Multiancestry analysis of the HLA locus in Alzheimer's and Parkinson's diseases uncovers a shared adaptive immune response mediated by HLA-DRB1*04 subtypes.

Across multiancestry groups, we analyzed Human Leukocyte Antigen (HLA) associations in over 176,000 individuals with Parkinson's disease (PD) and Alzheimer's disease (AD) versus controls. We demonstrate that the two diseases share the same protective association at the HLA locus. HLA-specific fine-mapping showed that hierarchical protective effects of HLA-DRB1*04 subtypes best accounted for the association, strongest with HLA-DRB1*04:04 and HLA-DRB1*04:07, and intermediary with HLA-DRB1*04:01 and HLA-DRB1*04:03. The same signal was associated with decreased neurofibrillary tangles in postmortem brains and was associated with reduced tau levels in cerebrospinal fluid and to a lower extent with increased Aß42. Protective HLA-DRB1*04 subtypes strongly bound the aggregation-prone tau PHF6 sequence, however only when acetylated at a lysine (K311), a common posttranslational modification central to tau aggregation. An HLA-DRB1*04-mediated adaptive immune response decreases PD and AD risks, potentially by acting against tau, offering the possibility of therapeutic avenues.

Genome-wide association study identifies three novel loci for type 2 diabetes.

Although over 60 loci for type 2 diabetes (T2D) have been identified, there still remains a large genetic component to be clarified. To explore unidentified loci for T2D, we performed a genome-wide association study (GWAS) of 6 209 637 single-nucleotide polymorphisms (SNPs), which were directly genotyped or imputed using East Asian references from the 1000 Genomes Project (June 2011 release) in 5976 Japanese patients with T2D and 20 829 nondiabetic individuals. Nineteen unreported loci were selected and taken forward to follow-up analyses. Combined discovery and follow-up analyses (30 392 cases and 34 814 controls) identified three new loci with genome-wide significance, which were MIR129-LEP [rs791595; risk allele = A; risk allele frequency (RAF) = 0.080; P = 2.55 × 10(-13); odds ratio (OR) = 1.17], GPSM1 [rs11787792; risk allele = A; RAF = 0.874; P = 1.74 × 10(-10); OR = 1.15] and SLC16A13 (rs312457; risk allele = G; RAF = 0.078; P = 7.69 × 10(-13); OR = 1.20). This study demonstrates that GWASs based on the imputation of genotypes using modern reference haplotypes such as that from the 1000 Genomes Project data can assist in identification of new loci for common diseases.

A functional SNP in FLT1 increases risk of coronary artery disease in a Japanese population.

Coronary artery disease (CAD) including myocardial infarction is one of the leading causes of death in many countries. Similar to other common diseases, its pathogenesis is thought to result from complex interactions among multiple genetic and environmental factors. Recent large-scale genetic association analysis for CAD identified 15 new loci. We examined the reproducibility of these previous association findings with 7990 cases and 6582 controls in a Japanese population. We found a convincing association of rs9319428 in FLT1, encoding fms-related tyrosine kinase 1 (P=5.98 × 10(-8)). Fine mapping using tag single-nucleotide polymorphisms (SNPs) at FLT1 locus revealed that another SNP (rs74412485) showed more profound genetic effect for CAD (P=2.85 × 10(-12)). The SNP, located in intron 1 in FLT1, enhanced the transcriptional level of FLT1. RNA interference experiment against FLT1 showed that the suppression of FLT1 resulted in decreased expression of inflammatory adhesion molecules. Expression of FLT1 was observed in endothelial cells of human coronary artery. Our results indicate that the genetically coded increased expression of FLT1 by a functional SNP implicates activation in an inflammatory cascade that might eventually lead to CAD.

A genome-wide association study of nephrolithiasis in the Japanese population identifies novel susceptible Loci at 5q35.3, 7p14.3, and 13q14.1.

Nephrolithiasis is a common nephrologic disorder with complex etiology. To identify the genetic factor(s) for nephrolithiasis, we conducted a three-stage genome-wide association study (GWAS) using a total of 5,892 nephrolithiasis cases and 17,809 controls of Japanese origin. Here we found three novel loci for nephrolithiasis: RGS14-SLC34A1-PFN3-F12 on 5q35.3 (rs11746443; P = 8.51×10⁻¹², odds ratio (OR) = 1.19), INMT-FAM188B-AQP1 on 7p14.3 (rs1000597; P = 2.16×10⁻¹4, OR = 1.22), and DGKH on 13q14.1 (rs4142110; P = 4.62×10⁻9, OR = 1.14). Subsequent analyses in 21,842 Japanese subjects revealed the association of SNP rs11746443 with the reduction of estimated glomerular filtration rate (eGFR) (P = 6.54×10⁻8), suggesting a crucial role for this variation in renal function. Our findings elucidated the significance of genetic variations for the pathogenesis of nephrolithiasis.

