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A genome wide association study reported that the T allele of rs2294008 in a cancer-related gene, PSCA, is a risk allele for diffuse-type gastric cancer. This allele has the highest frequency (0.63) in Japanese in Tokyo (JPT) among 26 populations in the 1000 Genomes Project database. FST &asymp, 0.26 at this single nucleotide polymorphism is one of the highest between JPT and the genetically close Han Chinese in Beijing (CHB). To understand the evolutionary history of the alleles in PSCA, we addressed: (i) whether the C non-risk allele at rs2294008 is under positive selection, and (ii) why the mainland Japanese population has a higher T allele frequency than other populations. We found that haplotypes harboring the C allele are composed of two subhaplotypes. We detected that positive selection on both subhaplotypes has occurred in the East Asian lineage. However, the selection on one of the subhaplotypes in JPT seems to have been relaxed or ceased after divergence from the continental population, this may have caused the elevation of T allele frequency. Based on simulations under the dual structure model (a specific demography for the Japanese) and phylogenetic analysis with ancient DNA, the T allele at rs2294008 might have had high frequency in the Jomon people (one of the ancestral populations of the modern Japanese), this may explain the high T allele frequency in the extant Japanese.

To understand the genetics of steroid-sensitive nephrotic syndrome (SSNS), we conducted a genome-wide association study in 987 childhood SSNS patients and 3,206 healthy controls with Japanese ancestry. Beyond known associations in the HLA-DR/DQ region, common variants in NPHS1-KIRREL2 (rs56117924, P=4.94E-20, odds ratio (OR) =1.90) and TNFSF15 (rs6478109, P=2.54E-8, OR=0.72) regions achieved genome-wide significance and were replicated in Korean, South Asian and African populations. Trans-ethnic meta-analyses including Japanese, Korean, South Asian, African, European, Hispanic and Maghrebian populations confirmed the significant associations of variants in NPHS1-KIRREL2 (P(meta)=6.71E-28, OR=1.88) and TNFSF15 (P(meta)=5.40E-11, OR=1.33) loci. Analysis of the NPHS1 risk alleles with glomerular NPHS1 mRNA expression from the same person revealed allele specific expression with significantly lower expression of the transcript derived from the risk haplotype (Wilcox test p=9.3E-4). Because rare pathogenic variants in NPHS1 cause congenital nephrotic syndrome of the Finnish type (CNSF), the present study provides further evidence that variation along the allele frequency spectrum in the same gene can cause or contribute to both a rare monogenic disease (CNSF) and a more complex, polygenic disease (SSNS).

Background & Aims Primary biliary cholangitis (PBC) is a chronic and cholestatic liver disease that eventually leads to cirrhosis and hepatic failure. We recently identified several susceptibility genes included NFKB1 and MANBA for PBC in the Japanese population by genome-wide association study. However, the primary functional variants in the NFKB1/MANBA region and the molecular mechanism for conferring disease susceptibility to PBC have not yet been clarified. Methods We performed high-density association mapping based on a single-nucleotide polymorphism (SNP) imputation analysis, using data from a whole-genome sequence reference panel of 1070 Japanese individuals and the previous genome-wide association study (1389 PBC patients, 1508 healthy controls). Among SNPs (P < 5.0 × 10-7) in the NFKB1/MANBA region, putative primary functional variants and the molecular mechanism for conferring disease susceptibility to PBC were identified by in silico/in vitro functional analysis. Results Among the SNPs in the NFKB1/MANBA region, rs17032850 and rs227361, which changed the binding of transcription factors lymphoid enhancer-binding factor 1 (LEF-1) and retinoid X receptor α (RXRα), respectively, were identified as putative primary functional variants that regulate gene expression. In addition, expression-quantitative trait locus data and gene editing using a clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 system supported the potential role of rs17032850 and rs227361 in regulating NFKB1 and MANBA expression, respectively. Conclusions We identified independent putative primary functional variants in NFKB1/MANBA and showed the distinct molecular mechanism by which each putative primary functional variant conferred susceptibility to PBC. Our approach was useful to dissect the pathogenesis not only of PBC, but also other digestive diseases in which NFKB1/MANBA has been reported as a susceptibility locus., Graphical abstract

The Eurasian house mouseMus musculusis useful for tracing prehistorical human movement related to the spread of farming. We determined whole mitochondrial DNA (mtDNA) sequences (ca. 16,000 bp) of 98 wild-derived individuals of two subspecies,M.m.musculus(MUS) andM.m.castaneus(CAS). We revealed directional dispersals reaching as far as the Japanese Archipelago from their homelands. Our phylogenetic analysis indicated that the eastward movement of MUS was characterised by five step-wise regional extension events: (1) broad spatial expansion into eastern Europe and the western part of western China, (2) dispersal to the eastern part of western China, (3) dispersal to northern China, (4) dispersal to the Korean Peninsula and (5) colonisation and expansion in the Japanese Archipelago. These events were estimated to have occurred during the last 2000–18,000 years. The dispersal of CAS was characterised by three events: initial divergences (ca. 7000–9000 years ago) of haplogroups in northernmost China and the eastern coast of India, followed by two population expansion events that likely originated from the Yangtze River basin to broad areas of South and Southeast Asia, including Sri Lanka, Bangladesh and Indonesia (ca. 4000–6000 years ago) and to Yunnan, southern China and the Japanese Archipelago (ca. 2000–3500). This study provides a solid framework for the spatiotemporal movement of the human-associated organisms in Holocene Eastern Eurasia using whole mtDNA sequences, reliable evolutionary rates and accurate branching patterns. The information obtained here contributes to the analysis of a variety of animals and plants associated with prehistoric human migration.

