Remco G. A. Erkens

Objective: Macrophage activation syndrome (MAS) is a rare, life‐threatening hyperinflammatory condition that complicates rheumatologic diseases, most commonly Still disease. Despite increasing awareness and emerging recommendations, MAS treatment remains heterogeneous worldwide. The METAPHOR project aimed to describe real‐life therapeutic strategies for MAS, exploring differences across underlying diseases, geographic regions, physician subspecialties, and drug availability. Methods: An international web‐based survey was developed by a multidisciplinary expert panel together with a patient representative following a systematic literature review. The survey was anonymously completed through Paediatric Rheumatology European Society/Paediatric Rheumatology International Trial Organisation and the Histiocyte Society networks (October to December 2023). Treatment strategies for MAS across different rheumatologic contexts were explored. Responses were analyzed descriptively and stratified by geography, subspecialty, clinical experience, and underlying disease. Results: A total of 287 physicians from 64 countries completed the survey, the majority being pediatric rheumatologists with more than five years of experience. Glucocorticoids, particularly intravenous methylprednisolone, were consistently identified as first‐line therapy across all MAS subtypes (for 44% of clinicians as monotherapy). Anakinra, ciclosporin, and intravenous Ig were the most frequently combined agents. However, their use varied markedly by underlying disease and geographic region, with anakinra preferentially used in Still disease–associated MAS. Striking global disparities in drug availability, particularly for anakinra and emapalumab, significantly influenced treatment choices and were associated with increased reliance on steroid monotherapy. Conclusion: MAS management remains highly heterogeneous worldwide. Although glucocorticoids remain foundational, reducing steroid reliance through equitable access to immunomodulatory therapies and advancing mechanism‐based strategies represent critical priorities in optimizing MAS care. image

Objective: Still disease (SD) is an autoinflammatory syndrome characterized by innate immune dysregulation. Although complement can drive inflammation, its involvement in SD remains to be defined. Thus, we aimed to assess complement activation in SD. Methods: Complement was assessed using transcriptomic, proteomic, and in vitro approaches. RNA sequencing of monocytes was performed in healthy donors (n = 15), those with nonsystemic juvenile idiopathic arthritis (JIA; n = 8), patients with SD at onset (n = 19) and remission (n = 18), and those with macrophage activation syndrome (n = 2). Whole‐blood NanoString analysis of complement and interferon (IFN)‐related gene expression was conducted in patients with SD (active n = 41, inactive n = 33) and JIA (n > 600). Complement products and inflammatory mediators were measured by Luminex and enzyme‐linked immunosorbent assay. Functional complement activity was evaluated in SD (active n = 30, inactive n = 67) and JIA sera (n = 12). In vitro assays examined monocytic C1q induction and complement‐mediated CD8 ⁺ T cell activation. Results: Transcriptomic analysis of monocytes from patients with SD at onset revealed enrichment of the complement cascade compared with patients in remission (adjusted P = 3.7 × 10 ⁻³⁶ ), ranking among the top 10 up‐regulated pathways. Classical complement genes ( C1QB / C1QC ) were markedly up‐regulated in onset SD compared with patients with remission SD and JIA. Patients with active SD showed increased C1q, C3a, C5a, and terminal complement complex protein levels, with enhanced functional classical complement activity. Whole‐blood C1QB / C1QC expression correlated with IFN‐related markers, including interleukin‐18, CXCL9, and CXCL10. Recombinant IFN‐γ induced monocytic C1q, whereas C1q enhanced IFN‐γ production by CD8 ⁺ T cells, supporting a feed‐forward loop. Conclusion: SD is characterized by complement activation with marked up‐regulation of C1q, which is closely linked to IFN‐γ/type II signaling. image

