Donor regulatory T cells rapidly adapt to recipient tissues to control murine acute graft-versus-host disease.

The adoptive transfer of regulatory T cells is a promising strategy to prevent graft-versus-host disease after allogeneic bone marrow transplantation. Here, we use a major histocompatibility complex-mismatched mouse model to follow the fate of in vitro expanded donor regulatory T cells upon migration to target organs. Employing comprehensive gene expression and repertoire profiling, we show that they retain their suppressive function and plasticity after transfer. Upon entering non-lymphoid tissues, donor regulatory T cells acquire organ-specific gene expression profiles resembling tissue-resident cells and activate hallmark suppressive and cytotoxic pathways, most evidently in the colon, when co-transplanted with graft-versus-host disease-inducing conventional T cells. Dominant T cell receptor clonotypes overlap between organs and across recipients and their relative abundance correlates with protection efficacy. Thus, this study reveals donor regulatory T cell selection and adaptation mechanisms in target organs and highlights protective features of Treg to guide the development of improved graft-versus-host disease prevention strategies.

D-2-hydroxyglutarate supports a tolerogenic phenotype with lowered major histocompatibility class II expression in non-malignant dendritic cells and acute myeloid leukemia cells.

D-2-hydroxyglutarate (D-2-HG) accumulates in primary acute myeloid leukemia (AML) patients with mutated isocitrate dehydrogenase (IDH) and other malignancies. D-2-HG suppresses antitumor T cell immunity but little is known about potential effects on non-malignant myeloid cells. Here we show that D-2-HG impairs human but not murine dendritic cell (DC) differentiation, resulting in a tolerogenic phenotype with low major histocompatibility (MHC) class II expression. In line, IDH-mutated AML blasts exhibited lower expression of HLA-DP and were less susceptible to lysis by HLA-DP-specific T cells. Interestingly, D-2-HG reprogrammed metabolism towards increased lactate production in DCs and AML besides its expected impact on DNA demethylation. Vitamin C accelerated DNA demethylation, but only the combination of vitamin C and glycolytic inhibition lowered lactate levels and supported MHC class II expression. Our results indicate an unexpected link between the immunosuppressive metabolites 2-HG and lactic acid and suggest a potentially novel therapeutic strategy with combinations of anti-glycolytic drugs and epigenetic modulators (hypomethylating agents) or other therapeutics for the treatment of AML.

The effector program of human CD8 T cells supports tissue remodeling.

CD8 T lymphocytes are classically viewed as cytotoxic T cells. Whether human CD8 T cells can, in parallel, induce a tissue regeneration program is poorly understood. Here, antigen-specific assay systems revealed that human CD8 T cells not only mediated cytotoxicity but also promoted tissue remodeling. Activated CD8 T cells could produce the epidermal growth factor receptor (EGFR)-ligand amphiregulin (AREG) and sensitize epithelial cells for enhanced regeneration potential. Blocking the EGFR or the effector cytokines IFN-γ and TNF could inhibit tissue remodeling. This regenerative program enhanced tumor spheroid and stem cell-mediated organoid growth. Using single-cell gene expression analysis, we identified an AREG+, tissue-resident CD8 T cell population in skin and adipose tissue from patients undergoing abdominal wall or abdominoplasty surgery. These tissue-resident CD8 T cells showed a strong TCR clonal relation to blood PD1+TIGIT+ CD8 T cells with tissue remodeling abilities. These findings may help to understand the complex CD8 biology in tumors and could become relevant for the design of therapeutic T cell products.

Tumor heterogeneity and tumor-microglia interactions in primary and recurrent IDH1-mutant gliomas.

