Genetically Predicted Higher Levels of Caffeic Acid Are Protective Against Ulcerative Colitis: A Comprehensive Metabolome Analysis.

BACKGROUND: It is crucial to pinpoint the metabolites that cause Crohn's disease (CD) and ulcerative colitis (UC) to comprehend their pathogenesis and identify possible targets for therapy. To achieve this goal, we performed the first metabolome-wide Mendelian randomization (MR) study of Japanese patients with CD and UC. METHODS: As exposure datasets, genetic instruments with blood-circulating metabolites were obtained from the Tohoku Medical Megabank Organization, which includes 204 metabolites from the genome-wide association study data of 7843 Japanese individuals. As outcome datasets, we enrolled Japanese patients with CD (n = 1803), Japanese patients with UC (n = 1992), and healthy controls (n = 2022). The main analysis utilized the inverse variance-weighted method, while stability of the findings was evaluated through sensitivity analyses. RESULTS: After single nucleotide polymorphism (SNP) filtering, 169 SNPs for 45 metabolites were available for MR. Genetically predicted elevated circulating trans-glutaconic acid and tryptophan were associated with a lower CD risk (odds ratio [OR], 0.68; P = 5.95 × 10-3; and OR, 0.64; P = 1.90 × 10-2, respectively). Genetically predicted elevated caffeic acid was associated with a lower UC risk (OR, 0.67; P = 4.2 × 10-4), which remained significant after multiple testing correction. We identified a causal link between UC and 3-hydroxybutyrate (OR, 2.21; P = 1.41 × 10-2), trans-glutaconic acid (OR, 0.72; P = 1.77 × 10-2), and 2-hydroxyvaleric acid (OR, 1.31; P = 4.23 × 10-2). There was no evidence of pleiotropy or reverse causal effects for these candidate metabolites. CONCLUSIONS: In our metabolome-wide MR study, we discovered a notable protective effect of caffeic acid against UC.

This metabolome-wide study using Japanese cohorts found that caffeic acid significantly reduces the risk of ulcerative colitis, while 3-hydroxybutyrate and 2-hydroxyvaleric acid increase it. Trans-glutaconic acid and tryptophan reduce the risk of Crohn's disease.

Identification of the hybrid gene LILRB5-3 by long-read sequencing and implication of its novel signaling function.

Leukocyte immunoglobulin (Ig)-like receptors (LILRs) on human chromosome 19q13.4 encode 11 immunoglobulin superfamily receptors, exhibiting genetic diversity within and between human populations. Among the LILR genes, the genomic region surrounding LILRB3 and LILRA6 has yet to be fully characterized due to their significant sequence homology, which makes it difficult to differentiate between them. To examine the LILRB3 and LILRA6 genomic region, a tool named JoGo-LILR CN Caller, which can call copy number from short-read whole genome sequencing (srWGS) data, was applied to an extensive international srWGS dataset comprising 2,504 samples. During this process, a previously unreported loss of both LILRB3 and LILRA6 was detected in three samples. Using long-read sequencing of these samples, we have discovered a novel large deletion (33,692 bp) in the LILRB3 and LILRA6 genomic regions in the Japanese population. This deletion spanned three genes, LILRB3, LILRA6, and LILRB5, resulting in LILRB3 exons 12-13 being located immediately downstream of LILRB5 exons 1-12 with the loss of LILRA6, suggesting the potential expression of a hybrid gene between LILRB5 and LILRB3 (LILRB5-3). Transcription and subsequent translation of the LILRB5-3 hybrid gene were also verified. The hybrid junction was located within the intracellular domain, resulting in an LILRB5 extracellular domain fused to a partial LILRB3 intracellular domain with three immunoreceptor tyrosine-based inhibitory motifs (ITIMs), suggesting that LILRB5-3 acquired a novel signaling function. Further application of the JoGo-LILR tool to srWGS samples suggested the presence of the LILRB5-3 hybrid gene in the CEU population. Our findings provide insight into the genetic and functional diversity of the LILR family.

A genome-wide association study identified PTPN2 as a population-specific susceptibility gene locus for primary biliary cholangitis.

