Human Leukocyte Antigen Profile Predicts Severity of Autoimmune Liver Disease in Children of European Ancestry.

BACKGROUND AND AIMS: Genetic predisposition to autoimmune hepatitis (AIH) in adults is associated with possession of human leukocyte antigen (HLA) class I (A*01, B*08) and class II (DRB1*03, -04, -07, or -13) alleles, depending on geographic region. Juvenile autoimmune liver disease (AILD) comprises AIH-1, AIH-2, and autoimmune sclerosing cholangitis (ASC), which are phenotypically different from their adult counterparts. We aimed to define the relationship between HLA profile and disease course, severity, and outcome in juvenile AILD. APPROACH AND RESULTS: We studied 236 children of European ancestry (152 female [64%], median age 11.15 years, range 0.8-17), including 100 with AIH-1, 59 with AIH-2, and 77 with ASC. The follow-up period was from 1977 to June 2019 (median 14.5 years). Class I and II HLA genotyping was performed using PCR/sequence-specific primers. HLA B*08, -DRB1*03, and the A1-B8-DR3 haplotype impart predisposition to all three forms of AILD. Homozygosity for DRB1*03 represented the strongest risk factor (8.8). HLA DRB1*04, which independently confers susceptibility to AIH in adults, was infrequent in AIH-1 and ASC, suggesting protection; and DRB1*15 (DR15) was protective against all forms of AILD. Distinct HLA class II alleles predispose to the different subgroups of juvenile AILD: DRB1*03 to AIH-1, DRB1*13 to ASC, and DRB1*07 to AIH-2. Possession of homozygous DRB1*03 or of DRB1*13 is associated with fibrosis at disease onset, and possession of these two genes in addition to DRB1*07 is associated with a more severe disease in all three subgroups. CONCLUSIONS: Unique HLA profiles are seen in each subgroup of juvenile AILD. HLA genotype might be useful in predicting responsiveness to immunosuppressive treatment and course.

Increasing Levels of Serum Heat Shock Protein 70 Precede the Development of AIDS-Defining Non-Hodgkin Lymphoma Among Carriers of HLA-B8-DR3.

BACKGROUND: We hypothesized that carriage of presumably high Hsp70-producing gene variants on a specific human major histocompatibility complex haplotype, the 8.1 ancestral haplotype (8.1AH), may predispose HIV-infected individuals to AIDS-non-Hodgkin lymphoma (NHL). SETTING: We compared serum Hsp70 levels in the years preceding the diagnosis of AIDS-NHL in a matched case-control study (n = 151 pairs) nested in the Multicenter AIDS Cohort Study. METHODS: We tested the impact of 8.1AH-specific single-nucleotide polymorphism (SNP) and joint SNP-human leukocyte antigen extended haplotypes previously associated with AIDS-NHL in the Multicenter AIDS Cohort Study on the circulating Hsp70 levels in mixed linear models. RESULTS: We report elevated serum levels of Hsp70 in the 4 years preceding the diagnosis of AIDS-NHL in cases that carry 8.1AH, but not in noncarrier cases and not in carrier- or non-carrier-matched controls. The strongest predictor of higher serum Hsp70 was the haplotype A-G-A-C formed by SNPs rs537160(A) and rs1270942(G) in the complement factor CFB gene cluster, and rs2072633(A) and rs6467(C) in nearby RDBP and CYP21A2 located 70 Kb apart from the Hsp70 gene cluster. The association with A-G-A-C haplotype (beta = 0.718; standard error = 0.182; P = 0.0002) and with other 8.1AH-specific haplotypes including the high-producing tumor necrosis factor-alpha haplotype rs909253(G)-rs1800629(A) (beta = 0.308; standard error = 0.140; P = 0.032) were observed only with NHL identified as an AIDS-defining condition, but not as a post-AIDS condition, nor in combined AIDS and post-AIDS cases. CONCLUSION: Our combined genetic and functional approach suggests that the altered level of Hsp70 is a correlate of 8.1AH-mediated AIDS-NHL. Further investigation of the Hsp70 gene cluster and nearby loci that are tagged by A-G-A-C could better elucidate the genetic determinants of the malignancy.

