to play a fundamental role in almost every aspect of plant life, including normal growth, physiology, and developmental responses. However, how specific signaling pathways integrate PD-mediated cell-to-cell communication is not well understood. Here, we present experimental evidence showing that the Arabidopsis thaliana plasmodesmata-located protein 5 (PDLP5; also known as HOPW1-1-INDUCED GENE1) mediates crosstalk between PD regulation and salicylic acid-dependent defense responses. PDLP5 was found to localize at the central region of PD channels and associate with PD pit fields, acting as an inhibitor to PD trafficking, potentially through its capacity to modulate PD callose deposition. As a regulator of PD, PDLP5 was also essential for conferring enhanced innate immunity against bacterial pathogens in a salicylic acid-dependent manner. Based on these findings, a model is proposed illustrating that the regulation of PD closure mediated by PDLP5 constitutes a crucial part of coordinated control of cell-to-cell communication and defense signaling.     

Human pressure perception values for constant and moving one- and two-point discrimination.

Despite the need to evaluate sensibility for accurate diagnosis and the need to record the degree of sensation achieved in the postoperative period, the clinician has been without the ability to measure human pressure perception accurately. Traditionally, the Semmes-Weinstein monofilaments were used to measure the static one-point discrimination threshold. A new sensory testing instrument, the Pressure-Specifying Sensory Device, was used to obtain normative data from the index and little finger of the dominant hand in 35 people ranging in age from 16 to 83 with no known neurologic impairment. Pressure perceptions for static one- and two-point discrimination (s1PD, s2PD) and moving one- and two-point discrimination (m1PD, m2PD) were recorded. The mean values (+/- SD) were 0.13 +/- 0.06, 0.24 +/- 0.12, 0.22 +/- 0.10, and 0.26 +/- 0.13 gm/mm2 for s1PD, s2PD, m1PD, and m2PD, respectively, on the index finger and 0.07 +/- 0.05, 0.16 +/- 0.12, 0.17 +/- 0.07, and 0.21 +/- 0.14 gm/mm2 for s1PD, s2PD, m1PD, and m2PD, respectively, for the little finger. The little finger was significantly more sensitive than the index finger (p less than 0.001). There was no significant change in pressure perception with increasing age.     

Lower vascular tone and larger plasma volume in Parkinson's disease with orthostatic hypotension

The pathophysiology of orthostatic hypotension in Parkinson's disease (PD) is incompletely understood. The primary focus has thus far been on failure of the baroreflex, a central mediated vasoconstrictor mechanism. Here, we test the role of two other possible factors: 1) a reduced peripheral vasoconstriction (which may contribute because PD includes a generalized sympathetic denervation); and 2) an inadequate plasma volume (which may explain why plasma volume expansion can manage orthostatic hypotension in PD). We included 11 PD patients with orthostatic hypotension (PD + OH), 14 PD patients without orthostatic hypotension (PD - OH), and 15 age-matched healthy controls. Leg blood flow was examined using duplex ultrasound during 60° head-up tilt. Leg vascular resistance was calculated as the arterial-venous pressure gradient divided by blood flow. In a subset of 9 PD + OH, 9 PD - OH, and 8 controls, plasma volume was determined by indicator dilution method with radiolabeled albumin ((125)I-HSA). The basal leg vascular resistance was significantly lower in PD + OH (0.7 ± 0.3 mmHg·ml(-1)·min) compared with PD - OH (1.3 ± 0.6 mmHg·ml(-1)·min, P < 0.01) and controls (1.3 ± 0.5 mmHg·ml(-1)·min, P < 0.01). Leg vascular resistance increased significantly during 60° head-up tilt with no significant difference between the groups. Plasma volume was significantly larger in PD + OH (3,869 ± 265 ml) compared with PD - OH (3,123 ± 377 ml, P < 0.01) and controls (3,204 ± 537 ml, P < 0.01). These results indicate that PD + OH have a lower basal leg vascular resistance in combination with a larger plasma volume compared with PD - OH and controls. Despite the increase in leg vascular resistance during 60° head-up tilt, PD + OH are unable to maintain their blood pressure.     