A genome-wide association study identifies locus at 10q22 associated with clinical outcomes of adjuvant tamoxifen therapy for breast cancer patients in Japanese.

Although many association studies of polymorphisms in candidate genes with the clinical outcomes of breast cancer patients receiving adjuvant tamoxifen therapy have been reported, genetic factors determining individual response to tamoxifen are not fully understood. To identify genetic polymorphisms associated with clinical outcomes of patients with tamoxifen treatment, we conducted a genome-wide association study (GWAS). We studied 462 Japanese patients with hormone receptor-positive, invasive breast cancer receiving adjuvant tamoxifen therapy. Of them, 240 patients were analyzed by genome-wide genotyping using the Illumina Human610-Quad BeadChips, and two independent sets of 105 and 117 cases were used for replication studies. In the GWAS, we detected significant associations with recurrence-free survival at 15 single-nucleotide polymorphisms (SNPs) on nine chromosomal loci (1p31, 1q41, 5q33, 7p11, 10q22, 12q13, 13q22, 18q12 and 19p13) that satisfied a genome-wide significant threshold (log-rank P= 2.87 × 10(-9)-9.41 × 10(-8)). Among them, rs10509373 in C10orf11 gene on 10q22 was significantly associated with recurrence-free survival in the replication study (log-rank P= 2.02 × 10(-4)) and a combined analysis indicated a strong association of this SNP with recurrence-free survival in breast cancer patients treated with tamoxifen (log-rank P= 1.26 × 10(-10)). Hazard ratio per C allele of rs10509373 was 4.51 [95% confidence interval (CI), 2.72-7.51; P= 6.29 × 10(-9)]. In a combined analysis of rs10509373 genotype with previously identified genetic makers, CYP2D6 and ABCC2, the number of risk alleles of these three genes had cumulative effects on recurrence-free survival among 345 patients receiving tamoxifen monotherapy (log-rank P= 2.28 × 10(-12)). In conclusion, we identified a novel locus associated with recurrence-free survival in Japanese breast cancer patients receiving adjuvant tamoxifen therapy.

A single-nucleotide polymorphism in ANK1 is associated with susceptibility to type 2 diabetes in Japanese populations.

To identify a novel susceptibility locus for type 2 diabetes, we performed an imputation-based, genome-wide association study (GWAS) in a Japanese population using newly obtained imputed-genotype data for 2 229 890 single-nucleotide polymorphisms (SNPs) estimated from previously reported, directly genotyped GWAS data in the same samples (stage 1: 4470 type 2 diabetes versus 3071 controls). We directly genotyped 43 new SNPs with P-values of <10(-4) in a part of stage-1 samples (2692 type 2 diabetes versus 3071 controls), and the associations of validated SNPs were evaluated in another 11 139 Japanese individuals (stage 2: 7605 type 2 diabetes versus 3534 controls). Combined meta-analysis using directly genotyped data for stages 1 and 2 revealed that rs515071 in ANK1 and rs7656416 near MGC21675 were associated with type 2 diabetes in the Japanese population at the genome-wide significant level (P < 5 × 10(-8)). The association of rs515071 was also observed in European GWAS data (combined P for all populations = 6.14 × 10(-10)). Rs7656416 was in linkage disequilibrium to rs6815464, which had recently been identified as a top signal in a meta-analysis of East Asian GWAS for type 2 diabetes (r(2) = 0.76 in stage 2). The association of rs7656416 with type 2 diabetes disappeared after conditioning on rs6815464. These results indicate that the ANK1 locus is a new, common susceptibility locus for type 2 diabetes across different ethnic groups. The signal of association was weaker in the directly genotyped data, so the improvement in signal indicates the importance of imputation in this particular case.

A genome-wide association study identifies 2 susceptibility Loci for Crohn's disease in a Japanese population.