Objective: Still disease (SD) is an autoinflammatory disorder characterized by remarkably high interleukin‐18 (IL‐18) levels. Increasing evidence suggests that adaptive immunity also contributes to its pathogenesis, particularly in refractory courses. Macrophage activation syndrome (MAS), one of SD's most severe complications, is associated with further IL‐18 elevation and expansion of CD38 ⁺ HLA‐DR ⁺ CD8 ⁺ T cells. However, whether and when this population emerges during SD remains unknown. We therefore examined CD38 ⁺ HLA‐DR ⁺ CD8 ⁺ T cells across disease stages. Methods: CD38 ⁺ HLA‐DR ⁺ CD8 ⁺ T cells were analyzed longitudinally: at SD onset (therapy‐naive; n = 19), during clinically inactive disease (n = 15), and before or at MAS occurrence (n = 12). Spectral flow cytometry and bulk RNA sequencing were performed. Correlations with clinical and laboratory parameters were assessed. In vitro cytokine stimulations of healthy donor (HD)–derived peripheral blood mononuclear cells and CD8 ⁺ lymphocytes were performed to explore mechanisms driving differentiation of these cells. Results: CD38 ⁺ HLA‐DR ⁺ CD8 ⁺ T cells were expanded from SD onset and peaked in MAS (median: 4.61% and 31.7%, respectively, versus 0.87% in HDs). Transcriptomic profiling of these cells revealed enrichment of activation, cytotoxicity, and proliferation programs, which were confirmed at the protein level. Frequencies correlated with markers of systemic inflammation, T cell activation, and serum IL‐18 levels. Sustained IL‐18 stimulation in vitro induced robust and persistent expansion of CD38 ⁺ HLA‐DR ⁺ CD8 ⁺ cells, recapitulating key ex vivo phenotypic features. Conclusion: CD38 ⁺ HLA‐DR ⁺ CD8 ⁺ cells, previously primarily associated with MAS, are detectable early in SD. Persistent IL‐18 exposure promotes their expansion. These findings identify CD38 ⁺ HLA‐DR ⁺ CD8 ⁺ T cells as a potential mechanistic link between SD and MAS and highlight this subset as a candidate therapeutic target. image

Alternative splicing is a key mechanism for expanding transcriptomic and proteomic complexity, yet its role in innate immune activation remains incompletely understood. Here, we applied Oxford Nanopore native RNA-sequencing to generate an isoform-level transcriptome of primary human monocytes before and after activation with lipopolysaccharide. We identify over 24,000 expressed isoforms, including thousands of previously unannotated variants. Activation induced widespread isoform-specific expression changes, leading to extensive isoform switching events, which were validated using matched short-read RNA-Seq. These activation-induced shifts enhanced transcript immune-regulatory functions: activated monocytes preferentially express longer, coding-competent isoforms with complete open reading frames, fewer retained introns, and increased domain complexity. By integrating matched Ribo-seq and proteomic data, we demonstrate that these isoform modulations are associated with enhanced translation of immune effector proteins. Together, our findings position alternative splicing as a dynamic and functional regulator of monocyte activation, emphasizing the need for isoform-level resolution to fully understand immune cell function.

Objective: To evaluate the applicability of the 2016 European Alliance of Associations for Rheumatology (EULAR)/American College of Rheumatology (ACR)/Paediatric Rheumatology International Trials Organisation (PRINTO) macrophage activation syndrome (MAS) classification criteria in patients with Still disease and systemic juvenile idiopathic arthritis (sJIA‐SD) treated with interleukin‐1 (IL‐1)–targeted therapy and to assess the incidence of MAS in this context. Methods: We analyzed retrospective and prospective data from Dutch patients with sJIA‐SD (diagnosis 2008–2017, n = 54) and data from a nationwide prospective Dutch cohort and intervention study (diagnosis 2017–2022, n = 66). From these cohorts, MAS episodes developing in patients with sJIA‐SD treated with IL‐1–targeted therapy (anakinra or canakinumab) with at least two years of follow‐up were selected. Clinical and laboratory data were extracted from the electronic patient files. Results: A total of 22 patients experienced 29 MAS episodes while receiving IL‐1–targeted treatment. Seven patients had recurrent MAS episodes (not all while receiving IL‐1 blockade). The 2016 criteria for MAS in sJIA‐SD were met for 28 of 29 MAS episodes (97%). Within the prospective nationwide cohort starting anakinra as first‐line monotherapy, the incidence rate of MAS in the first two years of disease was 18% (12 of 66 patients, with 11 of 12 while receiving IL‐1 inhibition). This incidence is comparable to that observed in historical glucocorticoid‐treated patients. Half of MAS episodes occurred within three months after diagnosis, and Epstein‐Barr virus was the most common identifiable trigger. Conclusion: Although first‐line anakinra in new‐onset sJIA‐SD has demonstrated high response rates, our data suggest the incidence of MAS in the first two years of disease is not reduced. Patients appear to be particularly at risk early in disease. Importantly, our data show that the EULAR/ACR/PRINTO 2016 MAS classification criteria remain applicable to patients receiving IL‐1–targeted therapy.

