Non-classical Human Leukocyte Antigen Research Articles

The Prognostic Value of Immune Factors in the Tumor Microenvironment of Penile Squamous Cell Carcinoma.

The host’s immune system plays a pivotal role in many tumor types, including squamous cell carcinomas (SCCs). We aim to identify immunological prognosticators for lymph node metastases (LNM) and disease-specific survival (DSS) in penile SCC. For this retrospective observational cohort study, penile SCC patients (n = 213) treated in the Netherlands Cancer Institute, were selected if sufficient formalin-fixed, paraffin-embedded tumor material was available. Analysis included previously described high-risk human papilloma virus (hrHPV) status, immunohistochemical scores for classical and non-classical human leukocyte antigen (HLA) class I, programmed death ligand-1 (PD-L1) expression, and novel data on tumor-infiltrating macrophages and cytotoxic an regulatory T-cells. Clinicopathological characteristics and extended follow-up were also included. Regression analyses investigated relationships of the immune parameters with LNM and DSS. In the total cohort, diffuse PD-L1 tumor-cell expression, CD163+ macrophage infiltration, non-classical HLA class I upregulation, and low stromal CD8+ T-cell infiltration were all associated with LNM. In the multivariable model, only tumor PD-L1 expression remained a significant predictor for LNM (odds ratio (OR) 2.8, p = 0.05). hrHPV negativity and diffuse PD-L1 tumor-cell expression were significantly associated with poor DSS and remained so upon correction for clinical parameters [hazard ratio (HR) 9.7, p < 0.01 and HR 2.8, p = 0.03]. The only immune factor with different expression in HPV+ and HPV− tumors was PD-L1, with higher PD-L1 expression in the latter (p = 0.03). In the HPV− cohort (n = 158), LNM were associated with diffuse PD-L1 tumor-cell expression, high intratumoral CD163+ macrophage infiltration, and low number of stromal CD8+ T-cells. The first two parameters were also linked to DSS. In the multivariable regression model, diffuse PD-L1 expression remained significantly unfavorable for DSS (HR 5.0, p < 0.01). These results emphasize the complexity of the tumor microenvironment in penile cancer and point toward several possible immunotherapy targets. Here described immune factors can aid risk-stratification and should be evaluated in clinical immunotherapy studies to ultimately lead to patient tailored treatment.

Prognostic Value of HLA-G in Malignant Liver and Pancreas Lesions.

Human leukocyte antigen (HLA)-G is a nonclassical HLA class I molecule with modulatory effects on NK and T cells. Because HLA-G expression is frequently detected in different solid tumors, it may be involved in tumor immune evasion. This study was designed to elucidate the prognostic value of HLA-G in hepatocellular carcinoma (HCC) and pancreatic adenocarcinoma (PADC). The influence of hepatitis B virus (HBV) infection on HLA-G expression was also evaluated in patients with HCC. HLA-G expression was investigated in tumor tissues from patients with HCC (n=74) or PADC (n=42) with immunohistochemical techniques. The presence of HBV genome was also examined in HCC tumor tissues by PCR. HLA-G expression was detected in 66% of PADC and in 31% of HCC samples. In contrast to HCC, HLA-G overexpression was associated with advanced stages and grades in PADC. HBV genome was detected in 31% of HCC samples but we found no correlation between HLA-G expression and the presence of HBV genome in these tumors. Our findings showed that HLA-G overexpression in tumor tissue correlated with poor prognosis in PADC. HLA-G expression is apparently affected by the patient's genetic background and other epigenetic factors rather than by HBV infection.

