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Modular engineering of Tregs for the treatment of inflammatory and autoimmune diseases 2259867
Abstract Introduction Tregs play a key role in establishing and maintaining immune tolerance and homeostasis but despite progress, current Treg therapies face challenges of unstable phenotypes, lack of IL-2 support, and tissue specificity. Methods We present a strategy to address these challenges by harnessing a gene editing platform with dual AAV to engineer human regulatory T cells (EngTreg) from bulk CD4+ T cells resulting in the stable expression of FOXP3 and a chemically inducible cytokine signaling complex providing IL-2 signaling support. Additionally, a novel hypoimmune approach improves allogeneic cell persistence by preventing T, B, and NK cell mediated rejection, making possible an off-the-shelf approach. This modular engineering allows for indication specific, interchangeable tissue specific targeting including TCRs, CARs, or inflammation tuned alarmin receptors. Results We present transcriptomic and flow cytometry analyses showing EngTregs express higher levels of core Treg and FOXP3 synergy genes, stability markers and tolerogenic proteins compared to cultured Tregs. EngTregs have been tailored for multiple indications: GNTI-122 targets IGRP for T1D, showing strong bystander suppression and disease prevention in mouse models. CAR19 EngTregs target B-cell driven autoimmune diseases with better safety compared to CAR-T cells. Gut epithelium targeting EngTregs ameliorate disease in murine IBD models. Tissue EngTregs target alarmin mediated inflammation and show efficacy in models of lung injury, AKI, and stroke. Conclusion Together, these data support EngTregs as a potent, stable, and versatile cell therapy platform for autoimmune, inflammatory, and ischemic diseases. Funding Source NIH-SBIR, venture capital, foundation funding Topic Categories Translational and Interventional Immunology (TI)
Contact‐Electrification‐Regulated Amino Acid for Improving Materials' Triboelectric Properties
ABSTRACT The dual sustainability of energy and materials, especially through the use of green molecular building blocks, is of critical importance for next‐generation self‐powered systems. Here, we propose a contact‐electrification (CE)‐regulated molecular‐engineering strategy to actively manipulate interfacial charge configurations in amino acid‐based triboelectric films. Amino acid molecules intrinsically contain both electron‐donating amino groups and electron‐accepting carboxyl groups; however, a dominant exposure of amino groups is more favorable for positive triboelectric properties. By exploiting CE‐driven interfacial electrostatic fields, preferential outward enrichment of amino groups and simultaneous interfacial anchoring of carboxyl groups and side chains to the underlying metal electrode are achieved, thereby maximizing interfacial charge transfer efficiency with a 1.51‐fold enhancement. These findings demonstrate that molecular‐level interfacial regulation, rather than material substitution alone, represents a fundamentally new paradigm for advancing triboelectric energy conversion. Notably, this CE‐regulated molecular orientation mechanism originates from the amino and carboxyl groups shared by amino acids. This finding provides a molecular perspective for understanding and regulating interfacial charge transfer in a conserved donor–acceptor framework.
Organized mucosal associated lymphoid tissue is present in the nurse shark spiral valve 2306433
Abstract Introduction Organized mucosal associated lymphoid tissue (O-MALT), such as Peyers Patches in the mammalian gut, play a critical role in immune protection by populating the mucosa with antigen-experienced B cells. Peyer’s Patches are also reported in the gut of birds, while organized lymphocyte aggregates have been described in the gut of both reptiles and amphibians. However, bony fishes appear to lack O-MALT, instead having a diffuse network of immune cells scattered throughout the gut mucosal epithelium and lamina propria. Thus, current data suggests an increase in gut lymphoid tissue organization in a common tetrapod ancestor. Methods To test this hypothesis, we used RNAscope in situ hybridization to study immune cell organization in the spiral valve (small intestine equivalent) of the nurse shark, a member of the oldest extant vertebrate lineage to possess antibody-based adaptive immunity, the cartilaginous fishes. Results We identified O-MALT in the form of AID+  T cells and AID+ B cell follicles with strikingly morphological similarity to mammalian Peyer’s Patches in the nurse shark spiral valve. We also mapped Ig bright (plasma) cells, T cells, and MHCII+ antigen presenting cells to the spiral valve lamina propria and villi. Finally, we detected antibody in spiral valve mucus and showed it coats the gut microbiota. Conclusion Together our data indicate that the emergence of O-MALT in gut occurred much earlier than previously thought, likely in a jawed vertebrate common ancestor. Funding Source TMH was supported by NIH training grant T32AI095190. The work was partly funded by the IMET Innovations in Science Program, sponsored by the G. Unger Vetlesen Foundation, awarded to HD, and the IMET director’s fund. Topic Categories Veterinary and Comparative Immunology (VET)