Land plants produce diverse flavonoids for growth, survival, and reproduction. Chalcone synthase is the first committed enzyme of the flavonoid biosynthetic pathway and catalyzes the production of 2′,4,4′,6′-tetrahydroxychalcone (THC). However, it also produces other polyketides, including p-coumaroyltriacetic acid lactone (CTAL), because of the derailment of the chalcone-producing pathway. This promiscuity of CHS catalysis adversely affects the efficiency of flavonoid biosynthesis, although it is also believed to have led to the evolution of stilbene synthase and p-coumaroyltriacetic acid synthase. In this study, we establish that chalcone isomerase-like proteins (CHILs), which are encoded by genes that are ubiquitous in land plant genomes, bind to CHS to enhance THC production and decrease CTAL formation, thereby rectifying the promiscuous CHS catalysis. This CHIL function has been confirmed in diverse land plant species, and represents a conserved strategy facilitating the efficient influx of substrates from the phenylpropanoid pathway to the flavonoid pathway., Chalcone synthase is the first committed enzyme in the plant flavonoid biosynthesis pathway, yet shows low product specificity in vitro. Here Waki et al. show that chalcone isomerase-like proteins bind to and reduce the catalytic promiscuity of chalcone synthase, ensuring efficient flavonoid production in planta.

In time-series analysis, state-space models (SSMs) have been widely used to estimate the conditional probability distributions of hidden variables and parameter values, as well as to understand structures that can generate the data. For example, the Kalman filter is used to analytically calculate the conditional probability distribution on linear SSMs in terms of minimizing the variance, and several extensions, such as the unscented Kalman filter and particle filter, have been applied to calculate the approximate distribution on nonlinear SSMs. Recently, the approximate Bayesian computation (ABC) has been applied to such time-series filtering to handle intractable likelihoods; however, it remains problematic with respect to consistently achieving a reduction of the estimation bias, evaluating the validity of the models, and dealing with replicated observations. To address these problems, in this paper, we propose a novel method combined with the kernel ABC to perform filtering, parameter estimation, and the model evaluation in SSMs. Simulation studies show that the proposed method produces inference that is comparable to other ABC methods, with the advantage of not requiring a careful calibration of the ABC threshold. In addition, we evaluate the performance of the model-selection capability using true and competitive models on synthetic data from nonlinear SSMs. Finally, we apply the proposed method to real data in rat circadian oscillations, and demonstrated the usefulness in practical situations.

The ninth Japan Bioanalysis Forum symposium took place at tower hall Funabori, Tokyo, Japan, between 6 and 8 February, 2018. Bioanalytical scientists from the pharmaceutical industry, CROs, academia and regulatory bodies had many meaningful and relevant discussions on current topics of interest in bioanalysis. The 3-day symposium featured updated perspectives and experiences on regulated bioanalysis of small and large molecules, biomarker measurement and assessment of immunogenicity, as well as new areas of bioanalytical validation such as quantitative polymerase chain reaction(qPCR) and flow cytometry. There were over 260 participants from six countries, with 23 oral and 11 poster presentations, including the outcomes of Japan Bioanalysis Forum discussion groups. This report summarizes the major discussion topics from the conference.

Background Nephrotic syndrome is the most common cause of chronic glomerular disease in children. Most of these patients develop steroid-sensitive nephrotic syndrome (SSNS), but the loci conferring susceptibility to childhood SSNS are mainly unknown.Methods We conducted a genome-wide association study (GWAS) in the Japanese population; 224 patients with childhood SSNS and 419 adult healthy controls were genotyped using the Affymetrix Japonica Array in the discovery stage. Imputation for six HLA genes (HLA-A, -C, -B, -DRB1, -DQB1, and -DPB1) was conducted on the basis of Japanese-specific references. We performed genotyping for HLA-DRB1/-DQB1 using a sequence-specific oligonucleotide-probing method on a Luminex platform. Whole-genome imputation was conducted using a phased reference panel of 2049 healthy Japanese individuals. Replication was performed in an independent Japanese sample set including 216 patients and 719 healthy controls. We genotyped candidate single-nucleotide polymorphisms using the DigiTag2 assay.Results The most significant association was detected in the HLA-DR/DQ region and replicated (rs4642516 [minor allele G], combined Pallelic=7.84×10-23; odds ratio [OR], 0.33; 95% confidence interval [95% CI], 0.26 to 0.41; rs3134996 [minor allele A], combined Pallelic=1.72×10-25; OR, 0.29; 95% CI, 0.23 to 0.37). HLA-DRB1*08:02 (Pc=1.82×10-9; OR, 2.62; 95% CI, 1.94 to 3.54) and HLA-DQB1*06:04 (Pc=2.09×10-12; OR, 0.10; 95% CI, 0.05 to 0.21) were considered primary HLA alleles associated with childhood SSNS. HLA-DRB1*08:02-DQB1*03:02 (Pc=7.01×10-11; OR, 3.60; 95% CI, 2.46 to 5.29) was identified as the most significant genetic susceptibility factor.Conclusions The most significant association with childhood SSNS was detected in the HLA-DR/DQ region. Further HLA allele/haplotype analyses should enhance our understanding of molecular mechanisms underlying SSNS.