Identification of the novel HLA-B*08:181 allele in a volunteer donor for hematopoietic stem cells.

HLA-B*08:181 differs from HLA-B*08:01:01:01 in codon 94, 95, 97 and 99 of exon 3.

HLA-B gene somatic insertion/deletion mutations in patients with acute myelogenous leukaemia.

Loss of heterozygosity is considered to be the most common type of tumour-specific somatic mutation of the human leucocyte antigens (HLA) genes in patients with haematological malignancies. Nevertheless, subtle DNA sequence changes, namely short insertions/deletions, may also abolish the expression of HLA molecules and interfere with routine HLA typing. Two male patients with acute myelogenous leukaemia (AML) were indicated for the search of a suitable donor for allogeneic haematopoietic stem cell transplantation (aHSCT). The patients and their relatives were initially HLA typed by serological and DNA techniques at a low-resolution level. The HLA high-resolution (HR) type was obtained by means of sequencing-based typing (SBT). In both cases, anomalous frameshifts in the sequence were observed in the HLA-B gene, namely in exon 3 (Case 1, heterozygous deletion of two bases) and exon 4 (Case 2, heterozygous insertion of two bases). In the second case, the insertion variant was associated with a loss of HLA-B8 expression. To reveal whether these sequence patterns may be caused by somatic mutations in the malignant cells, blood sample in remission (Case 1) and buccal swab sample (Case 2) were collected from the patients. In an important manner, the SBT in these germline samples revealed common HLA-B*07:02,*15:01 (Case 1) and HLA-B*08:01,*35:02 (Case 2) types with no evidence for the sequence alteration observed in the initial samples. In conclusion, the insertion/deletion sequence variants of the HLA-B gene in two patients were limited to the initial blood samples with a substantial proportion of AML cells and thus may be attributed to the somatic mutation in the malignant cells. HLA somatic mutations should be taken into account in patients with haematological malignancies to prevent HLA mistyping and inappropriate selection of an aHSCT donor.

Novel genetic loci associated HLA-B*08:01 positive myasthenia gravis.

OBJECTIVE: To identify potential causative markers involved in the development of early-onset myasthenia gravis (EOMG) in the MHC and non-MHC regions that may interact with the HLA-B*08:01 allele. METHODS: We analyzed 583 MG patients and identified 5 patients homozygous for the disease-associated ancestral haplotype 8.1 (HLA-A*01:01, B*08:01, DRB1*03:01, DQB1*02:01). We also analyzed more than 9000 controls and selected 24 for further investigation. We subsequently conducted a fine mapping analysis through high-throughput sequencing of the MHC region (from upstream of the GPα5 gene to downstream of the ZBTB9 gene). For the interaction analysis we analyzed a total of 150,090 SNPs equally distributed throughout the genome in the individuals that were homozygous for the main susceptibility HLA allele HLA-B*08:01 and investigated the expression of the genes located close to the observed susceptibility variants. RESULTS: The overall coverage of the 4.79 Mb MHC region ranged between 96.57% and 97.41%. We identified 705 new variants in the MHC region (673 SNPs and 32 InDels). However, no significant differences were found between patients and controls within the MHC region of the ancestral 8.1 haplotype. As the susceptibility gene is considered to be located close to the HLA-B locus, complete sequencing of the surrounding 200 kb was carried out in the 5 patients and 24 controls. No significant differences where observed, suggesting that the HLA-B molecule itself is the susceptibility factor for EOMG. We also observed two new susceptibility loci specific for MG HLA*08:01 patients (P < 3.33 × 10-7). These loci map to an intronic OVCH1 variant (rs10492374; P = 1.90 × 10-8) and a 5' downstream CNPY2 variant (rs10783780; P = 3.33 × 10-7) on chromosome 12. Individuals heterozygous for GA*rs10492374 showed an increased expression of the OVCH1 gene. The rs10783780 genotypes were not associated with CNPY2 mRNA levels, but the MG HLA*08:01 patients present a lower expression of this gene than the healthy controls. CONCLUSION: Our results showed that when we control for the influence of the ancestral haplotype 8.1, no polymorphism was demonstrated to be associated with EOMG development within the MHC region suggesting that the HLA-B*08:01 allele is the unique genetic factor within the HLA region responsible for EOMG development in patients who carry the ancestral haplotype 8.1. Our study also identified two novel polymorphisms as risk factors for MG HLA-B*08:01 positive patients which regulate the expression of the OVCH1 and CNYP2 genes.