TLR7/9—IFNα—LDHB Checkpoint Rewires NET Subtypes in Lupus 2254935
Abstract Introduction Excessive NET deposition inflames SLE pathology, yet patients remain unusually susceptible to invasive bacterial infection–a paradox implying qualitative defects in neutrophil effector programs. Building on our prior work, we asked whether nucleic-acid sensing and type I IFN signaling enforce a neutrophil state that skews NET subtype output and undermines antibacterial defense. Methods Neutrophils from MRL/lpr mice and SLE patients were profiled using flow-based NET phenotyping, mitochondrial ROS assays, LDHB protein/mRNA analysis, calcium influx/degranulation readouts, and bactericidal assays. In vivo S. aureus bacteremia models tested hydroxychloroquine (HCQ) and IFNAR blockade. Ex vivo perturbations in human SLE neutrophils assessed rescue of LDHB, mitochondrial ROS, and NET programs. Results Chronic TLR7/9 stimulation repressed mitochondrial LDHB, blocking lactate-triggered mitochondrial ROS and bactericidal suicidal NETosis. Conversely, staphylococcal α-hemolysin induced calcium influx and rapid degranulation that released preformed IFNα and drove vital (non-lytic) NETs. The programs were antagonistic: inducing vital NETs suppressed subsequent suicidal NETosis and produced NETs with reduced antibacterial protein content and poor bactericidal activity. In vivo, combined HCQ and IFNAR blockade restored LDHB, rebalanced NET-subtype composition, and reduced organ bacterial burdens; human SLE neutrophils showed parallel rescue ex vivo. Conclusion Persistent TLR7/9—IFNα signaling enforces an LDHB-low, vital-NET—biased state that fuels inflammation yet impairs host defense.Consequently, NETs are not monolithic–suicidal and vital NETs arise via distinct triggers, carry different proteomic cargo, and diverge in bactericidal capacity. These findings establish a mechanistic checkpoint controlling NET subtype allocation and provide a rationale and biomarker framework for normalizing neutrophil function in SLE using HCQ and IFNAR blockade. Funding Source 1R35GM154838 Topic Categories Basic Autoimmunity (BA)
In Vivo Dynamics of Memory CD8+ T Cell Activation Using a Fluorescent Timer Protein of TCR Signaling 2257541
Abstract Introduction T cell activation is a dynamic process governed by the strength and duration of antigen recognition via the T cell receptor (TCR). The timing of in vivo TCR signaling— when and how strongly memory CD8+ T cells become reactivated in the inflamed brain following antigen exposure— is a fundamental mechanism that remains unclear. The Nr4a3-Tocky reporter (Tocky) mouse employs a fluorescent timer that shifts from blue to red after TCR-driven Nr4a3 activation, which allows researchers to track activation kinetics through two parameters: the Timer Angle (recency of T cell activation), and the Timer Intensity (strength of signaling). Methods To understand CD8+ T cell activation during this neuroinflammation, we generated memory CD8+ T cells by intracranial Theiler’s murine encephalomyelitis virus infection of these Tocky mice. After viral clearance, we reactivated memory CD8+ T cells by intravenously injecting their cognate antigen. Tissues were collected 0-72 hours post-injection and profiled by flow cytometry. An R pipeline was constructed to transform event-level flow data to quantify the TCR activation. Data were analyzed with a Metropolis—Hastings MCMC algorithm to model activation dynamics and estimate probabilistic transitions between biologically defined states (newly activated, persistent, arrested states). Results These empirical transition probabilities enabled Bayesian comparison of antigen-specific versus nonspecific CD8+ T cells. It was revealed that antigen-specific CD8+ T cells rapidly transition to a high-intensity activation state within 24 hours and maintain persistent signaling compared to nonspecific populations. Conclusion This establishes a biologically grounded, in vivo framework for modeling and comparing TCR signaling during neuroinflammation. Funding Source R01 - NS103212 Topic Categories Neuroimmunology (NEUR)
In vitro modeling for the in-depth characterization of mRNA vaccines 2309820