Background Despite NUDT15 variants showing significant association with thiopurine-induced adverse events (AEs) in Asians, it remains unclear which variants of NUDT15 or whether additional genetic variants should be tested to predict AEs. To clarify the best pharmacogenetic test to be used clinically, we performed association studies of NUDT15 variants and haplotypes with AEs, genome-wide association study (GWAS) to discover additional variants, and ROC analysis to select the model to predict severe AEs. Methods Overall, 2630 patients with inflammatory bowel disease (IBD) were enrolled and genotyped for NUDT15 codon 139; 1291 patients were treated with thiopurines. diplotypes were analyzed in 970 patients, and GWASs of AEs were performed with 1221 patients using population-optimized genotyping array and imputation. Results We confirmed the association of NUDT15 p.Arg139Cys with leukopenia and alopecia (p = 2.20E−63, 1.32E−69, OR = 6.59, 12.1, respectively), and found a novel association with digestive symptoms (p = 6.39E−04, OR = 1.89). Time to leukopenia was significantly shorter, and when leukopenia was diagnosed, thiopurine doses were significantly lower in Arg/Cys and Cys/Cys than in Arg/Arg. In GWASs, no additional variants were found to be associated with thiopurine-induced AEs. Despite strong correlation of leukopenia frequency with estimated enzyme activities based on the diplotypes (r2 = 0.926, p = 0.0087), there were no significant differences in the AUCs of diplotypes from those of codon 139 to predict severe AEs (AUC = 0.916, 0.921, for acute severe leukopenia, AUC = 0.990, 0.991, for severe alopecia, respectively). Conclusions Genotyping of NUDT15 codon 139 was sufficient to predict acute severe leukopenia and alopecia in Japanese patients with IBD. Electronic supplementary material The online version of this article (10.1007/s00535-018-1486-7) contains supplementary material, which is available to authorized users.

Background and aim Patients with inflammatory bowel disease (IBD) are at a high risk of low bone mineral density (BMD). Reportedly, clinical and genetic factors cause low BMD in Caucasians; however, studies in non-Caucasian populations remain scarce. Methods Clinical risk factors for low BMD were investigated in 266 Japanese patients with IBD, and a genome-wide association analysis (GWAS) was performed using linear regression with associated clinical factors as covariates. Genotyping was performed using a population-optimized genotyping array (Japonica array® ). After quality control, the genotype data of 4 384 682 single-nucleotide polymorphisms (SNPs) from 254 patients with IBD were used for GWAS. Results Body mass index, age, and disease duration were independently associated with the BMD of the femoral neck (P = 1.41E - 13, 1.04E - 5, and 1.58E - 3, respectively), and body mass index and sex were associated with the BMD of the lumbar spine (P = 6.90E - 10 and 6.84E - 3, respectively). In GWAS, 118 and 42 candidate SNPs of the femoral neck and lumbar spine, respectively, were identified. Among 118, 111 candidate SNPs of the femoral neck were located within the SLC22A23 gene, which is a known IBD susceptibility gene (minimum P = 1.42E - 07). Among 42, 18 candidate SNPs of the lumbar spine were located within the MECOM gene, which is associated with osteopenia (minimum P = 5.86E - 07). Interestingly, none of the known loci showed a significant association with BMD. Conclusions Although clinical risk factors for low BMD in IBD were similar to those in the general population, genetic risk factors were rather different.

Approximately 10–20% of patients with primary biliary cholangitis (PBC) progress to jaundice stage regardless of treatment with ursodeoxycholic acid and bezafibrate. In this study, we performed a GWAS and a replication study to identify genetic variants associated with jaundice-stage progression in PBC using a total of 1,375 patients (1,202 early-stage and 173 jaundice-stage) in a Japanese population. SNP rs13720, which is located in the 3′UTR of cathepsin Z (CTSZ), showed the strongest association (odds ratio [OR] = 2.15, P = 7.62 × 10−7) with progression to jaundice stage in GWAS. High-density association mapping at the CTSZ and negative elongation factor complex member C/D (NELFCD) loci, which are located within a strong linkage disequilibrium (LD) block, revealed that an intronic SNP of CTSZ, rs163800, was significantly associated with jaundice-stage progression (OR = 2.16, P = 8.57 × 10−8). In addition, eQTL analysis and in silico functional analysis indicated that genotypes of rs163800 or variants in strong LD with rs163800 influence expression levels of both NELFCD and CTSZ mRNA. The present novel findings will contribute to dissect the mechanism of PBC progression and also to facilitate the development of therapies for PBC patients who are resistant to current therapies.

Two different types of data acquisition methods, "averaging mode" and "ion-counting mode", have been used in a time-of-flight (TOF) mass spectrometry. The most common method is an averaging mode that sums waveform signals obtained from each flight cycle. While it is possible to process many ions arriving at the same TOF in one flight cycle, low-abundance ions are difficult to measure because ion signals are overwhelmed by noises from the detection system. An ion-counting mode is suitable for the detection of such low-concentration ions, but counting loss occurs when two or more ions arrive at the detector within the dead time of the acquisition system. In this study, we introduce a technique that combines two methods to measure target ions with a high concentration difference, i.e., averaging mode and ion-counting mode are used simultaneously for high abundant and trace ions, respectively. By processing waveforms concurrently during data acquisition, one can choose to analyze either or both types of data to achieve a highly quantitative mass spectrum over a wide range of sample concentrations. The result of the argon isotope analysis shows that this method provides a more accurate determination of the isotope ratio compared to averaging mode alone at one-twentieth of the analysis time required by ion-counting alone. Graphical Abstract ᅟ.