The isocitrate dehydrogenase (IDH) gene is recurrently mutated in adult diffuse gliomas. IDH-mutant gliomas are categorized into oligodendrogliomas and astrocytomas, each with unique pathological features. Here, we use single-nucleus RNA and ATAC sequencing to compare the molecular heterogeneity of these glioma subtypes. In addition to astrocyte-like, oligodendrocyte progenitor-like, and cycling tumor subpopulations, a tumor population enriched for ribosomal genes and translation elongation factors is primarily present in oligodendrogliomas. Longitudinal analysis of astrocytomas indicates that the proportion of tumor subpopulations remains stable in recurrent tumors. Analysis of tumor-associated microglia/macrophages (TAMs) reveals significant differences between oligodendrogliomas, with astrocytomas harboring inflammatory TAMs expressing phosphorylated STAT1, as confirmed by immunohistochemistry. Furthermore, inferred receptor-ligand interactions between tumor subpopulations and TAMs may contribute to TAM state diversity. Overall, our study sheds light on distinct tumor populations, TAM heterogeneity, TAM-tumor interactions in IDH-mutant glioma subtypes, and the relative stability of tumor subpopulations in recurrent astrocytomas.

Changes in adenoviral chromatin organization precede early gene activation upon infection.

Within the virion, adenovirus DNA associates with the virus-encoded, protamine-like structural protein pVII. Whether this association is organized, and how genome packaging changes during infection and subsequent transcriptional activation is currently unclear. Here, we combined RNA-seq, MNase-seq, ChIP-seq, and single genome imaging during early adenovirus infection to unveil the structure- and time-resolved dynamics of viral chromatin changes as well as their correlation with gene transcription. Our MNase mapping data indicates that the adenoviral genome is arranged in precisely positioned nucleoprotein particles with nucleosome-like characteristics, that we term adenosomes. We identified 238 adenosomes that are positioned by a DNA sequence code and protect about 60-70 bp of DNA. The incoming adenoviral genome is more accessible at early gene loci that undergo additional chromatin de-condensation upon infection. Histone H3.3 containing nucleosomes specifically replaces pVII at distinct genomic sites and at the transcription start sites of early genes. Acetylation of H3.3 is predominant at the transcription start sites and precedes transcriptional activation. Based on our results, we propose a central role for the viral pVII nucleoprotein architecture, which is required for the dynamic structural changes during early infection, including the regulation of nucleosome assembly prior to transcription initiation. Our study thus may aid the rational development of recombinant adenoviral vectors exhibiting sustained expression in gene therapy.

A multilocus genetic risk score for obesity: Association with BMI and metabolic alterations in a cohort with severe obesity.

Genome wide association studies have identified numerous single nucleotide polymorphisms (SNPs) associated with obesity, yet effect sizes of individual SNPs are small. Therefore, the aim of our study was to investigate whether a genetic risk score (GRS) comprising risk alleles of SNPs identified in the GIANT consortium meta-analyses shows association with body mass index (BMI) and other BMI related metabolic alterations in a cohort with an extreme phenotype. Genotyping of 93 SNPs was performed in 314 obese individuals (mean BMI 40.5 ± 7.8 kg/m², aged 45 ± 12 years), participating in a standardized weight reduction program, and in 74 lean controls (mean BMI 24.6 ± 3.3 kg/m², aged 41.7 ± 13.4 years). Allele numbers of all 93 SNPs were added to a GRS. Anthropometric parameters, parameters of glucose/insulin and lipid metabolism were assessed standardized after a 12 hours fast. GRS was significantly different between controls and obese individuals (unweighted GRS: 86.6 vs 89.0, P = .002; weighted GRS: 84.9 vs 88.3, P = .005). Furthermore, linear regression analysis showed significant associations of GRS with BMI ( P < .0001), weight ( P = .0005), waist circumference ( P = .0039), fat mass ( P < .0001) and epicardial fat thickness ( P = .0032), yet with small effect sizes ( r ² < 0.06). In conclusion, in our study GRS could differentiate between extreme obese and lean individuals, and was associated with BMI and its related traits, yet with small effect sizes.

Glutamine synthetase expression rescues human dendritic cell survival in a glutamine-deprived environment.