The novel HLA-B*08:183 allele identified by sequence-based typing in a Caucasian leukemia patient.

HLA-B*08:183 differs from HLA-B*08:01:01 by a single substitution in exon 5.

Identification of a novel HLA-B allele, HLA-B*08:177, in a Hungarian patient and her HLA identical sibling.

The newly detected HLA-B*08:177 differs from HLA-B*08:01:01:01 by 1 single nucleotide substitution at position 365 of exon 3.

Gliadin-Specific CD8+ T Cell Responses Restricted by HLA Class I A*0101 and B*0801 Molecules in Celiac Disease Patients.

Initial studies associated the HLA class I A*01 and B*08 alleles with celiac disease (CD) susceptibility. Subsequent analyses showed a primary association with HLA class II alleles encoding for the HLA DQ2.5 molecule. Because of the strong linkage disequilibrium of A*01 and B*08 alleles with the DR3-DQ2.5 haplotype and a recent genome-wide association study indicating that B*08 and B*39 are predisposing genes, the etiologic role of HLA class I in CD pathogenesis needs to be addressed. We screened gliadin proteins (2α-, 2ω-, and 2γ-gliadin) using bioinformatic algorithms for the presence of peptides predicted to bind A*0101 and B*0801 molecules. The top 1% scoring 9- and 10-mer peptides (N = 97, total) were synthesized and tested in binding assays using purified A*0101 and B*0801 molecules. Twenty of ninety-seven peptides bound B*0801 and only 3 of 97 bound A*0101 with high affinity (IC50 < 500 nM). These 23 gliadin peptides were next assayed by IFN-γ ELISPOT for recognition in peripheral blood cells of CD patients and healthy controls carrying the A*0101 and/or B*0801 genes and in A*0101/B*0801- CD patients. Ten of the twenty-three peptides assayed recalled IFN-γ responses mediated by CD8+ T cells in A*0101/B*0801+ patients with CD. Two peptides were restricted by A*0101, and eight were restricted by B*0801. Of note, 50% (5/10) of CD8+ T cell epitopes mapped within the γ-gliadins. Our results highlight the value of predicted binding to HLA molecules for identifying gliadin epitopes and demonstrate that HLA class I molecules restrict the anti-gluten T cell response in CD patients.

Clinical and genetic associations of radiographic sacroiliitis and its different patterns in psoriatic arthritis.

OBJECTIVES: We aimed to 1) identify clinical and genetic associations of sacroiliitis (SI) in patients with psoriatic arthritis (PsA), and 2) describe the different radiographic patterns of SI in PsA and their clinical and genetic associations. METHODS: 283 PsA patients, fulfilling CASPAR criteria, underwent detailed skin and rheumatologic assessments. In addition, HLA-B*27 and B*080101 status was recorded, which have been shown as the key genetic markers of radiographic SI in PsA. Grade 2 Unilateral or bilateral radiographic changes of SI were required for inclusion and involvement was further defined as asymmetrical or symmetrical. RESULTS: 70 patients (25%) had radiographic SI; all either with a present or past history of backache. Regression analysis demonstrated a significant association of SI with peripheral joint erosions (p=0.043), PASI maximum (p=0.041), younger age of PsA onset (p=<0.001), presence of HLA-B*0801 (p=0.002) and only marginal significance with HLA-B*2705 (p=0.059). Asymmetrical SI was noted in 51 patients (73%). In striking contrast to those patients with symmetrical SI, patients with asymmetrical SI were more likely to be female (p=0.04), have a trend towards more severe nail disease (p=0.08) and peripheral joint erosions (p=0.08), more osteolysis (p=0.01), more HLA-B*0801 positivity (p=0.001) and much less HLA-B*270502 positivity (p=<0.001). CONCLUSIONS: PsA developing at a younger age, severe skin disease, peripheral joint erosions, and HLA-B*0801 are significantly associated with SI, and there was only a marginal trend towards significance for HLA-B*2705. HLA-B*27 positive Axial-PsA patients resemble AS, while HLA-B*0801 positive Axial-PsA patients have asymmetrical and/or unilateral SI, which are typical of PsA.