BACKGROUND AIMS: Previous genome-wide association studies (GWAS) have indicated the involvement of shared (population-non-specific) and non-shared (population-specific) susceptibility genes in the pathogenesis of primary biliary cholangitis (PBC) among European and East-Asian populations. Although a meta-analysis of these distinct populations has recently identified more than 20 novel PBC susceptibility loci, analyses of population-specific genetic architecture are still needed for a more comprehensive search for genetic factors in PBC. APPROACH RESULTS: Protein tyrosine phosphatase non-receptor type 2 (PTPN2) was identified as a novel PBC susceptibility gene locus through a GWAS and subsequent genome-wide meta-analysis involving 2,181 cases and 2,699 controls from the Japanese population (GWAS-lead variant: rs8098858, p=2.6×10-8). In-silico and in-vitro functional analyses indicated that the risk allele of rs2292758, which is a primary functional variant, decreases PTPN2 expression by disrupting Sp1 binding to the PTPN2 promoter in T follicular helper cells (Tfh) and plasmacytoid dendritic cells (pDCs). Infiltration of PTPN2-positive T-cells and pDCs were confirmed in the portal area of the PBC-liver by immunohistochemistry. Furthermore, transcriptomic analysis of PBC-liver samples indicated the presence of a compromised negative feedback loop in-vivo between PTPN2 and IFNG in patients carrying the risk allele of rs2292758. CONCLUSIONS: PTPN2, a novel susceptibility gene for PBC in the Japanese population, may be involved in the pathogenesis of PBC via an insufficient negative feedback loop caused by the PTPN2 risk allele of rs2292758 in IFN signaling. This suggests that PTPN2 could be a potential molecular target for PBC treatment.

rs10924104 in the expression enhancer motif of CD58 confers susceptibility to human autoimmune diseases.

CD58 plays roles in cell adhesion and co-stimulation with antigen presentation from major histocompatibility complex class II on antigen-presenting cells to T-cell antigen receptors on naïve T cells. CD58 reportedly contributes to the development of various human autoimmune diseases. Recently, genome-wide association studies (GWASs) identified CD58 as a susceptibility locus for autoimmune diseases such as systemic lupus erythematosus (SLE), multiple sclerosis (MS), and primary biliary cholangitis (PBC). However, the primary functional variant and molecular mechanisms of susceptibility to autoimmune diseases in the CD58 locus were not clarified. Here, rs10924104, located in the ZNF35-binding motif within the gene expression regulatory motif, was identified as the primary functional variant for SLE, MS, and PBC among genetic variants showing stronger linkage disequilibrium (LD) with GWAS-lead variants in the CD58 locus. Expression-quantitative trait locus (e-QTL) data for each distinct blood cell type and in vitro functional analysis using the CRISPR/Cas9 system corroborated the functional role of rs10924104 in the upregulation of CD58 transcription by the disease-risk allele. Additionally, the strength of disease susceptibility observed in the CD58 locus could be accounted for by the strength of LD between rs10924104 and each GWAS-lead variant. In conclusion, the present study demonstrated for the first time the existence of a shared autoimmune disease-related primary functional variant (i.e., rs10924104) that regulates the expression of CD58. Clarifying the molecular mechanism of disease susceptibility derived from such a shared genetic background is important for understanding human autoimmune diseases and human immunology.

Exploring the genetic diversity of the Japanese population: Insights from a large-scale whole genome sequencing analysis.

The Japanese archipelago is a terminal location for human migration, and the contemporary Japanese people represent a unique population whose genomic diversity has been shaped by multiple migrations from Eurasia. We analyzed the genomic characteristics that define the genetic makeup of the modern Japanese population from a population genetics perspective from the genomic data of 9,287 samples obtained by high-coverage whole-genome sequencing (WGS) by the National Center Biobank Network. The dataset comprised populations from the Ryukyu Islands and other parts of the Japanese archipelago (Hondo). The Hondo population underwent two episodes of population decline during the Jomon period, corresponding to the Late Neolithic, and the Edo period, corresponding to the Early Modern era, while the Ryukyu population experienced a population decline during the shell midden period of the Late Neolithic in this region. Haplotype analysis suggested increased allele frequencies for genes related to alcohol and fatty acid metabolism, which were reported as loci that had experienced positive natural selection. Two genes related to alcohol metabolism were found to be 12,500 years out of phase with the time when they began to increase in the allele frequency; this finding indicates that the genomic diversity of Japanese people has been shaped by events closely related to agriculture and food production.