Introduction: Glutamine deficiency is a well-known feature of the tumor environment. Here we analyzed the impact of glutamine deprivation on human myeloid cell survival and function. Methods: Different types of myeloid cells were cultured in the absence or presence of glutamine and/or with L-methionine-S-sulfoximine (MSO), an irreversible glutamine synthetase (GS) inhibitor. GS expression was analyzed on mRNA and protein level. GS activity and the conversion of glutamate to glutamine by myeloid cells was followed by 13C tracing analyses. Results: The absence of extracellular glutamine only slightly affected postmitotic human monocyte to dendritic cell (DC) differentiation, function and survival. Similar results were obtained for monocyte-derived macrophages. In contrast, proliferation of the monocytic leukemia cell line THP-1 was significantly suppressed. While macrophages exhibited high constitutive GS expression, glutamine deprivation induced GS in DC and THP-1. Accordingly, proliferation of THP-1 was rescued by addition of the GS substrate glutamate and 13C tracing analyses revealed conversion of glutamate to glutamine. Supplementation with the GS inhibitor MSO reduced the survival of DC and macrophages and counteracted the proliferation rescue of THP-1 by glutamate. Discussion: Our results show that GS supports myeloid cell survival in a glutamine poor environment. Notably, in addition to suppressing proliferation and survival of tumor cells, the blockade of GS also targets immune cells such as DCs and macrophages.

Biosensors for inflammation as a strategy to engineer regulatory T cells for cell therapy.

Engineered regulatory T cell (Treg cell) therapy is a promising strategy to treat patients suffering from inflammatory diseases, autoimmunity, and transplant rejection. However, in many cases, disease-related antigens that can be targeted by Treg cells are not available. In this study, we introduce a class of synthetic biosensors, named artificial immune receptors (AIRs), for murine and human Treg cells. AIRs consist of three domains: (a) extracellular binding domain of a tumor necrosis factor (TNF)-receptor superfamily member, (b) intracellular costimulatory signaling domain of CD28, and (c) T cell receptor signaling domain of CD3-ζ chain. These AIR receptors equip Treg cells with an inflammation-sensing machinery and translate this environmental information into a CD3-ζ chain-dependent TCR-activation program. Different AIRs were generated, recognizing the inflammatory ligands of the TNF-receptor superfamily, including LIGHT, TNFα, and TNF-like ligand 1A (TL1A), leading to activation, differentiation, and proliferation of AIR-Treg cells. In a graft-versus-host disease model, Treg cells expressing lymphotoxin ß receptor-AIR, which can be activated by the ligand LIGHT, protect significantly better than control Treg cells. Expression and signaling of the corresponding human AIR in human Treg cells prove that this concept can be translated. Engineering Treg cells that target inflammatory ligands leading to TCR signaling and activation might be used as a Treg cell-based therapy approach for a broad range of inflammation-driven diseases.

Postmitotic differentiation of human monocytes requires cohesin-structured chromatin.

Cohesin is a major structural component of mammalian genomes and is required to maintain loop structures. While acute depletion in short-term culture models suggests a limited importance of cohesin for steady-state transcriptional circuits, long-term studies are hampered by essential functions of cohesin during replication. Here, we study genome architecture in a postmitotic differentiation setting, the differentiation of human blood monocytes (MO). We profile and compare epigenetic, transcriptome and 3D conformation landscapes during MO differentiation (either into dendritic cells or macrophages) across the genome and detect numerous architectural changes, ranging from higher level compartments down to chromatin loops. Changes in loop structures correlate with cohesin-binding, as well as epigenetic and transcriptional changes during differentiation. Functional studies show that the siRNA-mediated depletion of cohesin (and to a lesser extent also CTCF) markedly disturbs loop structures and dysregulates genes and enhancers that are primarily regulated during normal MO differentiation. In addition, gene activation programs in cohesin-depleted MO-derived macrophages are disturbed. Our findings implicate an essential function of cohesin in controlling long-term, differentiation- and activation-associated gene expression programs.

