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The data presented in this article are related to the research article entitled "The autoimmune risk gene ZMIZ1 is a vitamin D responsived marker of a molecular phenotype of multiple sclerosis" Fewings et al. (2017) [1]. Here we identify the set of genes correlated with ZMIZ1 in multiple cohorts, provide phenotypic details on those cohorts, and identify the genes negatively correlated with ZMIZ1 and the cells predominantly expressing those genes. We identify the metabolic pathways in which the molecular phenotype genes are over-represented. Finally, we present the flow cytometry gating strategy we have used to identify the immune cells from blood which are producing ZMIZ1 and RPS6.

Measurable residual disease (MRD) is useful for prognostication and for monitoring response to treatment in patients with acute leukaemia. MRD by multiparametric flow cytometry (MFC-MRD) utilises the leukaemia-associated immunophenotype (LAIP) and difference from normal (DfN) strategies to identify the leukaemic clone. Difficulties arise when the LAIP overlaps with normal regeneration, there is clonal evolution, or when the abnormal clone population is exceptionally small e.g., <0.01% of CD45 + cells. Such cases are reported as 'indeterminate'; however, there is little international consensus on this reporting. The relationship between clinical outcomes and indeterminate MFC-MRD is unknown. Here we determine the rate of indeterminate MFC-MRD reporting, its relationship to concurrent molecular MRD results when available, and to clinical outcomes to 12 months. We performed an internal audit of all adult testing for MFC-MRD between January and December 2021. A total of 153 consecutive patients with a diagnosis of acute leukaemia were included. Successive MFC-MRD results and clinical outcomes were recorded over a 12-month period from time of inclusion into the study. In total, 460 MFC-MRD tests from 153 patients were reviewed and 73 (16%) MFC-MRD tests from 54 (35%) patients were reported as indeterminate. The majority (70%) were at low levels between 0.01-0.1% of CD45 + cells. Compared to patients with a negative result, acute myeloid leukaemia (AML) was more frequent in patients who had an indeterminate MFC-MRD (70% vs 36%), and B-cell acute lymphoblastic leukaemia was less common (20% vs 55%). In patients with indeterminate MFC-MRD results, one-third had received either chemotherapy or allogeneic haemopoietic stem cell transplant (aHSCT) within the preceding 3 months. Agreement between MFC and molecular MRD testing was low. Patients with indeterminate MFC-MRD had leukaemia relapse rates below patients with a positive MFC-MRD, but greater than those with negative MFC-MRD (positive 33% vs indeterminate 21% vs negative 8%, p = 0.038). Overall, these findings indicate that indeterminate MFC-MRD results are more common in adults with AML and also in those who have received chemotherapy or aHSCT within the previous 3 months. We report for the first time that indeterminate MFC-MRD is a finding of potential clinical significance, which associates with a numerically higher median relapse rate within 12 months when compared to a negative MFC-MRD result., (Copyright © 2024 Royal College of Pathologists of Australasia. All rights reserved.)

Measurable residual disease (MRD) detected by flow cytometry (FC) is well established in paediatric B- lymphoblastic leukaemia (B-ALL) and adult chronic lymphocytic leukaemia (CLL), but its utility in adult B-ALL and adult acute myeloid leukaemia (AML) is less clear. In this prospective MRD study, one of the largest in Australia to date, we examined consecutive bone marrow aspirates from adult participants with B-ALL ( n = 47) and AML ( n = 87) sent for FC-MRD testing at a quaternary referral hospital in Sydney. FC-MRD results were correlated to corresponding Mol-MRD testing where available and clinical outcomes at three-month intervals over 1 year. B-ALL showed a moderate positive correlation (r s = 0.401, p < 0.001), while there was no correlation between FC-MRD and Mol-MRD for AML (r s = 0.13, p = 0.237). Five FC-MRD patterns were identified which had significant associations with relapse (X 2 (4) = 31.17(4), p > 0.001) and survival (X 2 (4) = 13.67, p = 0.008) in AML, but not in B-ALL. The three-month MRD results were also strongly associated with survival in AML, while the association in B-ALL was less evident. There was a moderate correlation between FC-MRD and Mol-MRD in B-ALL but not AML. The association of FC-MRD with relapse and survival was stronger in AML than in B-ALL. Overall, these findings suggest divergent utilities of FC-MRD in AML and B-ALL.

