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Summary Wheat-dependent exercise-induced anaphylaxis (WDEIA) is a life-threatening food allergy triggered by wheat in combination with the second factor such as exercise. The identification of potential genetic risk factors for this allergy might help high-risk individuals before consuming wheat-containing food. We aimed to identify genetic variants associated with WDEIA. A genome-wide association study was conducted in a discovery set of 77 individuals with WDEIA and 924 control subjects via three genetic models. The associations were confirmed in a replication set of 91 affected individuals and 435 control individuals. Summary statistics from the combined set were analyzed by meta-analysis with a random-effect model. In the discovery set, a locus on chromosome 6, rs9277630, was associated with WDEIA in the dominant model (OR = 3.95 [95% CI, 2.31−6.73], p = 7.87 × 10−8). The HLA-DPB1∗02:01:02 allele displayed the most significant association with WDEIA (OR = 4.51 [95% CI, 2.66−7.63], p = 2.28 × 10−9), as determined via HLA imputation following targeted sequencing. The association of the allele with WDEIA was confirmed in replication samples (OR = 3.82 [95% CI, 2.33−6.26], p = 3.03 × 10−8). A meta-analysis performed in the combined set revealed that the HLA-DPB1∗02:01:02 allele was significantly associated with an increased risk of WDEIA (OR = 4.13 [95% CI, 2.89−5.93], p = 1.06 × 10−14). Individuals carrying the HLA-DPB1∗02:01:02 allele have a significantly increased risk of WDEIA. Further validation of these findings in independent multiethnic cohorts is needed.

Gitelman syndrome (GS) is an autosomal recessive disorder characterized by alkalosis, hypokalemia, and hypomagnesemia. Although hundreds of genetic variants associated with GS have been reported, many of them are categorized as of uncertain significance in ClinVar. Here, we describe a pediatric GS patient from a three-generation family whose mother and maternal grandmother were asymptomatic. The proband was a 16-year-old Japanese girl with muscle weakness and continuous hypokalemic metabolic alkalosis. The patient, her mother, and her maternal grandmother were compound heterozygous for, and each expressing a different combination of, previously reported SLC12A3variants in GS patients. The mother and the maternal grandmother had no symptoms related to GS, and blood gas tests showed that the blood potassium levels and venous pH were within normal limits; however, the venous blood HCO3- levels were slightly elevated. The phenotypic effect of missense mutations is difficult to evaluate, and accumulation of genotypic data with accurate phenotyping, including those of “healthy” and “asymptomatic” individuals in various ethnic populations, will improve the genetic diagnosis of GS.

Background Human and animal research indicates that oxytocin (OT) plays a key role in the cross-generational transmission of parental bonding, and human studies suggest that allelic variations on the oxytocin receptor gene (OXTR) and circulating OT levels interact with patterns of parental care to shape children's social-affiliative competencies. Yet, no study to date has tested the joint contribution of OT and parental care across three generations. Methods The study included 345 participants comprising 115 family lines of grandmothers, mothers, and their infants. Salivary OT and allelic variations on the OXTR (rs53576 and rs2254298) and CD38 (rs3796863) single nucleotide polymorphisms (SNPs), which have been previously associated with parental bonding, were assessed in all participants. Parental care was measured from grandmothers to mothers and from mothers to their infants. Results Mothers receiving parenting characterized by high overprotection from grandmothers showed more rejection toward their infants only when carrying the G allele on the OXTRrs53576 (AG/GG). These mothers of highly overprotective grandmothers also had lower oxytocin levels. Infants who were OXTRrs2254298 A carriers (AA/AG) and whose mothers reported more rejection toward their infants had higher oxytocin levels. Grandmothers receiving higher overprotection from great-grandmothers showed poorer parenting style compared to grandmothers experiencing lower parental overprotection only when carrying the OXTRrs2254298 GG genotype. Conclusions Our findings are the first to demonstrate how genetic and peripheral markers on the oxytocin system interact with experienced parenting to shape bonding across three generations. Results have important implications for specifying the biological and behavioral determinants associated with the continuity of adaptive versus maladaptive patterns of attachment across generations.

Background: Rhabdomyosarcoma (RMS) is one of the most common soft tissue sarcomas in children. Germline mutations in cancer predisposition genes have been detected in approximately 10% of pediatric cancers. However, the genetic background of RMS is still unclear, especially in Asian children. Procedure: DNA was extracted from the peripheral blood of children with RMS and cancer-associated genes analyzed using targeted re-sequencing. Results: Twenty patients participated in this study. The median age at diagnosis was 5.0 years. The male-to-female ratio was 3:2. There were three deaths due to RMS. One patient developed a second neoplasm. Nine patients had long-term co-morbidities. Five pathogenic variants were found in four patients: one nonsense variant of DICER1, one exon deletion of TP53, and three missense variants of LIG4 and MEN1. Two of the four patients had a family history of cancer. Two patients with missense variants of LIG4 had long-term co-morbidities of drug-induced cardiomyopathy. This is the first report of germline cancer-related gene variants with detailed clinical information in Japanese children with RMS. The missense variants of LIG4, essential for DNA double-strand break repair, were detected in two unrelated patients. Conclusions: When this is the first report of the germline genetic analysis of Japanese children with RMS, the frequency of the variant was almost equivalent to that of previous reports from Western countries. Unbiased exon sequencing may be useful to clarify the pathogenesis of RMS in children and in predicting the clinical course of these patients.

Allergic rhinitis (AR) is one of the most common diseases in Japan. However, several AR patients do not seek optimal treatments at clinics/hospitals. This may affect the patient's quality of life and labor productivity. In this study, we assessed the characteristics of the outpatients' AR and factors associated with their hospital visit, using the dataset obtained from a nation-wide survey in Japan.In this cross-sectional study, we used data from the nation-wide 2013 and 2016 Comprehensive Survey of Living Conditions (CSLC) in Japan. We analyzed the data of AR outpatients through logistic regression, using the outcome as the dependent variable, and age groups, sex, household size, educational status, smoking history, alcohol use, household expenditure, psychological distress, quality of sleep, asthma and atopic dermatitis outpatients as explanatory variables.Among the data of 317,984 outpatients aged between 20 and 79 years in 2016 CSLC survey, the proportion of AR outpatients was significantly less among current smokers (odds ratio (OR); 0.47, 95% confidence interval (CI); 0.43-0.51, P 0.001) and those with large household sizes (OR; 0.80, 95% CI; 0.72-0.89, P 0.001). Conversely, the proportion of AR outpatients was significantly more among subjects with a past smoking habit (OR; 1.19, 95% CI; 1.08-1.31, P 0.001), insufficient sleep (OR; 2.93, 95% CI; 2.52-3.42, P 0.001), psychological distress (OR; 1.71, 95% CI; 1.62-1.80, P 0.001), high household expenditures (OR; 1.68, 95% CI; 1.56-1.80, P 0.001), and asthma and atopic dermatitis outpatients (OR; 8.97, 95% CI; 8.13-9.89 P 0.001 for asthma and OR; 7.61, 95% CI; 6.76-8.58 P 0.001 for atopic dermatitis). We observed the same trend using the dataset of 2013 CLSC survey.This study revealed that smoking habit, psychological distress, insufficient sleep, high household expenditures and outpatients with other allergic diseases are the factors associated with AR outpatient visit.