Genome-wide association analysis reveals the associations of NPHP4, TYW1-AUTS2 and SEMA6D for Behçet's disease and HLA-B*46:01 for its intestinal involvement.

BACKGROUND: Intestinal involvement in Behçet's disease (BD) is associated with poor prognosis and is more prevalent in East Asian than in Mediterranean populations. Identifying the genetic causes of intestinal BD is important for understanding the pathogenesis and for appropriate treatment of BD patients. METHODS: We performed genome-wide association studies (GWAS) and imputation/replication genotyping of human leukocyte antigen (HLA) alleles for 1,689 Korean and Turkish patients with BD (including 379 patients with intestinal BD) and 2,327 healthy controls, followed by replication using 593 Japanese patients with BD (101 patients with intestinal BD) and 737 healthy controls. Stratified cross-phenotype analyses were performed for 1) overall BD, 2) intestinal BD, and 3) intestinal BD without association of overall BD. RESULTS: We identified three novel genome-wide significant susceptibility loci including NPHP4 (rs74566205; P=1.36 × 10-8), TYW1-AUTS2 (rs60021986; P=1.14 × 10-9), and SEMA6D (rs4143322; P=5.54 × 10-9) for overall BD, and a new association with HLA-B*46:01 for intestinal BD (P=1.67 × 10-8) but not for BD without intestinal involvement. HLA peptide binding analysis revealed that Mycobacterial peptides, have a stronger binding affinity to HLA-B*46:01 compared to the known risk allele HLA-B*51:01. CONCLUSIONS: HLA-B*46:01 is associated with the development of intestinal BD; NPHP4, TYW1-AUTS2, and SEMA6D are susceptibility loci for overall BD.

Novel HLA allele associations with susceptibility, staging, symptomatic state, autoimmune hepatitis and hepatocellular carcinoma events for primary biliary cholangitis in the Japanese population.

Primary biliary cholangitis (PBC) is a rare autoimmune disease with a clear predisposition for human leukocyte antigen (HLA)-DR/DQ-associated loss of immune tolerance for the E2 component of the pyruvate dehydrogenase complex. Three-field-resolution HLA imputation of 1,670 Japanese PBC patients and 2,328 healthy controls was conducted using Japanese population-specific HLA reference panels. Eighteen previously reported Japanese PBC-associated HLA alleles were confirmed and extended to 3-field-resolution, including HLA-DRB1*08:03 to HLA-DRB1*08:03:02, HLA-DQB1*03:01 to HLA-DQB1*03:01:01, HLA-DQB1*04:01 to HLA-DQB1*04:01:01 and HLA-DQB1*06:04 to HLA-DQB1*06:04:01. In addition, additional significant novel HLA alleles were identified, including 3 novel susceptible HLA-DQA1 alleles: HLA-DQA1*03:03:01, HLA-DQA1*04:01:01, HLA-DQA1*01:04:01 and 1 novel protective HLA-DQA1 allele, HLA-DQA1*05:05:01. In addition, PBC patients carrying HLA-DRB1*15:01:01 and HLA-DQA1*03:03:01 would have a higher predisposition toward developing concomitant autoimmune hepatitis (AIH). Further, late-stage and symptomatic PBC shared the same susceptible HLA alleles of HLA-A*26:01:01, HLA-DRB1*09:01:02 and HLA-DQB1*03:03:02. Lastly, HLA-DPB1*05:01:01 was identified as a potential risk HLA allele for development of hepatocellular carcinoma (HCC) in PBC patients. In conclusion, we have extended the current knowledge of HLA allele associations to 3-field resolution and identified novel HLA allele associations with predisposition risk, staging, symptomatic state, and AIH and HCC events for Japanese PBC patients.

Prediction Model with HLA-A*33:03 Reveals Number of Days to Develop Liver Cancer from Blood Test.

The development of liver cancer in patients with hepatitis B is a major problem, and several models have been reported to predict the development of liver cancer. However, no predictive model involving human genetic factors has been reported to date. For the items incorporated in the prediction model reported so far, we selected items that were significant in predicting liver carcinogenesis in Japanese patients with hepatitis B and constructed a prediction model of liver carcinogenesis by the Cox proportional hazard model with the addition of Human Leukocyte Antigen (HLA) genotypes. The model, which included four items-sex, age at the time of examination, alpha-fetoprotein level (log10AFP) and presence or absence of HLA-A*33:03-revealed an area under the receiver operating characteristic curve (AUROC) of 0.862 for HCC prediction within 1 year and an AUROC of 0.863 within 3 years. A 1000 repeated validation test resulted in a C-index of 0.75 or higher, or sensitivity of 0.70 or higher, indicating that this predictive model can distinguish those at high risk of developing liver cancer within a few years with high accuracy. The prediction model constructed in this study, which can distinguish between chronic hepatitis B patients who develop hepatocellular carcinoma (HCC) early and those who develop HCC late or not, is clinically meaningful.