BACKGROUND & AIMS: Crohn's disease is an inflammatory bowel disease induced by multiple genetic and environmental factors. Genome-wide association studies have identified genetic factors that affect the risk for Crohn's disease in European populations, but information from other ethnic groups is scarce. We therefore investigated genetic factors associated with Crohn's disease in the Japanese population. METHODS: We performed a genome-wide association study with 372 individuals with Crohn's disease (cases) and 3389 controls, all from the Japanese population. To confirm identified associations, we performed a replication study with an independent panel of 1151 Crohn's disease cases and 15,800 controls. We also performed an association analysis using genome-wide genotype imputation in the discovery cohort. RESULTS: We confirmed associations of Crohn's disease with variants in MHC (rs7765379, P = 2.11 × 10(-59)), TNFSF15 (rs6478106, P = 3.87 × 10(-45)), and STAT3 (rs9891119, P = 2.24 × 10(-14)). We identified 2 new susceptibility loci: on chromosome 4p14 (rs1487630, P = 2.40 × 10(-11); odds ratio, 1.33), and in the SLC25A15-ELF1-WBP4 region on 13q14 (rs7329174 in ELF1, P = 5.12 × 10(-9); odds ratio, 1.27). CONCLUSIONS: In a genome-wide association study, we identified 2 new susceptibility loci for Crohn's disease in a Japanese population. These findings could increase our understanding of the pathogenesis of Crohn's disease.

Association of a novel long non-coding RNA in 8q24 with prostate cancer susceptibility.

Recent genome-wide association studies reported strong and reproducible associations of multiple genetic variants in a large "gene-desert" region of chromosome 8q24 with susceptibility to prostate cancer (PC). However, the causative or functional variants of these 8q24 loci and their biological mechanisms associated with PC susceptibility remain unclear and should be investigated. Here, focusing on its most centromeric region (so-called Region 2: Chr8: 128.14-128.28 Mb) among the multiple PC loci on 8q24, we performed fine mapping and re-sequencing of this critical region and identified SNPs (single nucleotide polymorphisms) between rs1456315 and rs7463708 (chr8: 128,173,119-128,173,237 bp) to be most significantly associated with PC susceptibility (P = 2.00 × 10(-24) , OR = 1.74, 95% confidence interval = 1.56-1.93). Importantly, we show that this region was transcribed as a ∼13 kb intron-less long non-coding RNA (ncRNA), termed PRNCR1 (prostate cancer non-coding RNA 1), and PRNCR1 expression was upregulated in some of the PC cells as well as precursor lesion prostatic intraepithelial neoplasia. Knockdown of PRNCR1 by siRNA attenuated the viability of PC cells and the transactivation activity of androgen receptor, which indicates that PRNCR1 could be involved in prostate carcinogenesis possibly through androgen receptor activity. These findings could provide a new insight in understanding the pathogenesis of genetic factors for PC susceptibility and prostate carcinogenesis.

Ethnic and trans-ethnic genome-wide association studies identify new loci influencing Japanese Alzheimer's disease risk.

Alzheimer's disease (AD) has no cure, but early detection and risk prediction could allow earlier intervention. Genetic risk factors may differ between ethnic populations. To discover novel susceptibility loci of AD in the Japanese population, we conducted a genome-wide association study (GWAS) with 3962 AD cases and 4074 controls. Out of 4,852,957 genetic markers that passed stringent quality control filters, 134 in nine loci, including APOE and SORL1, were convincingly associated with AD. Lead SNPs located in seven novel loci were genotyped in an independent Japanese AD case-control cohort. The novel locus FAM47E reached genome-wide significance in a meta-analysis of association results. This is the first report associating the FAM47E locus with AD in the Japanese population. A trans-ethnic meta-analysis combining the results of the Japanese data sets with summary statistics from stage 1 data of the International Genomics of Alzheimer's Project identified an additional novel susceptibility locus in OR2B2. Our data highlight the importance of performing GWAS in non-European populations.

Empirical Bayes Estimation of Semi-parametric Hierarchical Mixture Models for Unbiased Characterization of Polygenic Disease Architectures.

Genome-wide association studies (GWAS) suggest that the genetic architecture of complex diseases consists of unexpectedly numerous variants with small effect sizes. However, the polygenic architectures of many diseases have not been well characterized due to lack of simple and fast methods for unbiased estimation of the underlying proportion of disease-associated variants and their effect-size distribution. Applying empirical Bayes estimation of semi-parametric hierarchical mixture models to GWAS summary statistics, we confirmed that schizophrenia was extremely polygenic [~40% of independent genome-wide SNPs are risk variants, most within odds ratio (OR = 1.03)], whereas rheumatoid arthritis was less polygenic (~4 to 8% risk variants, significant portion reaching OR = 1.05 to 1.1). For rheumatoid arthritis, stratified estimations revealed that expression quantitative loci in blood explained large genetic variance, and low- and high-frequency derived alleles were prone to be risk and protective, respectively, suggesting a predominance of deleterious-risk and advantageous-protective mutations. Despite genetic correlation, effect-size distributions for schizophrenia and bipolar disorder differed across allele frequency. These analyses distinguished disease polygenic architectures and provided clues for etiological differences in complex diseases.