Neoantigens have attracted attention as biomarkers or therapeutic targets. However, accurate prediction of neoantigens is still challenging, especially in terms of its accuracy and cost. Variant detection using RNA sequencing (RNA‐seq) data has been reported to be a low‐accuracy but cost‐effective tool, but the feasibility of RNA‐seq data for neoantigen prediction has not been fully examined. In the present study, we used whole‐exome sequencing (WES) and RNA‐seq data of tumor and matched normal samples from six breast cancer patients to evaluate the utility of RNA‐seq data instead of WES data in variant calling to detect neoantigen candidates. Somatic variants were called in three protocols using: (i) tumor and normal WES data (DNA method, Dm); (ii) tumor and normal RNA‐seq data (RNA method, Rm); and (iii) combination of tumor RNA‐seq and normal WES data (Combination method, Cm). We found that the Rm had both high false‐positive and high false‐negative rates because this method depended greatly on the expression status of normal transcripts. When we compared the results of Dm with those of Cm, only 14% of the neoantigen candidates detected in Dm were identified in Cm, but the majority of the missed candidates lacked coverage or variant allele reads in the tumor RNA. In contrast, about 70% of the neoepitope candidates with higher expression and rich mutant transcripts could be detected in Cm. Our results showed that Cm could be an efficient and a cost‐effective approach to predict highly expressed neoantigens in tumor samples., We evaluated the utility of RNA‐seq data instead of WES data in variant calling to detect neoantigen candidates. We found that the method combining tumor RNA‐seq data and normal WES data (Combination method) could detect neoantigen candidates that have higher expression and rich variant transcripts, and this method may be an efficient and cost‐effective strategy alternative to the conventional method (DNA method).

Murine models to elucidate the pathogenesis of pollen food allergy syndrome (PFAS), characterized by oral hypersensitivity symptoms induced by specific foods in patients previously sensitized with a pollen, are lacking. The study aimed to examine PFAS pathogenesis in a novel murine model. Birch pollen–immunized mice were orally administered apple extract, and oral symptoms were evaluated based on oral rubbing frequency following the challenge. The birch pollen–immunized mice orally challenged with apple extract exhibited PFAS-like symptoms, including oral rubbing and positive reaction of swelling by the prick test. The apple extract administered with a protease inhibitor reduced the oral rubbing frequency, which was also significantly reduced in the immunized Fcer1a−/− and mast cell–deficient mice compared with the immunized control mice. The oral rubbing frequency, serum IgE levels, and Th2-cytokine production by the cervical lymph node cells were significantly reduced in the immunized Il-33−/− and thymic stromal lymphopoietin receptor–deficient (Crlf2−/−) mice as compared with the immunized wild-type mice. IL-33 and thymic stromal lymphopoietin involve the pathogenesis of PFAS. The apple-extract stimulation did not lead to increased Th2-cytokine production in the oral mucosa or number of group 2 innate lymphoid cells or eosinophils. PFAS involves an early-phase response by mast cell degranulation via IgE signaling after the cross-reactivity of Bet v 1–specific IgE and the food allergen, and exacerbation of allergic symptom via proteases in food; PFAS does not involve a late phase with local Th2/eosinophilic inflammation in the oral mucosa. This novel murine model might be used for elucidating the pathogenesis and assessing new therapeutic strategies for PFAS.

Background Despite recent findings that epithelial cell adhesion molecule (EPCAM) deletions can cause Lynch syndrome (LS), its clinical characteristics are still unknown. We present the first case of ileum cancer in a patient with germline EPCAM gene deletion, which was discovered during ovarian tumor surgery. Case presentation A 59-year-old woman presented with a history of colon cancer occurring at 38 and 55 years old. Five of her siblings had a history of colon cancer, and an elder sister had confirmed LS. As imaging examination revealed an ovarian tumor, and we performed hysterectomy and bilateral salpingo-oophorectomy. Careful observation during surgery revealed a cherry-sized tumor in the ileum, prompting partial ileal resection. Pathological examination showed the ovarian tumor to be a metastasis of ileum cancer. Genetic testing with blood-relative information using multiplex ligation-dependent probe amplification showed EPCAM exons 8 and 9 deletions, confirming LS. The patient received adjuvant chemotherapy with CAPOX (capecitabine and oxaliplatin) and has remained disease-free for 24 months. Conclusions We were fortunate to identify ileum cancer that would have been difficult to find preoperatively through careful observation during ovarian tumor surgery and successfully treated the patient by using surgical resection and CAPOX chemotherapy. When treating patients with hereditary cancer syndromes including LS, we should keep all associated cancers in mind.

Type I allergy is an immunological disorder triggered by allergens and causes significant health problems. The major allergen of birch pollen is Bet v 1, which belongs to the pathogen-related protein 10 (PR-10) family. Here, we established a rapid and robust method for the production of Bet v 1 in Nicotiana benthamiana leaves, with binding activity to allergic patients’ IgE. The Bet v 1 allergen was expressed in N. benthamiana using a strong agroinfiltration-based transient protein expression system, which consists of a deconstructed geminiviral vector system with a double terminator. Five days post-infiltration, the allergen concentration in N. benthamiana leaves was 1.2 mg/g of fresh mass, being this the maximum yield of Bet v 1 in plants reported up to now. A part of plant-derived Bet v 1 was glycosylated. Bet v 1 purified from N. benthamiana or Brevibacillus brevis was used to carry out enzyme-linked immunoassays; both recombinant allergens were found to have comparable binding properties to the IgE of allergic patients. These results suggest that our plant expression system allows rapid and robust production of the allergen, which keeps the immunogenicity.