Abstract Introduction Pre-clinical evaluation of mRNA vaccine formulations requires ranking and de-risking diverse lipid nanoparticles (LNPs). The Modular Immune In vitro Construct Peripheral Tissue Equivalent (MIMIC PTE) system is a state-of-the-art, fully human platform that enables high-throughput evaluation of vaccine reactogenicity within a physiologically relevant environment. Methods The MIMIC PTE system is a three-dimensional construct composed of endothelial cells layered on a collagen matrix. Peripheral blood mononuclear cells (PBMCs) from individual human donors are introduced into the system, allowing antigen presenting cells (APCs) to migrate through the endothelial layer. Following vaccine treatment, the construct is incubated to enable cellular uptake, PBMC maturation and differentiation, and cytokine expression. Subsequent analyses include bead-based cytokine assays, flow cytometry, and transcriptomic profiling. Results This enhanced system successfully ranks mRNA-LNPs formulations across a variety of ionizable and cationic lipid formulations. Using this platform, we have deepened our understanding of LNP mechanisms of action, identifying a substantial population of cells that produce pro-inflammatory cytokines without expressing the vaccine antigen, indicative of pleiotropic activation. Cell depletion experiments demonstrated that CD14+ and/or CD11b+ myeloid cells, despite representing < 25% of all PBMCs, are the primary drivers of the global cytokine profile elicited by mRNA vaccines. In addition to inflammasome-mediated IL-1β production, we observed robust activation of TLR3 and TLR9 pathways, suggesting potential DAMP signaling via self-nucleic acids. Conclusion The fully human MIMIC PTE platform substantially improves our capacity to investigate mechanisms of action and predict vaccine safety profiles within physiologically relevant immune cell populations, thereby accelerating the development of safer, more effective vaccines and mitigating clinical trial risks. Funding Source Sanofi Topic Categories Vaccines and Immunotherapy (VAC)
Antigen specific T cell responses and cytokine production in healthy subjects following a recombinant hepatitis E vaccine 2256837
Abstract Introduction Hepatitis E virus (HEV) is a major cause of acute hepatitis in adults and has a particularly high mortality in pregnant women. HEV vaccines are known to induce protective antibodies; however, cellular immune responses following these have not been studied. We therefore evaluated T cell responses to a recombinant HEV capsid protein vaccine candidate. Methods Healthy adults (100; 61 males, mean age 34.9±9.6 years) were randomized 3:1 to receive three intramuscular doses of a HEV ORF2-based vaccine (30 α¼g/dose; n = 76) or a hepatitis-B vaccine (n = 24) at 0, 1 and 6 months. Eight subjects were lost to follow-up. Antigen-induced T cell proliferation was measured using Ki67 expression at flow cytometry. In addition, TNF-α, IL-6, IFN-γ, IL-10, IL-2 and IL-4 were measured using ELISA in culture supernatants from the proliferation assay. Results The vaccinees showed a significant increase in antigen-specific CD3 T-cell proliferation from baseline to 7 months (0.55±0.49 vs 3.00±3.59). The increase was more pronounced in CD4 T cells (1.37±4.48 vs 4.59±6.01) than in CD8 T cells (0.57±0.65 vs 1.09±1.06). By comparison, the controls showed only a minor increase in proliferation of CD3 T cells (0.54±0.31 vs 1.80±2.52). Among the cytokines, significant increase at 7 months was observed only in TNF-α (78.53±134.1 vs 234.9±301.7) and IL-6 (228.2±263.8 vs 314.2±281.9) in the test group. Conclusion In conclusion, administration of the HEV vaccine induced good antigen-specific T cell responses in the vaccinated individuals. Funding Source n/a Topic Categories Vaccines and Immunotherapy (VAC)
Helminth trained nerve and airway associated interstitial macrophages are indispensable for mitigating pathology during respiratory viral infection 2302249
Abstract Introduction Disease severity following respiratory viral infection often poorly correlates with viral burden, implicating dysfunction of inflammation regulation in host. Lung-resident macrophages are central to pulmonary homeostasis, yet how distinct subsets mediate disease tolerance is unclear. We identified CD169+ nerve- and airway-associated interstitial macrophages (NAMs) that expand early after infection and hypothesized that NAMs acquire local trained immunity to promote disease tolerance during viral challenge. Methods Mice were transiently infected with the lung-migrating helminth Nippostrongylus brasiliensis and challenged 4—5 weeks later with lethal H1N1 influenza. Survival, weight loss, lung pathology, pulmonary function, and viral burden were assessed. Immune responses were analyzed by flow cytometry, cytokine profiling, and confocal microscopy. NAM dependency was tested using a NAM-DTR model to deplete trained NAMs and replace them with untrained cells. Single-cell RNA-seq and ATAC-seq were performed to define transcriptional and epigenetic programs. Results Prior helminth exposure induced a durable trained state that protected mice from influenza-induced