Group 3 innate lymphoid cells (ILC3s) exhibit dynamic plasticity, with their differentiation and function orchestrated by epigenetic mechanisms, including histone modifications and DNA methylation. We identify the histone demethylases UTX and JMJD3 as pivotal regulators of ILC3 specialization. Their deficiency disrupts subset balance: NKp46+ ILC3s are depleted with impaired IL-22 production, whereas CCR6+ ILC3s expand and exhibit enhanced IL-17A-mediated antifungal immunity. Single-cell profiling reveals that UTX/JMJD3 ablation epigenetically restricts double-negative (DN)-to-NKp46+ differentiation while potentiating CCR6+ polarization, rewiring ILC3 lineage trajectories through chromatin remodeling. Cleavage under targets and tagmentation (CUT&Tag) analysis demonstrates that UTX directly occupies enhancer regions upstream of Tcf7, where it catalyzes H3K27me3 demethylation to maintain an open chromatin state. Retroviral Tcf7 reconstitution rescues the NKp46+ ILC3 deficit and normalizes cytokine production, positioning TCF7 as the key effector downstream of UTX. These findings establish UTX/JMJD3 as central epigenetic gatekeepers of mucosal immunity, offering therapeutic avenues for inflammatory disorders driven by ILC3 dysregulation.

Abstract Immune checkpoint blockade therapy has shown limited efficacy in gastric cancer, with most patients developing resistance through mechanisms that remain incompletely defined. Here, we generate a comprehensive single-cell RNA sequencing atlas of 526,583 cells from gastric cancer patients treated with anti-PD-1 plus chemotherapy, analyzing paired pre- and post-treatment samples to capture dynamic resistance mechanisms. We identify two distinct resistance pathways that emerge during treatment. First, CEACAM5/6+ cancer cells are markedly enriched in pre-treatment non-responders and predict treatment failure. These CEACAM5/6+ epithelial cells show the highest tumor scores and correlate with increased regulatory T cell infiltration expressing CEACAM1, establishing an alternative checkpoint axis that bypasses PD-1/PD-L1 blockade. External validation in independent cohorts confirms CEACAM5/6 expression as a robust predictor of anti-PD-1 resistance. Second, we uncover a macrophage-driven inflammatory cascade central to treatment resistance. IL-1β+ macrophages serve as the primary source of NF-κB pathway activation across the tumor microenvironment, triggering downstream IL-6 production, Th17 cell differentiation, chronic inflammation and epithelial-mesenchymal transition. This macrophage module is significantly enriched in post-treatment non-responders, with TNF-high expressing monocyte-macrophages absent in responders but prevalent in resistant tumors. The resulting inflammatory milieu drives PD-L1 upregulation across multiple cell types, creating a self-reinforcing immunosuppressive niche. Collectively, these findings nominate CEACAM5/6+ epithelial cells and IL-1β+ inflammatory macrophages as actionable therapeutic targets for overcoming anti-PD-1 resistance in gastric cancer, providing a mechanistic framework and rational blueprint for next-generation combination immunotherapy strategies. Citation Format: Liudeng Zhang, Jian Chen, Yikai Luo, Lie Wang, Han Liang. CEACAM5/6+ cancer cell and IL1B+ macrophage-mediated resistance in anti-PD-1 treated gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 7746.

The histone H3K4me3 broad domain (BD) is a unique epigenetic feature marking cell identity-associated genes, but its physiological role in lineage differentiation remains unclear. Here we show that CFP1, an integral component of the Setd1A/B methyltransferase complex, is critically required for functional H3K4me3-BD installation and B cell fate determination. Cfp1 deletion impairs H3K4me3-BDs on a subset of B lineage genes and redistributes H3K4me3 from active genes to bivalent promoters. Transcription of subsets of the H3K4me3-BD-bound genes is diminished in part due to reduced RNA polymerase II at the promoter and gene body. Specifically, CFP1 ablation abolishes H3K4me3-BD at the recombination center and distal PAIR elements in the immunoglobulin heavy chain (IgH) gene, severely compromises its locus contraction for efficient V-to-DJ recombination, and consequently arrests cells at the pro-B stage. Moreover, Cfp1-deficient pro-B cells up-regulate a panel of progenitor and myeloid-specific genes with elevated H3K4me3 marks and can trans-differentiate into various myeloid cell types. Therefore, our study reveals pivotal roles for CFP1-mediated H3K4me3-BDs in the transcriptional regulation of cell lineage fate specification and commitment.