Background: An orosomucoid-like 3 (ORMDL3)/gasdermin B (GSDMB) gene locus on chromosome 17q is consistently associated with childhood-onset asthma, which is highly atopic. As some evidence suggests the relationship between asthma and allergic sensitization reflects asthma patient susceptibility to augmented IgE responses driven by common environmental allergens rather than an increased asthma risk after allergen exposure, we aimed to determine any relationships between this locus region and childhood-onset adult asthma with regard to serum total IgE levels or allergic sensitization. Methods: We conducted a case–control association study using three independent Japanese populations (3869 total adults) and analyzed the ORs for association of rs7216389, an expression quantitative trait locus for ORMDL3/GSDMB, with adult asthma according to onset age. Additionally, associations between the rs7216389 genotype and total serum IgE levels or allergic sensitization was examined. Results: Rs7216389 was associated with both childhood-onset adult asthma (OR for asthmatic patients afflicted at the age of 10 years or younger = 1.61, p = 0.00021) and asthmatic patients with higher levels of total serum IgE (OR for asthmatic patients with IgE ≥1000IU/mL = 1.55, p = 0.0033). In both healthy controls and in the combined healthy and asthmatic individuals, rs7216389 was correlated with increased total serum IgE levels (p 0.1). Conclusions: ORMDL3/GSDMB is an important susceptibility gene for childhood-onset adult asthma in Japanese populations and this association is linked to elevated total serum IgE levels but not to allergic sensitization.

Immunodeficiency, centromeric instability, and facial anomalies (ICF) syndrome is a rare autosomal recessive primary immunodeficiency. Hypogammaglobulinemia is a major manifestation of ICF syndrome, but immunoglobulin replacement therapy does not seem to be effective for some ICF patients. Therefore, we aimed to reassess the immunological characteristics of this syndrome. Eleven Japanese patients with ICF syndrome were enrolled. We performed whole-exome sequencing in four cases and homozygosity mapping using SNP analysis in two. We evaluated their clinical manifestations and immunological status. We newly diagnosed six ICF patients who had tentatively been diagnosed with common variable immunodeficiency. We identified two novel mutations in the DNMT3B gene and one novel mutation in the ZBTB24 gene. All patients showed low serum IgG and/or IgG2 levels and were treated by periodic immunoglobulin replacement therapy. Three of the six patients showed worse results of the mitogen-induced lymphocyte proliferation test. Analyses of lymphocyte subpopulations revealed that CD19+CD27+ memory B cells were low in seven of nine patients, CD3+ T cells were low in three patients, CD4/8 ratio was inverted in five patients, CD31+ recent thymic emigrant cells were low in two patients, and CD19+ B cells were low in four patients compared with those in the normal controls. ICF2 patients showed lower proportions of CD19+ B cells and CD16+56+ NK cells and significantly higher proportions of CD3+ T cells than ICF1 patients. T cell receptor excision circles were undetectable in two patients. Despite being treated by immunoglobulin replacement therapy, three patients died of influenza virus, fatal viral infection with persistent Epstein–Barr virus infection, or JC virus infection. One of three dead patients showed normal intelligence with mild facial anomaly. Two patients presented with autoimmune or inflammatory manifestations. Infectious episodes decreased in three patients who were started on trimethoprim–sulfamethoxazole and/or antifungal drugs in addition to immunoglobulin replacement therapy. These patients might have suffered from T cell immunodeficiency. These results indicate that patients with ICF syndrome have a phenotype of combined immunodeficiency. Thus, to achieve a better prognosis, these patients should be treated as having combined immunodeficiency in addition to receiving immunoglobulin replacement therapy.

Aim Gingivitis commonly progresses to periodontitis in permanent dentition but rarely in deciduous teeth. Little is known about the biochemical differences between gingiva of deciduous and permanent teeth. Here we compared the protein profiles of gingival crevicular fluids (GCF) from the gingiva of deciduous and permanent teeth. Materials and Methods Forty children with mixed dentition (Hellman's dental age IIIA) were selected and GCF samples were collected from deciduous cuspids and central incisors in the maxilla. Pairs of GCF samples were labelled using isobaric tags to permit quantitative comparison of protein abundance in the samples using liquid chromatography-electron spray ionization-tandem mass spectrometry. Results Sixty-two proteins were upregulated in deciduous teeth GCF and 54 in permanent teeth GCF. In particular, neutrophil-derived proteins, including myeloperoxidase and lactoferrin, were repeatedly higher in deciduous teeth GCF than in permanent teeth GCF. These differences were verified using ELISA (P < 0.01). In contrast, immunoglobulin components were upregulated in permanent teeth GCF. Conclusions Neutrophil-related proteins were enriched in deciduous teeth GCF and immunoglobulins in permanent teeth GCF. This suggests that neutrophil accumulation plays a protective role in innate immunity against bacterial infection in gingival tissue of deciduous teeth. This article is protected by copyright. All rights reserved.

Background and objective Periodontal disease is a chronic inflammatory disease caused by bacterial infection that can lead to tooth loss. Gingival crevicular fluid can be collected easily and noninvasively. We previously discovered the presence of apolipoprotein B (apoB), the main constituent of low-density lipoprotein, and oxidized low-density lipoprotein (oxLDL) in the gingival crevicular fluid of healthy subjects. In this study, we investigated whether periodontal conditions affect the levels of apoB and oxLDL in gingival crevicular fluid. Material and methods The study population comprised 11 patients with chronic periodontitis. A pair of gingival crevicular fluid samples was collected from each patient at a healthy site and at a site with periodontitis (baseline samples). Thereafter, gingival crevicular fluid samples were collected from the same patients again at 4 and 8 wk after scaling and root planing (SRP). The levels of apoB, oxLDL, protein and cytokines in gingival crevicular fluid, in addition to gingival crevicular fluid volume, were measured. Results At baseline, the levels of apoB and oxLDL in gingival crevicular fluid were higher at the sites with periodontitis than at the healthy sites. The levels of apoB and oxLDL at periodontal sites decreased after SRP. The level of oxLDL in gingival crevicular fluid correlated well with the probing pocket depth. The oxLDL : apoB ratio in gingival crevicular fluid was significantly higher than that in plasma. Conclusions The levels of apoB and oxLDL in gingival crevicular fluid change according to the periodontal tissue conditions.