weight loss and mortality without altering viral burden, indicating disease tolerance. Trained mice showed reduced lung pathology and improved gas exchange. Protection was associated with elevated type-2 cytokines and increased eosinophils, whereas untrained mice developed excessive neutrophilia and NET formation. NAMs expanded following helminth exposure and were essential for protection, as depletion and replacement of trained NAMs with untrained cells abolished survival. ScRNA-seq and ATAC-seq analyses revealed stable reprogramming of NAMs toward inflammation control, lung repair, and type-2 immunity. Conclusion These findings establish NAMs as critical mediators of pulmonary disease tolerance and demonstrate that lung-resident macrophages can acquire local trained immunity that preserves tissue integrity during severe viral infection. Funding Source 5F32HL154598 (P.D.Y.), 5T3-A1100853 (P.D.Y.) R01AI188824, 1R01AI143861 (K.M.K.), 3R01AI143861-02S1 (K.M.K.), NYU Cardiovascular Research Center pilot award (K.M.K.) Topic Categories Innate Immune Responses and Host Defense: Cellular Mechanisms (INC)
Synthetic Sufficiency Models of Cell Death for Immunology 2260195
Abstract Introduction Regulated cell death is essential for tissue homeostasis and host defense. Many death pathways culminate in the loss of plasma membrane integrity which is orchestrated by distinct executioner proteins. We aim to define the cell-intrinsic programs that determine a single cell’s decision between survival and death downstream of executioners for apoptosis, pyroptosis, and necroptosis. We are also interested in regulation of the terminal NINJ1-mediated membrane rupture. Methods We engineered a suite of synthetic sufficiency models that allow inducible and temporal control of constitutively active death executioners. These executioners include truncated BID (tBID) to trigger mitochondrial apoptosis, the N terminal fragment of gasdermin D (NT-GSDMD) for pyroptosis, a phosphomimetic mutant of MLKL for necroptosis, and overexpressed wild-type NINJ1 to model terminal lytic rupture. Using chemogenetic induction or transient transfection, we precisely initiate each program and quantify cell fate outcomes by live-cell imaging, flow cytometry, and bulk enzymatic assays. Results Phenotypic analysis reveals distinct and expected membrane behaviors across models. Annexin V staining and DRAQ7 uptake delineate apoptotic cells that expose phosphatidylserine (Annexin V+/DRAQ7⁻) before secondary necrosis, while pyroptotic, necroptotic, and NINJ1-driven cells display early DRAQ7 positivity consistent with rapid membrane perforation or rupture. LDH release assays confirm minimal lysis during apoptosis but high lysis in lytic death models. ATP quantification shows apoptotic cells maintain intermediate energetic levels, whereas pyroptotic, necroptotic, and NINJ1-driven cells rapidly lose energetic viability. Conclusion Together, these synthetic death tools provide a modular framework to dissect the distinct kinetics, molecular features, and immunogenic potential of each cell death program. Funding Source K.R.R supported by the Catherine Othieno Sempa Post-Baccalaureate Fellowship, C.L.E. supported by the Ragon Early independence Fellowship, Karin Grunebaum Cancer Research Foundation Faculty Research Fellowship, and NIH grant R35GM159938 Topic Categories Immune Response Regulation: Molecular Mechanisms (IRM)
Self‐Cooperative RNA Vaccine Mitigates Dendritic Cell–Mediated Acquired Immune Resistance to Potentiate Cell Therapy for Solid Tumors
ABSTRACT Conventional mRNA cancer vaccines are designed to maximize antigen potency but often overlook vaccination‐induced immune resistance. In this study, we identified a negative immune regulatory mechanism, whereby mRNA vaccination induces programmed death‐ligand 1 (PD‐L1) expression in dendritic cells (DCs) through type I interferon (IFN‐I) signaling. Elevated PD‐L1 expression impairs T‐cell priming in lymph nodes through engagement of programmed death receptor 1 (PD‐1) on T lymphocytes. To address this challenge, we developed a self‐cooperative RNA vaccine (SCORV) strategy by co‐delivering antigen‐encoding RNA and small interfering RNA against PD‐L1 (siPD‐L1) within a single lipid nanoparticle (LNP). Through iterative screening of >300 ionizable lipids, we optimized a DC‐targeted LNP formulation with high RNA delivery efficiency and minimal immunotoxicity. SCORV simultaneously suppresses PD‐L1–mediated immune resistance during antigen presentation and enhances T cell priming while alleviating T cell exhaustion. Importantly, SCORV potentiates the tumor reactivity of adoptively transferred tumor‐infiltrating lymphocytes and elicits robust antitumor immunity in murine melanoma and hepatocellular carcinoma models. This work highlights a rational design principle for mRNA vaccines that self‐correct vaccination‐induced immune resistance.