Identification of 3 novel HLA-B alleles: B*08:173, B*18:72:03 and B*53:05:02.

A total of 3 novel human leukocyte antigen-B (HLA-B) alleles were detected by next generation sequencing and confirmed by monoallelic sequencing.

Cell surface expression level variation between two common Human Leukocyte Antigen alleles, HLA-A2 and HLA-B8, is dependent on the structure of the C terminal part of the alpha 2 and the alpha 3 domains.

Constitutive cell surface expression of Human Leukocyte Antigen (HLA) class I antigens vary extremely from tissue to tissue and individual antigens may differ widely in expression levels. Down-regulation of class I expression is a known immune evasive mechanism used by cancer cells and viruses. Moreover, recent observations suggest that even minor differences in expression levels may influence the course of viral infections and the frequency of complications to stem cell transplantation. We have shown that some human multipotent stem cells have high expression of HLA-A while HLA-B is only weakly expressed, and demonstrate here that this is also the case for the human embryonic kidney cell line HEK293T. Using quantitative flow cytometry and quantitative polymerase chain reaction we found expression levels of endogenous HLA-A3 (median 71,204 molecules per cell) 9.2-fold higher than the expression of-B7 (P = 0.002). Transfection experiments with full-length HLA-A2 and -B8 encoding plasmids confirmed this (54,031 molecules per cell vs. 2,466, respectively, P = 0.001) independently of transcript levels suggesting a post-transcriptional regulation. Using chimeric constructs we found that the cytoplasmic tail and the transmembrane region had no impact on the differential cell surface expression. In contrast, ~65% of the difference could be mapped to the six C-terminal amino acids of the alpha 2 domain and the alpha 3 domain (amino acids 176-284), i.e. amino acids not previously shown to be of importance for differential expression levels of HLA class I molecules. We suggest that the differential cell surface expression of two common HLA-A and-B alleles is regulated by a post-translational mechanism that may involve hitherto unrecognized molecules.

The autoimmune spectrum of myasthenia gravis: a Swedish population-based study.

OBJECTIVES: To determine the prevalence of myasthenia gravis (MG) and the rate of concurrent autoimmune diseases in patients with MG. DESIGN AND SETTING: Using the Swedish health and population registers, during the period 2005-2010, we conducted a nested case-control study of patients with MG (n = 2045) with five age- and sex-matched population-based controls per case. Register-based MG diagnosis was validated against the Stockholm MG Cohort. Similar nested case-control studies were conducted in patients with multiple sclerosis (MS), as a neuroinflammatory disease control, and siblings of patients with MG. MAIN OUTCOME MEASURE: Odds ratios (ORs) and their 95% confidence intervals (CIs) were calculated as a measure of the association between MG and other autoimmune diseases. RESULTS: The prevalence of MG was 24.8/100,000, and patients with MG had an increased risk of another autoimmune disease compared to controls (22.0% vs. 8.9%; OR: 2.82, 95% CI: 2.49-3.20); this risk was stronger amongst younger persons and women. Polymyositis/dermatomyositis, systemic lupus erythematosus and Addison's disease, three conditions regulated by the HLA-B8-DR3 haplotype, were most strongly associated with MG, especially early-onset disease. HLA typing in the Stockholm MG Cohort showed that early-onset MG was indeed dominated by HLA-B8-DR3. The risk of another autoimmune disease was increased in both patients with MS and siblings of patients with MG, compared to their respective controls, but to a lesser extent than in patients with MG. CONCLUSIONS: Our results suggest that MG shares risk factors with other autoimmune diseases, to a greater degree than MS, with a particular role of the HLA-B8-DR3 haplotype, especially amongst younger and female patients.