House dust mite (HDM) is a major allergen that causes allergic diseases such as atopic dermatitis. However, the regulatory mechanisms of HDM-induced immune responses are incompletely understood. NC/Nga mice are an inbred strain that is more susceptible to HDM and develops more severe dermatitis than other strains. Using whole-exome sequencing, we found that NC/Nga mice carry a stop-gain mutation in Clec10a, which encodes a C-type lectin receptor, Clec10a (MGL1/CD301a). The repair of this gene mutation using the CRISPR-Cas9 system ameliorated HDM-induced dermatitis, indicating that the Clec10a mutation is responsible for hypersensitivity to HDM in NC/Nga mice. Similarly, Clec10a−/− mice on the C57BL/6J background showed exacerbated HDM-induced dermatitis. Clec10a expressed on skin macrophages inhibits HDM-induced Toll-like receptor 4 (TLR4)–mediated inflammatory cytokine production through the inhibitory immunoreceptor tyrosine activating motif in its cytoplasmic portion. We identified asialoglycoprotein receptor 1 (Asgr1) as a functional homolog of mouse Clec10a in humans. Moreover, we found that a mucin-like molecule in HDM is a ligand for mouse Clec10a and human Asgr1. Skin application of the ligand ameliorated a TLR4 ligand-induced dermatitis in mice. Our findings suggest that Clec10a in mice and Asgr1 in humans play an important role in skin homeostasis against inflammation associated with HDM-induced dermatitis.

Background Asthma is a complex chronic inflammatory disease of the airways. Association studies between HLA and asthma were first reported in the 1970s, and yet, the precise role of HLA alleles in asthma is not fully understood. Numerous genome-wide association studies were recently conducted on asthma, but were always limited to simple genetic markers (single nucleotide polymorphisms) and not complex HLA gene polymorphisms (alleles/haplotypes), therefore not capturing the biological relevance of this complex locus for asthma pathogenesis. Objective To run the first HLA-centric association study with asthma and specific asthma-related phenotypes in a large cohort of African-ancestry individuals. Methods We collected high-density genomics data for the Consortium on Asthma among African-ancestry Populations in the Americas (N = 4993) participants. Using computer-intensive machine-learning attribute bagging methods to infer HLA alleles, and Easy-HLA to infer HLA 5-gene haplotypes, we conducted a high-throughput HLA-centric association study of asthma susceptibility and total serum IgE (tIgE) levels in subjects with and without asthma. Results Among the 1607 individuals with asthma, 972 had available tIgE levels, with a mean tIgE level of 198.7 IU/mL. We could not identify any association with asthma susceptibility. However, we showed that HLA-DRB1∗09:01 was associated with increased tIgE levels (P = 8.5 × 10−4; weighted effect size, 0.51 [0.15-0.87]). Conclusions We identified for the first time an HLA allele associated with tIgE levels in African-ancestry individuals with asthma. Our report emphasizes that by leveraging powerful computational machine-learning methods, specific/extreme phenotypes, and population diversity, we can explore HLA gene polymorphisms in depth and reveal the full extent of complex disease associations.

Background The chitinase-like protein YKL-40 plays a major role in inhibiting the inflammasome. Deregulation of inflammasome activation is emerging as a key modulator of pathologic airway inflammation in patients with asthma. We determined whether cis-expression quantitative trait loci (eQTLs) of the gene that encodes YKL-40, chitinase 3-like 1 (CHI3L1), are involved in the onset of asthma or in specific asthma phenotypes. Methods This case-control study, which was conducted at the University of Tsukuba, Japan, included a total of 2709 adults from the Tsukuba genome-wide association study (GWAS) cohort (734 healthy volunteers and 237 asthma patients), the Tsukuba replication cohort (375 healthy adult volunteers and 381 adult asthma patients), and the Hokkaido replication cohort (554 healthy adult volunteers and 428 adult asthma patients). Among 34 cis-eQTLs in CHI3L1 in the lung, rs946261 was associated with adult asthma in these Japanese cohorts. The genetic impact of rs946261 on asthma was also examined according to the age at onset and adult asthma clusters. Results In the Tsukuba GWAS cohort, the C allele at rs946261 was significantly associated with reduced expression of CHI3L1 mRNA in the lung and with development of asthma (odds ratio (OR) 1.27; P = 0.036). The association was also observed following analysis of the three Japanese cohorts (OR 1.16; P = 0.013). A stronger association was found with late-onset asthma that developed at 41 years of age or later (OR 1.24; 95% confidence interval (CI) 1.07–1.45; P = 0.0058) and with a specific asthma phenotype characterized by late onset, less atopy, and mild airflow obstruction (OR 1.29; 95% CI 1.03–1.61; P = 0.027). Conclusions The genotype consisting of the cis-eQTL allele that reduces expression of CHI3L1 was specifically associated with late-onset adult asthma. Given the important role of YKL-40 in many pathophysiological processes, including cell growth, migration, chemotaxis, reorganization, and tissue remodeling, it may be involved in an important pathogenic role in the establishment of inflammation and remodeling in asthmatic airways. Our findings may indicate the presence of a specific endotype related to exaggerated activation of YKL-40 in the pathogenesis of late-onset adult asthma.

e18049 Background: Anaplastic thyroid carcinoma (ATC) is one of the worst prognostic malignancies with a 1-year survival rate of 5-20%. ATC is thought to be originated from differentiated thyroid carcinoma (DTC) caused by undifferentiated transformation because ATCs frequently have preexisting or coexisting region of differentiated thyroid carcinoma (co-DTC), but the detailed mechanism is unclear. In this study, we analyzed mRNA expression among ATCs, DTCs and co-DTCs, and compared their molecular characteristics. Methods: Twelve patients with ATC and eight patients with DTC, who were diagnosed at Tsukuba University Hospital between 2009 to 2020, were included in this study. Of 12 ATCs, four cases had co-DTCs. Total RNA was extracted from formalin-fixed paraffin-embedded (FFPE) samples of ATCs, DTCs and co-DTCs. We analyzed mRNA expression profiles among three groups using nCounter PanCancer IO 360 Panel (NanoString Technologies) and nCounter Advanced Analysis (version 2.0.134). The p-values were adjusted using Benjamini-Hochberg method and adjusted p-value < 0.05 was considered as significant. Results: The 1-year survival rate of the 12 ATC cases in this study was 37.5%. Heatmap of pathway revealed that most ATCs showed up-regulation of almost all tumor-related pathways included in the panel except for autophagy while three of ATCs didn’t show immune-related pathways up-regulation, and one co-DTC sample showed similar behavior of pathways to ATCs. The top 10 up-regulated gene in co-DTCs compare to DTCs were BATF3, TDO2, PRF1, BIRC5, MKI67, ANLN, FCGR3A/B, PPARGC1B, C2 and PFKFB3, and gene set enrichment analysis revealed that the genes with differential expression were enriched in gene set of cell proliferation. The top 10 genes with differential expression in ATCs compare to co-DTCs were CXCL5, BRCA2, MMP1, CSCL6, CXCL1, DKK1 and CXCL8, for up-regulation and ID4, ARID1A and EPM2AIP1 for down-regulation. In ATCs, gene sets of cell proliferation, cytokine and chemokine signaling, and myeloid compartment were especially up-regulated. Immune cell profiling showed that ATC had abundant inflammatory cells, especially TILs, neutrophils and macrophages, while co-DTC showed intermediate characteristics between ATC and DTC. Of note, three case of ATCs had lower inflammatory cell scores than the other ATCs except for macrophages and neutrophils, and were clustered as “DTC-like”. Conclusions: These data revealed that co-DTCs showed up-regulation of cell proliferation related genes compare to DTCs, and some ATCs had different immune cell profiles. co-DTCs showed an intermediate immune cell profile between ATC and DTC. Biological profiling of ATCs, DTCs and co-DTCs may help to understand the developmental mechanism of ATCs. In this presentation, we will report the detailed results of mRNA expression analysis with discussion of the previous reports.