The transcription co-regulator OCA-B/Pou2af1 differentially controls autoimmunity and tumor immunity 2307240
Abstract Introduction Expression of the transcription coregulator OCA-B in T cells is dispensable for primary immune responses but critical for CD4+ memory recall and autoimmune disease. OCA-B deletion in T cells also protects non-obese diabetic (NOD) mice from spontaneous type 1 diabetes (T1D) and blunts T1D in a subset of other more acute models. However, the roles of OCA-B in checkpoint blockade-induced diabetes and anti-tumor immunity have not been studied. Different from primary tumor, tumor relapse/rechallenge shares some similarities with memory responses. The roles of OCA-B in these rarer tumor responses have not been investigated before. Methods We used OCA-B T cell conditional mice to study the role of OCA-B in autoimmunity and anti-tumor immunity. Results Here we show that OCA-B deletion in T cells completely protects 8-week-old and partially protects 12-week-old NOD mice against PD-1 blockade-induced diabetes. Protection was associated with a failure of progenitor exhausted CD8+ cells to mature into pathogenic effectors. In parallel, we show that OCA-B T cell loss does not compromise anti-tumor immunity induced by PD-1 blockade in different immune-hot tumor models. We develop a proof-of-concept small molecule inhibitor of Oct1/OCA-B transcription complexes and show that the administration of this compound into NOD mice completely protects 8-week-old NOD mice from PD-1 blockade-induced diabetes. Using OCA-B reporter mice, we identify a small population of antigen-experienced OCA-B-expressing CD4+ T cells that are CD44-intermediate but otherwise naïve-like. We term these cells Tprogenitor-OCA-B (Tp-oca). Tp-oca cells express SLAMF7 and accumulate during MC38 and chronic infection. Conclusion These findings support OCA-B as a druggable target that could be used for the treatment of autoimmune diseases and checkpoint blockade-induced immune-related adverse events (irAEs). These findings also suggest OCA-B as a novel marker for antigen-experienced naïve-like CD4+ T cells with memory potential. Funding Source n/a Topic Categories Tumor Immunology: Checkpoints, Prevention, and Treatment (TIPT)
A Novel Dual-targeting OX40L and IL-13 Bispecific Antibody with Synergistic Effect for Atopic Dermatitis Treatment 2255160
Abstract Introduction Atopic dermatitis (AD) is a chronic inflammatory skin disorder. While the immune response in AD is predominantly mediated by T-helper 2 (Th2) cells, contributions from Th1, Th17, and Th22 pathways are also significant. Interleukin-13 (IL-13), a key Th2 cytokine, promotes B-cell proliferation and goblet cell metaplasia. Concurrently, the OX40 ligand (OX40L) and its receptor OX40 function as critical co-stimulatory immune checkpoints regulating T-cell activation. Both the anti-IL-13 antibody lebrikizumab and the anti-OX40L antibody amlitelimab have shown clinical benefits in clinical. Dual targeting of IL-13 and OX40L may thus provide enhanced Th2 suppression and induce broad inhibition across T-helper subsets. Methods Binding affinity was quantified using ForteBio, while functional inhibition of downstream signaling was assessed via STAT6 and NF-κB luciferase reporter assays. Additionally, Mixed lymphocyte reaction (MLR) assay was employed to measure the secretion of IL-2, thymus and activation-regulated chemokine (TARC), IL-17A, and IL-22, serving as indicators of Th1, Th2, Th17, and Th22 activity, respectively. Finally, the in vivo efficacy was evaluated in a murine AD model. Results The bispecific exhibited high binding affinities to both IL-13 and OX40L. It also potently inhibited both STAT6 and NF-κB pathway activation. In MLR assays, the bispecific and the anti-OX40L antibody similarly suppressed the release of IL-2, IL-17A, and IL-22. Notably, the bispecific agent demonstrated superior inhibition of TARC release compared to either parental antibody alone. In vivo, the bispecific showed the highest therapeutic efficacy. Conclusion The anti—OX40/IL-13 bispecific exhibited enhanced suppression of Th1, Th2, Th17, and Th22 inflammatory responses both in vitro and in vivo, indicating synergistic immunomodulation through dual target engagement. These findings underscore its strong potential for future clinical development in the treatment of AD. Funding Source Nanjing Leads Biolabs Co., Ltd Topic Categories Therapeutic Approaches to Autoimmunity (THER)
MHC-II-restricted neoantigen expression drives tumor infiltration, activation, and maintenance of nonspecific T cells in lung cancer 2251169
Abstract Introduction Cancer mutations can result in the expression of tumor-specific “neoantigens” that are targeted by T cells. Neoantigen expression drives robust T cell infiltration of tumors across cancer models, yet most of these T cells are not specific for the neoantigens and contain both pathogenic and immunosuppressive subsets. We sought to investigate how neoantigen-specific CD4+ T cells orchestrate nonspecific “Passenger” T cell responses within tumors and determine their impact on tumor progression. Methods Our studies used an autochthonous murine model of lung adenocarcinoma (KrasG12D/+; tp53-/-) in which tumors are synchronously initiated via lentiviral delivery of Cre recombinase with neoantigen, and employed flow cytometry, histological analyses, single-cell transcriptomics, and temporal immune cell perturbations. Results Expression of a single MHC-II-restricted neoantigen (mITGB1) unexpectedly promoted tumor development. mITGB1 expression drove robust intratumoral enrichment of CD4+ and CD8+ Passenger T cells compared to mITGB1-neg controls. A fraction of Passenger T cells in mITGB1+ tumors uniquely displayed hallmarks of antigen experience, clonal expansion, and readily formed aggregates with B cells. Passenger T cells persisted in mITGB1+ tumors for at least three weeks, while lymphocytes in mITGB1-neg tumors rapidly vanished if cells could not traffic from the periphery. Critically, long-term maintenance of Passenger T cells within tumors required continuous expression of the neoantigen. Conclusion Our study reveals an unappreciated role for neoantigen-specific CD4+ T cells in promoting tumorigenesis and actively coordinating non-specific Passenger T cell responses within tumors. Paradoxically, neoantigen expression may furnish the tumor microenvironment with novel T cell specificities and unlock cell states that support tumor growth alongside populations that work to combat progression. Funding Source Jane Coffin Childs Fund, Hope Funds for Cancer Research Topic Categories Lymphocyte Differentiation and Peripheral Maintenance (LYM)