Although genetic and epidemiological evidence indicates vitamin D insufficiency contributes to multiple sclerosis (MS), and serum levels of vitamin D increase on treatment with cholecalciferol, recent metanalyses indicate that this vitamin D form does not ameliorate disease. Genetic variation in genes regulating vitamin D, and regulated by vitamin D, affect MS risk. We evaluated if the expression of vitamin D responsive MS risk genes could be used to assess vitamin D response in immune cells. Peripheral blood mononuclear cells (PBMCs) were isolated from healthy controls and people with MS treated with dimethyl fumarate. We assayed changes in expression of vitamin D responsive MS risk (VDRMS) genes in response to treatment with 25 hydroxy vitamin D in the presence or absence of inflammatory stimuli. Expression of CYP24A1 and other VDRMS genes was significantly altered in PBMCs treated with vitamin D in the homeostatic and inflammatory models. Gene expression in MS samples had similar responses to controls, but lower initial expression of the risk genes. Vitamin D treatment abrogated these differences. Expression of CYP24A1 and other MS risk genes in blood immune cells indicate vitamin D response and could enable assessment of immunological response to vitamin D in clinical trials and on therapy., (© 2021. Crown.)

Up to 50% of multiple sclerosis (MS) patients do not respond to interferon-beta (IFN-β) treatment and determination of response requires lengthy clinical follow-up of up to 2 years. Response predictive genetic markers would significantly improve disease management. We aimed to identify IFN-β treatment response genetic marker(s) by performing a two-stage genome-wide association study (GWAS). The GWAS was carried out using data from 151 Australian MS patients from the ANZgene/WTCCC2 MS susceptibility GWAS (responder (R)=51, intermediate responders=24 and non-responders (NR)=76). Of the single-nucleotide polymorphisms (SNP) that were validated in an independent group of 479 IFN-β-treated MS patients from Australia, Spain and Italy (R=273 and NR=206), eight showed evidence of association with treatment response. Among the replicated associations, the strongest was observed for FHIT (Fragile Histidine Triad; combined P-value 6.74 × 10 - 6 ) and followed by variants in GAPVD1 (GTPase activating protein and VPS9 domains 1; combined P-value 5.83 × 10 -5 ) and near ZNF697 (combined P-value 8.15 × 10 -5 ).

Multiple Sclerosis (MS) is a neurological condition driven in part by immune cells from the peripheral circulation, the targets for current successful therapies. The autoimmune and MS risk gene ZMIZ1 is underexpressed in blood in people with MS. We show that, from three independent sets of transcriptomic data, expression of ZMIZ1 is tightly correlated with that of hundreds of other genes. Further we show expression is partially heritable (heritability 0.26), relatively stable over time, predominantly in plasmacytoid dendritic cells and non-classical monocytes, and that levels of ZMIZ1 protein expression are reduced in MS. ZMIZ1 gene expression is increased in response to calcipotriol (1,25 Vitamin D3) (p < 0.0003) and associated with Epstein Barr Virus (EBV) EBNA-1 antibody titre (p < 0.004). MS therapies fingolimod and dimethyl fumarate altered blood ZMIZ1 gene expression compared to untreated MS. The phenotype indicates susceptibility to MS, and may correspond with clinical response and represent a novel clinical target., (Copyright © 2016. Published by Elsevier Ltd.)

The data presented in this article are related to the research article entitled "The autoimmune risk gene ZMIZ1 is a vitamin D responsived marker of a molecular phenotype of multiple sclerosis" Fewings et al. (2017) [1]. Here we identify the set of genes correlated with ZMIZ1 in multiple cohorts, provide phenotypic details on those cohorts, and identify the genes negatively correlated with ZMIZ1 and the cells predominantly expressing those genes. We identify the metabolic pathways in which the molecular phenotype genes are over-represented. Finally, we present the flow cytometry gating strategy we have used to identify the immune cells from blood which are producing ZMIZ1 and RPS6.

