Immune Remodeling Research Articles

Transcriptomic features of Epstein-Barr virus-associated gastric cancer based on PD-L1 status.

489 Background: Epstein-Barr virus (EBV)-associated gastric cancer (EBVaGC) accounts for 5-10% of all gastric cancers and has a relatively favorable prognosis compared to other subtypes. While the genomic profile of EBVaGC has been previously reported to exhibit extensive CpG island methylation, elevated levels of PD-L1/2, and an absence of TP53 mutations, transcriptomic features remain less explored. The response to immunotherapy of EBVaGC has been various. This study comprehensively investigates the transcriptomic characteristics of EBVaGC in relation to PD-L1 status. Methods: RNA sequencing was conducted on formalin-fixed, paraffin-embedded samples from 27 EBV-positive and 12 EBV-negative patients with HER2-negative gastric cancer. Gene expression was quantified using the RSEM package. Predict-IO, a machine learning-based predictive algorithm developed at Auristone Pte Ltd based on initial work done at the Genome Institute of Singapore (GIS), assessed the likelihood of a patient responding to immunotherapy based on transcriptome expression data. PD-L1 immunohistochemistry was performed using the 22C3 pharmDx assay, and status was classified by CPS. Results: A total of 200 differentially expressed genes (DEGs; log2FC > 2, FDR < 0.05) were identified in EBV-positive versus negative samples, with 157 genes down-regulated and 43 genes up-regulated. Upregulated genes included immune response related genes such as DMBT1, IDO1, PLA2G2A, and IL13RA2, while downregulated genes were primarily associated with cell adhesion, including LAMA1 and CCN6. In the EBV positive group, immune response pathways were enriched; however, within the PD-L1-Low group (CPS < 1), activated signaling pathways such as MYC, E2F, and MTORC1 were observed. Conversely, the PD-L1-High group (CPS ≥ 5) exhibited upregulated snoRNAs. In the EBV negative group, the PD-L1-Low cohort displayed suppressed immune response and ECM remodeling factors, whereas the PD-L1-High cohort showed enrichment of ECM remodeling factors. The Predict-IO Score, which predicts immunotherapy response, was significantly higher in EBV-positive compared to EBV-negative samples ( p = 0.003), and scores were elevated in the PD-L1-High group compared to the PD-L1-Low group. Conclusions: In summary, EBVaGC includes subsets of different PD-L1 expression patterns with distinct activated signaling pathways, differences in ECM remodeling factors, and varying Predict-IO Scores. With these information, we could properly select the responsive patients to immunotherapy among EBVaGC.

NFE2-driven neutrophil polarization promotes pancreatic cancer liver metastasis progression.

Pancreatic cancer liver metastasis is an important factor leading to dismal prognoses. The details of adaptive immune remodeling in liver metastasis, especially the role of neutrophils, remain elusive. Here, combined single-cell sequencing with spatial transcriptomics results revealed that liver metastases exhibit more aggressive transcriptional characteristics and higher levels of immunosuppression compared with the primary tumor. We identified neutrophils_S100A12 (S100 calcium binding protein A12) cells as the pivotal pro-metastatic cluster, specifically distributed at the invasive front of the metastatic lesions. Mechanistically, our findings indicated that nuclear factor erythroid 2 (NFE2) is a key transcription factor regulating neutrophil phenotypic polarization. Metastatic tumors produce transforming growth factor β to activate the SMAD3 pathway within neutrophils, inducing NFE2-driven polarization. NFE2 promotes the transcription of peptidylarginine deiminase 4 by binding to its promoter, leading to the generation of neutrophil extracellular traps at the invasive front. Collectively, our data demonstrate that NFE2-driven neutrophil polarization is a potential target for anti-metastatic therapy.

Epithelial cell diversity and immune remodeling in bladder cancer progression: insights from single-cell transcriptomics.