This study investigated rare variants associated with atopic dermatitis. We performed exome analyses on 37 patients who were diagnosed with atopic dermatitis by board-certified dermatologists and had total serum IgE levels greater than 1000 IU/ml. The exome analysis identified seven variants with1% allele frequency in Asian (ASN) population of 1000 Genomes Project phase 1 data and5% allele frequency in the atopic dermatitis exome samples. We then conducted a replication study using 469 atopic dermatitis patients with total serum IgE ≥1000 IU/ml and 935 Japanese controls to assess the presence of these 7 candidate variants. The replication study confirmed that CYP27A1 rs199691576 (A/G) was associated with atopic dermatitis with high serum IgE levels (P = 0.012, odds ratio = 2.1). CYP27A1 is involved in the metabolism of vitamin D3, which plays important roles in modulating immune function. Previous studies have reported polymorphisms in vitamin D pathway genes that are associated with allergy-related phenotypes. Our data confirm the importance of genes regulating the vitamin D pathway in the development of atopic dermatitis.

The pharmacokinetics among children has been altered dynamically. The difference between children and adults is caused by immaturity in things such as metabolic enzymes and transport proteins. The periods when these alterations happen vary from a few days to some years after birth. We hypothesized that the effect of gene polymorphisms associated with the dose of medicine could be influenced by age. In this study, we analyzed 51 patients with childhood acute lymphoblastic leukemia (ALL) retrospectively. We examined the associations between the polymorphism in NUDT15 and clinical data, especially the dose of 6-mercaptopurine (6-MP). Ten of the patients were heterozygous for the variant allele in NUDT15. In patients under 7 years old with NUDT15 variant allele, the average administered dose of 6-MP was lower than that for the patients homozygous for the wild-type allele (P=0.04). Genotyping of NUDT15 could be a beneficial to estimate the tolerated dose of 6-MP for patients with childhood ALL, especially at a preschool age in Japan. Furthermore, the analysis with stratification by age might be useful in pharmacogenomics among children.

Background Food allergy is a growing health problem worldwide because of its increasing prevalence, life-threatening potential, and shortage of effective preventive treatments. In an outbreak of wheat allergy in Japan, thousands of patients had allergic reactions to wheat after using soap containing hydrolyzed wheat protein (HWP). Objectives The aim of the present study was to investigate genetic variation that can contribute to susceptibility to HWP allergy. Methods We conducted a genome-wide association study of HWP allergy in 452 cases and 2700 control subjects using 6.6 million genotyped or imputed single nucleotide polymorphisms. Replication was assessed by genotyping single nucleotide polymorphisms in independent samples comprising 45 patients with HWP allergy and 326 control subjects. Results Through the genome-wide association study, we identified significant associations with the class II HLA region on 6p21 (P = 2.16 × 10−24 for rs9271588 and P = 2.96 × 10−24 for HLA-DQα1 amino acid position 34) and with the RBFOX1 locus at 16p13 (rs74575857, P = 8.4 × 10−9). The associations were also confirmed in the replication data set. Both amino acid polymorphisms (HLA-DQβ1 amino acid positions 13 and 26) located in the P4 binding pockets on the HLA-DQ molecule achieved the genome-wide significance level (P Conclusions Our data provide the first demonstration of genetic risk for HWP allergy and show that this genetic risk is mainly represented by multiple combinations of HLA variants.

BackgroundNasal polyps (NP) are characterized by pseudocysts derived from stromal tissue edema and cause persistent infections in patients with chronic rhinosinusitis (CRS). A low level of tissue-type plasminogen activator (gene name PLAT) is considered a cause of stromal tissue edema because of insufficient plasmin activation in NP; however, the mechanism regulating PLAT gene expression levels is still unclear. The epigenetic mechanism regulating the PLAT gene expression has been studied in other tissues.ObjectiveWe aimed to investigate the methylation levels in the proximal PLAT promoter and their effects on gene expression in NP tissue.MethodsWe investigated the methylation levels at 3 CpG sites in the proximal PLAT promoter regions (−618, −121, and −105 with respect to the transcription initiation site) by bisulfite pyrosequencing and their effects on the gene expression by quantitative real-time polymerase chain reaction (qPCR) in 20 paired samples of NP and inferior turbinate tissue (IT) from patients with CRS.ResultsThe DNA methylation levels at all CpG sites were higher ( P ConclusionsHypermethylation of PLAT promoter may downregulate the gene expression in NP, leading to excessive fibrin deposition by aberrant coagulation cascade. DNA methylation of proximal PLAT promoter may contribute to NP growth and have a potential as a new therapeutic target.

Background: TYRO3 is a member of the TAM (TYRO3, AXL, MERTK) receptor tyrosine kinase family and functions to limit type 2 immune responses implicated in allergic sensitization. Recent studies have shown that multiple intronic variants of TYRO3 were associated with asthma, implying that genetic variation could contribute to errant immune activation. We therefore hypothesized that expression quantitative trait loci (eQTLs) of the TYRO3 gene influence the development of allergic diseases (including asthma and allergic rhinitis) in Japanese populations. Methods: We performed a candidate gene case–control association study of 8 eQTLs of TYRO3 on atopy, asthma, and allergic rhinitis using 1168 unrelated Japanese adults who had GWAS genotyping. We then examined the genetic impact of rs2297377 (TYRO3) on atopy and allergic rhinitis in 2 other independent Japanese populations. Results: A meta-analysis of 3 Japanese populations (a total of 2403 Japanese adults) revealed that rs2297377 was associated with atopy and allergic rhinitis (OR = 1.29 and 1.31; P = 0.00041 and 0.0010, respectively). The risk allele at rs2297377 correlated with decreased expression of TYRO3 mRNA. The gene–gene interaction between HLA-DPB1 and TYRO3 was not significant with regard to sensitization. The estimated proportion of atopy and allergic rhinitis cases attributable to the risk genotype was 14% and 16%, respectively. Conclusions: Our study identified TYRO3 as an important susceptibility gene to atopy and allergic rhinitis in Japanese. Keywords: Allergic rhinitis, Asthma, Atopy, eQTL, TYRO3