Performance comparison of four commercial human whole-exome capture platforms.

Whole exome sequencing (WXS) is widely used to identify causative genetic mutations of diseases. However, not only have several commercial human exome capture platforms been developed, but substantial updates have been released in the past few years. We report a performance comparison for the latest release of four commercial platforms, Roche/NimbleGen's SeqCap EZ Human Exome Library v3.0, Illumina's Nextera Rapid Capture Exome (v1.2), Agilent's SureSelect XT Human All Exon v5 and Agilent's SureSelect QXT, using the same DNA samples. Agilent XT showed the highest target enrichment efficiency and the best SNV and short indel detection sensitivity in coding regions with the least amount of sequencing. Agilent QXT had slightly inferior target enrichment than Agilent XT. Illumina, with additional sequencing, detected SNVs and short indels at the same quality as Agilent XT, and showed the best performance in coverage of medically interesting mutations. NimbleGen detected more SNVs and indels in untranslated regions than the others. We also found that the platforms, which enzymatically fragment the genomic DNA (gDNA), detected more homozygous SNVs than those using sonicated gDNA. We believe that our analysis will help investigators when selecting a suitable exome capture platform for their particular research.

Genome-Wide Association Study of Peripheral Arterial Disease in a Japanese Population.

Characteristics of peripheral arterial disease (PAD) are the occlusion or stenosis of multiple vessel sites caused mainly by atherosclerosis and chronic lower limb ischemia. To identify PAD susceptible loci, we conducted a genome-wide association study (GWAS) with 785 cases and 3,383 controls in a Japanese population using 431,666 single nucleotide polymorphisms (SNP). After staged analyses including a total of 3,164 cases and 20,134 controls, we identified 3 novel PAD susceptibility loci at IPO5/RAP2A, EDNRA and HDAC9 with genome wide significance (combined P = 6.8 x 10-14, 5.3 x 10-9 and 8.8 x 10-8, respectively). Fine-mapping at the IPO5/RAP2A locus revealed that rs9584669 conferred risk of PAD. Luciferase assay showed that the risk allele at this locus reduced expression levels of IPO5. To our knowledge, these are the first genetic risk factors for PAD.

A combination of targeted enrichment methodologies for whole-exome sequencing reveals novel pathogenic mutations.

Whole-exome sequencing (WES) is a useful method to identify disease-causing mutations, however, often no candidate mutations are identified using commonly available targeted probe sets. In a recent analysis, we also could not find candidate mutations for 20.9% (9/43) of our pedigrees with congenital neurological disorder using pre-designed capture probes (SureSelect V4 or V5). One possible cause for this lack of candidates is that standard WES cannot sequence all protein-coding sequences (CDS) due to capture probe design and regions of low coverage, which account for approximately 10% of all CDS regions. In this study, we combined a selective circularization-based target enrichment method (HaloPlex) with a hybrid capture method (SureSelect V5; WES), and achieved a more complete coverage of CDS regions (~97% of all CDS). We applied this approach to 7 (SureSelect V5) out of 9 pedigrees with no candidates through standard WES analysis and identified novel pathogenic mutations in one pedigree. The application of this effective combination of targeted enrichment methodologies can be expected to aid in the identification of novel pathogenic mutations previously missed by standard WES analysis.

The construction of risk prediction models using GWAS data and its application to a type 2 diabetes prospective cohort.

Recent genome-wide association studies (GWAS) have identified several novel single nucleotide polymorphisms (SNPs) associated with type 2 diabetes (T2D). Various models using clinical and/or genetic risk factors have been developed for T2D risk prediction. However, analysis considering algorithms for genetic risk factor detection and regression methods for model construction in combination with interactions of risk factors has not been investigated. Here, using genotype data of 7,360 Japanese individuals, we investigated risk prediction models, considering the algorithms, regression methods and interactions. The best model identified was based on a Bayes factor approach and the lasso method. Using nine SNPs and clinical factors, this method achieved an area under a receiver operating characteristic curve (AUC) of 0.8057 on an independent test set. With the addition of a pair of interaction factors, the model was further improved (p-value 0.0011, AUC 0.8085). Application of our model to prospective cohort data showed significantly better outcome in disease-free survival, according to the log-rank trend test comparing Kaplan-Meier survival curves (p--value 2:09 x 10(-11)). While the major contribution was from clinical factors rather than the genetic factors, consideration of genetic risk factors contributed to an observable, though small, increase in predictive ability. This is the first report to apply risk prediction models constructed from GWAS data to a T2D prospective cohort. Our study shows our model to be effective in prospective prediction and has the potential to contribute to practical clinical use in T2D.