Progress in the pathogenesis and genetics of Parkinson's disease

Recent progresses in the pathogenesis of sporadic Parkinson's disease (PD) and genetics of familial PD are reviewed. There are common molecular events between sporadic and familial PD, particularly between sporadic PD and PARK1-linked PD due to alpha-synuclein (SNCA) mutations. In sporadic form, interaction of genetic predisposition and environmental factors is probably a primary event inducing mitochondrial dysfunction and oxidative damage resulting in oligomer and aggregate formations of alpha-synuclein. In PARK1-linked PD, mutant alpha-synuclein proteins initiate the disease process as they have increased tendency for self-aggregation. As highly phosphorylated aggregated proteins are deposited in nigral neurons in PD, dysfunctions of proteolytic systems, i.e. the ubiquitin-proteasome system and autophagy-lysosomal pathway, seem to be contributing to the final neurodegenerative process. Studies on the molecular mechanisms of nigral neuronal death in familial forms of PD will contribute further on the understanding of the pathogenesis of sporadic PD.     

Expression and biochemical characterization of beta-primeverosidase and application of beta-primeverosylamidine to affinity purification

Beta-primeverosidase (PD) is a family 1 glycosidase catalyzing the hydrolysis of beta-primeverosides (6-O-beta-D-xylopyranosyl-beta-D-glucopyranosides) to release a disaccharide primeverose. To investigate how PD recognizes the disaccharide moiety of beta-primeverosides, the recombinant PD was expressed by a baculovirus-insect cell system. The recombinant PD was secreted from High Five cells and was properly modified with N-glycosylation and correct cleavage at the N-terminal signal peptide. The recombinant PD exhibited high substrate specificity to beta-primeverosides in terms of the glycone moiety, consistently with the substrate specificity of native PD from Camellia sinensis. Next, beta-glycosylamidines were synthesized as substrate analog inhibitors. Beta-primeverosylamidine strongly inhibited PD activity, but beta-glucosylamidine did not. Hence beta-primeverosylamidine is an ideal chemical tool for probing disaccharide recognition in the active site of PD. An affinity adsorbent for PD was prepared using beta-primeverosylamidine as a ligand. Affinity chromatography gave large amounts of PD with high purity, permitting crystallographic study.     

Chronic prenatal ethanol exposure-induced decrease of guinea pig hippocampal CA1 pyramidal cell and cerebellar Purkinje cell density

The brain is a key target of ethanol teratogenicity, in which ethanol can produce neurodegeneration in selected areas, including the hippocampus and cerebellum. The research objective was to test the hypothesis that chronic prenatal ethanol exposure, via maternal ethanol administration, produces differential time course of decreased linear density of hippocampal CA1 pyramidal cells and cerebellar Purkinje cells. Timed pregnant guinea pigs received chronic oral administration of ethanol, isocaloric-sucrose/pair-feeding, or water throughout gestation (term, about gestational day (GD) 68), and the offspring were studied at GD 62 (near-term fetus), postnatal day (PD) 1 (neonate), PD 5, and PD 12 (early postnatal life). Ethanol treatment, compared with isocaloric-sucrose/pair-feeding and water treatments, decreased brain, hippocampal, and cerebellar weights at GD 62, PD 1, PD 5, and PD 12. Hippocampal CA1 pyramidal cell linear density and cerebellar Purkinje cell linear density were unaffected at GD 62. Ethanol treatment produced 25, 30, and 30% decreases in linear density of hippocampal CA1 pyramidal cells at PD 1, PD 5, and PD 12, respectively, and a 30% decrease in linear density of cerebellar Purkinje cells at PD 12 only. At PD 5, Purkinje cell profile linear density remained unaffected; however, ethanol treatment appeared to increase linear density of apoptotic Purkinje cell nuclei, as determined by a modified TUNEL method. The data demonstrate that chronic prenatal ethanol exposure produces apparent differential time course of decreased linear density of hippocampal CA1 pyramidal cells and cerebellar Purkinje cells in the developing guinea pig.     

Insights into links between familial and sporadic Parkinson's disease: physical relationship between UCH-L1 variants and chaperone-mediated autophagy

Ubiquitin C-terminal hydrolase L1 (UCH-L1) is expressed abundantly in neurons and has been reported to be a major target of oxidative/carbonyl damage associated with sporadic Parkinson's disease (PD). The I93M mutation in UCH-L1 is also associated with familial PD. We recently reported that UCH-L1 physically interacts with LAMP-2A, the lysosomal receptor for chaperone-mediated autophagy (CMA), and Hsc70 and Hsp90, both of which can function as components of the CMA pathway. We found that the levels of these interactions were aberrantly increased by the I93M mutation, and that expression of I93M UCH-L1 in cells induced the CMA inhibition-associated increase in the amount of alpha-synuclein, a risk factor for PD. The interactions of UCH-L1 with LAMP-2A, Hsc70 and Hsp90 were also abnormally enhanced by carbonyl modification of UCH-L1. We propose that aberrant interactions of UCH-L1 variants with CMA machinery, at least partly, underlie the pathogenesis of I93M UCH-L1-associated PD, and possibly of sporadic PD. Our findings may provide novel insights into the links between familial and sporadic PD.     