AB029. Physiological expression of HLA-G and pregnancy

: Human leukocyte antigen (HLA)-G is a nonclassical major histocompatibility complex (MHC) class I molecule with immune-modulatory properties. Such molecule predominantly possesses tolerogenic and anti-inflammatory functions. Until now, it has been verified the existence of seven isoforms of HLA-G, being that four are membrane-bound isoforms and the other three are soluble isoforms. During physiological pregnancy, it should be noted that HLA-G is expressed in high levels by extravillous cytotrophoblasts. It has been proposed that in the course of pregnancy period HLA-G induces fetus immune tolerance through direct binding to the inhibitory receptors immunoglobulin-like transcript (ILT)-2 present on lymphoid and myelomonocytic cells, and ILT-4 expressed by dendritic cells, macrophages and monocytes. In addition, in this context, the killer cell immunoglobulin-like receptor expressed by natural killer (NK) cells is also an HLA-G-specific receptor. Therefore, HLA-G induces the maintenance of maternal-fetal tolerance at different stages of the immune response, such as in differentiation, proliferation, cytolysis and cytokine secretion, given that it can directly interact with different immune cell subpopulations. Furthermore, by indirect mechanisms HLA-G can express the nonclassical HLA class I molecule HLA-E, which directly binds peptides derived from HLA-G. Consequently, HLA-E can interact with the inhibitory receptor CD94/NKG2A (present on NK cells and T lymphocytes) resulting in inhibition of cytolysis. However, despite this some controversies can be found in the literature about this subject. We expect that the original results of our ongoing research can provide new and relevant explanations concerning the role of HLA-G during pregnancy. We aim to publish them soon in a scientific journal.

Structural Analysis and Epitope Prediction of MHC Class-1-Chain Related Protein-A for Cancer Vaccine Development

Major histocompatibility complex class 1 chain-related gene sequence A is a polymorphic gene found at about 46.6 kb centromeric to HLA-B. It encodes a transmembrane protein, which is a non-classical human leukocyte antigen whose expression is normally induced by stress conditions like cancer and viral infections. The expression of MIC-A leads to the activation of NKG2D receptors of natural killer and T cells, leading to the generation of innate immune response that can easily eliminate/cleanse tumour cells and other cells that express the protein. Several bioinformatics and immunoinformatics tools were used to analyse the sequence and structure of the MIC-A protein. These tools were used in building and evaluating modelled structure of MIC-A, and to predict several antigenic determinant sites on the protein. The MIC-A protein structure generated an average antigenic propensity of 1.0289. Additionally, the hydrophilic regions on the surface of the MIC-A protein where antibodies can be attached were revealed. A total of fourteen antigenic epitopes were predicted, with six found in the transmembrane protein topology, and are predicted to play a role in the development of vaccines that can reactivate the functionalities of the MIC-A protein on the surface of cancer cells in order to elicit a desired immune response.

Natural Killer Cell Interactions with Classical and Non-Classical Human Leukocyte Antigen Class I in HIV-1 Infection.

Natural killer (NK) cells are effector lymphocytes of the innate immune system that are able to mount a multifaceted antiviral response within hours following infection. This is achieved through an array of cell surface receptors surveilling host cells for alterations in human leukocyte antigen class I (HLA-I) expression and other ligands as signs of viral infection, malignant transformation, and cellular stress. This interaction between HLA-I ligands and NK-cell receptor is not only important for recognition of diseased cells but also mediates tuning of NK-cell-effector functions. HIV-1 alters the expression of HLA-I ligands on infected cells, rendering them susceptible to NK cell-mediated killing. However, over the past years, various HIV-1 evasion strategies have been discovered to target NK-cell-receptor ligands and allow the virus to escape from NK cell-mediated immunity. While studies have been mainly focusing on the role of polymorphic HLA-A, -B, and -C molecules, less is known about how HIV-1 affects the more conserved, non-classical HLA-I molecules HLA-E, -G, and -F. In this review, we will focus on the recent progress in understanding the role of non-classical HLA-I ligands in NK cell-mediated recognition of HIV-1-infected cells.

Human Leukocyte Antigen (HLA) and Immune Regulation: How Do Classical and Non-Classical HLA Alleles Modulate Immune Response to Human Immunodeficiency Virus and Hepatitis C Virus Infections?

The genetic factors associated with susceptibility or resistance to viral infections are likely to involve a sophisticated array of immune response. These genetic elements may modulate other biological factors that account for significant influence on the gene expression and/or protein function in the host. Among them, the role of the major histocompatibility complex in viral pathogenesis in particular human immunodeficiency virus (HIV) and hepatitis C virus (HCV), is very well documented. We, recently, added a novel insight into the field by identifying the molecular mechanism associated with the protective role of human leukocyte antigen (HLA)-B27/B57 CD8+ T cells in the context of HIV-1 infection and why these alleles act as a double-edged sword protecting against viral infections but predisposing the host to autoimmune diseases. The focus of this review will be reexamining the role of classical and non-classical HLA alleles, including class Ia (HLA-A, -B, -C), class Ib (HLA-E, -F, -G, -H), and class II (HLA-DR, -DQ, -DM, and -DP) in immune regulation and viral pathogenesis (e.g., HIV and HCV). To our knowledge, this is the very first review of its kind to comprehensively analyze the role of these molecules in immune regulation associated with chronic viral infections.