The progression of bladder cancer (BC) from non-muscle-invasive bladder cancer (NMIBC) to muscle-invasive bladder cancer (MIBC) significantly increases disease severity. Although the tumor microenvironment (TME) plays a pivotal role in this process, the heterogeneity of tumor cells and TME components remains underexplored. We characterized the transcriptomes of single cells from 11 BC samples, including 4 NMIBC, 4 MIBC, and 3 adjacent normal tissues. Bulk RNA-seq data were used to validate the clinical features of characteristic cells, and protein levels of these cells were further confirmed through immunohistochemistry (IHC) and multiplex immunofluorescence. Bladder cancer progression was associated with distinct transcriptomic features in the TME. Tumor cells in MIBC displayed enhanced glycolytic activity and downregulation of chemokines and MHC-II molecules, reducing immune cell recruitment and facilitating immune evasion. This highlights glycolysis as a potential therapeutic target for disrupting tumor progression. We identified a T cell exhaustion pathway from naive CD8 + T cells (CD8 + TCF7) to terminally exhausted CD8 + STMN1 cells, with progressively declining immune surveillance. Targeting intermediate exhaustion states may restore T cell function and improve anti-tumor immunity. Macrophages polarized toward a pro-tumorigenic phenotype, while VEGFA + mast cells promoted angiogenesis in early-stage BC, suggesting their role as potential targets for therapeutic intervention in NMIBC. Furthermore, conventional dendritic cells (DCs) transformed into LAMP3 + DCs, contributing to an immunosuppressive microenvironment and enabling immune evasion. This study reveals dynamic changes in the TME during BC progression, including enhanced glycolysis, T cell exhaustion, and immune cell remodeling, which contribute to immune evasion and tumor progression. These findings identify critical pathways and cell populations as potential therapeutic targets, offering new strategies to improve treatment outcomes in BC patients.

SEC61G Facilitates Brain Metastases via Antagonizing PGAM1 Ubiquitination and Immune Microenvironment Remodeling in Non-Small Cell Lung Cancer

SEC61G Facilitates Brain Metastases via Antagonizing PGAM1 Ubiquitination and Immune Microenvironment Remodeling in Non-Small Cell Lung Cancer

Predictive signatures of immune response to vaccination and implications of the immune setpoint remodeling.

In 2020, I featured two articles in the "mSphere of Influence" commentary series that had profound implications for the field of immunology and helped shape my research perspective. These articles were "Global Analyses of Human Immune Variation Reveal Baseline Predictors of Postvaccination Responses" by Tsang et al. (Cell 157:499-513, 2014, https://doi.org/10.1016/j.cell.2014.03.031) and "A crowdsourced analysis to identify ab initio molecular signatures predictive of susceptibility to viral infection" by Fourati et al. (Nat Commun 9:4418, 2018, https://doi.org/10.1038/s41467-018-06735-8). From these topics, the identification of signatures predictive of immune responses to vaccination has greatly advanced and pivoted our understanding of how the immune state at the time of vaccination predicts (and potentially determines) vaccination outcomes. While most of this work has been done using influenza vaccination as a model, pan-vaccine signatures have been also identified. The key implications are their potential use to predict who will respond to vaccinations and to inform strategies for fine-tuning the immune setpoint to enhance immune responses. In addition, investigations in this area led us to understand that immune perturbations, such as acute infections and vaccinations, can remodel the baseline immune state and alter immune responses to future exposures, expanding this exciting field of research. These processes are likely epigenetically encoded, and some examples have already been identified and are discussed in this minireview. Therefore, further research is essential to gain a deeper understanding of how immune exposures modify the epigenome and transcriptome, influence the immune setpoint in response to vaccination, and define its exposure-specific characteristics.

Accumulation of CD38 in Hybrid Epithelial/Mesenchymal Cells Promotes Immune Remodeling and Metastasis in Breast Cancer.

Triple-negative breast cancer (TNBC) is a highly metastatic subtype of breast cancer. The epithelial-to-mesenchymal transition is a nonbinary process in the metastatic cascade that generates tumor cells with both epithelial and mesenchymal traits known as hybrid EM cells. Recent studies have elucidated the enhanced metastatic potential of cancers featuring the hybrid EM phenotype, highlighting the need to uncover molecular drivers and targetable vulnerabilities of the hybrid EM state. Here, we discovered that hybrid EM breast tumors are enriched in CD38, an immunosuppressive molecule associated with worse clinical outcomes in liquid malignancies. Altering CD38 expression in tumor cell impacted migratory, invasive, and metastatic capabilities of hybrid EM cells. Abrogation of CD38 expression stimulated an antitumor immune response, thereby preventing the generation of an immunosuppressive microenvironment in hybrid EM tumors. CD38 levels positively correlated with PD-L1 expression in samples from patients with TNBC. Moreover, targeting CD38 potentiated the activity of anti-PD-L1, eliciting strong antitumor immunity, with reduced tumor growth in hybrid EM models. Overall, this research exposes upregulation of CD38 as a specific survival strategy utilized by hybrid EM breast tumors to suppress immune cell activity and sustain metastasis, with strong implications in other carcinomas that have hybrid EM properties. Significance: Hybrid cells co-featuring epithelial and mesenchymal traits in triple-negative breast cancer express elevated levels of CD38 to induce immunosuppression and metastasis, indicating CD38 inhibition as potential strategy for treating breast cancer.