Multiple Sclerosis (MS) is an autoimmune disease treated by therapies targeting peripheral blood cells. We previously identified that expression of two MS-risk genes, the transcription factors EOMES and TBX21 (ET), was low in blood from MS and stable over time. Here we replicated the low ET expression in a new MS cohort (p<0.0007 for EOMES, p<0.028 for TBX21) and demonstrate longitudinal stability (p<10(-4)) and high heritability (h(2)=0.48 for EOMES) for this molecular phenotype. Genes whose expression correlated with ET, especially those controlling cell migration, further defined the phenotype. CD56+ cells and other subsets expressed lower levels of Eomes or T-bet protein and/or were under-represented in MS. EOMES and TBX21 risk SNP genotypes, and serum EBNA-1 titres were not correlated with ET expression, but HLA-DRB1*1501 genotype was. ET expression was normalised to healthy control levels with natalizumab, and was highly variable for glatiramer acetate, fingolimod, interferon-beta, dimethyl fumarate., (Crown Copyright © 2016. Published by Elsevier Inc. All rights reserved.)

Objective: We examined a cohort of adults with aquaporin-4 (AQP4) antibody-negative neuromyelitis optica/neuromyelitis optica spectrum disorder (NMO/NMOSD) for antibodies to myelin oligodendrocyte glycoprotein (MOG)., Methods: We performed a flow cytometry cell-based assay using live human lentivirus-transduced cells expressing full-length surface MOG. Serum was tested in 23 AQP4 antibody-negative NMO/NMOSD patients with bilateral and/or recurrent optic neuritis (BON, n = 11), longitudinally extensive transverse myelitis (LETM, n = 10), and sequential BON and LETM (n = 2), as well as in patients with multiple sclerosis (MS, n = 76) and controls (n = 52)., Results: MOG antibodies were detected in 9/23 AQP4 antibody-negative patients with NMO/NMOSD, compared to 1/76 patients with MS and 0/52 controls (p < 0.001). MOG antibodies were detected in 8/11 patients with BON, 0/10 patients with LETM, and 1/2 patients with sequential BON and LETM. Six of 9 MOG antibody-positive patients had a relapsing course. MOG antibody-positive patients had prominent optic disc swelling and were more likely to have a rapid response to steroid therapy and relapse on steroid cessation than MOG antibody-negative patients (p = 0.034 and p = 0.029, respectively). While 8/9 MOG antibody-positive patients had good follow-up visual acuity, one experienced sustained visual impairment, 3 had retinal nerve fiber layer thinning, and one had residual spinal disability., Conclusions: MOG antibodies have a strong association with BON and may be a useful clinical biomarker. MOG antibody-associated BON is a relapsing disorder that is frequently steroid responsive and often steroid dependent. Failure to recognize the disorder early and institute immunotherapy promptly may be associated with sustained impairment., Classification of Evidence: This study provides Class II evidence that MOG antibodies are associated with AQP4 antibody-negative BON (sensitivity 69%, 95% confidence interval [CI] 42%-87%; specificity 99%, 95% CI 93.7%-99.8%).

Background: Multiple Sclerosis (MS) is an immune-mediated disease of the central nervous system which responds to therapies targeting circulating immune cells., Objective: Our aim was to test if the T-cell activation gene expression pattern (TCAGE) we had previously described from whole blood was replicated in an independent cohort., Methods: We used RNA-seq to interrogate the whole blood transcriptomes of 72 individuals (40 healthy controls, 32 untreated MS). A cohort of 862 control individuals from the Brisbane Systems Genetics Study (BSGS) was used to assess heritability and seasonal expression. The effect of interferon beta (IFNB) therapy on expression was evaluated., Results: The MS/TCAGE association was replicated and rationalized to a single marker, ribosomal protein S6 (RPS6). Expression of RPS6 was higher in MS than controls (p<0.0004), and lower in winter than summer (p<4.6E-06). The seasonal pattern correlated with monthly UV light index (R=0.82, p<0.002), and was also identified in the BSGS cohort (p<0.0016). Variation in expression of RPS6 was not strongly heritable. RPS6 expression was reduced by IFNB therapy., Conclusions: These data support investigation of RPS6 as a potential therapeutic target and candidate biomarker for measuring clinical response to IFNB and other MS therapies, and of MS disease heterogeneity.

