Citations

Abstract Description Generation of T cells requires dynamic epigenetic alteration of histones to control gene expression at each developmental stage. The Class I deacetylases HDAC1, HDAC2, and HDAC3 have been shown to have 3 functions: adaptor, decrotonylase, and deacetylase activity. The relative importance of each function in T cells is not known. CD2icre HDAC3 cKO mice develop severe colitis and demonstrate that HDAC3 is required during thymocyte development for positive selection, DP survival, and CD4 lineage commitment. Recently, a mutation was identified in HDAC3 that eradicated deacetylase activity (“VRPP”). We generated a novel mouse model with Cre-mediated deletion of endogenous HDAC3 and linked re-expression of HDAC3 VRPP, which maintained adaptor function but lacks both deacetylase and decrotonylase enzymatic activity. Similar to CD2icre HDAC3 cKO mice, HDAC3 VRPP mice have a block in positive selection due to failure to down regulate RORγt and decreased DP cell survival due to increased expression of P2RX7, demonstrating that HDAC3 enzymatic activity is required for these functions. However, when the positive selection defect is bypassed in HDAC3 VRPP mice, CD4+ lineage commitment is restored. Strikingly, this mouse does not develop spontaneous colitis even up to one year of age, although T cell development is still impaired. Thus, there are different roles for HDAC3 enzymatic vs. adaptor function during T cell development which are made evident in HDAC3 VRPP mice. Funding Sources NIAID RO1 AI150100; T32 AI170478; Mayo Clinic Graduate School of Biomedical Sciences Topic Categories Lymphocyte Differentiation and Peripheral Maintenance (LYM)

Abstract Background Heavy ion radiotherapy offers distinct advantages over conventional treatments due to its superior dose distribution and enhanced biological effectiveness. However, the microdosimetric characteristics of high linear energy transfer (LET) ions, particularly near the Bragg peak, remain poorly characterized. Most experimental microdosimetry has been conducted at lower LETs, leaving a critical gap in high‐LET microdosimetric data. Purpose This study aims to extend microdosimetry to several different high‐LET ions (700–2500 keV/µm) using the novel ENCORE detector. Methods Microdosimetry was performed at the Florida State University John D. Fox Accelerator Laboratory using the ENCORE detector, a multi‐sampling ionization chamber filled with low‐pressure tissue‐equivalent gas capable of resolving energy deposition from approximately 200 keV to 3 MeV. 12 C, 16 O, and 28 Si beams were accelerated to 17–55 MeV and delivered to the segmented detector. Energy deposition patterns near the Bragg peak were recorded, and microdosimetric spectra were reconstructed. Monte Carlo was used to model the experimental setup. From each measured and simulated spectra, dose‐mean lineal energy () was calculated and compared. Results The 16 strips of the ENCORE corresponded to a tissue‐equivalent thickness of 17.6 µm, enabling resolution of energy deposition immediately proximal and distal to the Bragg peak. In six measurement configurations, the ion beam stopped within this range, allowing precise mapping of the profile around the peak; in the remaining configurations, the beam fully traversed the detector. Across all beams, difference between measured and simulated averaged 12%, based on one measurement per strip. Peak measured reached 920 ± 220, 1240 ± 206, and 2460 ± 620 keV/µm for 12 C, 16 O, and 28 Si beams, respectively. Conclusion This study demonstrated that ENCORE can resolve microdosimetric spectra with high spatial and LET resolution across a range of ion energies. Measured values reasonably agreed with Monte Carlo simulations, validating the potential of ENCORE for position‐resolved microdosimetry and model benchmarking in particle therapy.