Fibroblasts in rheumatoid arthritis: novel roles in joint inflammation and beyond

High-throughput technologies in human and animal studies have revealed novel molecular and cellular pathways involved in tissue inflammation of rheumatoid arthritis (RA). Fibroblasts have been in the forefront of research for several decades. Subpopulations with specific phenotypic and functional properties have been characterized both in mouse models and human disease. Data supporting the active involvement of fibroblasts in immune responses and tissue remodeling processes, as well as their central role in promoting clinical relapses and contributing to treatment resistance, have clearly reshaped their role in disease evolution. The lung is an important non-synovial component of RA both from a clinical and an immunopathogenic aspect. Interstitial lung disease (ILD) is a significant contributor to disease burden affecting morbidity and mortality. Although our knowledge of ILD has progressed, significant gaps in both basic and clinical science remain, posing hurdles to efficient diagnosis, prediction of disease course and its effective treatment. The specific role and contribution of fibroblasts to this process has not been clearly defined. The focus of this review is on fibroblasts and their contribution to RA and RA-ILD, presenting data on genetics and immune responses associated with RA-ILD in humans and animal models.

Modulation of post‐surgery fibrosis in glaucoma filtration surgery by targeting retinoic acid signaling

Aims/Purpose: Glaucoma filtration surgery (GFS) aims to alleviate intraocular pressure by improving fluid drainage from the eye. However, post‐surgery scarring, particularly around the surgical site, can impede fluid flow and compromise surgical outcomes. Dysregulated inflammatory responses and abnormal extracellular matrix (ECM) remodeling contribute to excessive scarring. Retinoic acid signaling is known to modulate immune responses, ECM remodeling, and fibrosis in various organs. This study seeks to investigate the influence of key retinoic acid signaling components on post‐surgery scarring in the eye.Methods: Expression levels and patterns of selected retinoic acid signaling pathway components were assessed in conjunctival tissues of C57BL/6 mice undergoing GFS using RT‐PCR and Western blot analysis. Mice with global deletion of specific retinoic acid signaling pathway components underwent GFS to evaluate their role in subconjunctival fibrosis. Inflammatory responses were evaluated by immunofluorescence staining for the pan‐myeloid marker CD11b, and fibrotic tissue was visualized using Masson's trichrome stain. Subconjunctival tissues from knockout and wild‐type littermate controls were analyzed for inflammatory cytokines, chemokines, and fibrotic markers via Western blot analysis.Results: GFS induced the expression of different components of the retinoic acid signaling pathway in mice. Deletion of specific retinoic acid signaling pathway components resulted in alterations in GFS‐induced recruitment of myeloid cells and ECM remodeling.Conclusions: Modulating retinoic acid signaling presents a promising approach for controlling post‐surgery fibrosis in the eye, offering an alternative avenue to enhance the outcomes of glaucoma filtration surgery.

The Transcriptomic and Gene Fusion Landscape of Pleomorphic Salivary Gland Adenomas.

Pleomorphic adenoma (PA) is the most common salivary gland tumor. PAs are characterized by chromosomal rearrangements of 8q12 and 12q14-15, leading to gene fusions involving the PLAG1 and HMGA2 oncogenes. Here, we performed the first comprehensive study of the transcriptomic and gene fusion landscape of 38 cytogenetically characterized PAs. RNA-seq identified PLAG1 or HMGA2 fusions in 33/38 cases (87%), of which 15 were novel fusions. Fusions were found also in tumors with normal karyotype, demonstrating that they are generated by cryptic rearrangements. PLAG1 was mainly activated by promoter swapping and HMGA2 by truncation of its 3'-part. RNA-seq revealed upregulation of genes involved in extracellular matrix production, WNT-signaling, and epithelial-mesenchymal transition in PA compared to normal salivary tissue. Principal component analysis identified two PA subclusters characterized by PLAG1- and HMGA2-activation, respectively, that differed in expression of genes involved in the immune system, cell adhesion, and microenvironment remodeling. Moreover, comparative analyses of PA and salivary carcinomas revealed that PA resembles myoepithelial carcinoma. Our study reveals new oncogenic gene fusions and expands our knowledge about the molecular underpinnings of PA.