Epidemiological, Clinical, and Molecular Study of a Cohort of Italian Parkinson Disease Patients: Association with Glutathione-S-Transferase and DNA Repair Gene Polymorphisms

Parkinson’s disease (PD) is one of the most common neurodegenerative disorders whose etiology is multifactorial including both hereditary and environmental factors. Currently, pathogenic mutations in at least five genes have been implicated in familial PD generally accounting for less than 10 % of all PD cases in most populations. It has been suggested that polymorphisms in other genes such as those encoding enzymes involved in oxidative metabolism and detoxification could be involved in predisposition to PD since oxidative stress in dopaminergic neurons is thought to be of central importance in the pathogenesis of the disease. The aim of our work was to study the association of genetic polymorphisms in genes involved in oxidative metabolism and detoxification mechanism, namely GSTM1, GSTT1, GSTP1, and those involved in DNA damage repair, OGG1 and XRCC1, in an Italian cohort of sporadic PD patients. We did not detect any association between GSTT1 and GTTM1 null polymorphisms and PD, whereas the 104GSTP1 polymorphism was associated with PD in male patients but not in females. Furthermore, we detected a protective effect of wild type genotype of XRCC1 in women.     

Characterization of the Xylella fastidiosa PD1311 gene mutant and its suppression of Pierce's disease on grapevines

Xylella fastidiosa causes Pierce's disease (PD) on grapevines, leading to significant economic losses in grape and wine production. To further our understanding of X. fastidiosa virulence on grapevines, we examined the PD1311 gene, which encodes a putative acyl‐coenzyme A (acyl‐CoA) synthetase, and is highly conserved across Xylella species. It was determined that PD1311 is required for virulence, as the deletion mutant, ΔPD1311, was unable to cause disease on grapevines. The ΔPD1311 strain was impaired in behaviours known to be associated with PD development, including motility, aggregation and biofilm formation. ΔPD1311 also expressed enhanced sensitivity to H₂O₂ and polymyxin B, and showed reduced survival in grapevine sap, when compared with wild‐type X. fastidiosa Temecula 1 (TM1). Following inoculation, ΔPD1311 could not be detected in grape shoots, which may be related to its altered growth and sensitivity phenotypes. Inoculation with ΔPD1311 2 weeks prior to TM1 prevented the development of PD in a significant fraction of vines and eliminated detectable levels of TM1. In contrast, vines inoculated simultaneously with TM1 and ΔPD1311 developed disease at the same level as TM1 alone. In these vines, TM1 populations were distributed similarly to populations in TM1‐only inoculated plants. These findings suggest that, through an indirect mechanism, pretreatment of vines with ΔPD1311 suppresses pathogen population and disease.     

PD‐L1 expression in melanoma shows marked heterogeneity within and between patients: implications for anti‐PD‐1/PD‐L1 clinical trials

This study evaluated the expression of PD‐L1 in immunotherapy‐naïve metastatic melanoma patients to determine longitudinal intrapatient concordance and correlate PD‐L1 status with clinicopathologic characteristics and outcome. PD‐L1 expression was assessed by immunohistochemistry in 58 patients (43 primary tumors, 96 metastases). Seventy‐two percent of patients had at least one specimen expressing PD‐L1 in ≥1% of tumor cells. Median positive tumor cell count overall was low (8% in nonzero specimens). PD‐L1 expression was frequently discordant between primary tumors and metastases and between intrapatient metastases, such that 23/46 longitudinal patient specimens were discordant. PD‐L1 was associated with higher TIL grade but not with other known prognostic features. There was a positive univariate association between PD‐L1 expression in locoregional metastases and melanoma‐specific survival, but the effect was not observed for primary melanoma. In locoregional lymph node metastasis, PD‐L1+/TIL+ patients had the best outcome, and PD‐L1+/TIL? patients had poor outcome.