Immunohistochemical investigations on the expression of programmed cell death ligand 1, human leukocyte antigens G and E, and granzyme B in intraoral mucoepidermoid carcinoma

ObjectiveTo identify the expression of nonclassical human leukocyte antigen G and E (HLA-G and -E), programmed cell death ligand-1 (PD-L1) and granzyme B (GB) in intraoral mucoepidermoid carcinomas (MECs), and to assess whether such expressions are related to metastasis, survival, staging, tumor grade and number of GB-positive cells. DesignFor this cross-sectional study, samples of MEC (n=30) were selected and classified as low-grade (LG), intermediate-grade (IG) or high-grade (HG), according to the WHO grading system. HLA-G, -E and PD-L1 were identified by immunohistochemistry and quantified as the proportion of positive neoplastic cells. The density of GB+ cells was also evaluated. The Kruskal-Wallis test was used with a 5% significance level. ResultsExpressions of HLA-G, -E and PD-L1 were identified in the majority of epidermoid, intermediate and clear cells, but not in the mucous cells of the MECs. The quantitative analysis of the total percentage of positive neoplastic cells showed overexpression of this set of proteins in all MEC samples. The expression of these proteins and histological grading were positively correlated [HLA-G (LG=79% positive cells, IG=96%, HG=99%; p=0.0004), HLA-E (LG=70%, IG=96%, HG=99%; p<0.0001) and PD-L1 (LG=34%, IG=79%, HG=80%; p=0.01)]. No relationship was observed between the immunosuppressive proteins and other clinicopathological parameters. Low GB density was found in all MEC samples. ConclusionsThe augmented expression of HLA-G, -E and PD-L1 in the intraoral MEC might suggest a role of these molecules in the scape of neoplastic cells from immunosurveillance.

From genetic single candidate gene studies to complex genomics of GvHD

Graft-versus-host disease (GvHD) is a serious complication affecting the recipients of allogeneic haematopoietic stem cells. In this present review we attempt to summarize the current knowledge on the effect of the donor and recipient genotypes on GvHD, starting from human leucocyte antigen (HLA) matching for an optimal donor selection, typing of non-classical HLA and minor histocompatibility antigens through the polymorphic variations in genes coding for non-HLA proteins contributing to the development of GvHD and response to treatment. The results of recent Candidate Gene Studies (CGS) and Genome-Wide Association Studies (GWAS) are presented and discussed.

Expression and differential regulation of HLA-G isoforms in the retinal pigment epithelial cell line, ARPE-19

The purpose of this study was to examine if HLA-G is expressed in the retinal pigment epithelium (RPE) cells of the eye. The RPE comprises the outer most layer of the retina and as such defines the interface to the blood and contributes to the immune privilege in the posterior part of the eye. One way the RPE might be regulating the immune system could be by expressing the non-classical human leukocyte antigen (HLA) molecule, HLA-G. We therefore sought to define if the RPE cell line, ARPE-19, expressed HLA-G and analyse the regulation as a response to pro-inflammatory cytokines. This was done by digital droplet PCR, measuring the gene expression of HLA-G in total RNA. The protein expression was analysed by immunohistochemistry and by immunofluorescence followed by confocal microscopy and the expression of the HLA-G isoforms was explored by fragment analysis. In the current study, we show that HLA-G is expressed by ARPE-19 cells and is upregulated as a response to pro-inflammatory cytokines. Moreover, we are the first to describe a differential regulation of the HLA-G isoforms as a direct response to stimulation. These results might indicate that HLA-G can be part of the immune privilege of the posterior part of the eye, but further experiments on primary RPE cells are needed.

Circulating and renal expression of HLA-G prevented chronic renal allograft dysfunction in Japanese recipients.