Organic anion transporter 10 (OAT10), also known as SLC22A13, has hitherto been identified as a urate transporter by in vitro analyses.1 Despite the reported expression of OAT10 on the apical membrane of the renal proximal tubular cells,1 the physiological impact of OAT10 on urate handling in humans remains to be elucidated. Accumulating evidence suggests that functional variants of already-characterised, physiologically important urate transporters—URAT1/SLC22A12, GLUT9/SLC2A9, BCRP/ABCG2 and NPT1/SLC17A1—affect serum uric acid (SUA) levels and susceptibility of gout,2–6 the most common form of inflammatory arthritis. However, there are no reports on the association between OAT10 gene and either hyperuricaemia or gout. Here, for the first time, we reveal that a dysfunctional variant of OAT10 decreases both gout risk and SUA levels, suggesting OAT10 to be physiologically involved in urate reabsorption in the human kidney, as described below. To explore exonic variants in OAT10 potentially associated with gout susceptibility, we sequenced all exons of OAT10 in 480 gout cases and 480 controls of Japanese male6 and conducted an association analysis (see online supplementary tables S1 and S2), followed by a replication study on 924 gout cases and 2113 controls (see online supplementary figure S1). In two identified OAT10 variants with minor allele frequency (MAF) >0.5%, only rs117371763 (c.1129C>T; p.Arg377Cys [R377C]) was significantly associated with gout susceptibility after Bonferroni correction (p=0.014). The significant association between rs117371763 and gout susceptibility was replicated, and our meta-analysis showed a significant protective effect of rs117371763 on gout susceptibility (OR=0.67; 95% CI 0.53 to 0.85; pmeta …

Primary biliary cholangitis (PBC) is a chronic cholestatic autoimmune liver disease that appears to be strongly influenced by genetic factors. Recently, an international meta-analysis of genome-wide association studies (GWAS) identified CC-Motif Chemokine Receptor-6 (CCR6) and FGFR1 Oncogene-Partner (FGFR1OP) as PBC-susceptibility genes. However, the lead single nucleotide polymorphisms (SNPs) of CCR6/FGFR1OP showed low linkage disequilibrium with each other in East Asian and European populations. Additionally, the primary functional variants and the molecular mechanisms responsible for PBC-susceptibility remain unclear. Here, among the PBC-susceptibility SNPs identified by high-density association mapping in our previous meta-GWAS (Patients: n = 10,516; healthy controls: n = 20,772) within the CCR6/FGFR1OP locus, rs9459874 and rs1012656 were identified as primary functional variants. These functional variants accounted for the effects of GWAS-identified lead SNPs in CCR6/FGFR1OP. Additionally, the roles of rs9459874 and rs1012656 in regulating FGFR1OP transcription and CCR6 translation, respectively, were supported by expression quantitative trait loci (eQTL) analysis and gene editing technology using the CRISPR/Cas9 system. Immunohistochemistry showed higher expression of CCR6 protein in the livers of patients with PBC than in those of a non-diseased control. In conclusion, we identified primary functional variants in CCR6/FGFR1OP and revealed the molecular mechanisms by which these variants confer PBC-susceptibility in an eQTL-dependent or -independent manner. The approach in this study is applicable for the elucidation of the pathogenesis of other autoimmune disorders in which CCR6/FGFR1OP is known as a susceptibility locus, as well as PBC.

Clarifying allele frequencies of disease-related genetic variants in a population is important in genomic medicine; however, such data is not yet available for the Japanese population. To estimate frequencies of actionable pathogenic variants in the Japanese population, we examined the reported pathological variants in genes recommended by the American College of Medical Genetics and Genomics (ACMG) in our reference panel of genomic variations, 2KJPN, which was created by whole-genome sequencing of 2049 individuals of the resident cohort of the Tohoku Medical Megabank Project. We searched for pathogenic variants in 2KJPN for 57 autosomal ACMG-recommended genes responsible for 26 diseases and then examined their frequencies. By referring to public databases of pathogenic variations, we identified 143 reported pathogenic variants in 2KJPN for the 57 ACMG recommended genes based on a classification system. At the individual level, 21% of the individuals were found to have at least one reported pathogenic allele. We then conducted a literature survey to review the variants and to check for evidence of pathogenicity. Our results suggest that a substantial number of people have reported pathogenic alleles for the ACMG genes, and reviewing variants is indispensable for constructing the information infrastructure of genomic medicine for the Japanese population.

Primary open-angle glaucoma (POAG) can develop even within normal ranges of intraocular pressure, and this type of glaucoma (so-called ‘normal-tension glaucoma [NTG]’) is highly prevalent in East Asia including Korea and Japan. We conducted exome chip analysis to identify low-frequency and rare variants associated with POAG from the primary cohort (309 POAG patients and 5,400 control, all Koreans). For replication, Korean (310 POAG patients and 5,612 controls) and Japanese (565 POAG patients and 1,104 controls) cohorts were further investigated by targeted genotyping. SNP rs116121322 in LRRC27 showed nominally significant association with POAG in the discovery cohort (OR = 29.85, P = 2E–06). This SNP was validated in the Korean replication cohort but only in the NTG subgroups (OR = 9.86, P = 0.007). Japanese replication cohort did not show significant association with POAG (P .00.44). However, the meta-analysis in the entire cohort revealed significant association of rs116121322 with POAG (ORcombined = 10.28, Pcombined = 1.4E–07). The LRRC27 protein expression was confirmed from human trabecular meshwork cells. For gene-based testing, METTL20 showed a significant association in POAG (Pcombined = 0.002) and in the subgroup of NTG (Pcombined = 0.02), whereas ZNF677 were significantly associated with only in the subgroup of high-tension glaucoma (Pcombined = 1.5E–06). Our findings may provide further genetic backgrounds into the pathogenesis of POAG, especially for the patients who have lower baseline intraocular pressures.