Salt-inducible kinase 3 protects tumor cells from cytotoxic T-cell attack by promoting TNF-induced NF-κB activation.

BACKGROUND: Cancer immunotherapeutic strategies showed unprecedented results in the clinic. However, many patients do not respond to immuno-oncological treatments due to the occurrence of a plethora of immunological obstacles, including tumor intrinsic mechanisms of resistance to cytotoxic T-cell (TC) attack. Thus, a deeper understanding of these mechanisms is needed to develop successful immunotherapies. METHODS: To identify novel genes that protect tumor cells from effective TC-mediated cytotoxicity, we performed a genetic screening in pancreatic cancer cells challenged with tumor-infiltrating lymphocytes and antigen-specific TCs. RESULTS: The screening revealed 108 potential genes that protected tumor cells from TC attack. Among them, salt-inducible kinase 3 (SIK3) was one of the strongest hits identified in the screening. Both genetic and pharmacological inhibitions of SIK3 in tumor cells dramatically increased TC-mediated cytotoxicity in several in vitro coculture models, using different sources of tumor and TCs. Consistently, adoptive TC transfer of TILs led to tumor growth inhibition of SIK3-depleted cancer cells in vivo. Mechanistic analysis revealed that SIK3 rendered tumor cells susceptible to tumor necrosis factor (TNF) secreted by tumor-activated TCs. SIK3 promoted nuclear factor kappa B (NF-κB) nuclear translocation and inhibited caspase-8 and caspase-9 after TNF stimulation. Chromatin accessibility and transcriptome analyses showed that SIK3 knockdown profoundly impaired the expression of prosurvival genes under the TNF-NF-κB axis. TNF stimulation led to SIK3-dependent phosphorylation of the NF-κB upstream regulators inhibitory-κB kinase and NF-kappa-B inhibitor alpha on the one side, and to inhibition of histone deacetylase 4 on the other side, thus sustaining NF-κB activation and nuclear stabilization. A SIK3-dependent gene signature of TNF-mediated NF-κB activation was found in a majority of pancreatic cancers where it correlated with increased cytotoxic TC activity and poor prognosis. CONCLUSION: Our data reveal an abundant molecular mechanism that protects tumor cells from cytotoxic TC attack and demonstrate that pharmacological inhibition of this pathway is feasible.

Physiological levels of 25-hydroxyvitamin D3 induce a suppressive CD4+ T cell phenotype not reflected in the epigenetic landscape.

1,25-dihydroxyvitamin D3 (1,25(OH)2 D3 ), the active metabolite of vitamin D3 has a strong impact on the differentiation and function of immune cells. Here we analysed the influence of its precursor 25-hydroxyvitamin D3 (25(OH)D3 ) on the differentiation of human CD4+ T cells applying physiological concentrations in vitro. Our data show that 25(OH)D3 is converted to its active form 1,25(OH)2 D3 by T cells, which in turn supports FOXP3, CD25 and CTLA-4 expression and inhibits IFN-γ production. These changes were not reflected in the demethylation of the respective promoters. Furthermore, we investigated the impact of vitamin D3 metabolites under induced Treg (iTreg) polarization conditions using TGF-ß. Surprisingly, no additive effect but a decreased percentage of FOXP3 expressing cells was observed. However, the combination of 25(OH)D3 or 1,25(OH)2 D3 together with TGF-ß further upregulated CD25 and CTLA-4 and significantly increased soluble CTLA-4 and IL-10 secretion whereas IFN-γ expression of iTreg was decreased. Our data suggest that physiological levels of 25(OH)D3 act as potent modulator of human CD4+ T cells and autocrine or paracrine production of 1,25(OH)2 D3 by T cells might be crucial for the local regulation of an adaptive immune response. However, since no epigenetic changes are detected by 25(OH)D3 a rather transient phenotype is induced.

Aberrant DNA methylation patterns in microsatellite stable human colorectal cancers define a new marker panel for the CpG island methylator phenotype.