Recruitment of circulating monocytes and neutrophils to infection sites is essential for host defense against infections. Here, we identified a previously unannotated gene that encodes an immunoglobulin-like receptor, designated CD300H, which is located in the CD300 gene cluster. CD300H has a short cytoplasmic tail and associates with the signaling adaptor proteins, DAP12 and DAP10. CD300H is expressed on CD16(+) monocytes and myeloid dendritic cells. Ligation of CD300H on CD16(+) monocytes and myeloid dendritic cells with anti-CD300H monoclonal antibody induced the production of neutrophil chemoattractants. Interestingly, CD300H expression varied among healthy subjects, who could be classified into two groups according to "positive" and "negative" expression. Genomic sequence analysis revealed a single-nucleotide substitution (rs905709 (G → A)) at a splice donor site on intron 1 on either one or both alleles. The International HapMap Project database has demonstrated that homozygosity for the A allele of single nucleotide polymorphism (SNP) rs905709 ("negative" expression) is highly frequent in Han Chinese in Beijing, Japanese in Tokyo, and Europeans (A/A genotype frequencies 0.349, 0.167, and 0.138, respectively) but extremely rare in Sub-Saharan African populations. Together, these results suggest that CD300H may play an important role in innate immunity, at least in populations that carry the G/G or G/A genotype of CD300H.

Background Chronic rhinosinusitis (CRS) can be classified into CRS with nasal polyps (CRSwNP) and CRS without nasal polyps (CRSsNP). CRSwNP displays more intense eosinophilic infiltration and the presence of Th2 cytokines. Mucosal eosinophilia is associated with more severe symptoms and often requires multiple surgeries because of recurrence; however, even in eosinophilic CRS (ECRS), clinical course is variable. In this study, we wanted to set objective clinical criteria for the diagnosis of refractory CRS. Methods This was a retrospective study conducted by 15 institutions participating in the Japanese Epidemiological Survey of Refractory Eosinophilic Chronic Rhinosinusitis (JESREC). We evaluated patients with CRS treated with endoscopic sinus surgery (ESS), and risk of recurrence was estimated using Cox proportional hazard models. Multiple logistic regression models and receiver operating characteristics curves were constructed to create the diagnostic criterion for ECRS. Results We analyzed 1716 patients treated with ESS. To diagnose ECRS, the JESREC scoring system assessed unilateral or bilateral disease, the presence of nasal polyps, blood eosinophilia, and dominant shadow of ethmoid sinuses in computed tomography (CT) scans. The cutoff value of the score was 11 points (sensitivity: 83%, specificity: 66%). Blood eosinophilia (>5%), ethmoid sinus disease detected by CT scan, bronchial asthma, aspirin, and nonsteroidal anti-inflammatory drugs intolerance were associated significantly with recurrence. Conclusion We subdivided CRSwNP in non-ECRS, mild, moderate, and severe ECRS according to our algorithm. This classification was significantly correlated with prognosis. It is notable that this algorithm may give useful information to clinicians in the refractoriness of CRS before ESS or biopsy.

Background Management of the adverse effects of chemotherapy is essential to improve outcome of children with leukemia. Some genetic polymorphisms can predict treatment-related toxicity, and be used individually in dose modification of 6-mercaptopurine (6-MP) and methotrexate (MTX) in maintenance therapy for childhood acute lymphoblastic leukemia (ALL). We investigated associations between clinical course and candidate gene polymorphisms less evaluated in Japanese patients. Methods Fifty-three children who received maintenance chemotherapy were enrolled in this study. The scheduled dose of oral 6-MP was 40 mg/m2 daily and that of oral MTX was 25 mg/m2 weekly. The doses were adjusted according to white blood cell count (target range, 2.5–3.5 × 109/L) and aspartate aminotransferase and alanine aminotransferase level ( C than in the patients with wild homozygous genotype. The other analyzed polymorphisms were not associated with toxicity, 6-MP, or MTX dose. Conclusions Polymorphism of SLCO1B1 c.521 T > C could be a strong predictor of 6-MP dose reduction in maintenance chemotherapy in childhood ALL.

Middle interhemispheric variant of holoprosencephaly (MIH) is a rare brain malformation; hemispheric fusion does not occur at the rostral forebrain, but rather across the posterior frontal region. Barkovich and Quint1 first described and proposed MIH as part of the holoprosencephaly (HPE) spectrum in 1993. Although classic HPE and MIH share several similarities, they are related to different embryological mechanisms: classic HPE is caused by a defect in the formation of the embryonic floor plate, whereas MIH occurs after a disturbance to the roof plate formation.2 The gene ZIC2 is important for the differentiation of the roof plate in the dorsal midline of the neural tube of the developing embryo. In humans, ZIC2 mutations have been identified in 3%–4% of HPE cases, including individuals with MIH, thus confirming that MIH is a variant of HPE.3

We previously reported that chronic rhinosinusitis with nasal polyps (CRSwNP) was subdivided into four chronic rhinosinusitis (CRS) subtypes using the JESREC scoring system. We sought to identify the gene expression profile and biomarkers related with CRSwNP by RNA-sequence. RNA-sequencing was performed to identify differentially expressed genes between nasal polyps (NPs) and inferior turbinate mucosa from 6 patients with CRSwNP, and subsequently, quantitative real-time PCR was performed to verify the results. ELISA was performed to identify possible biomarkers for postoperative recurrence. In the RNA-sequencing results, periostin (POSTN) expression was the highest in NP. We focused on POSTN and investigated the protein level of POSTN by immunohistochemistry and ELISA. POSTN was diffusely expressed in moderate and severe eosinophilic CRS using immunohistochemistry, and its staining pattern was associated with the severity of the phenotype of the CRSwNP (P P = 0.0072). Our data suggests that the protein expression level of POSTN was associated with the severity of CRSwNP, and serum POSTN can be a novel biomarker for postoperative recurrence of CRSwNP.