Nontuberculous mycobacteria (NTM) are environmental mycobacteria that can cause a chronic progressive lung disease. Although epidemiological data indicate potential genetic predisposition, its nature remains unclear.We aimed to identify host susceptibility loci forThis genome-wide association study (GWAS) was conducted in Japanese patients with pulmonary MAC and healthy controls, followed by genotyping of candidate single-nucleotide polymorphisms (SNPs) in another Japanese cohort. For verification by Korean and European ancestry, we performed SNP genotyping.The GWAS discovery set included 475 pulmonary MAC cases and 417 controls. Both GWAS and replication analysis of 591 pulmonary MAC cases and 718 controls revealed the strongest association with chromosome 16p21, particularly with rs109592 (p=1.64×10We identified rs109592 in the

Background To clarify the genetic background of ulcerative colitis (UC) in the Japanese population, we conducted a genome-wide association study (GWAS) using a population-specific single nucleotide polymorphism (SNP) array. Methods We performed a GWAS and replication study including 1676 UC patients and 2381 healthy controls. The probability of colectomy was compared between genotypes of rs117506082, the top hit SNP at HLA loci, by the Kaplan-Meier method. We studied serum expression of miR-622, a newly identified candidate gene, from 32 UC patients and 8 healthy controls by quantitative reverse-transcription polymerase chain reaction. Results In the GWAS, only the HLA loci showed genome-wide significant associations with UC (rs117506082, P = 6.69E-28). Seven nominally significant regions included 2 known loci, IL23R (rs76418789, P = 6.29E-7) and IRF8 (rs16940202, P = 1.03E-6), and 5 novel loci: MIR622 (rs9560575, P = 8.23E-7), 14q31 (rs117618617, P = 1.53E-6), KAT6B (rs12260609, P = 1.81E-6), PAX3-CCDC140-SGPP2 (rs7589797, P = 2.87E-6), and KCNA2 (rs118020656, P = 4.01E-6). Combined analysis revealed that IL23R p.G149R (rs76418789, P = 9.03E-11; odds ratio [OR], 0.51) had genome-wide significant association with UC. Patients with GG genotype of rs117506082 had a significantly lower probability of total colectomy than those with the GA+AA genotype (P = 1.72E-2). Serum expression of miR-622 in patients with inactive UC tended to be higher than in healthy controls and patients with active UC (inactive UC vs healthy controls, P = 3.03E-02; inactive UC vs active UC, P = 6.44E-02). Conclusions IL23R p.G149R is a susceptibility locus for UC in Japanese individuals. The GG genotype of rs117506082 at HLA loci may predict a better clinical course.

Gout is a common arthritis caused by monosodium urate crystals. The heritability of serum urate levels is estimated to be 30–70%; however, common genetic variants account for only 7.9% of the variance in serum urate levels. This discrepancy is an example of “missing heritability.” The “missing heritability” suggests that variants associated with uric acid levels are yet to be found. By using genomic sequences of the ToMMo cohort, we identified rare variants of the SLC22A12 gene that affect the urate transport activity of URAT1. URAT1 is a transporter protein encoded by the SLC22A12 gene. We grouped the participants with variants affecting urate uptake by URAT1 and analyzed the variance of serum urate levels. The results showed that the heritability explained by the SLC22A12 variants of men and women exceeds 10%, suggesting that rare variants underlie a substantial portion of the “missing heritability” of serum urate levels.

Background: Kawasaki disease (KD) is a systemic vasculitis which may be associated with coronary artery aneurysms. A notable risk factor for the development of coronary artery aneurysms is resistance to intravenous immunoglobulin (IVIG) therapy, which comprises standard treatment for the acute phase of KD. The cause of IVIG resistance in KD is largely unknown; however, the contribution of genetic factors, especially variants in immune-related genes, has been suspected. Methods: To explore genetic variants related to IVIG-unresponsiveness, we designated KD patients who did not respond to both first and second courses of IVIG therapy as IVIG-unresponsive patients. Using genomic DNA from 30 IVIG-unresponsive KD patients, we performed pooled genome sequencing targeting 39 immune-related cytokine receptor genes. Results: The single nucleotide variant (SNV), rs563535954 (located in the IL4R locus), was concentrated in IVIG-unresponsive KD patients. Individual genotyping showed that the minor allele of rs563535954 was present in 4/33 patients with IVIG-unresponsive KD, compared with 20/1063 individuals in the Japanese genome variation database (odds ratio = 7.19, 95% confidence interval 2.43-21.47). Furthermore, the minor allele of rs563535954 was absent in 42 KD patients who responded to IVIG treatment (P = 0.0337), indicating that a low-frequency variant, rs563535954, is associated with IVIG-unresponsiveness in KD patients. Although rs563535954 is located in the 3'-untranslated region of IL4R, there was no alternation in IL4R expression associated with the mior allele of rs563535954. However, IVIG-unresponsive patients that exhibited the minor allele of rs563535954 tended to be classified into the low-risk group (based on previously reported risk scores) for prediction of IVIG-resistance. Therefore, IVIG-unresponsiveness associated with the minor allele of rs563535954 might differ from IVIG-unresponsiveness associated with previous risk factors used to evaluate IVIG-unresponsiveness in KD. Conclusion: These findings suggest that the SNV rs563535954 could serve as a predictive indicator of IVIG-unresponsiveness, thereby improving the sensitivity of risk scoring systems, and may aid in prevention of coronary artery lesions in KD patients., Article, PEDIATRIC RHEUMATOLOGY.17:34(2019)