A distinct group of colorectal carcinomas (CRCs) referred to as the "CpG island methylator phenotype" (CIMP) shows an extremely high incidence of de novo DNA methylation and may share common pathological, clinical or molecular features. However, there is limited consensus about which CpG islands (CGIs) define a CIMP, particularly in microsatellite stable (MSS) carcinomas. To study this phenotype in a systematic manner, we analyzed genome-wide CGI DNA methylation profiles of 19 MSS CRC using methyl-CpG immunoprecipitation (MCIp) and hybridization on 244K CGI oligonucleotide microarrays, determined KRAS and BRAF mutation status and compared disease-related DNA methylation changes to chromosomal instability as detected by microarray-based comparative genomic hybridization. Results were validated using mass spectrometry analysis of bisulfite-converted DNA at a subset of 76 individual CGIs in 120 CRC and 43 matched normal tissue samples. Both genome-wide profiling and CpG methylation fine mapping segregated a group of CRC showing pronounced and frequent de novo DNA methylation of a distinct group of CGIs that only partially overlapped with previously established classifiers. The CIMP group defined in our study revealed significant association with colon localization, either KRAS or BRAF mutation, and mostly minor chromosomal losses but no association with known histopathological features. Our data provide a basis for defining novel marker panels that may enable a more reliable classification of CIMP in all CRCs, independently of the MS status.

Metabolic imbalance of T cells in COVID-19 is hallmarked by basigin and mitigated by dexamethasone.

Metabolic pathways regulate immune responses and disrupted metabolism leads to immune dysfunction and disease. Coronavirus disease 2019 (COVID-19) is driven by imbalanced immune responses, yet the role of immunometabolism in COVID-19 pathogenesis remains unclear. By investigating 87 patients with confirmed SARS-CoV-2 infection, 6 critically ill non-COVID-19 patients, and 47 uninfected controls, we found an immunometabolic dysregulation in patients with progressed COVID-19. Specifically, T cells, monocytes, and granulocytes exhibited increased mitochondrial mass, yet only T cells accumulated intracellular reactive oxygen species (ROS), were metabolically quiescent, and showed a disrupted mitochondrial architecture. During recovery, T cell ROS decreased to match the uninfected controls. Transcriptionally, T cells from severe/critical COVID-19 patients showed an induction of ROS-responsive genes as well as genes related to mitochondrial function and the basigin network. Basigin (CD147) ligands cyclophilin A and the SARS-CoV-2 spike protein triggered ROS production in T cells in vitro. In line with this, only PCR-positive patients showed increased ROS levels. Dexamethasone treatment resulted in a downregulation of ROS in vitro and T cells from dexamethasone-treated patients exhibited low ROS and basigin levels. This was reflected by changes in the transcriptional landscape. Our findings provide evidence of an immunometabolic dysregulation in COVID-19 that can be mitigated by dexamethasone treatment.

Methyl donor micronutrients, CD40-ligand methylation and disease activity in systemic lupus erythematosus: A cross-sectional association study.

OBJECTIVE: Hypomethylation of CD40-ligand (CD40L) in T-cells is associated with increased disease activity in systemic lupus erythematosus (SLE). We therefore investigated possible associations of dietary methyl donors and products with CD40L methylation status in SLE. METHODS: Food frequency questionnaires were employed to calculate methyl donor micronutrients in 61 female SLE patients (age 45.7 ± 12.0 years, disease duration 16.2 ± 8.4 years) and compared to methylation levels of previously identified key DNA methylation sites (CpG17 and CpG22) within CD40L promotor of T-cells using quantitative DNA methylation analysis on the EpiTYPER mass spectrometry platform. Disease activity was assessed by SLE Disease Activity Index (SLEDAI). Linear regression modelling was used. P values were adjusted according to Benjamini & Hochberg. RESULTS: Amongst the micronutrients assessed (g per day), methionine and cysteine were associated with methylation of CpG17 (ß = 5.0 (95%CI: 0.6-9.4), p = 0.04; and ß = 2.4 (0.6-4.1), p = 0.02, respectively). Methionine, choline, and cysteine were additionally associated with the mean methylation of the entire CD40L (ß = 9.5 (1.0-18.0), p = 0.04; ß = 1.6 (0.4-3.0), p = 0.04; and ß = 4.3 (0.9-7.7), p = 0.02, respectively). Associations of the SLEDAI with hypomethylation were confirmed for CpG17 (ß=-32.6 (-60.6 to -4.6), p = 0.04) and CpG22 (ß=-38.3 (-61.2 to -15.4), p = 0.004), but not the mean methylation of CD40L. Dietary products with the highest impact on methylation included meat, ice cream, white bread, and cooked potatoes. CONCLUSIONS: Dietary methyl donors may influence DNA methylation levels and thereby disease activity in SLE.