Although the risk of acute rejection has been studied in renal transplanted patients, there is little data about the long-term renal survival effects of non-classical human leukocyte antigen class I (HLA-G) in Japanese patients. We investigated the changes in the estimated glomerular filtration rate (eGFR) for Japanese, and factors affecting the eGFR in 141 adult Japanese subjects whose allografts had survived for at least 1 year. Clinical background data, gender, HLA matching status, the total ischemic time, ABO incompatibility, immunosuppressive therapy, and the serum soluble(s) HLA-G5 level were examined. In addition, renal biopsy specimens from 32 cases, which were obtained before, or 2-4 weeks or one year after the transplant were also evaluated for HLA-G1/5 expression using monoclonal anti-HLA-G antibodies (clone 87G or 4H84). The rates of change per year in the median eGFR (ΔeGFR) and sHLA-G5 were -1.5 ml/min/1.73 m2/year and 11.8 ng/ml, respectively. A positive correlation was detected between the ΔeGFR and sHLA-G5 (r = 0.188, p = 0.025). In multivariate regression analysis, sHLA-G5 and HLA-matching were significant predictors of an improvement in eGFR (beta for sHLA-G: 0.374, p = 0.009; beta for mismatching: -1.135, p = 0.045). The renal tubular epithelial cells (TEC) in 11 cases showed a perinuclear HLA-G1/5 expression after renal transplantation. The renal HLA-G1/5-positive patients displayed much better ΔeGFR (p < 0.05). In conclusion, the sHLA-G5 level and HLA matching status are independent predictors of renal allograft function, as determined by the ΔeGFR, in Japanese patients. HLA-G1/5 was also detected on TEC in the patients with favorable renal function.

New insights in HLA-E polymorphism by refined analysis of the full-length gene.

Human leukocyte antigen (HLA)-E is a non-classical HLA class I molecule that plays a role in both the innate and the adaptive immune response through interaction with receptors on natural killer- and T-cells. The HLA-E gene is characterized by limited polymorphism compared with the classical HLA loci on chromosome 6. At the start of this study, only 13 variable sites had been identified (IPD-IMGT/HLA Database v3.18.0). While most previous studies focused on polymorphism in exons 2 and 3 or specific gene regions, polymorphism in the other exons and introns could influence protein expression and function as well. Studies that investigate extended HLA-E polymorphism are therefore needed to better understand the functional relevance of HLA-E in health and disease. The aim of this study was to examine the variability of the full-length HLA-E gene region in individuals originating from different populations. A total of 7 new HLA-E alleles were identified using full-length HLA-E sequencing of 123 individuals from Asian, Dutch or Hunan Han origin. Furthermore, genome variation analysis of the third phase of the 1000 genomes database showed 107 new variable sites in 2504 individuals originating from 26 different populations. Our study demonstrates that the nucleotide variability of the HLA-E gene is much higher than previously known, albeit in only a limited number of individuals. Overall only 2 variants, HLA-E*01:01 and *01:03, are frequently present worldwide, suggesting that balancing selection is acting on HLA-E.

Genetic Engineering of Pluripotent Cells for the Continuous Derivation of Off-the-Shelf Effector Lymphocytes with Enhanced Therapeutic Persistence By Overcoming the Host Histocompatibility Barrier