Host Lipid and Mitochondrial Metabolism Networks as Functional Targets for Novel Antimicrobials against Intracellular Bacteria 2309839
Abstract Introduction Host lipid metabolism has been implicated in viral and bacterial pathogenesis. Here we identify that temporal manipulation of host cell lipid synthesis pathways via the targeting of a master regulator of lipid biogenesis by unique lipids present in the virulent intracellular bacterium Francisella tularensis (FT) is a critical element of this bacterium’s virulence. Methods We utilized both in vitro and in vivo models to establish the pathways associated with Franicsella lipid (FTL) modulation of host cell function. Results Pretreatment of cells with FTL triggered lipid droplet formation and significantly increased replication of FT. We observed that FTL and FT triggered rapid and transient upregulation of genes associated with lipid biogenesis. Specifically, increased expression of the master regulator PGC-1a, ACOX-1, and FASN was observed within the first 8 hours of exposure to FTL or FT infection. PGC-1a controls lipid droplet biogenesis and mitochondrial function, both of which are critical for optimal replication of FT. Inhibition of PGC-1a dramatically impaired formation of lipid droplets in untreated and FTL treated cells. Importantly, impairment of PGC1-a also completely controlled FT replication in a fashion that was independent of the generation of reactive oxygen species. Conclusion Our data identify the unique lipids associated with FT play an important role in pathogenesis via their ability to control multiple metabolic nodes within the host cell. Specifically, their ability to rapidly activate host molecules that drive lipid droplet biogenesis and mitochondrial function greatly improved FT replication and spread. Our data also show that targeting of these pathways, e.g. inhibiting PGC-1a, with drugs that are well tolerated in other models of inflammation may be an effective alternative to or supplemental treatment for antibiotic intervention. Funding Source Division of Intramural Research, NIAID,NIH Topic Categories Innate Immune Responses and Host Defense: Molecular Mechanisms (INM)
Shield or Showcase: Unraveling How BCR Binding Shapes MHCII Peptide Presentation for CD4+ T Cell Activation 2256684
Abstract Introduction Effective cooperation between B cells and CD4+ T cells is central to durable antibody responses. This interaction is mediated by B cells presenting MHC class II (MHCII) peptides to activate CD4+ T cells. However, how B cells select the optimal MHCII peptide for display remains unresolved. Two competing models seek to explain this process. Model 1 suggests the B cell receptor (BCR) displays MHCII peptides derived broadly from the internalized immunogen. Model 2 states that BCR binding protects its contact region, thereby selecting MHCII peptides from the shielded region. Resolving this mechanistic question is fundamental to faster immunogen design and improved vaccine efficacy. Methods We developed a computational framework integrating structural, proteolytic, and immunogenetic features. Antigen—antibody complexes were analyzed using modeling tools to map BCR—epitope contact sites. Patient-specific MHCII alleles and immunogen sequences were evaluated with NetMHCIIpan to predict high-affinity candidate MHCII peptides. Custom Python pipelines are being developed to model endolysosomal protease cleavage and assess antibody influence on peptide stability. Results Preliminary analysis of spatial proximity of candidate MHCII peptides to the antibody interface and modelling of MHCII—peptide interactions by molecular docking indicates a possible co-relation between BCR proximity and predicted MHCII peptide affinity. Assessment of how antibody engagement influences peptide degradation is ongoing. Altogether, these results will develop a user-accessible tool that can predict optimal MHCII peptides within vaccine immunogens that elicit greater antibody responses. Conclusion This framework provides a powerful lens to visualize BCR engagement with the MHCII peptides using computational modelling. Insights from these studies will advance understanding of T—B cell cooperation and guide rational design of next-generation vaccines that more effectively recruit CD4+ T cell help and drive durable antibody responses Funding Source LOS ALAMOS NATIONAL LABORATORY Topic Categories Immune Response Regulation: Molecular Mechanisms (IRM)
The antigen-specific immunotherapy, AKS-107, selectively depletes anti-insulin B lymphocytes in the pancreas and reduces insulitis in T1D-prone mice 2259549