Figure S1 shows model performance and clinical stratification metrics

Figure S4 shows interaction between CD8+ TRM and activated B cells in m-TLSs

OBJECTIVE:Autoimmune diseases, such as systemic lupus erythematosus (SLE), are associated with pulmonary arterial hypertension (PAH), a condition that can lead to heart failure. However, whether T cells also contribute to the occurrence of PAH in SLE has not been clarified. The objective of this study was to elucidate the role of Activin A and activated receptor signaling in SLE-PAH. METHODS:Mass Cytometry (CyTOF) analysis was performed to identify the major affected immune cell population in patients with SLE-PAH. Serum Activin A and interleukin-17 (IL-17) levels in patients with SLE-PAH, patients with SLE, and healthy donors were determined by enzyme-linked immunosorbent assay. Cocultures of Th17 cells with pulmonary microvascular endothelial cells (PMECs) and relevant rodent models were used to identify the converged target. RESULTS:The reduced CD4+ T cell number was detected in patients with SLE-PAH after treatment with immunosuppressant and vasodilator. Increased Th17 cells population and higher serum Activin A and IL-17 levels were found in patients with SLE-PAH compared to patients with SLE only or donors. Activin A signals via activin receptor-like kinase 4 (ALK4) in both Th17 cells and PMECs. Overexpressing ALK4 in Th17 cells increased IL-6 and endothelial-mesenchymal transition (EndoMT) marker levels in cocultured PMECs. We found severe SLE-pulmonary hypertension (PH) in mice by overexpressing ALK4, and alleviated hemodynamic changes in CD4+ T cells depletion mice. ALK4 inhibitor vactosertib (TEW-7197) effectively treated SLE-PH mice by repressing connective tissue growth factor (CTGF) transcription, which was induced by ALK4 activated pSmad2 and pSTAT3. CONCLUSION:Our findings suggest that Activin A activates ALK4 in Th17 cells, thereby inducing IL-17 secretion. Concurrently, activated ALK4 induces EndoMT in human PMECs (hPMECs) via CTGF up-regulation. It suggests that ALK4 is a promising therapeutic target for SLE-PAH.

Table S2 shows characteristics summary of patients in the GC external validation cohort

Intestinal immunosenescence, a hallmark of organismal aging, has emerged as a critical biological process impacting the health of elderly individuals. This review systematically examines the core mechanisms underlying intestinal immunosenescence, including immune cell dysfunction, imbalances in immune-microbiota interactions, and impaired barrier function. We analyze its associations with infectious diseases, chronic inflammation, and neurodegenerative disorders, summarizing recent advances in dietary interventions, microecological therapy, and other emerging strategies. By integrating cutting-edge technologies, we prospect the development of precision interventions aimed at delaying intestinal immunosenescence, thereby providing a theoretical basis for improving the healthspan of the aging population.

Figure S3 shows biological patterns of TRM in TLSs in gastric cancer

FOXP3-expressing regulatory T (Treg) cells play a pivotal role in maintaining immune homeostasis and tolerance, with their activation being crucial for preventing various inflammatory responses. However, the mechanisms governing the epigenetic program in Treg cells during their dynamic activation remain unclear. In this study, we demonstrate that CXXC-finger protein 1 (CXXC1) interacts with the transcription factor FOXP3 and facilitates the regulation of target genes by modulating H3K4me3 deposition. Cxxc1 deletion in Treg cells leads to severe inflammatory disease and spontaneous T cell activation, with impaired immunosuppressive function. As a transcriptional regulator, CXXC1 promotes the expression of key Treg functional markers under steady-state conditions, which are essential for the maintenance of Treg cell homeostasis and their suppressive functions. Epigenetically, CXXC1 binds to the genomic regulatory regions of Treg program genes in mouse Treg cells, overlapping with FOXP3-binding sites. Given its critical role in Treg cell homeostasis, CXXC1 presents itself as a promising therapeutic target for autoimmune diseases.