Aging-induced immune microenvironment remodeling fosters melanoma in male mice via γδ17-Neutrophil-CD8 axis

Aging-induced immune microenvironment remodeling fosters melanoma in male mice via γδ17-Neutrophil-CD8 axis

Unveiling chemotherapy-induced immune landscape remodeling and metabolic reprogramming in lung adenocarcinoma by scRNA-sequencing.

Chemotherapy is widely used to treat lung adenocarcinoma (LUAD) patients comprehensively. Considering the limitations of chemotherapy due to drug resistance and other issues, it is crucial to explore the impact of chemotherapy and immunotherapy on these aspects. In this study, tumor samples from nine LUAD patients, of which four only received surgery and five received neoadjuvant chemotherapy, were subjected to scRNA-seq analysis. In vitro and in vivo assays, including flow cytometry, immunofluorescence, Seahorse assay, and tumor xenograft models, were carried out to validate our findings. A total of 83,622 cells were enrolled for subsequent analyses. The composition of cell types exhibited high heterogeneity across different groups. Functional enrichment analysis revealed that chemotherapy drove significant metabolic reprogramming in tumor cells and macrophages. We identified two subtypes of macrophages: Anti-mac cells (CD45+CD11b+CD86+) and Pro-mac cells (CD45+CD11b+ARG +) and sorted them by flow cytometry. The proportion of Pro-mac cells in LUAD tissues increased significantly after neoadjuvant chemotherapy. Pro-mac cells promote tumor growth and angiogenesis and also suppress tumor immunity. Moreover, by analyzing the remodeling of T and B cells induced by neoadjuvant therapy, we noted that chemotherapy ignited a relatively more robust immune cytotoxic response toward tumor cells. Our study demonstrates that chemotherapy induces metabolic reprogramming within the tumor microenvironment of LUAD, particularly affecting the function and composition of immune cells such as macrophages and T cells. We believe our findings will offer insight into the mechanisms of drug resistance and provide novel therapeutic targets for LUAD in the future.

Unveiling chemotherapy-induced immune landscape remodeling and metabolic reprogramming in lung adenocarcinoma by scRNA-sequencing

Chemotherapy is widely used to treat lung adenocarcinoma (LUAD) patients comprehensively. Considering the limitations of chemotherapy due to drug resistance and other issues, it is crucial to explore the impact of chemotherapy and immunotherapy on these aspects. In this study, tumor samples from nine LUAD patients, of which four only received surgery and five received neoadjuvant chemotherapy, were subjected to scRNA-seq analysis. In vitro and in vivo assays, including flow cytometry, immunofluorescence, Seahorse assay, and tumor xenograft models, were carried out to validate our findings. A total of 83,622 cells were enrolled for subsequent analyses. The composition of cell types exhibited high heterogeneity across different groups. Functional enrichment analysis revealed that chemotherapy drove significant metabolic reprogramming in tumor cells and macrophages. We identified two subtypes of macrophages: Anti-mac cells (CD45+CD11b+CD86+) and Pro-mac cells (CD45+CD11b+ARG +) and sorted them by flow cytometry. The proportion of Pro-mac cells in LUAD tissues increased significantly after neoadjuvant chemotherapy. Pro-mac cells promote tumor growth and angiogenesis and also suppress tumor immunity. Moreover, by analyzing the remodeling of T and B cells induced by neoadjuvant therapy, we noted that chemotherapy ignited a relatively more robust immune cytotoxic response toward tumor cells. Our study demonstrates that chemotherapy induces metabolic reprogramming within the tumor microenvironment of LUAD, particularly affecting the function and composition of immune cells such as macrophages and T cells. We believe our findings will offer insight into the mechanisms of drug resistance and provide novel therapeutic targets for LUAD in the future.