Abstract High-grade gliomas (HGG) have significant spatial heterogeneity, complicating therapeutic approaches. While multiparametric MRI (mpMRI) enables noninvasive assessment of this heterogeneity, most predictive models lack biological interpretability when used to classify or predict outcomes. Here, we developed an unsupervised framework that identifies novel mpMRI feature patterns through dimensional reduction, linking them to clinically relevant molecular signatures. We analyzed 18 advanced mpMRI features from 73 stereotactic biopsies across 20 HGG patients. Imaging data were normalized, followed by dimensional reduction using an iterative UMAP across varying parameters. Density-based clustering was used to generate stable image-derived clusters through consensus co-clustering. RNA-seq data from matched biopsies were analyzed using DESeq2 and GSEA to identify differentially expressed genes and enriched pathways. We then constructed a network linking specific imaging features to these transcriptional signatures. Validation was performed in an independent cohort, allowing us to extend clusters across full brain mpMRI. Unsupervised clustering of mpMRI features produced nine stable image-based clusters. These clusters were not fully explained by contrast enhancement, metabolic class, or tumor purity, shown by intraclass correlation (ICC) analyses, suggesting they capture novel biological variation. Transcriptomic analysis showed each cluster was associated with distinct gene expression patterns and enriched biological pathways, including neuronal differentiation, immune chemotaxis, and chromatin remodeling. Key imaging features (sRCBV, MTT, K2)—combined in cluster-specific patterns—enabled construction of an interpretable imaging–transcriptomic network. This was validated in an independent cohort. We then applied to full brain mpMRI, mapping spatially distinct regions to biological trends and patient outcomes. Addressing a research gap in imaging HGG, we present an interpretable network that connects specific patterns of mpMRI to clinically meaningful molecular signatures. This approach advances our understanding of tumor heterogeneity and supports future translational work in precision patient care.

Abstract BACKGROUND Glioblastoma (GBM) is a molecularly and phenotypically heterogeneous tumor, with regional invasion patterns driving treatment resistance and universal recurrence. In infiltrative regions, GBM cells adopt at least two predominant invasive phenotypes: Neuronal-enriched (NEU), with synaptic and neurodevelopment programs, and Glycolytic/Plurimetabolic (GPM), with heightened immune and metabolic activity. These subtypes differ in therapeutic vulnerability and microenvironmental interactions; features not captured by conventional contrast-enhanced (T1+C) MRI due to minimal blood-brain barrier (BBB) disruption. Our group sought to develop predictive models to distinguish variability of NEU/GPM subtypes within infiltrative GBM, utilizing graph convolutional networks (GCN), advanced multiparametric MRI (mpMRI), and spatially-matched image-localized biopsies with transcriptomic profiling. METHODS We analyzed 18 mpMRI features —including conventional, diffusion-weighted, and Dynamic Susceptibility Contrast (DSC) perfusion MRI— of 101 image-localized biopsies from MRI non-enhancing infiltrative margins of 64 GBM patient tumors. Features were hierarchically clustered per modality; each cluster was represented as a node in a Neo4j graph. Edges connected nodes sharing overlapping samples. Nodes were annotated with NEU or GPM subtype proportion. GCNs were trained to classify subtype-enriched nodes using graph topology and node features. RESULTS GCNs achieved high predictive performance on held-out test data (AUC = 0.95 for NEU, 0.88 for GPM; 70:30 train-test split) with similar 5-fold cross-validation performance and generalized well to an independent cohort (AUC > 0.85, n = 45). Importantly, performance was robust without relying on T1+C imaging, enabling prediction beyond BBB-disrupted regions. GCNs outperformed traditional classifiers (logistic regression, random forest). CONCLUSION Our results demonstrate that GCN-modeling of mpMRI features enables non-invasive prediction of invasive GBM subtypes at the tumor margin. Unlike convolutional neural networks (CNN), which rely on spatial continuity, GCNs leverage relational structure between heterogeneous imaging features, making them ideal for modeling sparse, non-grid data. This may support non-invasive regional vulnerability detection, surgical targeting, and improved response assessment.