Objective: Human leukocyte antigen (HLA)‐DRB1*15:01 has been recently associated with interstitial lung disease (LD), eosinophilia, and drug reactions in systemic juvenile idiopathic arthritis (sJIA). Additionally, genetic variants in IL1RN have been linked to poor response to anakinra. We sought to reproduce these findings in a prospective cohort study of patients with new‐onset sJIA treated with anakinra as first‐line therapy.Methods: HLA and IL1RN risk alleles were identified via whole‐genome sequencing. Treatment responses and complications were compared between carriers versus noncarriers.Results: Seventeen of 65 patients (26%) carried HLA‐DRB1*15:01, comparable with the general population, and there was enrichment for HLA‐DRB1*11:01, a known risk locus for sJIA. The rates of clinical inactive disease (CID) at 6 months, 1 year, and 2 years were generally high, irrespective of HLA‐DRB1 or IL1RN variants, but significantly lower in carriers of an HLA‐DRB1*11:01 allele. One patient, an HLA‐DRB1*15:01 carrier, developed sJIA‐LD. Of the three patients with severe drug reactions to biologics, one carried HLA‐DRB1*15:01. The prevalence of eosinophilia did not significantly differ between HLA‐DRB1*15:01 carriers and noncarriers at disease onset (6.2% vs 14.9%, P = 0.67) nor after the start of anakinra (35.3% vs 37.5% in the first 2 years of disease).Conclusion: We observed high rates of CID using anakinra as first‐line treatment irrespective of HLA‐DRB1 or IL1RN variants. Only one of the 17 HLA‐DRB1*15:01 carriers developed sJIA‐LD, and of the three patients with drug reactions to biologics, only one carried HLA‐DRB1*15:01. Although thorough monitoring for the development of drug hypersensitivity and refractory disease courses in sJIA, including sJIA‐LD, remains important, our data support the early start of biologic therapy in patients with new‐onset sJIA irrespective of HLA‐DRB1 background or IL1RN variants. image

Purpose: Liver metastasis develops in >50% of patients with colorectal cancer (CRC), and is a leading cause of CRC-related mortality. We aimed to identify which inhibitory immune checkpoint pathways can be targeted to enhance functionality of intra-tumoral T-cells in mismatch repair-proficient liver metastases of colorectal cancer (LM-CRC). Methodology: Intra-tumoral expression of multiple inhibitory molecules was compared among mismatch repair-proficient LM-CRC, peritoneal metastases of colorectal cancer (PM-CRC) and primary CRC. Expression of inhibitory molecules was also analyzed on leukocytes isolated from paired resected metastatic liver tumors, tumor-free liver tissues, and blood of patients with mismatch repair-proficient LM-CRC. The effects of blocking inhibitory pathways on tumor-infiltrating T-cell responses were studied in ex vivo functional assays. Results: Mismatch repair-proficient LM-CRC showed higher expression of inhibitory receptors on intra-tumoral T-cells and contained higher proportions of CD8⁺ T-cells, dendritic cells and monocytes than mismatch repair-proficient primary CRC and/or PM-CRC. Inhibitory receptors LAG3, PD-1, TIM3 and CTLA4 were higher expressed on CD8⁺ T-cells, CD4⁺ T-helper and/or regulatory T-cells in LM-CRC tumors compared with tumor-free liver and blood. Antibody blockade of LAG3 or PD-L1 increased proliferation and effector cytokine production of intra-tumoral T-cells isolated from LM-CRC in response to both polyclonal and autologous tumor-specific stimulations. Higher LAG3 expression on intra-tumoral CD8⁺ T-cells associated with longer progression-free survival of LM-CRC patients. Conclusion: Mismatch repair-proficient LM-CRC may be more sensitive to immune checkpoint inhibitors than mismatch repair-proficient primary CRC. Blocking LAG3 enhances tumor-infiltrating T-cell responses of mismatch repair-proficient LM-CRC, and therefore may be a new promising immunotherapeutic target for LM-CRC.