Tumor Immune Microenvironment and Its Regulatory Mechanism

Gastric cancer (GC) is a highly heterogeneous tumor, and TME plays a key role in tumorigenesis and progression. TME is composed of immune cells, CAF and ECM, which regulate the growth, invasion and metastasis of GC through complex interactions. Immune escape mechanisms, such as the activation of the PD-1/PD-L1 pathway, weaken the body's anti-tumor immune response and lead to further proliferation of tumor cells. In addition, high density of immunosuppressive cells and active CAF are strongly associated with poor prognosis in patients with GC. In recent years, targeted therapeutic strategies for TME, such as immune checkpoint inhibitors (ICIs), CAF inhibitors, and ECM remodeling modulators, have shown potential therapeutic effects and become a new way to improve the survival rate of GC patients. However, due to the complexity and heterogeneity of TME, future studies still need to further explore the specific mechanism of action of TME in different stages of GC, and develop more accurate personalized treatment plans. Multidisciplinary combined treatment strategies are expected to play an important role in improving the prognosis of patients with GC.

Multimodal Spatial Profiling Reveals Immune Suppression and Microenvironment Remodeling in Fallopian Tube Precursors to High-Grade Serous Ovarian Carcinoma.

High-Grade Serous Ovarian Cancer (HGSOC) originates from fallopian tube (FT) precursors. However, the molecular changes that occur as precancerous lesions progress to HGSOC are not well understood. To address this, we integrated high-plex imaging and spatial transcriptomics to analyze human tissue samples at different stages of HGSOC development, including p53 signatures, serous tubal intraepithelial carcinomas (STIC), and invasive HGSOC. Our findings reveal immune modulating mechanisms within precursor epithelium, characterized by chromosomal instability, persistent interferon (IFN) signaling, and dysregulated innate and adaptive immunity. FT precursors display elevated expression of MHC-class I, including HLA-E, and IFN-stimulated genes, typically linked to later-stage tumorigenesis. These molecular alterations coincide with progressive shifts in the tumor microenvironment, transitioning from immune surveillance in early STICs to immune suppression in advanced STICs and cancer. These insights identify potential biomarkers and therapeutic targets for HGSOC interception and clarify the molecular transitions from precancer to cancer.

Integrated Analysis of Gene Expression and Immune Cell Infiltration Reveals Dysregulated Genes and miRNAs in Acute Kidney Injury.

Acute Kidney Injury (AKI) is a multifaceted condition characterised by rapid deterioration of renal function, often precipitated by diverse etiologies. A comprehensive understanding of the molecular underpinnings of AKI is pivotal for identifying potential diagnostic markers and therapeutic targets. This study utilised bioinformatics to elucidate gene expression and immune infiltration in AKI. Publicly available mRNA and miRNA datasets were harnessed to discern differentially expressed genes (DEGs) and miRNAs in AKI. The CIBERSORT algorithm was employed to quantify immune cell infiltration in AKI samples. Functional enrichment analyses were conducted to unravel the implicated biological processes. Furthermore, the expression of identified genes and miRNAs was validated by quantitative real-time PCR in an AKI model. Our study revealed significant dysregulation of three genes (Aspn, Clec2h, Tmigd1) and two miRNAs (mmu-miR-21a-3p, mmu-miR-223-3p) in AKI, each with p < 0.0001. These molecular markers are implicated in immune responses, tissue remodelling, and inflammation. We observed notable disturbances in specific immune cells, including activated and immature dendritic cells, M1 macrophages, and subsets of T cells (Treg, Th1, Th17). These alterations correlated significantly with AKI pathology, with dendritic cells and M1 macrophages showing p < 0.01, and T cell subsets demonstrating p < 0.05. These results highlight the intricate involvement of the immune system in AKI and indicate significant enrichment of pathways related to immune response, inflammation, and tissue remodelling, pointing to their pivotal roles in AKI pathophysiology. Our study underscored the significance of immune cell infiltration and dysregulated gene and miRNA expression in AKI. The identified genes (Clec2h, Aspn, and Tmigd1) and miRNAs (mmu-miR-21a-3p and mmu-miR-223-3p) offer potential diagnostic markers and therapeutic avenues for AKI. Subsequent investigations targeting these genes and miRNAs, along with the elucidated pathways, may augment the clinical management and outcomes for AKI patients.