Abstract Glioma stem cells (GSCs) are key drivers of glioblastoma (GBM) progression, therapy resistance, and recurrence, making them critical therapeutic targets. While the lineage-defining transcription factor OLIG2 is critical for GSC maintenance, the molecular co-regulators and downstream pathways it engages remain poorly defined. Here, we identify a previously unrecognized, functionally significant interaction between OLIG2 and STAT5B, uncovering a novel regulatory axis in GSCs. Using proximity ligation assay (PLA), we demonstrate that OLIG2 interacts with STAT5 in human and mouse GSC models, particularly in EGFRvIII-mutant cells. We find that OLIG2 preferentially partners with STAT5B and not STAT5A. Knockdown studies in GSCs reveal that OLIG2 and STAT5B co-regulate critical oncogenes BIRC5 and NUSAP1, both of which are required for sphere formation and migration capacity of EGFRvIII positive GSCs. In orthotopic xenograft models, knockdown of either STAT5B or OLIG2 significantly prolongs survival reduces expression of stemness and invasion markers, SOX2, VIMENTIN, and NUSAP1. In contrast, STAT5A knockdown fails to recapitulate these phenotypes, underscoring the role of OLIG2-STAT5B signaling in GSCs. These findings reveal a novel OLIG2-STAT5B transcriptional circuit that plays a role in GSC stemness maintenance, invasion, and tumor growth, particularly in EGFRvIII-driven gliomas. Targeting this axis may offer a new avenue for GBM treatment.

Abstract Glioblastoma (GBM) invasion is shaped by dynamic clonal evolution within the spatial and cellular architecture of the brain. To investigate this process, we integrated multiparametric MRI, whole-exome sequencing (WES), bulk RNA-seq and single-nucleus RNA-seq (snRNA-seq) from 69 IDH-wildtype GBM patients, using spatially resolved biopsies from contrast-enhancing (CE) core tumor and non-enhancing (NE) invasive regions. Phylogenetic analysis revealed distinct clonal architectures. CE-enriched clones carried truncal mutations in conanical drivers of cell cycle and RTK signaling (TP53, PTEN, EGFR, PDGFRA/B), and private alterations in DNA repair genes (CDH11, PAX7, RAD51B). In contrast, NE-enriched clones exhibited private mutations in MAPK and Hippo pathways components (STAG2, BRAF, MAP2K3, CTNNA1), suggesting region-specific evolutionary trajectories. Furthermore, snRNAseq cell-cell communication analysis uncovered enriched ligand-receptor signaling in NE regions between malignant cells and brain-resident populations, including PTN—PTPRZ1 (myeloid), NRG-ERBB4 and SLIT-ROBO (neuronal, astrocytic, and oligodendrocytic). Bulk RNAseq deconvolution confirmed increased representation of neurons and glial cells in biopsies harboring NE-specific clones. These findings define a unique population of GBM clones localized at the infiltrative margin, marked by private MAPK/Hippo mutations and enhanced intracellular communication with the surrounding brain microenvironment. The recurrent engagement of neural-glial signaling pathways suggests that these NE-associated clones co-opt normal cellular networks to support invasion, immune evasion, and persistence following surgical resection. This study establishes a mechanistic link between spatial clonal evolution and tumor-host communication at the GBM infiltrative region and provide a framework for targeting cell-cell signaling circuits that sustain invasive tumor ecosystems and escape therapeutic treatment.

