Theranostics

Glycoproteins lining the luminal endothelial surface form the glycocalyx, composing the tripartite blood brain barrier. We explored the glycocalyx as a source of danger signals for complement lectin pathway after ischemic stroke. Our data indicate that hypoxic microvascular cells increased -D-mannosyl and N-acetylglucosaminyl exposure after re-oxygenation, favoring mannose binding lectin (MBL) pathogenic deposition, and overexpression of inflammatory genes (ICAM-1 and MMP-2). The hypoxia-conditioned medium induced neuronal damage (reduced MAP-2), microglia and astrocytic reactivity (increased/thickened ramifications) when applied to induced pluripotent stem cell-derived neurons, astrocytes and microglia co-cultures. All these effects were counteracted by mannose-capped gold nanoparticles (Man-GNPs), shown to bind and sequester MBL from the medium. We then tested the Man-GNPs in vivo, in an ischemic stroke model using humanized mice, knocked-in for human MBL. The ischemic mice (males:females 1:1) treated with Man-GNPs (3h after the ischemic onset) exhibited less anxiety at the elevated plus maze and reduced neuronal loss at 8d after ischemia compared to vehicle-treated. Thus, multivalent Man-GNPs represent a promising approach to take MBL away from its glycoproteic targets on the ischemic endothelium, hence preventing downstream pathogenesis. Moreover, these data support circulating MBL as a druggable pharmacological target to prevent the thrombo-inflammatory events following acute brain injury.

ObjectivesFutile recanalization (FR) after endovascular thrombectomy (EVT) is driven by neutrophil-mediated thromboinflammation, yet conventional neuroprotectants fail to address the spatiotemporal complexity of stroke injury. This study aimed to identify the clinical link between circSCMH1, traditionally viewed as a late-stage repair molecule, and FR, while evaluating its hyperacute anti-inflammatory potential via a neutrophil-targeted delivery system. MethodsWe analyzed thrombi and plasma from LVO-AIS patients stratified by functional outcomes. We engineered neutrophil-targeted lipid nanoparticles (circSCMH1@pepLNP) for delivery in a mouse transient middle cerebral artery occlusion model. Therapeutic outcomes were evaluated through infarct volume measurement, histological assessment of NETosis, and intravital two-photon microscopy to monitor real-time neutrophil dynamics and capillary stalling. ResultsClinical analyses showed circSCMH1 was significantly downregulated in FR patients, with its levels negatively correlated with NET burden. Furthermore, circSCMH1-negative neutrophils exhibited a higher propensity for NETosis within the thrombus microenvironment. In mice, circSCMH1@pepLNP achieved specific neutrophil delivery, significantly reduced infarct volume, and suppressed the expression of citrullinated histone H3 and neutrophil elastase. Intravital two-photon and laser speckle imaging further confirmed that this intervention attenuated neutrophil adhesion, increased migratory velocity, and alleviated capillary stalling, thereby restoring cortical microvascular perfusion. ConclusionsOur findings associate neutrophil-specific circSCMH1 downregulation with clinical FR in LVO-AIS. Targeted intracellular delivery reveals an acute anti-thromboinflammatory function that complements established reparative properties, proposing a novel adjunctive approach for EVT to address the spatiotemporal complexity of stroke and improve clinical outcomes.