Table S7 shows characteristics of NSCLC patients in the flow cytometry cohort

Table S5 shows characteristics of colorectal cancer patients in the flow cytometry cohort

Table S1 shows clinicopathological correlations of m-TLSs in the GC H&E cohort

Figure S5 shows representative gating strategy for co-culture experiments

Recent advances in single-cell technology enable the simultaneous capture of T cell receptor (TCR) sequences and gene expression (GEX), providing an integrated view of T cell function. However, linking TCRαβ information and T cell phenotypes at the population level to elucidate their disease association remains an unaddressed gap. Here, by constructing a large-scale reference of paired single-cell RNA/TCR sequencing (scRNA/TCR-seq) comprising more than 2 million T cells from 70 studies, 1017 biological samples, 583 individuals, and 46 disease conditions, along with their single-cell transcriptome, full-length paired TCR, and human leukocyte antigen (HLA) genotypes, we revealed the intrinsic features of germline-encoded TCR-major histocompatibility complex (MHC) restriction in CD4+/CD8+ lineages. We also observed widely existing public TCRαβs across the population, associated with higher clonal expansion levels and shared HLA alleles. The most publicly shared TCRs are likely to target epitopes from common viruses, such as Epstein-Barr virus (EBV), cytomegalovirus (CMV), and influenza A virus (IAV). Furthermore, we introduced TCR-DeepInsight, a computational framework to identify HLA-shared and disease-associated TCRαβ clusters that exhibit similar TCR sequence and GEX profiles, extensible for researchers to incorporate their data with our reference and characterize potentially functional TCRs. In summary, our work presents a panoramic scTCRαβ reference and computational methods for TCR study.

Table S6 shows characteristics of liver cancer patients in the flow cytometry cohort

Tertiary lymphoid structures (TLS) are critical components of the tumor microenvironment in gastric cancer, but clinical assessment of TLSs is challenging. The development of automated annotation tools for histopathologic slide analysis could facilitate the identification of TLSs and enhance our understanding of the mechanisms driving TLS maturation. In this study, we generated a transformer-based deep learning model that enables quantitative characterization of TLS maturity from whole-slide images. Application of the model to a large gastric cancer cohort (n = 253) showed that higher TLS maturity correlated with improved patient survival. Integration of single-cell RNA sequencing data from 17 patients with gastric cancer combined with multiplex IHC, flow cytometry, and functional coculture assays identified a key immune circuit in mature TLSs involving CD8+ tissue-resident memory T cells, which recruit activated B cells via the CXCL13-CXCR5 axis to enhance tissue-resident memory T-cell cytotoxicity through granzyme B upregulation. Overall, this study established a clinically applicable artificial intelligence tool and uncovered key immune interactions that regulate TLS maturation and antitumor immunity in gastric cancer. SIGNIFICANCE:A deep learning model demonstrates that higher tertiary lymphoid structure maturity predicts improved gastric cancer patient survival and identifies a key immune circuit, offering a clinically applicable tool that could guide treatment. This article is part of a special series: Driving Cancer Discoveries with Computational Research, Data Science, and Machine Learning/AI.

Group 3 innate lymphoid cells (ILC3s) serve as critical guardians of mucosal immunity. However, the transcriptional networks governing their function remain incompletely characterized. Here, we demonstrate that interferon regulatory factor 4 (IRF4) is essential for maintaining intestinal ILC3 homeostasis and function. IRF4-deficient mice exhibit reduced NKp46+ ILC3s, expanded precursor-like NKp46-CCR6- ILC3s, and impaired interleukin-22 (IL-22)/IL-17A production, increasing susceptibility to infections. Furthermore, IRF4 loss disrupted major histocompatibility complex (MHC)-class II-associated transcriptional signatures in ILC3s, particularly in CCR6+ ILC3s, accompanied by downregulation of MHC class II protein expression. This perturbation consequently diminished ILC-mediated apoptosis of effector CD4+ T cells. Sequencing and trajectory analysis link IRF4 to NKp46+ ILC3 maintenance and Tbx21 regulation. ATAC-seq/CUT&Tag reveal direct IRF4 binding to Batf, Tbx21, Il22, Il17a, and MHC II loci. Overexpression of T-bet partially rescued the differentiation defects in intestinal ILC3s, whereas Batf overexpression partially restored functional impairments and significantly enhanced MHC class II expression in ILC3s.

In this article for the Highlights of 2024 Series, we discuss research on Group 3 innate lymphoid cells (ILC3s), which revealed their complex roles in mucosal immunity and inflammation. ILC3s can migrate from the gut to the kidneys, contributing to renal fibrosis. Their functions are metabolically regulated, with proteins such as nucleophosmin 1 and Tox2 influencing oxidative phosphorylation and glycolysis, respectively. ILC3s also express immune checkpoint molecules (e.g. cytotoxic T-lymphocyte antigen 4 and programmed cell death1), which modulate inflammation. These findings highlight the tissue-specific roles of ILC3s and the need for targeted immunotherapies.