Background Recent studies have demonstrated that a coding SNP (rs6967330, Cys529→Tyr) in cadherin-related family member 3 ( CDHR3 ), which was previously associated with wheezing illness and hospitalizations in infancy, could support efficient human rhinovirus C (RV-C) entry and replication. Here, we sought to examine the genetic contribution of this variant to the development of adult asthma. Methods We performed a candidate gene case–control association study of 2 independent Japanese populations (a total of 3366 adults). The odds ratios (ORs) for association of the A allele at rs6967330 with adult asthma were calculated according to age at onset of asthma. In addition, the effect of the CDHR3 genotype on the development of specific asthma phenotypes was examined. Results The A allele was associated with asthma (OR = 1.56; Mantel–Haenszel p = 0.0040) when the analysis was limited to patients with early-onset adult asthma. In addition, when the analysis was limited to atopic individuals, a stronger association of the CDHR3 variant with early-onset asthma was found, and interaction of the CDHR3 genotype with atopy was demonstrated. Finally, a significant association of this variant was specifically found with a phenotype of asthma characterized by atopy, early-onset, and lower lung function. Conclusions Our study supports the concept that the CDHR3 variant is an important susceptibility factor for severe adult asthma in individuals who develop the disease in early life. The interaction between the CDHR3 variant and atopy indicates that genetic predisposition to early respiratory viral infection is combined with atopy in promoting asthma.

Key Clinical Message The FOXP protein family (FOXP1-4) is a group of transcription factors that play important roles in embryological, immunological, hematological, and speech and language development. Here, we report FOXP1 de novo mutation and severe speech delay in an individual belonging to a non-Caucasian population.

SummaryBackground It is increasingly clear that asthma is not a single disease, but a disorder with vast heterogeneity in pathogenesis, severity, and treatment response. To date, 30 genomewide association studies (GWASs) of asthma have been performed, including by our group. However, most gene variants identified so far confer relatively small increments in risk and explain only a small proportion of familial clustering. Objective To identify additional genetic determinants of susceptibility to asthma using a selected Japanese population with reduced tobacco smoking exposure. Methods We performed a GWAS by genotyping a total of 480 098 single-nucleotide polymorphisms (SNPs) for a Japanese cohort consisting of 734 healthy controls and 240 patients with asthma who had smoked for no more than 10 pack-years. The SNP with the strongest association was genotyped in two other independent Japanese cohorts consisting of a total of 531 healthy controls and 418 patients with asthma who had smoked for no more than 10 pack-years. For the hyaluronan synthase 2 (HAS2) gene, we investigated SNP–gene associations using an expression quantitative trait loci (eQTL) database and also analysed its gene expression profiles in 13 different normal tissues. Results In the discovery GWAS, a SNP located upstream of HAS2, rs7846389, showed the strongest statistical significance (P = 1.43 × 10−7). In the two independent replication cohorts, rs7846389 was consistently associated with asthma (nominal P = 0.0152 and 0.0478 in the first and second replication cohorts, respectively). In the meta-analysis, association of rs7846389 with susceptibility to asthma reached the level of genomewide significance (P = 7.92 × 10−9). This variant was strongly correlated with HAS2 mRNA expression. The strongest expression of the gene was detected in the lung. Conclusions Our study identified HAS2 as a novel candidate gene for susceptibility to adult asthma.

The recently identified cell surface immunoreceptor MILR1 (mast cell immunoglobulin-like receptor 1; synonyms, Allergin-1) has been shown to suppress immunoglobulin E (IgE)-mediated, mast cell-dependent responses in both mice and humans. We performed a mutation search of MILR1 together with a genetic association study to determine whether polymorphisms in MILR1 are associated with atopy in human. Mutation screening of MILR1 was performed using DNA from 146 unrelated Japanese. Genotyping of the identified polymorphisms was done with 1505 individuals from the general Japanese adult population. Atopy, as defined by positive responses for specific IgEs against at least one of the 26 common allergens, was evaluated using MAST-26. Five polymorphisms (rs6504230, c.-170_-166delAGGAA, rs8071835, rs143526766 and rs12936887) and two rare missense variants (Val273Ala and Leu311Val) were identified by mutation screening. The C allele of rs6504230 had protective effects against atopy (P=0.002). A luciferase reporter assay using the promoter region of MILR1 revealed that the C allele of rs6504230 was associated with increased expression of MILR1, which was in accordance with the results of expression quantitative trait loci analysis using human leukocytes. Our data indicates that the rs6504230 polymorphism affects MILR1 expression levels in humans, leading to a susceptibility to producing specific IgE antibodies against common allergens.

In this study, we found Cystatin SN (CST1), a type 2 cystatin subfamily member, to be highly expressed in nasal polyps from patients with intractable chronic rhinosinusitis (CRS) with nasal polyps, using a whole-transcript analysis with next-generation sequencing. Eosinophilic CRS (ECRS) involves nasal polyps that are refractory and recur immediately after endoscopic sinus surgery. We hypothesized that CST1 may contribute to the pathogenesis of ECRS. We examined the expression of CST1 in nasal polyps from patients with ECRS by assessing mRNA expression levels using real-time PCR and immunohistochemistry. CST1 showed significantly greater expression in the epithelial cells of nasal polyps from patients with ECRS than in those from patients who did not have ECRS (non-ECRS). In particular, CST1 showed very strong expression in patients with severe ECRS. The expression of CST1 may be correlated with the recurrent and refractory nature of ECRS. We examined the function of CST1 using nasal epithelial cells and nasal fibroblasts. Stimulation by a combination of IL-4 plus double-stranded RNA plus CST1 significantly elevated mRNA expression levels and protein levels of TSLP in nasal epithelial cells. Stimulation by TSLP or IL-33 significantly elevated mRNA expression levels of CST1 in nasal epithelial cells. Stimulation of CST1 significantly elevated mRNA expression levels of CCL11 and POSTN in nasal fibroblasts. CST1 could amplify eosinophilic infiltration and T-helper cell type 2 inflammation by interacting with epithelial-derived cytokines and fibroblasts on nasal polyps. CST1 may be involved in the pathogenesis of ECRS, and may contribute to the severity and recurrence of CRS with nasal polyps after endoscopic sinus surgery.

