15.04.2022 (10:00 CET) - Immunogenetic studies in ME/CFS
by Riad Hajdarevic, Faculty of Medicine, University of Oslo & Oslo University Hospital, Norway
Link (Microsoft Teams): https://bit.ly/3E3or8v
Abstract: Myalgic encephalopathy/chronic fatigue syndrome (ME/CFS) is a chronic and debilitating disease that affects about 0.1-0.2% of the general population. The core symptoms are persistent debilitating fatigue, post-exertional malaise (PEM) and cognitive dysfunction. Most symptoms of ME/CFS are not disease specific. Additionally, there is a lack of both biomarkers and diagnostic tests for the disease, which makes accurate diagnosis difficult. More than 20 different patient classifications and diagnostic criteria have emerged over the last four decades. Due to this, the patient population can be quite heterogeneous in terms of clinical symptoms and the extent to which the disease impacts quality of life. There are several different theories that aim to explain the disease development of ME/CFS. In this thesis, we have taken as our starting point the growing evidence for an immunological background for ME/CFS pathogenesis. Several studies have pointed to altered NK cells, autoantibodies and T cell abnormalities in ME/CFS patients. In addition, several genetic studies reported significant associations in various immunologically relevant genes. Most of these previous studies have been suboptimal and included heterogeneous patient populations and/or few patients in total. Therefore, we aimed to gain a better understanding of the role of immunologically relevant genes and disease development of ME/CFS. To do this, we employed known strategies from genetic studies in autoimmune disease and applied them to ME/CFS. We used strict quality control and included, to the best of our knowledge, the largest cohort diagnosed with the Canadian consensus criteria.
We first followed up previously performed work by our group that reported associations between ME/CFS and HLA-C: 07: 04 and HLA-DQB1: 03: 03 alleles. The HLA (human leukocyte antigen) region consists a multitude of immunologically relevant genes in addition to the HLA genes, and there is extensive and complex linkage disequilibrium (LD) in the region. The previously observed association signals in the HLA region were fine-mapped by genotyping five additional classical HLA loci and 5,342 SNPs (single nucleotide variants) in 427 Norwegian ME/CFS patients, diagnosed according to the Canadian consensus criteria, and 480 healthy Norwegian controls. The analysis revealed two independent association signals (p ≤ 0.001) represented by the genetic variants rs4711249 in the HLA class I region and rs9275582 in the HLA class II region. The primary association signal in the HLA class II region was located in the vicinity of the HLA-DQ genetic region, most likely due to the HLA-DQB1 gene. In particular, amino acid position 57 (aspartic acid / alanine) in the peptide binding pit of HLA-DQB1, or an SNP upstream of HLA-DQB1 seemed to explain the association signal we observed in the HLA class II region. In the HLA class I region, the putative primary locus was not as clear and could possibly lie outside the classical HLA genes (the association signal spans several genes DDR1, GTF2H4, VARS2, SFTA2 and DPCR1) with expression levels influenced by the ME/CFS associated SNP genotypes. Interestingly, we also observed that > 60% of the patients who responded to cyclophosphamide treatment for ME/CFS had either the rs4711249 risk allele and/or DQB1* 03:03 versus 12% of the patients who did not respond to the treatment. Our findings suggest the involvement of the HLA region, and in particular the HLA-DQB1 gene, in ME/CFS. Although our study is the largest to date, it is still a relatively small study in the context of genetic studies. Our findings need to be replicated in much larger, statistically more representative, cohorts. In particular, it is necessary to investigate the involvement of HLA- 12 DQB1, a gene that contains alleles that increase the risk of several established autoimmune diseases such as celiac disease. Additionally, we aimed to investigate immunologically relevant genes using a genotyping array (iChip) targeting immunological gene regions previously associated with different autoimmune diseases. In addition to the Norwegian cohort of 427 ME/CFS patients (the Canadian consensus criteria), we also analyzed data from two replication cohorts, a Danish one of 460 ME/CFS patients (Canadian consensus criteria) and a data set from the UK Biobank of 2105 self-reported CFS patients. To the best of our knowledge, this is the first ME/CFS genetic association study of this magnitude and it included more than 2,900 patients in total (of whom 887 are diagnosed according to Canadian consensus criteria). We found no ME/CFS risk variants with a genome wide significance level (p <5x10-8), but we identified six gene regions (TPPP, LINC00333, RIN3. IGFBP/IGFBP3, IZUMO1/MAMSTR and ZBTB46/STMN3) with possible association with ME/CFS which require further follow-up in future studies in order to assess whether they are real findings or not. Interestingly, these genes are expressed in disease-relevant tissue, e.g. brain, nerve, skeletal muscle and blood, including immune cells (subgroups of T cells, B cells, NK cells and monocytes). Furthermore, several of the ME/CFS associated SNP genotypes are associated with differential expression levels of these genes. Although we could not identify statistically convincing associations with genetic variants across the three cohorts, we believe that our data sets and analysis represent an important step in the ME/CFS research field. Our study demonstrated that for the future understanding of the genetic architecture of ME/CFS much larger studies are required to established reliable associations. As last part of our study we wanted to investigate previous findings from a genome wide association study of 42 ME/CFS patients who reported significant association with two SNPs in the T cell receptor alpha (TRA) locus (P-value <5x10-8). In order to replicate these previously reported findings, we used a large Norwegian ME/CFS cohort (409 cases and 810 controls) and data from the UK Biobank (2105 cases and 4786 controls). We examined a number of SNPs in the TRA locus, including the two previous ME/CFS-associated variants, rs11157573 and rs17255510. No statistically significant associations were observed in either the Norwegian cohort or UK biobank cohorts. Nevertheless, other SNPs in the region showed weak signs of association (P-value <0.05) in the UK Biobank cohort and meta-analyzes of Norwegian and UK Biobank cohorts, but did not remain associated after applying correction for multiple testing. Thus, we could not confirm associations with genetic variants in the TRA locus in this study.