The development of advanced therapies for stroke, spinal cord injury or neurodegenerative diseases -main causes of death, disability and dementia- requires a profound understanding of the complex interactions established among excitotoxic neuronal death, aberrant neurotrophic-signaling, glial reactivity, and neuroinflammation. However, the master proteins coordinating these mechanisms have not been yet defined. Different evidence suggests that the truncated form of the neurotrophin tyrosine kinase receptor, TrkB-T1, might play such a key role. The levels of this TrkB isoform increase in stroke while those of the full-length pro-survival isoform (TrkB-FL) are reduced. Additionally, ischemic stroke and, specifically, excitotoxicity induce TrkB-T1 regulated intramembrane proteolysis (RIP), a process releasing a receptor ectodomain able to bind the brain-derived neurotrophic factor (BDNF) and leading to decreased BDNF-signaling. We hypothesize that the second RIP product, TrkB-T1 intracellular domain (TrkB-T1-ICD), might similarly contribute to neurotoxicity but also reactive gliosis and neuroinflammation. Herein, we first demonstrate migration of the cytoplasmic TrkB-T1-ICD to the nuclei of neurons undergoing excitotoxicity, suggesting a possible role in the transcriptional control induced by injury. Then, taking advantage of cell-penetrating peptides (CPPs), we produce a TrkB-T1-ICD mock peptide (Bio-LTT1Ct) containing the short TrkB-T1 intracellular region (23 amino acids) and test it in vitro and in vivo. Notably, this peptide migrates to the nucleus of both neurons and astrocytes cultured in vitro and provokes cell death. Additionally, Bio-LTT1Ct induces early transcriptional changes in neurons resembling those triggered by excitotoxicity such as the inhibition of the promoter activity of pro-survival transcription factors CREB and MEF2, and altered mRNA levels of their regulated genes. In vivo, Bio-LTT1Ct is accessible to the brain cortex after intranasal delivery, being efficiently distributed into cortical neurons and astrocytes of both hemispheres. Moreover, peptide administration is sufficient to promote important pathological hallmarks of stroke such as the imbalance of the TrkB isoforms, and the reactivity of astrocyte and microglia, cells that acquire proinflammatory profiles. Altogether, these results establish TrkB-T1 RIP as a central mechanism of ischemic damage and demonstrate that the receptor intracellular region is sufficient to recapitulate stroke-like effects on neurotoxicity, glial reactivity and neuroinflammation.

Current treatment of IDH-wildtype glioblastoma (GBM) relies on the first-line chemotherapy-temozolomide. Although MGMT methylation is routinely conducted to predict chemosensitivity, its efficacy is often compromised. Thus, there is an urgent need to discover more accurate prognostic biomarkers. Cholesteryl ester (CE) has been recently recognized as a key feature of GBM, however, its role in GBM prognosis remains poorly understood. We first employed label-free stimulated Raman scattering (SRS) imaging to quantitatively analyze CE level in intact tumor tissues obtained from IDH-wildtype GBM patients. Our result revealed significantly prolonged 2-year overall survival (OS) in patients with CE level [&ge;] 40% compared to those with CE level < 40%. CE outperformed MGMT methylation for 2-year OS prognosis (AUC: 0.836 vs. 0.763). Importantly, CE also achieved superior prognostic performance over MGMT methylation on an independent cohort, with higher sensitivity (0.856 vs. 0.667), specificity (0.833 vs. 0.583), NPV (1.00 vs. 0.667), PPV (0.833 vs. 0.583). Given synergistic effects between CE and MGMT methylation, we developed a prognostic model combining these two biomarkers. Specially, machine learning (XGBoost) model exhibited optimal performance in the training cohort (AUC: 0.920), and maintained its superior performance on the independent cohort (sensitivity: 0.946, specificity: 0.873, NPV: 1.00; PPV: 0.917). Mechanistically, integrative analysis of TCGA database linked poor prognosis to the coordinated upregulation of genes involved in cholesterol efflux, hydrolysis, transport, and inhibition of de novo synthesis, unraveling a possible underlying mechanism between poor prognosis and cholesterol metabolism. This work identified CE as a prognostic biomarker for IDH-wildtype GBM.

Intracerebral haemorrhage is the most severe subtype of stroke; however, pre-clinical investigation often fails to translate to the clinic. Cerebral organoids offer an adaptable, in vitro model of human brain tissue for pre-clinical investigation of disease. We recently demonstrated that the tissue can be successfully vascularised to mimic the cerebrovasculature. Cerebrovascular weakness was induced with atorvastatin to mimic damage observed in intracerebral haemorrhage and to replicate the diseases pathological features. We used atorvastatin to disrupt functional morphology in human brain microvascular endothelial cells in 2D and 3D model systems. Whole human blood was added to initiate damage to cerebral tissues. Vascularised cerebral organoids exhibited loss of vascular integrity when treated with atorvastatin. Tissue was vulnerable to injury from human whole blood, and an innate immune response was initiated, resulting in increased cell death. Here we show that vascularised cerebral organoids demonstrate a novel model platform for investigating pathology associated with human whole blood insult in intracerebral haemorrhage.