Abstract IDH-wildtype glioblastoma (GBM) is the most malignant primary adult brain tumor, with a five-year survival rate below 5%. Despite maximal treatment, including surgical resection, radiotherapy, and temozolomide (TMZ), recurrence is universal, driven by profound intratumoral heterogeneity, diffuse invasion, and therapy resistance. A hallmark of IDH-wildtype GBM is frequent alteration of epidermal growth factor receptor (EGFR), including amplification (50%), the EGFRvIII deletion variant (25%), and extracellular missense mutations such as G598V and A289V (10–15%). These missense mutations increase ligand affinity and receptor activity, yet their downstream signaling consequences and role in shaping the tumor phenotypes remain poorly defined. Recent pathway-based subclassifications identified four subtypes in IDH-wildtype GBM. Notably, neuronal (NEU) and glycolytic/plurimetabolic (GPM) subtypes predominate the invasive tumor margins. NEU programs are enriched in synaptic and neurodifferentiation signals, while GPM programs are associated with metabolic adaptation, angiogenesis, and immune evasion. Here, we show EGFR-G598V and EGFR-A289V mutations differentially regulate these invasive subtypes identities. Transcriptional analysis reveals EGFR G598V upregulates metabolic and inflammatory pathways consistent with the GPM phenotype, while EGFR A289V activates genes involved in neuronal differentiation and synaptic function, aligning with the NEU subtype. Phosphoproteomics and kinase-substrate analysis further uncovered distinct kinases to be functionally activated/inhibited in EGFR A289V-expressing cells and G598V cells. Mechanistic studies revealed EGFR A289V induces sustained EGFR phosphorylation at Y992, activating PLCγ2 via the SRC-family kinase LCK. This cascade drives calcium release, CAMK1 activation, and STAT5 phosphorylation, defining a novel EGFR A289V–LCK/PLCγ2/CAMK1/STAT5 axis. In addition, Y1068 phosphorylation is also constitutively active in EGFR A289V, resulting in increased BRAF activation. In contrast, EGFR G598V activates MAPK6 and CAMK2 inducing AKT2 activation, reinforcing GPM-associated signaling. These findings uncover mutation-specific signaling networks that drive subtype fate in invasive GBM and highlight unique pathway(s) as tractable targets for subtype-directed therapy in EGFR-mutant GBM.

Abstract High-grade gliomas represent the most common type of primary adult malignant brain tumor. IDH1/2 mutations are linked to lower-grade gliomas and better patient outcomes; however, molecular profiling has revealed additional alterations that drive aggression. CDKN2A/B loss in IDH-mutant astrocytoma is recognized as a key marker of grade 4 tumor biology, yet there remain no effective targeted therapies in aggressive gliomas. Here, we analyzed a cohort of patient-derived IDH-mutant astrocytoma with whole-exome and RNA sequencing data and identified several candidate genes and pathways concurrently regulated with CDKN2A/B loss, revealing an enhanced signature related to vascular processes. We sought to elucidate the angiogenic mechanisms and functional outcomes associated with high-grade behavior following CDKN2A/B loss in IDH-mutant astrocytoma to inform future therapeutic strategies. We used grade 4 IDH-mutant astrocytoma patient-derived xenograft (PDX) and established cell lines harboring homozygous CDKN2A/B deletion. We re-expressed the cell cycle regulators p14, p15, and p16 using a Tet-inducible lentiviral system and confirmed reduced proliferation compared to parental lines. Re-expression of p14, p15, or p16 also resulted in decreased glioma cell migration and invasion. Intriguingly, re-expressing these regulators reduced VEGF secretion. HUVEC cells cultured with conditioned media (CM) from the re-expression glioma lines decreased canonical activation of PI3K/Akt and Stat3 signaling and a diminished capacity to promote endothelial migration, invasion, and tube formation. Furthermore, in vivo Matrigel plug assays using CM from the re-expression lines showed a decreased ability to recruit endothelial cells, pericytes, and form vascular organized structures. We identified the WT1 transcription factor as an upstream regulator of angiogenesis. WT1 expression was inversely correlated with CDKN2A/B expression in patient data, and knockdown of WT1 in astrocytoma cells reduced VEGF secretion and impaired ability to promote endothelial migration. Taken together, our findings reveal a novel role for WT1 in VEGF-driven angiogenesis in IDH-mutant astrocytoma with CDKN2A/B homozygous deletion.