A new HLA allele, HLA-B*08:108, described in two unrelated Spanish individuals.

HLA-B*08:108 shows one nucleotide difference regarding B*08:01:01 at codon 109 (CTC>TTC, L109>F109).

Autoantibody and human leukocyte antigen profiles in children with autoimmune liver disease and their first-degree relatives.

OBJECTIVE: Familial clustering of juvenile autoimmune liver disease (AILD), including autoimmune hepatitis and autoimmune sclerosing cholangitis (ASC), is rare, despite a high prevalence of autoimmune disorders in AILD families. METHODS: To investigate this discrepancy, we measured autoantibodies diagnostic for AILD, anti-nuclear, anti-smooth muscle, anti-liver kidney microsomal type 1, anti-liver cytosol type 1, and anti-soluble liver antigen antibodies, and human leukocyte antigen profiles in 31 patients and 65 of their first-degree relatives (FDR). The autoantibody profile was compared with that of 42 healthy subjects (HS). RESULTS: Autoantibodies were detected in 71% (22/31) patients. Anti-nuclear antibody or anti-smooth muscle antibody were present in 4/65 FDR (6.2%). HS were negative for all autoantibodies. The frequencies of homozygous HLA DRB1*0301 (DR3) genes and haplotype A1-B8-DR3 were higher in the patients (25% and 43%) than in FDR (9% and 27%) and HS (0% and 16%). The frequencies of disease-protective genes DR4 and/or DR15 were lower in the patients (25%) than in FDR (42%) and HS (42%). Only 1 family contained 2 patients with AILD, 1 with ASC and 1 with primary sclerosing cholangitis. Both patients possessed A1-B8-DR3 genes, the ASC being homozygous and the primary sclerosing cholangitis heterozygous. Six FDR had nonhepatic autoimmune disorders, none being autoantibody positive. CONCLUSIONS: Homozygosity for DR3 plays a major role in the predisposition to juvenile AILD. Diagnostic autoantibodies for AILD are rare among patients' FDR and not linked to clinical manifestation of AILD.

Detection of a novel HLA-B allele, HLA-B*08:111, in an Italian bone marrow donor.

The newly detected HLA-B*08:111 allele shows two nucleotide differences from B*08:01:01 in codons 113 and 114.

Risk for myasthenia gravis maps to a (151) Pro→Ala change in TNIP1 and to human leukocyte antigen-B*08.

OBJECTIVE: The objective of this study is to comprehensively define the genetic basis of early onset myasthenia gravis (EOMG). METHODS: We have carried out a 2-stage genome-wide association study on a total of 649 North European EOMG patients. Cases were matched 1:4 with controls of European ancestry. We performed imputation and conditional analyses across the major histocompatibility complex, as well as in the top regions of association outside the human leukocyte antigen (HLA) region. RESULTS: We observed the strongest association in the HLA class I region at rs7750641 (p = 1.2 × 10(-92) ; odds ratio [OR], 6.25). By imputation and conditional analyses, HLA-B*08 proves to be the major associated allele (p = 2.87 × 10(-113) ; OR, 6.41). In addition to the expected association with PTPN22 (rs2476601; OR, 1.71; p = 8.2 × 10(-10) ), an imputed coding variant (rs2233290) at position 151 (Pro→Ala) in the TNFAIP3-interacting protein 1, TNIP1, confers even stronger risk than PTPN22 (OR, 1.91; p = 3.2 × 10(-10) ). INTERPRETATION: The association at TNIP1 in EOMG implies disease mechanisms involving ubiquitin-dependent dysregulation of NF-κB signaling. The localization of the major HLA signal to the HLA-B*08 allele suggests that CD8(+) T cells may play a key role in disease initiation or pathogenesis.

A structural basis for varied αß TCR usage against an immunodominant EBV antigen restricted to a HLA-B8 molecule.