Encouraging clinical outcomes in autologous cellular immunotherapy have garnered hope and excitement. However, considerable challenges and limitations of patient-derived cancer immunotherapies remain and need to be addressed in order to consistently deliver reliable and efficacious therapies with broadened applicability. Human induced pluripotent stem cells (hiPSCs) are a unique, renewable source for the continuous generation of cellular therapeutics for the treatment of hematological and non-hematological malignancies, and represent a highly promising approach for overcoming many of the limitations of autologous therapy. To advance the promise of hiPSC technology as an "off-the-shelf" source of cellular therapeutics, several considerations need to be addressed. Enabling cell transfer across histocompatibility barriers to permit persistence and therapeutic efficacy in an allogeneic setting is a key requirement. In addition to improving persistence, the ability to overcome histocompatibility barriers may facilitate multi-dosing regimens which may be a requirement in more advanced and complicated disease settings.Genetic incompatibilities between donor and recipient among the classical human leukocyte antigen (HLA) molecules is the leading cause of alloresponse by the host immune system and is currently mitigated by immunosuppressive strategies. Unfortunately, this treatment strategy is not only a stressful event for the patient but also damages the endogenous immune system, compromising the patient’s ability to continue to fight the disease and opportunistic infections. Genetic editing of the HLA genes to generate histocompatible universal cell products is a viable opportunity that is currently being investigated. In addition to selective editing of unique genes to avoid a T cell mediated alloresponse, additional considerations such as natural killer (NK) cell-mediated rejection will need to be addressed. We have previously demonstrated that our proprietary reprogramming platform supports efficient and rapid derivation of clonal hiPSC lines with properties indicative of the naïve state of pluripotency. In addition to maintaining a homogeneous renewable population of hiPSCs, our platform is amenable to precise multi-gene and multi-loci targeted engineering at the single cell level, in both nuclease -dependent and -independent strategies. Furthermore, we have shown through small molecule-guided differentiation protocols, these highly-stable pluripotent cell lines can be banked and repeatedly tapped to consistently produce homogenous populations of immune cells with enhanced effector properties.Here we demonstrate a multi-faceted and comprehensive approach for the generation of immune tolerant hiPSCs and hiPSC-derived immune effector cells. We successfully combined deletion of classical HLA molecules with enforced expression of robust immunosuppressive proteins, including non-classical HLA molecules, to generate clonal hiPSC lines with the ability to escape immune rejection for "off-the-shelf" (OTS-hiPSCs) cellular immunotherapy. Utilizing in vitro real-time quantitative live cell analysis we determined that OTS-hiPSCs elicit a significantly decreased cytotoxic response from both activated peripheral blood (PB)-NK cells and primed PB-T cells compared to wildtype controls. Furthermore we demonstrate that OTS-hiPSCs exhibit improved persistence in xenograft studies in vivo. Bilateral teratomas were formed in a non-conditioned, fully immune-competent recipient mice using luciferized wildtype and OTS-hiPSCs. Daily bioluminescence imaging over a period of 7 days revealed a significant increase (>50 fold difference) in persistence of OTS-hiPSCs compared to wildtype hiPSCs during the 60-144 hour post injection window. Lastly we demonstrate that OTS-hiPSCs can successfully differentiate into functional effector lymphocytes using our potent chemically-defined monolayer hematopoietic differentiation platform. Our current studies focus on the functional characterization of OTS-hiPSC-derived effector lymphocytes in humanized mouse models and generating increased potency of OTS-hiPSC-derived effector lymphocytes through precise genetic engineering of antigen targeting and costimulatory proteins to create and optimized source of "off-the-shelf" cell-based immunotherapies. DisclosuresBauer:Fate Therapeutics: Employment. Clarke:Fate Therapeutics: Employment. Sasaki:Fate Therapeutics: Employment. Groff:Fate Therapeutics: Employment. Lee:Fate Therapeutics: Employment. Lan:Fate Therapeutics: Employment. Abujarour:Fate Therapeutics: Employment. Bonello:Fate Therapeutics: Employment. Burrascano:Fate Therapeutics: Employment. Robinson:Fate Therapeutics: Employment. Bjordahl:Fate Therapeutics, Inc: Employment. Gaidarova:Fate Therapeutics: Employment. Abbot:Fate Therapeutics: Employment. Wolchko:Fate Therapeutics: Employment. Shoemaker:Fate Therapeutics: Employment, Equity Ownership. Valamehr:Fate Therapeutics, Inc: Employment.

Classical and non-classical HLA class I aberrations in primary cervical squamous- and adenocarcinomas and paired lymph node metastases

BackgroundTumors avoid destruction by cytotoxic T cells (CTL) and natural killer (NK) cells by downregulation of classical human leukocyte antigens (HLA) and overexpression of non-classical HLA. This is the first...

Association of high HLA-E expression during acute cellular rejection and numbers of HLA class I leader peptide mismatches with reduced renal allograft survival

Non-classical Human Leukocyte Antigen (HLA)-E preferentially presents leader peptides derived from classical HLA-class I molecules. HLA-E can trigger opposed immune responses by interacting with inhibitory NKG2A or by activating NKG2C receptors on NK and T-cells. We studied the impact of HLA-E on renal allograft survival during acute cellular rejection. HLA-E expression was up-regulated in acute cellular rejection (ACR) biopsies (n=12) compared to biopsies from 13 renal allografts with no rejection-signs. HLA-E up-regulation was correlated with numbers of HLA-class I leader peptide mismatches (p=0.04). CD8+ and CD56+ infiltrating cells correlated with HLA-E expression (p<0.0001 and p=0.0009, respectively). Activating NKG2C receptor dominated on effector cells in biopsies and peripheral blood during ACR potentially allowing HLA-E-mediated immune activation. Moreover, HLA-E expression correlated with deterioration in renal allograft function (p<0.008) and reduced allograft survival (p=0.002). Our findings provide evidence that during renal allograft rejection HLA-E along with high numbers of mismatched HLA-class I leader peptides might represent additional targets for immune-activating responses.