Abstract Background Clinical nutrition (CN) is becoming increasingly complex because of the rising prevalence of chronic illness, cancer, and malnutrition‐related conditions such as short bowel syndrome and refeeding syndrome. Despite its clinical significance, nutrition education among US physicians remains limited. Simultaneously, large language model (LLM)–based artificial intelligence assistants (AIAs) have emerged as tools to support complex clinical decision‐making but remain largely untested in CN. Methods This retrospective study evaluated four LLM‐based AIAs—ChatGPT (OpenAI), OpenEvidence (OpenEvidence Inc), Gemini (Google, Google DeepMind), and Copilot (Microsoft Corporation)—using five complex CN cases from our nutrition support service. Each AIA was queried with patient‐specific CN questions. Responses were blinded and reviewed by five physician CN experts using an eight‐item assessment tool evaluating clarity, relevance, evidence, and clinical utility. Results All AIAs produced clinically appropriate responses, with Gemini scoring highest in relevance (4.04) and clarity (4.16). Overall satisfaction scores ranged from 3.08 (Copilot) to 3.84 (Gemini). Citation quality and originality of insights varied and were generally limited, and no consistent differences in performance were observed across the five cases among the four AIAs. Conclusion LLM‐based AIAs can reliably replicate expert reasoning in CN. Although not yet a source of novel clinical insights, the true potential of this approach may lie in its application among physicians without specialized expertise in CN, helping to bridge existing knowledge gaps in nutrition care. Presenting full clinical cases, as shown in this study, could support AIA‐enabled e‐consultation in the future, thereby addressing gaps in CN education.

Introduction: Transthyretin (TTR) amyloid cardiomyopathy (ATTR-CM) arises from amyloidogenic aggregates of destabilized TTR protein in variant (ATTRv-CM) or acquired wild-type TTR (ATTRwt-CM). The p.V142I (V122I) variant is the most common pathogenic TTR allele in the USA and has higher mortality vs wild type. Acoramidis, an oral TTR stabilizer achieving ≥90% stabilization, is approved in USA, UK, Europe, and Japan for treating ATTR-CM in adults. In ATTRibute-CM (NCT03860935), acoramidis showed a reduction in all-cause mortality (ACM) or first cardiovascular-related hospitalization (CVH) vs placebo (PBO), with consistent benefit in ATTRwt-CM and ATTRv-CM (all variants). We report clinical outcomes in participants with p.V142I ATTRv-CM. Research Question: What was the efficacy of acoramidis for clinical outcomes (ACM, CVH) in participants with p.V142I ATTRv-CM in ATTRibute-CM? Methods: In ATTRibute-CM, 632 participants were randomized 2:1 to receive acoramidis HCl 800 mg or PBO twice daily for 30 months. All participants enrolled in open-label extension (OLE) received acoramidis only. Exploratory post hoc analyses were done in the p.V142I ATTRv-CM subgroup. Time-to-event analyses used a stratified Cox proportional hazards model using treatment group as an explanatory factor and baseline 6MWD as a covariate, stratified by randomization factors: serum NT-proBNP and eGFR. ACM was analyzed at Month 42 (ATTRibute-CM 30 months + 12 months OLE). Results: Of the 59 patients with ATTRv-CM reported at randomization, 35 (59.3%) had p.V142I (23 acoramidis, 12 PBO) with comparable baseline characteristics; 4 were homozygous for p.V142I (1 acoramidis, 3 PBO). Through Month 30, ACM/first CVH occurred in 43.5% (10/23) of acoramidis vs 83.3% (10/12) in PBO (HR 0.311; 95% CI 0.120–0.805; Figure 1 ). Through Month 42, ACM occurred in 26.1% (6/23) of continuous acoramidis vs 66.7% (8/12) in PBO to acoramidis (HR 0.307; 95% CI 0.097–0.973; Figure 2 ). Conclusions: In participants with p.V142I ATTRv-CM, acoramidis use was associated with 69% risk reduction in ACM/first CVH through Month 30 and ACM through Month 42 vs PBO. This is the first report of clinical benefit of this magnitude observed in this high-risk population. These findings have biologic plausibility and may reflect the near-complete stabilization observed experimentally with acoramidis in p.V142I ATTR fibrils. These observations require confirmation in larger cohort studies.