RationaleNontuberculous mycobacteria (NTM) are environmental mycobacteria that can cause a chronic progressive lung disease. Although epidemiological data indicate potential genetic predisposition, its nature remains unclear.ObjectivesWe aimed to identify host susceptibility loci for Mycobacterium avium complex (MAC), the most common NTM pathogen.MethodsThis genome-wide association study (GWAS) was conducted in Japanese patients with pulmonary MAC and healthy controls, followed by genotyping of candidate single-nucleotide polymorphisms (SNPs) in another Japanese cohort. For verification by Korean and European ancestry, we performed SNP genotyping.ResultsThe GWAS discovery set included 475 pulmonary MAC cases and 417 controls. Both GWAS and replication analysis of 591 pulmonary MAC cases and 718 controls revealed the strongest association with chromosome 16p21, particularly with rs109592 (p=1.64×10−13, OR 0.54), which is in an intronic region of the calcineurin-like EF-hand protein 2 (CHP2). Expression quantitative trait loci analysis demonstrated an association with lung CHP2 expression. CHP2 was expressed in the lung tissue in pulmonary MAC disease. This SNP was associated with the nodular bronchiectasis subtype. Additionally, this SNP was significantly associated with the disease in patients of Korean (p=2.18×10−12, OR 0.54) and European (p=5.12×10−03, OR 0.63) ancestry.ConclusionsWe identified rs109592 in the CHP2 locus as a susceptibility marker for pulmonary MAC disease.

Case 1: A 75-year-old man underwent distal gastrectomy with Billroth Ⅰ reconstruction and resection of the involved transverse mesocolon. Microscopic examination revealed adenocarcinoma(tub2, tub1), pT4b(SI)N3M0, pStageⅢc. Adjuvant chemotherapy with S-1 was performed for 6 months after the operation. One year later, CT revealed a localized dissemination in the transverse mesocolon; therefore, we performed transverse colectomy. Adjuvant chemotherapy with PTX was performed, and the patient remains free from recurrence 7 years after the initialoperation. Case 2: A 65-year-old man was diagnosed with gastric scirrhous carcinoma by esophagogastroduodenoscopy. CT and colonoscopy showed a tumorous lesion in the pelvis(Schnitzler's metastasis). Neo-adjuvant chemotherapy with S-1 plus CDDP was performed. After 6 courses, CT and endoscopy showed shrinkage of the tumors, and no other distant metastasis was detected by PET-CT. We performed totalgastrectomy(D2), splenectomy, and low anterior resection of the rectum simultaneously. Microscopic examination revealed adenocarcinoma(tub2, por2, sig), pT4a(SE)N0, and the histological response was Grade 1a. S-1 was administered, and the patient has had no recurrence in the 1 year 6 months after the operation. Dissemination of gastric cancer tends to be difficult to treat and has a poor prognosis. However, in some cases, the proper combination of chemotherapy and surgery might be beneficial for long survival.

Article, Scientific reports.9 :102(2019)

BACKGROUND Inflammatory bowel disease (IBD) has an unknown etiology; however, accumulating evidence suggests that IBD is a multifactorial disease influenced by a combination of genetic and environmental factors. The influence of genetic variants on DNA methylation in cis and cis effects on expression have been demonstrated. We hypothesized that IBD susceptibility single-nucleotide polymorphisms (SNPs) regulate susceptibility gene expressions in cis by regulating DNA methylation around SNPs. For this, we determined cis-regulated allele-specific DNA methylation (ASM) around IBD susceptibility genes in CD4+ effector/memory T cells (Tem) in lamina propria mononuclear cells (LPMCs) in patients with IBD and examined the association between the ASM SNP genotype and neighboring susceptibility gene expressions. METHODS CD4+ effector/memory T cells (Tem) were isolated from LPMCs in 15 Japanese IBD patients (ten Crohn's disease [CD] and five ulcerative colitis [UC] patients). ASM analysis was performed by methylation-sensitive SNP array analysis. We defined ASM as a changing average relative allele score ([Formula: see text]) >0.1 after digestion by methylation-sensitive restriction enzymes. Among SNPs showing [Formula: see text] >0.1, we extracted the probes located on tag-SNPs of 200 IBD susceptibility loci and around IBD susceptibility genes as candidate ASM SNPs. To validate ASM, bisulfite-pyrosequencing was performed. Transcriptome analysis was examined in 11 IBD patients (seven CD and four UC patients). The relation between rs36221701 genotype and neighboring gene expressions were analyzed. RESULTS We extracted six candidate ASM SNPs around IBD susceptibility genes. The top of [Formula: see text] (0.23) was rs1130368 located on HLA-DQB1. ASM around rs36221701 ([Formula: see text] = 0.14) located near SMAD3 was validated using bisulfite pyrosequencing. The SMAD3 expression was significantly associated with the rs36221701 genotype (p = 0.016). CONCLUSIONS We confirmed the existence of cis-regulated ASM around IBD susceptibility genes and the association between ASM SNP (rs36221701) genotype and SMAD3 expression, a susceptibility gene for IBD. These results give us supporting evidence that DNA methylation mediates genetic effects on disease susceptibility.

We report a family with progressive myoclonic epilepsy who underwent whole-exome sequencing but was negative for pathogenic variants. Similar clinical courses of a devastating neurodegenerative phenotype of two affected siblings were highly suggestive of a genetic etiology, which indicates that the survey of genetic variation by whole-exome sequencing was not comprehensive. To investigate the presence of a variant that remained unrecognized by standard genetic testing, PacBio long-read sequencing was performed. Structural variant (SV) detection using low-coverage (6×) whole-genome sequencing called 17,165 SVs (7,216 deletions and 9,949 insertions). Our SV selection narrowed down potential candidates to only five SVs (two deletions and three insertions) on the genes tagged with autosomal recessive phenotypes. Among them, a 12.4-kb deletion involving the CLN6 gene was the top candidate because its homozygous abnormalities cause neuronal ceroid lipofuscinosis. This deletion included the initiation codon and was found in a GC-rich region containing multiple repetitive elements. These results indicate the presence of a causal variant in a difficult-to-sequence region and suggest that such variants that remain enigmatic after the application of current whole-exome sequencing technology could be uncovered by unbiased application of long-read whole-genome sequencing.