Abstract Introduction Type 1 diabetes (T1D), results from T cell-mediated destruction of insulin-producing beta cells. Developing new immunotherapies that control islet autoimmunity without compromising protective immune responses is an important therapeutic goal. Insulin autoantibodies predict T1D in mice and humans, implicating B lymphocytes in islet attack. Anti-insulin B cells present autoantigen to T cells to promote T1D in non-obese diabetic (NOD) mice and are required for diabetes to develop. AKS-107 is an antigen-specific immunotherapy composed of a hormonally inactive insulin moiety that lacks a critical T cell epitope and which is fused to an Fc. AKS-107 specifically eliminates anti-insulin B cells and prevents diabetes in mice. However, it is unknown whether AKS-107 can deplete anti-insulin B cells in the pancreas or reduce islet inflammation. Methods VH125SD.NOD mice were treated either once or twice weekly with 0.4 mg/kg of AKS-107 for either 2 or 8 weeks. Pancreas, pancreatic lymph nodes, and spleen were harvested, and flow cytometry phenotyping was used to evaluate selective anti-insulin B cell depletion. NOD mice were treated twice weekly with 2 mg/kg of AKS-107 for 9 weeks after which pancreata were harvested, sectioned, and insulitis severity was blind scored. Results AKS-107 therapy selectively depletes anti-insulin B cells (but not other B cells) in the pancreas and pancreatic lymph nodes of VH125SD.NOD BCR transgenic mice, in which a 1-3% population of anti-insulin B cells can be readily tracked. This observed depletion is not caused by AKS-107 masking the detection of the anti-insulin B cells by biotinylated insulin. NOD mice treated with AKS-107 from 4-13 wks of age showed decreased insulitis severity relative to controls. Conclusion Our data highlight pancreas tissue actions of the antigen-specific immunotherapy, AKS-107 which supports its potential as a novel T1D immunotherapy. Funding Source This work was supported by Breakthrough T1D (formerly the Juvenile Diabetes Research Foundation) grants 3-SRA-2024-1562-S-B (RHB) and 3-PDF-2024-1495-A-N (DHM), and NIH grants F30 DK145130 (LMC), F31 DK141224 (LEB), T32 AR059039 (TWJ), and T32 GM007347 (LMC). We also acknowledge the Vanderbilt University Medical Center (VUMC) Flow Cytometry Shared Resource [supported by supported by the Vanderbilt Ingram Cancer Center (P30 CA068485) and the Vanderbilt Digestive Disease Research Center (P30 DK058404)], the VUMC Tissue Pathology Shared Resource [supported by NCI/NIH Cancer Center Support Grant, P30 CA068485], and the Islet and Pancreas Analysis Core [supported by the Vanderbilt Diabetes Research and Training Center (NIH grant P30 DK20593)]. Topic Categories Therapeutic Approaches to Autoimmunity (THER)
B-Cell Tolerance Checkpoint Fidelity in Patients with Myelin Oligodendrocyte Glycoprotein Antibody-Associated Disease (MOGAD) 2259996
Abstract Introduction Myelin oligodendrocyte glycoprotein antibody-associated disease (MOGAD) is an autoimmune demyelinating disorder of the central nervous system classically characterized by optic neuritis, transverse myelitis, and/or encephalitis. Although MOG-specific autoantibodies are a diagnostic hallmark, the cellular mechanisms underlying their production remain poorly understood. In several autoimmune diseases, impaired B-cell tolerance checkpoints allow autoreactive B cells to escape negative selection and may contribute to the production of pathogenic antibodies. Here, we investigate whether similar tolerance defects contribute to MOGAD pathogenesis. Methods A well-established tolerance assay was utilized to assess checkpoint fidelity by generating recombinant monoclonal antibodies (mAbs) from naïve B-cell subsets downstream of the central and peripheral tolerance checkpoints. Peripheral B cells were enriched by negative selection, and single transitional/new emigrant (CD19+CD21loCD10+IgMhi CD27⁻) and mature naïve (CD19+CD21+CD10⁻IgM+CD27⁻) B cells were sorted by flow cytometry. Ig heavy and light chain genes were amplified, and mAbs were expressed and tested for polyreactivity by ELISAs against dsDNA, insulin, and LPS and for autoreactivity against HEp-2 cell lysates. Additionally, single-cell 5’ gene expression data paired with B-cell receptor (BCR) sequencing were analyzed using a droplet-based single-cell RNA sequencing platform to define naïve B-cell expression profiles together with their corresponding immunoglobulin heavy and light chain genes. Results Preliminary analyses suggest that tolerance checkpoint fidelity in MOGAD differs from that observed in other autoimmune diseases. Ongoing studies will expand these analyses in larger cohorts to better delineate the integrity of central and peripheral tolerance checkpoints and their contribution to MOGAD immunopathology. Conclusion NA Funding Source This study was supported by the National Institute of Allergy and Infectious Diseases (NIAID) of the NIH under award numbers R01-AI114780 and R21-AI164590 (to KCO). GW is funded through Yale - Cambridge Collaborative Research Programme Topic Categories Immune Response Regulation: Cellular Mechanisms (IRC)
Ultrastrong and Tough Anisotropic Organogel via Alignment‐Induced Densification and Glycerol Plasticization
ABSTRACT Polymer gels, such as hydrogel and organogel, are widely used in tissue engineering, biomedicine, and flexible electronic devices due to their good compatibility and environmental stability. However, it is still a great challenge to achieve simultaneous high strength, toughness, and elongation to meet the demands of different applications. Herein, this work proposes a strategy involving freeze‐thawing, stretch‐drying and thermal annealing in glycerol to fabricate ultrastrong and tough polyvinyl alcohol (PVA) organogel with anisotropic microstructure. Freeze‐thawing cycles push PVA chains together through the formation of ice crystals, facilitating the entanglement of polymer chains and the formation of hydrogen bond network. Directional stretch‐drying process induces molecular chain orientation and enhances the intermolecular interactions. Thermal annealing (TA) in glycerol promotes chain rearrangement and densifies the cross‐linking of hydrogen bond network, which synergistically enhances strength and toughness. The prepared anisotropic PVA organogel exhibits extraordinary toughness of 416 , excellent strength of 61.14 MPa, and fracture strain of 961%. Simultaneously, the organogel has high optical transparency () and superior stability in sub‐zero conditions. This work provides a novel and effective strategy for fabricating ultrastrong and tough organogel, offering promising potential applications in bionic tendons for soft robotics and transparent protective shields.