Response to COVID-19 vaccine is reduced in patients with inflammatory bowel disease, but improved with additional dose.

BACKGROUND: Coronavirus disease 2019 (COVID-19) vaccination is recommended for patients with inflammatory bowel disease (IBD); however, suppressed immune responses have been reported for fully vaccinated patients under immunosuppressive therapy, mainly from Western countries. We prospectively analyzed antibody titers of IBD patients in Asia induced by two-dose and additional dose of messengerRNA COVID-19 vaccine. METHODS: After measuring high-affinity antibody titers, factors associated with antibody titers were identified by multiple regression analyses using the following covariates: sex, age (≥60 or <60 years), disease type (Crohn's disease or ulcerative colitis), vaccine type (BNT162b2 or mRNA-1273), time from second/third vaccination, molecular-targeted agent (anti-tumor necrosis factor [TNF] agents, ustekinumab, vedolizumab, tofacitinib, or no molecular-targeted agents), thiopurine, steroid, and 5-aminosalicylic acid. RESULTS: Among 409 patients analyzed, mean titer was 1316.7 U/mL (SD, 1799.3); 403 (98.5%) were judged to be seropositive (≥0.8 U/mL), and 389 (95.1%) had neutralizing antibodies (≥15 U/mL). After the third vaccination, mean titer raised up to 21 123.8 U/mL (SD, 23 474.5); all 179 were seropositive, and 178 (99.4%) had neutralizing antibodies. In 248 patients with genetic data, there was no difference in mean titer after two/third doses between carriers and non-carriers of HLA-A24 associated with severe disease during COVID-19 infection. A multiple regression analyses using covariates revealed that older age, vaccine type (BNT162b2), time from second/third dose, anti-TNF agent, tofacitinib, and thiopurine were independently associated with lower antibody titers. CONCLUSIONS: Our findings further support the recommendation for COVID-19 vaccination in patients under immunosuppressive therapy, especially additional third dose for patients receiving anti-TNF agents and/or thiopurine or tofacitinib.

The NBDC-DDBJ imputation server facilitates the use of controlled access reference panel datasets in Japan.

Accurate genotype imputation requires large-scale reference panel datasets. When conducting genotype imputation on the Japanese population, researchers can use such datasets under collaborative studies or controlled access conditions in public databases. We developed the NBDC-DDBJ imputation server, which securely provides users with a web user interface to execute genotype imputation on the server. Our benchmarking analysis showed that the accuracy of genotype imputation was improved by leveraging controlled access datasets to increase the number of haplotypes available for analysis compared to using publicly available reference panels such as the 1000 Genomes Project. The NBDC-DDBJ imputation server facilitates the use of controlled access datasets for accurate genotype imputation.

Intraocular human cytomegaloviruses of ocular diseases are distinct from those of viremia and are capable of escaping from innate and adaptive immunity by exploiting HLA-E-mediated peripheral and central tolerance.