EBV is a ubiquitous and persistent human pathogen, kept in check by the cytotoxic T cell response. In this study, we investigated how three TCRs, which differ in their T cell immunodominance hierarchies and gene usage, interact with the same EBV determinant (FLRGRAYGL), bound to the same Ag-presenting molecule, HLA-B8. We found that the three TCRs exhibit differing fine specificities for the viral Ag. Further, via structural and biophysical approaches, we demonstrated that the viral Ag provides the greatest energetic contribution to the TCR-peptide-HLA interaction, while focusing on a few adjacent HLA-based interactions to further tune fine-specificity requirements. Thus, the TCR engages the peptide-HLA with the viral Ag as the main glue, such that neighboring TCR-MHC interactions are recruited as a supportive adhesive. Collectively, we provide a portrait of how the host's adaptive immune response differentially engages a common viral Ag.

[Pathogenesis and diagnosis of Sjögren's syndrome]. / Pathogenese und Diagnostik des Sjögren-Syndroms.

Sjögren's syndrome is a common autoimmune disorder. Several genetic risk factors such as STAT-4, ILT6 and the haplotype HLA-B8/DR3 have been identified. In addition, there are environmental risk factors, possibly chronic viral infections. In the pathophysiology of Sjögren's syndrome T and B cells infiltrate the salivary and lacrimal glands. As a consequence of the destruction of glandular cells by cytotoxic T cells, production of cytokines and autoantibodies inhibiting glandular function, the production of saliva and tears is decreased. The feeling of dry eyes and mouth is frequently not noticed by the patients. Therefore, Sjögren's syndrome should also be considered when extraglandular manifestations such as vasculitis, polyneuropathy or arthritis occur, even when the patients do not complain of dry eyes and mouth. Establishing the diagnosis of Sjögren's syndrome requires verification of reduced glandular function, for example using Schirmer's test and the Saxon test. The confirmation of Sjögren's syndrome as a cause of sicca syndrome is subsequently performed by the detection of autoantibodies against Ro (SS-A) and La (SS-B) and/or by a salivary gland biopsy.

Analysis of TACI mutations in CVID & RESPI patients who have inherited HLA B*44 or HLA*B8.

BACKGROUND: Recent reports have suggested that Common Variable Immunodeficieny (CVID) can present as an autosomal dominant trait dependent on the inheritance of a set of uncommon mutations/alleles of TACI (transmembrane activator and calcium-modulator and cyclophilin ligand interactor) involving exons 3 or 4. Penetrance, however, appears to be incomplete. Among our clinic population, the greatest genetic linkage for CVID is to the major histocompatibility complex (MHC) on chromosome 6. The majority of our patients have inherited HLA *DQ2, *DR7, *DR3(17), *B8, and/or *B44. Of these, HLA*B44 was present in almost half of the patients and was thus the most common susceptibility allele. HLA *B44 was also found to be over-represented among patients who presented to our clinic with adult-onset recurrent sinopulmonary infections (RESPI) and normal serum immunoglobulin levels, a cohort that included first and second degree relatives of patients with CVID. One of the two original reports of the association between TACI and CVID also reported Human Leukocyte Antigen (HLA) haplotypes. Of 13 affected subjects, nine had inherited HLA *B8 and six had inherited HLA B44. This raised the possibility that TACI mutations might synergize with MHC class I alleles to enhance susceptibility to humoral immune deficiency. METHODS: We identified 63 CVID patients irrespective of HLA status and 13 RESPI patients who had inherited HLA*B44. To evaluate for mutations in the gene for TACI, we PCR amplified and sequenced TACI exons 3 and 4 from these patients. RESULTS: Of the 76 patients, eleven proved heterozygous for a previously reported, silent T->G polymorphism [rs35062843] at proline 97 in exon 3. However, none of the 13 RESPI patients and only one of the 63 CVID patients inherited a TACI allele previously associated with CVID. This patient was heterozygous for the TACI A181E allele (exon 4). She did not carry *DQ2, *DR7, *DR3(17), *B8, or *B44. CONCLUSION: These findings suggest that TACI mutations are unlikely to play a critical role in creating susceptibility to CVID among patients with previously recognized MHC class I and class II susceptibility alleles. Supported by NIH/USIDNET N01-AI30070, NIH R21 AI079741 and NIH M01-RR00032.