Background and Aims Genome-wide association studies [GWASs] of European populations have identified numerous susceptibility loci for Crohn’s disease [CD]. Susceptibility genes differ by ethnicity, however, so GWASs specific for Asian populations are required. This study aimed to clarify the Japanese-specific genetic background for CD by a GWAS using the Japonica array [JPA] and subsequent imputation with the 1KJPN reference panel. Methods Two independent Japanese case/control sets (Tohoku region [379 CD patients, 1621 controls] and Kyushu region [334 CD patients, 462 controls]) were included. GWASs were performed separately for each population, followed by a meta-analysis. Two additional replication sets [254 + 516 CD patients and 287 + 565 controls] were analysed for top hit single nucleotide polymorphisms [SNPs] from novel genomic regions. Results Genotype data of 4 335 144 SNPs from 713 Japanese CD patients and 2083 controls were analysed. SNPs located in TNFSF15 (rs78898421, Pmeta = 2.59 × 10−26, odds ratio [OR] = 2.10), HLA-DQB1 [rs184950714, pmeta = 3.56 × 10−19, OR = 2.05], ZNF365, and 4p14 loci were significantly associated with CD in Japanese individuals. Replication analyses were performed for four novel candidate loci [p Conclusions RAP1A is a novel susceptibility locus for CD in the Japanese population.

Breast cancer is the most common cancer and the second-leading cause of cancer-related death among women. Inconsistent findings for the relationship between melatonin levels, sleep duration and breast cancer have been reported. We investigated the association of sleep duration at cohort entry and its interaction with body mass index (BMI) with risk of developing breast cancer in the large population-based Multiethnic Cohort study. Among the 74,481 at-risk participants, 5,790 breast cancer cases were identified during the study period. Although we detected no significant association between sleep duration and breast cancer incidence, higher risk estimates for short (HR = 1.03; 95% CI: 0.97-1.09) and long sleep (HR = 1.05; 95% CI: 0.95-1.15) compared to normal sleep (7-8 hr) were found. The patterns for models stratified by age, BMI, ethnicity and hormone receptor status were similar but did not indicate significant interaction effects. When examining the combined sleep duration and BMI interaction effect, in comparison to the normal BMI-normal sleep group, risk estimates for underweight, overweight and obesity were similar across categories of sleep duration (≤6, 7-8, and ≥9 hr). The underweight-normal sleep group had lower breast cancer incidence (HR = 0.66, 95% CI: 0.50-0.86), whereas the overweight-short sleep, overweight-normal sleep group and all obese women experienced elevated breast cancer incidence. The respective HRs for short, normal and long sleep among obese women were 1.35 (95% CI: 1.20-1.53), 1.27 (95% CI: 1.15-1.42) and 1.46 (95% CI: 1.21-1.76). Future perspectives need to examine the possibility that sleep quality, variations in circadian rhythm and melatonin are involved in breast cancer etiology.

Background Genotype imputation from single-nucleotide polymorphism (SNP) genotype data using a haplotype reference panel consisting of thousands of unrelated individuals from populations of interest can help to identify strongly associated variants in genome-wide association studies. The Tohoku Medical Megabank (TMM) project was established to support the development of precision medicine, together with the whole-genome sequencing of 1070 human genomes from individuals in the Miyagi region (Northeast Japan) and the construction of the 1070 Japanese genome reference panel (1KJPN). Here, we investigated the performance of 1KJPN for genotype imputation of Japanese samples not included in the TMM project and compared it with other population reference panels. Results We found that the 1KJPN population was more similar to other Japanese populations, Nagahama (south-central Japan) and Aki (Shikoku Island), than to East Asian populations in the 1000 Genomes Project other than JPT, suggesting that the large-scale collection (more than 1000) of Japanese genomes from the Miyagi region covered many of the genetic variations of Japanese in mainland Japan. Moreover, 1KJPN outperformed the phase 3 reference panel of the 1000 Genomes Project (1KGPp3) for Japanese samples, and IKJPN showed similar imputation rates for the TMM and other Japanese samples for SNPs with minor allele frequencies (MAFs) higher than 1%. Conclusions 1KJPN covered most of the variants found in the samples from areas of the Japanese mainland outside the Miyagi region, implying 1KJPN is representative of the Japanese population’s genomes. 1KJPN and successive reference panels are useful genome reference panels for the mainland Japanese population. Importantly, the addition of whole genome sequences not included in the 1KJPN panel improved imputation efficiencies for SNPs with MAFs under 1% for samples from most regions of the Japanese archipelago. Electronic supplementary material The online version of this article (10.1186/s12864-018-4942-0) contains supplementary material, which is available to authorized users.

The associations of single nucleotide polymorphisms (SNPs) in PLA2R1 and HLA-DQA1, as well as HLA-DRB1*15:01-DQB1*06:02 haplotype with idiopathic membranous nephropathy (IMN) is well known. However, the primary associations of these loci still need to be determined. We used Japanese-specific SNP genotyping array and imputation using 2,048 sequenced Japanese samples to fine-map PLA2R1 region in 98 patients and 413 controls. The most significant SNPs were replicated in a separate sample set of 130 patients and 288 controls. A two-SNP haplotype of intronic and missense SNPs showed the strongest association. The intronic SNP is strongly associated with PLA2R1 expression in the Genotype-Tissue Expression (GTEx) database, and the missense SNP is predicted to alter peptide binding with HLA-DRB1*15:01 by the Immune Epitope Database (IEDB). In HLA region, we performed relative predispositional effect (RPE) tests and identified additional risk alleles in both HLA-DRB1 and HLA-DQB1. We collapsed the risk alleles in each of HLA-DRB1 and HLA-DQB1 into single risk alleles. Reciprocal conditioning of these collapsed risk alleles showed more residual significance for HLA-DRB1 collapsed risk than HLA-DQB1 collapsed risk. These results indicate that changes in the expression levels of structurally different PLA2R protein confer risk for IMN in the presence of risk HLA-DRB1 alleles.