Treg cells promote decidual vascular remodeling and modulate uterine NK cells in pregnant mice.

Regulatory T (Treg) cells are essential for maternal immune tolerance of the fetus and placenta. In preeclampsia, aberrant Treg cell tolerance is implicated, but how Treg cells affect the uterine vascular dysfunction thought to precede placental impairment and maternal vasculopathy is unclear. We used Foxp3-diphtheria toxin receptor mice to test the hypothesis that Treg cells are essential regulators of decidual spiral artery adaptation to pregnancy. Transient Treg cell depletion during early placental morphogenesis caused impaired remodeling of decidual spiral arteries, altered uterine artery function, and fewer Dolichos biflorus agglutinin+ uterine natural killer (uNK) cells, resulting in late-gestation fetal loss and fetal growth restriction. Replacing the Treg cells by transfer from wild-type donors mitigated the impact on uNK cells, vascular remodeling, and fetal loss. RNA sequencing of decidua revealed genes associated with NK cell function and placental extravillous trophoblasts were dysregulated after Treg cell depletion and normalized by Treg cell replacement. These data implicate Treg cells as essential upstream drivers of uterine vascular adaptation to pregnancy, through a mechanism likely involving phenotypic regulation of uNK cells and trophoblast invasion. The findings provide insight into mechanisms linking impaired adaptive immune tolerance and altered spiral artery remodeling, 2 hallmark features of preeclampsia.

Single-cell analysis of human peripheral blood reveals high immune response activity in successful ageing individuals

Beneficial remodeling of the immune system in successful ageing individuals (centenarians and supercentenarians) is critical for healthy ageing. However, mechanisms for dynamic regulation of immunity during ageing remain unclear. We use single-cell RNA sequencing (scRNA-seq) as an analytical strategy to study the dynamic regulation of immunity during aging and its molecular mechanisms at the single-cell level. We performed an integrative analysis of 87,215 peripheral blood mononuclear cells, from seven supercentenarians, three centenarians, and four elderly controls, generated by single-cell transcriptomics complemented with fluorescence-activated cell sorting. Animals experiments were also conducted to validate the makers of healthy aging found by our bioinformatic analysis and further explore the dynamic of immune changes during aging process. We found that CD8+ effector memory T cells and terminally differentiated B cells were enriched in the longevity group (centenarians and supercentenarians), whereas naïve T cells and Tregs were enriched in elderly controls. CD56dim NK cells in the longevity group activated Fc-γ receptor signaling. The higher antigen-presenting ability of CD14+ monocytes in the longevity group and the CellChat analysis indicated that CD14+ monocytes might assist active T and B cells. Here, we revealed the adaptive immune remodeling geromarkers of immunosenescence in centenarians and supercentenarians, which could be considered as biomarkers of healthy aging, and might help sustain immune responses and achieve exceptional longevity.

STING-activating photo-vaccination with glutamine metabolism reprogramming and cascade mitochondrial dysfunction for robust immune landscape remodeling

Autologous tumor cells hold great promise as personalized therapeutic vaccines. Nevertheless, the metabolic competition between tumor cells and the functional immune cells limits patient responses to autologous vaccine via fostering immunosuppressive microenvironment and facilitating immune escape. Herein, the STING-activating Photo-vaccination Inducer (BMA) is developed which can shift in immunological state from “cold” to “hot” by tumor metabolic reprogramming and the release of autologous tumor vaccines. BMA can form into peroxidase mimics in situ upon external energy stimulation,enabling continuous production of reactive oxygen species (ROS) through photodynamic effects and photosensitizer recycling. Tumor metabolic reprogramming with continually ROS generation facilitates maturation of dendritic cells (DCs) and reeducation of tumor immune landscapedue tospatiotemporallycascading mitochondrial and nucleus dysfunction, leading to in situ nucleic acid vaccine release and stimulator of the interferon genes (STING) pathway activation. As anticipated, results from Cytometry by Time-Of-Flight (CyTOF) results indicate that the tumor landscape has been altered, effectively eradicating primary tumors and inducing beneficial “Abscopal Effects” that counteract checkpoint blockade (ICB) resistance. Our work for the first time leverages a universal and novel strategy of free radical therapy and personalized therapeutic vaccine for tumor immunoscape re-education thereby enhancing αPD-L1 blockade, presenting a rational way to surmount immunotherapy barriers to available clinical treatments.