Tailoring Weakly Coordinating Electrolytes via Orbital‐Overlap‐Enhanced Dipole–dipole Interactions for Low‐Temperature Lithium‐Ion Batteries
ABSTRACT Lithium‐ion batteries (LIBs) suffer rapid capacity fade at low temperatures. Weakly coordinating electrolytes via adding low‐polarity or non‐coordinating co‐solvents (such as fluorinated ethers) have shown promise in rapid desolvation, yet these electrolytes often exhibit low ionic conductivity at low temperature, limiting the application of high‐energy density LIBs. Here, we design a weakly coordinating electrolyte by incorporating the non‐coordinating co‐solvent (pentafluoroethyl)trimethylsilane (PFTMS) into the coordinating solvent diethyl carbonate (DEC) via orbital‐overlap‐enhanced dipole–dipole interactions. The slight Si─O orbital overlap drives strong dipole–dipole interactions between PFTMS and DEC. This interaction lowers the negative electrostatic potential at the carbonyl oxygen of DEC, thereby weakening Li + ‐DEC coordination. Thus, by leveraging enhanced dipole–dipole interactions, this strategy realizes a weak Li + ‐solvent coordination through a lower content of PFTMS (10 vol%) compared to traditional fluorinated solvents. Besides, the designed electrolyte delivers sufficient ionic conductivity of 1.56 mS cm −1 at −40°C. Accordingly, the graphite || LiNi 0.8 Co 0.1 Mn 0.1 O 2 (NCM811) coin cell shows reversible capacity of 156.5mAh g −1 at −40°C. Notably, 4.7 Ah graphite || NCM811 pouch cell also demonstrates 219.8 Wh kg −1 at −20°C. This work advances the design of traditional weakly coordinating electrolytes via an orbital overlap strategy, which paves the way for application in extreme environments.
Immunotherapeutic control of MHC I-heterogeneous tumors to minimize therapy resistance 2307892
Abstract Introduction The anti-tumor activity of CD8 T cells, which recognize MHC I-presented tumor antigens, is enhanced by blocking checkpoint receptors. Longitudinal tumor biopsies from patients with acquired resistance to anti-PD-1 therapy often reveal genetic deficiencies in antigen presentation or MHC I expression. The role of CD8 T cells in selecting for MHC I-deficiency has been inferred but never demonstrated. Defining causes of MHC I loss and approaches to prevent it are essential for improving the outcomes of cancer immunotherapy. Methods To define immune cells responsible for selecting for outgrowth of MHC I-deficient tumors, we developed MHC I-mixed tumor models, applied immunotherapies, and depleted immune subsets. Tumor growth and composition (MHC I+ vs MHC I-) were determined. Results PD-1 blockade immunotherapy was poorly effective in mixed MHC I+/MHC I- tumor models and exerted potent selection mediated by CD8+ T cells for outgrowth of MHC I- cells. In contrast, IL-2 “superkines”, alone or in combination with STING agonists, were more effective and prevented outgrowth of MHC I- tumor cells in a manner dependent on multiple immune cell types. An alternative therapeutic approach in which Treg cells are depleted intratumorally also suppressed growth of both MHC I+ and MHC I- tumor cells, in this case largely independently of CD8 T cells. Conclusion We demonstrated for the first time in a controlled setting with mixed MHC I+/- tumors that anti-PD-1 therapy selects for outgrowth of MHC I-deficient tumor cells, dependent on CD8 T cells. Outgrowth of MHC I-deficient cells was minimized and mixed tumors were better controlled with IL-2 “superkine” therapy, alone or in combination with a STING agonist, and intratumoral Treg depletion. Tumor control in those cases was mediated by multiple effector cell types. Thus, immunotherapies that drive mixed responses of CD8+ and CD4+ T cells and NK cells have the potential to minimize acquired resistance resulting from MHC I loss. Funding Source NIH: R01CA270790 Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)