Reproducibility, performance, and clinical utility of a genetic risk prediction model for prostate cancer in Japanese.

Prostate specific antigen (PSA) is widely used as a diagnostic biomarker for prostate cancer (PC). However, due to its low predictive performance, many patients without PC suffer from the harms of unnecessary prostate needle biopsies. The present study aims to evaluate the reproducibility and performance of a genetic risk prediction model in Japanese and estimate its utility as a diagnostic biomarker in a clinical scenario. We created a logistic regression model incorporating 16 SNPs that were significantly associated with PC in a genome-wide association study of Japanese population using 689 cases and 749 male controls. The model was validated by two independent sets of Japanese samples comprising 3,294 cases and 6,281 male controls. The areas under curve (AUC) of the model were 0.679, 0.655, and 0.661 for the samples used to create the model and those used for validation. The AUCs were not significantly altered in samples with PSA 1-10 ng/ml. 24.2% and 9.7% of the patients had odds ratio <0.5 (low risk) or >2 (high risk) in the model. Assuming the overall positive rate of prostate needle biopsies to be 20%, the positive biopsy rates were 10.7% and 42.4% for the low and high genetic risk groups respectively. Our genetic risk prediction model for PC was highly reproducible, and its predictive performance was not influenced by PSA. The model could have a potential to affect clinical decision when it is applied to patients with gray-zone PSA, which should be confirmed in future clinical studies.

Variation in the DEPDC5 locus is associated with progression to hepatocellular carcinoma in chronic hepatitis C virus carriers.

Chronic viral hepatitis is the most important risk factor for progression to hepatocellular carcinoma (HCC). To identify genetic risk factors for progression to HCC in individuals with chronic hepatitis C virus (HCV), we analyzed 467,538 SNPs in 212 Japanese individuals with chronic HCV with HCC and 765 individuals with chronic HCV without HCC. We identified one intronic SNP in the DEPDC5 locus on chromosome 22 associated with HCC risk and confirmed the association using an independent case-control population (710 cases and 1,625 controls). The association was highly significant when we analyzed the stages separately as well as together (rs1012068, P(combined) = 1.27 × 10(-13), odds ratio = 1.75). The significance level of the association further increased after adjustment for gender, age and platelet count (P = 1.35 × 10(-14), odds ratio = 1.96). Our findings suggest that common variants within the DEPDC5 locus affect susceptibility to HCC in Japanese individuals with chronic HCV infection.

Identification of a set of genes associated with response to interleukin-2 and interferon-α combination therapy for renal cell carcinoma through genome-wide gene expression profiling.

Interleukin (IL)-2 and interferon (IFN)-α combination therapy for metastatic renal cell carcinoma (RCC) improves the prognosis for a subset of patients, while some patients suffer from severe adverse drug reactions with little benefit. To establish a method to predict responses to this combination therapy (approximately 30% response rate), the gene expression profiles of primary RCCs were analyzed using an oligoDNA microarray consisting of 38,500 genes or ESTs, after enrichment of the cancer cell population by laser micro-beam microdissection. The analysis of 10 responders and 18 non-responders identified 24 genes that exhibited significant differential expression between the two groups. In addition, the patients whose tumors did not express HLA-DQA1 or HLA-DQB1 molecules demonstrated poor clinical response. Exclusion of patients with tumors lacking either of these two genes is likely to improve the response rate to IL-2 and IFN-α combination therapy from 30 to 67%, indicating that a simple pretreatment test provides useful information with which to subselect patients with renal cancer in order to improve the efficacy of this treatment and reduce unnecessary medical costs.

Variation in TP63 is associated with lung adenocarcinoma susceptibility in Japanese and Korean populations.

Lung cancer is the most common cause of death from cancer worldwide, and its incidence is increasing in East Asian and Western countries. To identify genetic factors that modify the risk of lung adenocarcinoma, we conducted a genome-wide association study in a Japanese cohort, with replication in two independent studies in Japanese and Korean individuals, in a total of 2,098 lung adenocarcinoma cases and 11,048 controls. The combined analyses identified two susceptibility loci for lung adenocarcinoma: TERT (rs2736100, combined P = 2.91 × 10⁻¹¹), odds ratio (OR) = 1.27) and TP63 (rs10937405, combined P = 7.26 × 10⁻¹²), OR = 1.31). Fine mapping of the region containing TP63 showed that a SNP (rs4488809) in intron 1 of TP63 showed the most significant association. Our results suggest that genetic variation in TP63 may influence susceptibility to lung adenocarcinoma in East Asian populations.