Allele-specific expression of GATA2 due to epigenetic dysregulation in CEBPA double-mutant AML.

Transcriptional deregulation is a central event in the development of acute myeloid leukemia (AML). To identify potential disturbances in gene regulation, we conducted an unbiased screen of allele-specific expression (ASE) in 209 AML cases. The gene encoding GATA binding protein 2 (GATA2) displayed ASE more often than any other myeloid- or cancer-related gene. GATA2 ASE was strongly associated with CEBPA double mutations (DMs), with 95% of cases presenting GATA2 ASE. In CEBPA DM AML with GATA2 mutations, the mutated allele was preferentially expressed. We found that GATA2 ASE was a somatic event lost in complete remission, supporting the notion that it plays a role in CEBPA DM AML. Acquisition of GATA2 ASE involved silencing of 1 allele via promoter methylation and concurrent overactivation of the other allele, thereby preserving expression levels. Notably, promoter methylation was also lost in remission along with GATA2 ASE. In summary, we propose that GATA2 ASE is acquired by epigenetic mechanisms and is a prerequisite for the development of AML with CEBPA DMs. This finding constitutes a novel example of an epigenetic hit cooperating with a genetic hit in the pathogenesis of AML.

The epigenetic pioneer EGR2 initiates DNA demethylation in differentiating monocytes at both stable and transient binding sites.

The differentiation of human blood monocytes (MO), the post-mitotic precursors of macrophages (MAC) and dendritic cells (moDC), is accompanied by the active turnover of DNA methylation, but the extent, consequences and mechanisms of DNA methylation changes remain unclear. Here, we profile and compare epigenetic landscapes during IL-4/GM-CSF-driven MO differentiation across the genome and detect several thousand regions that are actively demethylated during culture, both with or without accompanying changes in chromatin accessibility or transcription factor (TF) binding. We further identify TF that are globally associated with DNA demethylation processes. While interferon regulatory factor 4 (IRF4) is found to control hallmark dendritic cell functions with less impact on DNA methylation, early growth response 2 (EGR2) proves essential for MO differentiation as well as DNA methylation turnover at its binding sites. We also show that ERG2 interacts with the 5mC hydroxylase TET2, and its consensus binding sequences show a characteristic DNA methylation footprint at demethylated sites with or without detectable protein binding. Our findings reveal an essential role for EGR2 as epigenetic pioneer in human MO and suggest that active DNA demethylation can be initiated by the TET2-recruiting TF both at stable and transient binding sites.

Single-cell chromatin accessibility landscape identifies tissue repair program in human regulatory T cells.