Human cytomegalovirus (HCMV) infections develop into CMV diseases that result in various forms of manifestations in local organs. CMV-retinitis is a form of CMV disease that develops in immunocompromised hosts with CMV-viremia after viruses in the peripheral circulation have entered the eye. In the HCMV genome, extensive diversification of the UL40 gene has produced peptide sequences that modulate NK cell effector functions when loaded onto HLA-E and are subsequently recognized by the NKG2A and NKG2C receptors. Notably, some HCMV strains carry UL40 genes that encode peptide sequences identical to the signal peptide sequences of specific HLA-A and HLA-C allotypes, which enables these CMV strains to escape HLA-E-restricted CD8+T cell responses. Variations in UL40 sequences have been studied mainly in the peripheral blood of CMV-viremia cases. In this study, we sought to investigate how ocular CMV disease develops from CMV infections. CMV gene sequences were compared between the intraocular fluids and peripheral blood of 77 clinical cases. UL40 signal peptide sequences were more diverse, and multiple sequences were typically present in CMV-viremia blood compared to intraocular fluid. Significantly stronger NK cell suppression was induced by UL40-derived peptides from intraocular HCMV compared to those identified only in peripheral blood. HCMV present in intraocular fluids were limited to those carrying a UL40 peptide sequence corresponding to the leader peptide sequence of the host's HLA class I, while UL40-derived peptides from HCMV found only in the peripheral blood were disparate from any HLA class I allotype. Overall, our analyses of CMV-retinitis inferred that specific HCMV strains with UL40 signal sequences matching the host's HLA signal peptide sequences were those that crossed the blood-ocular barrier to enter the intraocular space. UL40 peptide repertoires were the same in the intraocular fluids of all ocular CMV diseases, regardless of host immune status, implying that virus type is likely to be a common determinant in ocular CMV disease development. We thus propose a mechanism for ocular CMV disease development, in which particular HCMV types in the blood exploit peripheral and central HLA-E-mediated tolerance mechanisms and, thus, escape the antivirus responses of both innate and adaptive immunity.

Detection of the HLA-B*15:01:74 allele, an HLA-B*15 variant discovered in a Japanese individual.

HLA-B*15:01:74 differs from B*15:01:01:01 by one nucleotide in exon 2 at position 470.

Detection of the novel HLA-C*06:02:01:62 allele by next-generation sequencing in a Japanese individual.

HLA-C*06:02:01:62 differs from HLA-C*06:02:01:01 by one nucleotide in intron 3 at position 1413.

Genomic sequencing of the novel HLA-C*07:02:01:107 allele by next-generation sequencing in two Japanese individuals.

HLA-C*07:02:01:107 differs from HLA-C*07:02:01:01 by two nucleotides in 3' UTR at position 3789 and 3806.

Identification of the novel HLA-B*40:01:02:40 and HLA-B*40:01:02:41 alleles in a Japanese individual.

HLA-B*40:01:02:40 and -B*40:01:02:41 differ from B*40:01:02:04 by one and two nucleotides in the 3'UTR, respectively.

The HLA-B*15:02:01:05 allele identified by two next-generation sequencing methods in a Japanese individual.

HLA-B*15:02:01:05 differs from HLA-B*15:02:01:01 by four nucleotides in 3'UTR at positions 3435, 3457, 3472, and 3511.

Genome-Wide Association Study for Chronic Hepatitis B Infection in the Thai Population.

To identify novel host genetic variants that predispose to hepatitis B virus (HBV) persistence, we performed the first genome-wide association study in the Thai population involving 318 cases of chronic hepatitis B and 309 healthy controls after quality control measures. We detected the genome-wide significant association of the HLA class II region (HLA-DPA1/DPB1, rs7770370, p-value = 7.71 × 10-10, OR = 0.49) with HBV chronicity. Subsequent HLA allele imputation revealed HLA-DPA1*01:03 (Pc = 1.21 × 10-6, OR = 0.53), HLA-DPB1*02:01 (Pc = 2.17 × 10-3, OR = 0.50), and HLA-DQB1*06:09 (Pc = 2.17 × 10-2, OR = 0.07) as protective alleles, and HLA-DPA1*02:02 (Pc = 6.32 × 10-5, OR = 1.63), HLA-DPB1*05:01 (Pc = 1.13 × 10-4, OR = 1.72), HLA-DPB1*13:01 (Pc = 4.68 × 10-2, OR = 1.60), and HLA-DQB1*03:03 (Pc = 1.11 × 10-3, OR = 1.84) as risk alleles for HBV persistence. We also detected suggestive associations in the PLSCR1 (rs35766154), PDLIM5 (rs62321986), SGPL1 (rs144998273), and MGST1 (rs1828682) loci. Among single-nucleotide polymorphisms in the PLSCR1 locus, rs1061307 was identified as the primary functional variant by in silico/in vitro functional analysis. In addition to replicating the association of the HLA class II region, we detected novel candidate loci that provide new insights into the pathophysiology of chronic hepatitis B.

Discovery of a novel HLA-B*40 allele, HLA-B*40:02:01:30 in a Japanese individual.

HLA-B*40:02:01:30 differs from HLA-B*40:02:01:01 by one nucleotide in 3'UTR at position 2869.