Backgrounds. Outcome of childhood malignancy has been improved mostly due to the advances in diagnostic techniques and treatment strategies. While methotrexate (MTX) related polymorphisms have been under investigation in childhood malignancies, many controversial results have been offered. Objectives. To evaluate associations of polymorphisms related MTX metabolisms and clinical course in childhood lymphoid malignancies. Method. Eighty-two acute lymphoblastic leukemia and 21 non-Hodgkin’s lymphoma children were enrolled in this study. Four single nucleotide polymorphisms in 2 genes (MTHFR (rs1801133/c.677C>T/p.Ala222Val and rs1801131/c.1298A>C/p.Glu429Ala) and SLCO1B1 (rs4149056/c.521T>C/p.V174A and rs11045879/c.1865+4846T>C)) were genotyped by Taqman PCR method or direct sequencing. Clinical courses were reviewed retrospectively. Results. No patient who had the AC/CC genotype of rs1801131 (MTHFR) had relapsed or died, in which distribution was statistically different among the AA genotype of rs1801131 (P=0.004). Polymorphisms of SLCO1B1 (rs11045879 and rs4149056) were not correlated with MTX concentrations, adverse events, or disease outcome. Conclusions. Polymorphisms of MTHFR (rs1801131) could be the plausive candidate for prognostic predictor in childhood lymphoid malignancies.

Background Atopy is a phenotypically heterogeneous condition, and the extent to which atopy accounts for asthma is controversial. In this study, we aimed to identify the presence of distinct sensitization patterns to common inhaled allergens and their association with asthma, allergic rhinitis and TSLP genotypes. Methods We studied 1683 adults from Tsukuba, a city in central Japan and 297 adults from Kamishihoro, a cedar-free, birch-dominant town in northern Japan. Levels of total serum IgE and specific IgE antibodies towards 14 major inhaled allergens were measured. With the use of these measures, cluster analysis was applied to classify the subjects' sensitization patterns. We also examined the genetic effects of 2 TSLP functional SNPs on the development of each sensitization pattern. Results In the Tsukuba study, cluster analysis identified four clusters, including "Dust mite dominant", "Multiple pollen", "Cedar dominant", and "Low reactivity". In the Kamishihoro study, "Dust mite dominant", "Multiple pollen" and "Low reactivity" clusters were also identified, but a "Cedar dominant" cluster was not formed. The association with asthma was strongest for the "Dust mite dominant" cluster in both the Tsukuba and the Kamishihoro studies. In never smokers, both SNPs were associated with the "Dust mite dominant" cluster (OR > 1.2). In contrast, in current or past smokers, these alleles were inversely associated with the "Multiple pollen" cluster (OR Conclusions Cluster analysis identified the presence of distinct sensitization patterns to common inhaled allergens. TSLP may cause asthma by promoting innate allergic responses to indoor allergens and this contribution is significantly modified by smoking.

Background The number of patients with eosinophilic chronic rhinosinusitis (ECRS) has been increasing in recent years in Japan. In ECRS, nasal polyps recur immediately after endoscopic sinus surgery. The molecular biological mechanism underlying the refractoriness of ECRS is unclear. Methods Whole-transcriptome analysis with next-generation sequencing (RNA-seq) was conducted to investigate the molecular biological mechanism of ECRS. Real-time PCR, immunohistochemical staining, and immunofluorescence staining were performed to validate the results of RNA-seq. Results RNA-seq analysis revealed that in the nasal polyps of ECRS, the levels of 3 transcripts were elevated significantly and those of 7 transcripts were diminished significantly. Among the genes encoding these transcripts, TRPV3 (transient receptor potential cation channel, subfamily V, member 3) was identified as the only gene that is highly expressed in ECRS nasal polyps but this gene's expression was not previously detected using DNA microarray analysis in peripheral blood eosinophils. TRPV3 is newly identified here as a gene transcribed in ECRS. Our analysis also revealed that TRPV3 was highly expressed in the infiltrating eosinophils and mucosal epithelium of the nasal polyps of ECRS, and further that the more severe the refractoriness was after surgery, the higher the TRPV3 expression was in nasal polyps. Conclusions TRPV3 might play a role in the refractoriness of ECRS. Additional studies are required to evaluate the function of TRPV3 in ECRS.

Background & aims Genetic factors in class II human leukocyte antigen ( HLA ) have been reported to be associated with inter-individual variation in hepatitis B virus (HBV) vaccine response. However, the mechanism underlying the associations remains elusive. In particular, the broad linkage disequilibrium in HLA region complicates the localization of the independent effects of genetic variants. Thus, the present study aimed to identify the most probable causal variations in class II HLA loci involved in the immune response to HBV vaccine. Methods We performed a case-control study to assess whether HLA-DRB1 , - DQB1 , and -DPB1 4-digit alleles were associated with the response to primary HBV vaccination in 574 healthy Japanese students. To identify causative variants, we next assessed independently associated amino acid variants in these loci using conditional logistic regression analysis. Furthermore, to clarify the functional effects of these variants on HLA proteins, we performed computational structural studies. Results HLA-DRB1 ∗ 01:01 , HLA-DRB1 ∗ 08:03 , HLA-DQB1 ∗ 05:01 , and HLA-DPB1 ∗ 04:02 were significantly associated with sufficient response, whereas HLA-DPB1 ∗ 05:01 was associated with poor response. We then identified amino acids independently associated with sufficient response, namely, leucine at position 26 of HLA-DRβ1 and glycine-glycine-proline-methionine at positions 84–87 of HLA-DPβ1. These amino acids were located in antigen-binding pocket 4 of HLA-DR and pocket 1 of HLA-DP, respectively, which are important structures for selective binding of antigenic peptides. In addition, the detected variations in HLA-DP protein were responsible for the differences in the electrostatic potentials of the pocket, which can explain in part the sufficient/poor vaccine responses. Conclusion HLA-DRβ1 position 26 and HLA-DPβ1 positions 84–87 are independently associated with anti-HBs production against HBV vaccine. Our results suggest that HBsAg presentation through these HLA pocket structures plays an important role in the inter-individual variability of HBV vaccination.

Although our previous GWAS failed to identify SNPs associated with pulmonary function at the level of genomewide significance, it did show that the heritability for FEV1/FVC was 41.6% in a Japanese population, suggesting that the heritability of pulmonary function traits can be explained by the additive effects of multiple common SNPs. In addition, our previous study indicated that pulmonary function genes identified in previous GWASs in non-Japanese populations accounted for 4.3% to 12.0% of the entire estimated heritability of FEV1/FVC in a Japanese population. Therefore, given that many loci with individual weak effects may contribute to asthma risk, in this study, we created a quantitative score of genetic load based on 16 SNPs implicated in lower lung function in both Japanese and non-Japanese populations. This genetic risk score (GRS) for lower FEV1/FVC was consistently associated with the onset of asthma (P = 9.6 × 10(-4)) in 2 independent Japanese populations as well as with the onset of COPD (P = 0.042). Clustering of asthma patients based on GRS levels indicated that an increased GRS may be responsible for the development of a particular phenotype of asthma characterized by early onset, atopy, and severer airflow obstruction.