EPCO-02. HIGH CELLULAR PLASTICITY STATE OF HUMAN MEDULLOBLASTOMA LOCAL RECURRENCE AND DISTANT DISSEMINATION

Abstract Medulloblastoma (MB) is a heterogeneous pediatric brain tumor with variable clinical outcomes. Since current treatments facing limitations due to tumor resistance, less than 10% of relapsed patients survive beyond five years, with poorly understood recurrence mechanisms. To obtain cellular diversity and genetic characters in primary and recurrent MB, we conducted comprehensive analyses utilizing the single-cell/nucleus RNA sequencing (sc/snRNA-seq), snATAC-seq and spatial transcriptomics, complemented with bulk RNA sequencing and matched primary-recurrent whole exon profiles. SHH and Group_3 subgroups revealed distinct cellular subsets, including cycling, differentiated and quiescent tumor cells. Recurrent or disseminated MB displayed a dramatically different cellular ecosystem compared to primary tumors, with varying proportions of shared cell types. Local recurrence exhibited a higher enrichment of cycling tumor cells, while differentiated subset was notably present in disseminated lesions, indicating that local recurrences harbored the stronger proliferative capacity, whereas disseminations relatively well differentiated. Additionally, chromosomal alterations were assessed, reflecting distinct genetic subclones, such as chr7q gain and chr11 loss in Group_3 disseminations. Notably, the emergence of a subpopulation termed “high cellular plasticity (HCP)” during MB progression was noted, characterized by dynamic adaptability and stemness properties. As validated, HCP state was associated with increased chromatin accessibility, contributing to tumor recurrence and immune microenvironment remodeling. Functionally, inhibiting markers associated with HCP cells, such as PTPRZ1, efficiently suppressed MB metastasis in preclinical models. In conclusion, our findings fill the critical knowledge gaps in understanding the cellular diversity, chromosomal alterations and biological dynamics in MB recurrence and dissemination, offering potential therapeutic interventions.

EXTH-78. A NOVEL BISPECIFIC S100A4/TFR ANTIBODY TO REPROGRAM THE GLIOMA TUMOR MICROENVIRONMENT AND TARGET GLIOMA STEM CELLS

Abstract GBM (glioblastoma) is the most common and aggressive primary brain tumor in adults. While immunotherapy is a promising treatment modality for GBM, most clinical trials have thus far only provided a modest benefit to GBM patients. We propose that enhancing the efficacy of immunotherapy for GBM requires approaches that increase T cell infiltration and reprogram the myeloid cells to a more pro-inflammatory state. S100A4 is a major regulator of stemness, epithelial-mesenchymal transition, and a critical regulator of immune suppressive myeloid and T cell phenotypes in GBM. Targeting S100A4 in either glioma or stromal cells is sufficient to reprogram the tumor immune landscape and extend survival of glioma bearing mice, indicating that S100A4 is a promising immunotherapy target. We recently developed a S100A4 blocking monoclonal antibody; however, this antibody does not cross the blood brain barrier (BBB) efficiently to confer significant therapeutic benefit, limiting its efficacy in treating GBM. To overcome this challenge, we generated a bispecific S100A4-TfR antibody (BsA) that allows robust BBB penetration, and tested this in a pre-clinical syngeneic, orthotopic mouse model of GBM. Using ELISA, confocal immunofluorescent microscopy, flow cytometry, and bulk RNA sequencing, we report that systemic administration of the S100A4-TfR BsA results in increased immune cell infiltration with a preferential increase in CD4 T cell infiltration, and potent immune remodeling of the tumor microenvironment to a more pro-inflammatory state. Furthermore, the BsA is taken up by glioma cells intracellularly, which is correlated with depletion of intracellular S100A4 and decreased nuclear SOX2 localization, which would reduce stemness of glioma cells. Functionally, in vitro BsA treatment of primary glioma cell cultures shows decreased S100A4 expression, proliferation, and self-renewal. Our results suggest that S100A4-TfR BsA is a novel dual activity inhibitor to simultaneously reprogram the GBM immune landscape and reduce glioma stemness.