Genome-wide association studies have identified a linkage disequilibrium (LD) block on chromosome 12 associated with multiple sclerosis (MS), type 1 diabetes and other autoimmune diseases. This block contains CYP27B1, which catalyzes the conversion of 25 vitamin D3 (VitD3) to 1,25VitD3. Fine-mapping analysis has failed to identify which of the 17 genes in this block is most associated with MS. We have previously used a functional approach to identify the causal gene. We showed that the expression of several genes in this block in whole blood is highly associated with the MS risk allele, but not CYP27B1. Here, we show that CYP27B1 is predominantly expressed in dendritic cells (DCs). Its expression in these cells is necessary for their response to VitD, which is known to upregulate pathways involved in generating a tolerogenic DC phenotype. Here, we utilize a differentiation protocol to generate inflammatory (DC1) and tolerogenic (DC2) DCs, and show that for the MS risk allele CYP27B1 is underexpressed in DCs, especially DC2s. Of the other Chr12 LD block genes expressed in these cells, only METT21B expression was as affected by the genotype. Another gene associated with autoimmune diseases, CYP24A1, catabolizes 1,25 VitD3, and is predominantly expressed in DCs, but equally between DC1s and DC2s. Overall, these data are consistent with the hypothesis that reduced VitD pathway gene upregulation in DC2s of carriers of the risk haplotype of CYP27B1 contributes to autoimmune diseases. These data support therapeutic approaches aimed at targeting VitD effects on DCs.

We have identified a marked over-representation of transcription factors controlling differentiation of T, B, myeloid and NK cells among the 110 MS genes now known to be associated with multiple sclerosis (MS). To test if the expression of these genes might define molecular subtypes of MS, we interrogated their expression in blood in three independent cohorts of untreated MS (from Sydney and Adelaide) or clinically isolated syndrome (CIS, from San Francisco) patients. Expression of the transcription factors (TF) controlling T and NK cell differentiation, EOMES, TBX21 and other TFs was significantly lower in MS/CIS compared to healthy controls in all three cohorts. Expression was tightly correlated between these TFs, with other T/NK cell TFs, and to another downregulated gene, CCL5. Expression was stable over time, but did not predict disease phenotype. Optimal response to therapy might be indicated by normalization of expression of these genes in blood., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Type 1 diabetes (T1D) results from an autoimmune attack against the insulin-producing β-cells of the pancreas. The present study aimed to reverse T1D by gene therapy., Methods: We used a novel surgical technique, which involves isolating the liver from the circulation before the delivery of a lentiviral vector carrying furin-cleavable human insulin (INS-FUR) or empty vector to the livers of diabetic non-obese diabetic mice (NOD). This was compared with the direct injection of the vector into the portal circulation. Mice were monitored for body weight and blood glucose. Intravenous glucose tolerance tests were performed. Expression of insulin and pancreatic transcription factors was determined by the reverse transcriptase-polymerase chain reaction and immunohistochemistry and immunoelectron microscopy was used to localise insulin., Results: Using the novel surgical technique, we achieved long-term transduction (42% efficiency) of hepatocytes, restored normoglycaemia for 150 days (experimental endpoint) and re-established normal glucose tolerance. We showed the expression of β-cell transcription factors, murine insulin, glucagon and somatostatin, and hepatic storage of insulin in granules. The expression of hepatic markers, C/EBP-β, G6PC, AAT and GLUI was down-regulated in INS-FUR-treated livers. Liver function tests remained normal, with no evidence of intrahepatic inflammation or autoimmune destruction of the insulin-secreting liver tissue. By comparison, direct injection of INS-FUR reduced blood glucose levels, and no pancreatic transdifferentiation or normal glucose tolerance was observed., Conclusions: This gene therapy protocol has, for the first time, permanently reversed T1D with normal glucose tolerance in NOD mice and, as such, represents a novel therapeutic strategy for the treatment of T1D., (Copyright © 2013 John Wiley & Sons, Ltd.)