Murine regulatory T (Treg) cells in tissues promote tissue homeostasis and regeneration. We sought to identify features that characterize human Treg cells with these functions in healthy tissues. Single-cell chromatin accessibility profiles of murine and human tissue Treg cells defined a conserved, microbiota-independent tissue-repair Treg signature with a prevailing footprint of the transcription factor BATF. This signature, combined with gene expression profiling and TCR fate mapping, identified a population of tissue-like Treg cells in human peripheral blood that expressed BATF, chemokine receptor CCR8 and HLA-DR. Human BATF+CCR8+ Treg cells from normal skin and adipose tissue shared features with nonlymphoid T follicular helper-like (Tfh-like) cells, and induction of a Tfh-like differentiation program in naive human Treg cells partially recapitulated tissue Treg regenerative characteristics, including wound healing potential. Human BATF+CCR8+ Treg cells from healthy tissue share features with tumor-resident Treg cells, highlighting the importance of understanding the context-specific functions of these cells.

The IKZF1-IRF4/IRF5 Axis Controls Polarization of Myeloma-Associated Macrophages.

The bone marrow niche has a pivotal role in progression, survival, and drug resistance of multiple myeloma cells. Therefore, it is important to develop means for targeting the multiple myeloma bone marrow microenvironment. Myeloma-associated macrophages (MAM) in the bone marrow niche are M2 like. They provide nurturing signals to multiple myeloma cells and promote immune escape. Reprogramming M2-like macrophages toward a tumoricidal M1 phenotype represents an intriguing therapeutic strategy. This is especially interesting in view of the successful use of mAbs against multiple myeloma cells, as these therapies hold the potential to trigger macrophage-mediated phagocytosis and cytotoxicity. In this study, we observed that MAMs derived from patients treated with the immunomodulatory drug (IMiD) lenalidomide skewed phenotypically and functionally toward an M1 phenotype. Lenalidomide is known to exert its beneficial effects by modulating the CRBN-CRL4 E3 ligase to ubiquitinate and degrade the transcription factor IKAROS family zinc finger 1 (IKZF1). In M2-like MAMs, we observed enhanced IKZF1 levels that vanished through treatment with lenalidomide, yielding MAMs with a bioenergetic profile, T-cell stimulatory properties, and loss of tumor-promoting capabilities that resemble M1 cells. We also provide evidence that IMiDs interfere epigenetically, via degradation of IKZF1, with IFN regulatory factors 4 and 5, which in turn alters the balance of M1/M2 polarization. We validated our observations in vivo using the CrbnI391V mouse model that recapitulates the IMiD-triggered IKZF1 degradation. These data show a role for IKZF1 in macrophage polarization and can provide explanations for the clinical benefits observed when combining IMiDs with therapeutic antibodies.See related Spotlight on p. 254.

Associations of site-specific CD4+-T-cell hypomethylation within CD40-ligand promotor and enhancer regions with disease activity of women with systemic lupus erythematosus.

OBJECTIVE: To comprehensively assess associations of site-specific CD4+-T-cell hypomethylation of the CD40-Ligand gene (CD40L) with disease activity of women with systemic lupus erythematosus (SLE). METHODS: CpG-sites within the DNA of the promotor and two enhancer regions (n = 22) of CD40L were identified and numbered consecutively. The rate of methylated DNA in isolated CD4+-T-cells of women with SLE were quantified for each methylation site by MALDI-TOF. Disease activity was assessed by SLE Disease Activity Index (SLEDAI). Associations of site-specific methylation rates with the SLEDAI scores were assessed by linear regression modelling. P values were adjusted according to Bonferroni-Holm as indicated. RESULTS: 60 female SLE patients participated in the study (age 45.7 ± 11.1 years, disease duration 17.0 ± 8.3 years). Significant associations to the SLEDAI were noted for CpG22 hypomethylation of the promotor (ß = -40.1, p = 0.017, adjusted p = 0.027), trends were noted for CpG17 hypomethylation of the promotor (ß = -30.5, p = 0.032, adjusted p = 0.6), and for CpG11 hypermethylation of the second enhancer (ß = 15.0, p = 0.046, adjusted p = 0.8). CONCLUSION: Site-specific hypomethylation of the CD40L promotor in CD4+-T-cells show associations with disease activity in female SLE patients.

Author Correction: Mechanisms governing the pioneering and redistribution capabilities of the non-classical pioneer PU.1.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.