The Activating NKG2C Receptor Is Significantly Reduced in NK Cells after Allogeneic Stem Cell Transplantation in Patients with Severe Graft-versus-Host Disease.

Natural killer (NK) cells play a central role in the innate immune system. In allogeneic stem cell transplantation (alloSCT), alloreactive NK cells derived by the graft are discussed to mediate the elimination of leukemic cells and dendritic cells in the patient and thereby to reduce the risk for leukemic relapses and graft-versus-host reactions. The alloreactivity of NK cells is determined by various receptors including the activating CD94/NKG2C and the inhibitory CD94/NKG2A receptors, which both recognize the non-classical human leukocyte antigen E (HLA-E). Here we analyze the contribution of these receptors to NK cell alloreactivity in 26 patients over the course of the first year after alloSCT due to acute myeloid leukemia, myelodysplastic syndrome and T cell Non-Hodgkin-Lymphoma. Our results show that NK cells expressing the activating CD94/NKG2C receptor are significantly reduced in patients after alloSCT with severe acute and chronic graft-versus-host disease (GvHD). Moreover, the ratio of CD94/NKG2C to CD94/NKG2A was reduced in patients with severe acute and chronic GvHD after receiving an HLA-mismatched graft. Collectively, these results provide evidence for the first time that CD94/NKG2C is involved in GvHD prevention.

Molecular mechanisms of HLA class I-mediated immune evasion of human tumors and their role in resistance to immunotherapies.

Although the human immune system can recognize and eradicate tumor cells, tumors have also been shown to develop different strategies to escape immune surveillance, which has been described for the first time in different mouse models. The evasion of immune recognition was often associated with a poor prognosis and reduced survival of patients. During the last years the molecular mechanisms, which protect tumor cells from this immune attack, have been identified and appear to be more complex than initially expected. However, next to the composition of cellular, soluble and physical components of the tumor microenvironment, the tumor cells changes to limit immune responses. Of particular importance are classical and non-classical human leukocyte antigen (HLA) class I antigens, which often showed a deregulated expression in cancers of distinct origin. Furthermore, HLA class I abnormalities were linked to defects in the interferon signaling, which have both been shown to be essential for mounting immune responses and are involved in resistances to T cell-based immunotherapies. Therefore this review summarizes the expression, regulation, function and clinical relevance of HLA class I antigens in association with the interferon signal transduction pathway and its role in adaptive resistances to immunotherapies.

HLA-E expression and its clinical relevance in human renal cell carcinoma.

The non-classical human leukocyte antigen E (HLA-E) expression is frequently overexpressed in tumor diseases, transplants and virus-infected cells and represents an immunomodulatory molecule by binding to the receptors CD94/NKG2A, -B and –C on NK and T cells. Due to its immune suppressive features HLA-E expression might represent an important mechanism of tumors to escape immune surveillance.While an aberrant expression of the non-classical HLA-G antigen in human renal cell carcinoma (RCC) has been demonstrated to be associated with a worse outcome of patients and reduced sensitivity to immune effector cell-mediated cytotoxicity, the expression and function of HLA-E has not yet been analyzed in this tumor entity.Higher levels of HLA-E transcripts were detected in all RCC cell lines and tumor lesions, which were tested in comparison to normal kidney epithelium. Immunohistochemical staining of a tissue microarray (TMA) using the HLA-E-specific monoclonal antibody TFL-033 recognizing the cytoplasmic HLA-E α-chain as monomer revealed a heterogeneous HLA-E expression in RCC lesions with the highest frequency in chromophobe RCC when compared to other RCC subtypes. HLA-E expression did not correlate with the frequency of CD3+, CD4+, CD8+ and FoxP3+ immune cell infiltrations, but showed an inverse correlation with infiltrating CD56+ cells. In contrast to HLA-G, HLA-E expression in RCCs was not statistically significant associated with a decreased disease specific survival. These data suggest that HLA-E overexpression frequently occurs in RCC and correlates with reduced immunogenicity.