Forensic DNA typing is widely used to identify missing persons and plays a central role in forensic profiling. DNA typing usually uses capillary electrophoresis fragment analysis of PCR amplification products to detect the length of short tandem repeat (STR) markers. Here, we analyzed whole genome data from 1,070 Japanese individuals generated using massively parallel short-read sequencing of 162 paired-end bases. We have analyzed 843,473 STR loci with two to six basepair repeat units and cataloged highly polymorphic STR loci in the Japanese population. To evaluate the performance of the cataloged STR loci, we compared 23 STR loci, widely used in forensic DNA typing, with capillary electrophoresis based STR genotyping results in the Japanese population. Seventeen loci had high correlations and high call rates. The other six loci had low call rates or low correlations due to either the limitations of short-read sequencing technology, the bioinformatics tool used, or the complexity of repeat patterns. With these analyses, we have also purified the suitable 218 STR loci with four basepair repeat units and 53 loci with five basepair repeat units both for short read sequencing and PCR based technologies, which would be candidates to the actual forensic DNA typing in Japanese population.

Kinesin family member 20A (KIF20A), which is involved in cytokinesis and intracellular transportation, has been recently reported to be upregulated in several malignancies and may contribute to chemotherapeutic resistance. We examined the distribution and expression of KIF20A in clear-cell carcinoma (CCC) of the ovary to elucidate its clinical significance and molecular mechanism. Paraffin sections from ovarian CCC tissues (N=43) were immunostained with KIF20A antibody, and the staining intensities were semi-quantitatively evaluated. Furthermore, we investigated whether silencing of KIF20A contributes to the proliferation-inhibitory potential using CCC cells. During the observational period, 18 patients (41.9%) developed recurrence. The median time to recurrence was 11.5 months. Patients in the high KIF20A expression group showed poorer progression-free survival (PFS) and overall survival (OS) than those in the low expression group (P=0.0443 and P=0.0478, respectively). In multivariable analyses, KIF20A expression was also a significantly independent indicator of PFS and a marginally significant indicator of OS [PFS: HR (high vs. low), 5.488; 95% CI, 1.410–24.772 (P=0.0136); OS: HR, 2.835; 95% CI, 0.854–11.035, (P=0.0897)]. In in vitro studies, the ovarian CCC cell proliferation was significantly decreased by KIF20A silencing or in the presence of KIF20A inhibitor in CCC cells. Cell cycle G2/M arrest and a higher apoptosis-induced fraction were more frequently observed in si-KIF20A-transfected CCC cells than in the control cells. Although the present study was preliminary, these data indicate the possible involvement of KIF20A in the proliferation of CCC, suggesting that targeting this molecule may contribute to reversing the malignant potential consequently affecting the oncologic outcome of CCC patients.

The patient was a 78-year-old man with resection of distal pancreatectomy for pancreatic cancer with simultaneous multiple lung metastasis. They were papillary and tubular adenocarcinoma, Pt, TS3, infiltrative type, ly0, v0, pT3, CH0, DU0, S1, RP1, PV0, A1(Asp), PL0, OO0 and pN0, M1(PUL), pStage IV. He was received gemcitabine after the surgery. S-1 was added because of lung metastasis progression. Chemotherapy was continued for about 10 years from resection, and intra-abdominal recurrence was not observed and good performance status was maintained. 5-year survival rate of pancreatic cancer is as low as about 6.5%in Stage IV. There are cases where lung resection to isolated lung metastasis are performed after resection of pancreas and long-term-survival are obtained. A resected case of long-term-survival of pancreatic cancer with simultaneous multiple lung metastasis is rare, so we will report with a few literature considerations.

A 51-year-old man was admitted to our hospital because of anorexia and jaundice. Abdominal CT and MRCP taken after admission revealed tumor in the pancreatic head. Upper gastrointestinal endoscopy showed bleeding from the orifice of the major papilla. In angiography, an irregular image was found in the branch of the gastroduodenal artery, and a coil embolization surgery was performed mainly on the same part. We diagnosed pancreatic cancer and pancreaticoduodenectomy was performed. Pancreatic cancer with bleeding from the orifice of major papilla is very rare. We describe our case with known reports.

A 67-year-old woman underwent laparoscopy-assisted left hemicolectomy for early descending colon cancer(pTis, pN0, cH0, cM0, Stage 0).Her postoperative course was uneventful, without fever and/or tenderness at the anastomotic site.A month following discharge from the hospital, enhanced computed tomography revealed a liver abscess measuring 80mm in diameter at the lateral segment and a left adrenal abscess measuring 30mm in diameter.Although some free air and fluid collection was noted near the anastomotic site, there was no tenderness, and a gastrografin enema did not reveal leakage and/or pooling of the contrast agent near the anastomotic site.We administered antibiotics and performed percutaneous transhepatic abscess drainage following which imaging revealed shrinkage of her liver and adrenal abscesses and lowering of fever.However, enhanced computed tomography, performed a month later, revealed recurrence of the liver abscess, for which we performed a hepatic lateral segmentectomy.After undergoing the hepatectomy, she has shown no recurrence of the liver and adrenal abscesses.Several cases of liver abscess have been reported in association with colorectal cancer; however, an adrenal abscess occurring in association with colorectal cancer has not yet been reported.This case reveals that a minor leak could be associated with a liver and adrenal abscess.