Contribution of a Non-classical HLA Gene, HLA-DOA, to the Risk of Rheumatoid Arthritis

Despite the progress in human leukocyte antigen (HLA) causal variant mapping, independent localization of major histocompatibility complex (MHC) risk from classical HLA genes is challenging. Here, we conducted a large-scale MHC fine-mapping analysis of rheumatoid arthritis (RA) in a Japanese population (6,244 RA cases and 23,731 controls) population by using HLA imputation, followed by a multi-ethnic validation study including east Asian and European populations (n = 7,097 and 23,149, respectively). Our study identified an independent risk of a synonymous mutation at HLA-DOA, a non-classical HLA gene, on anti-citrullinated protein autoantibody (ACPA)-positive RA risk (p = 1.4× 10(-9)), which demonstrated a cis-expression quantitative trait loci (cis-eQTL) effect on HLA-DOA expression. Trans-ethnic comparison revealed different linkage disequilibrium (LD) patterns in HLA-DOA and HLA-DRB1, explaining the observed HLA-DOA variant risk heterogeneity among ethnicities, which was most evident in the Japanese population. Although previous HLA fine-mapping studies have identified amino acid polymorphisms of the classical HLA genes as driving genetic susceptibility to disease, our study additionally identifies the dosage contribution of a non-classical HLA gene to disease etiology. Our study contributes to theunderstanding of HLA immunology in human diseases and suggests the value of incorporating additional ancestry in MHC fine-mapping.

Soluble monomers, dimers and HLA-G-expressing extracellular vesicles: the three dimensions of structural complexity to use HLA-G as a clinical biomarker.

The HLA-G molecule belongs to the family of nonclassical human leukocyte antigen (HLA) class I. At variance to classical HLA class I, HLA-G displays (i) a low number of nucleotide variations within the coding region, (ii) a high structural diversity, (iii) a restricted peptide repertoire, (iv) a limited tissue distribution and (v) strong immune-suppressive properties. The physiological HLA-G surface expression is restricted to the maternal-fetal interface and to immune-privileged adult tissues. Soluble forms of HLA-G (sHLA-G) are detectable in various body fluids. Cellular activation and pathological processes are associated with an aberrant or a neo-expression of HLA-G/sHLA-G. Functionally, HLA-G and its secreted forms are considered to be key players in the induction of short- and long-term tolerance. Thus, its unique expression profile and tolerance-inducing functions render HLA-G/sHLA-G an attractive biomarker to monitor the systemic health/disease status and disease activity/progression for clinical approaches in disease management and treatments. Here, we place emphasis on (i) the current status of the tolerance-inducing functions by HLA-G/sHLA-G, (ii) the current complexity to implement this molecule as a meaningful clinical biomarker regarding the three dimensions of structural diversity (monomers, dimers and HLA-G-expressing extracellular vesicles) with its functional implications, and (iii) novel and future approaches to detect and quantify sHLA-G structures and functions.

HLA-G expression is regulated by miR-365 in trophoblasts under hypoxic conditions

IntroductionHypoxia occurs in the first trimester of placental development and is implicated in the regulation of trophoblast differentiation. Prolonged hypoxic conditions in the placenta are related to the development of preeclampsia. MicroRNAs (miRNAs) are noncoding, single-stranded RNAs that modulate gene expression by targeting messenger RNA. We hypothesized that, under hypoxic conditions, trophoblasts may have a unique miRNA profile that may play a critical role in placental development. MethodsTotal RNA was extracted from human trophoblast, HChEpC1b, exposed to normoxia (20% O2) or hypoxia (2% O2) for 24 h, and the miRNA expression profiles were investigated using a microRNA array. Several differential miRNAs were selected and validated using real-time reverse transcription PCR. We identified potential targets of these miRNAs using in silico analysis. We confirmed a potential target protein by western blot analysis and luciferase assays. ResultsThe expression of miR-365 was significantly upregulated under hypoxic conditions. In silico analysis showed that miR-365 targeted human leukocyte antigen (HLA)-G. Both hypoxic conditions and overexpression of miR-365 inhibited the expression of HLA-G proteins. The overexpression of miR-365 also decreased the activity of the luciferase reporter containing the 3′-untranslated region (UTR) of HLA-G with the predicted miR-365-binding site. DiscussionHLA-G is a non-classical HLA class-Ib molecule that is expressed mainly in extravillous trophoblasts and which plays a key role in maintaining immune tolerance at the maternal–fetal interface. Our results indicate that miR-365 targets the HLA-G 3′ UTR to repress its expression. The expression of miR-365 may play an important role in human placental development and in immunoprotection of the semiallogenic embryo.