Breast cancer is the most common malignancy in women, with triple-negative breast cancer (TNBC) representing the most aggressive subtype and carrying a poor metastatic prognosis. Metastasis requires tumor cells to cross the endothelial barrier, a process facilitated by tumor-derived extracellular vesicles (EVs), which can disrupt vascular integrity. Fluid shear stress (FSS), generated by blood flow, shapes endothelial physiology and may influence EV uptake, yet the mechanisms underlying TNBC-derived small EV (sEV) internalization remain unclear. Here, we investigated TNBC sEV-endothelial interactions using combined in silico and in vitro approaches. Human umbilical vein endothelial cells (HUVECs) were cultured under static or FSS conditions (20 dyn/cm{superscript 2}), followed by proteomic profiling and protein-protein interaction analyses with sEV proteomes. Uptake assays employed pharmacological inhibition (Dynasore, M{beta}CD, Pitstop2), Caveolin-1 (CAV-1) and Clathrin Heavy Chain (CLHC), siRNA-mediated knockdown, and junctional interaction analyses via confocal microscopy and co-immunoprecipitation. FSS downregulated proliferation- and angiogenesis-associated proteins while upregulating adhesion and cytoskeletal regulators assessed by proteomics. Network analysis identified clathrin- and caveolin-mediated endocytosis (CME and CavME), integrins, and early endosomes as central mediators of sEV uptake. Functionally, uptake was reduced by Pitstop2, M{beta}CD, and CAV-1/CLHC knockdown under static conditions, but silencing paradoxically enhanced uptake under FSS, suggesting compensatory flow-dependent pathways. Notably, under FSS, sEVs accumulated at endothelial junctions, colocalizing with VE-CAD and associating with CLDN5, indicating a potential disruption mechanism of adherens and tight junctions and consequent endothelial permeability. These findings identify CME and CavME as key uptake routes while underscoring FSS as a critical determinant of endothelial-tumor EV interactions. By revealing junctional targeting of sEVs, this work provides new mechanistic insight into vascular remodeling during metastasis and highlights EV pathways as potential therapeutic targets in TNBC. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=104 SRC="FIGDIR/small/721946v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@f91c5org.highwire.dtl.DTLVardef@2b4dc8org.highwire.dtl.DTLVardef@ff94f1org.highwire.dtl.DTLVardef@18b714b_HPS_FORMAT_FIGEXP M_FIG C_FIG Uptake and localization of sEVs on HUVEC under (a) static and (b) fluid shear-stress conditions. sEVs: Small Extracellular Vesicles. CME: Clathrin-mediated Endocytosis. CavME: Caveolin-mediated Endocytosis. CLDN5: Claudin-5. VE-CAD: Vascular Endothelial Cadherin. FSS: Fluid shear-stress.

Immune checkpoint inhibitors (ICI) have revolutionized cancer therapy, yet response rates remain suboptimal across many solid tumors, and resistance mechanisms, particularly those involving glycans, are not fully understood. Recent studies have identified sialic acid-containing glycans and their interactions with Siglec receptors on tumor-associated macrophages as an important contributor to immune suppression within the tumor microenvironment (TME). Targeting this sialic acid-Siglec axis by glycan engineering with sialidases and other glycosidases has shown therapeutic potential in preclinical models. However, safe and effective delivery of sialidases to tumors remains a challenge. Here, we present a novel approach using adeno-associated virus (AAV)-mediated therapy to deliver sialidases (AAVSia) and other glycosidases, including fucosidase, directly to the TME. Intratumoral administration of AAVSia in mouse models resulted in significant tumor growth reduction, enhanced survival, and robust systemic antitumor immunity through improved cross-presentation and dendritic cell activation. Furthermore, combining local sialidase expression with fucosidase treatment and classical PD-1 blockade allowed a synergistic effect, amplifying antitumor response. Our findings highlight the therapeutic promise of glycoengineering the TME using local delivery systems and support the development of combination strategies to overcome glycan-mediated resistance in cancer immunotherapy. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=129 SRC="FIGDIR/small/720097v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@dc9d72org.highwire.dtl.DTLVardef@1e4e455org.highwire.dtl.DTLVardef@4a8f93org.highwire.dtl.DTLVardef@11813a3_HPS_FORMAT_FIGEXP M_FIG C_FIG

Radiotherapy is widely used in the management of brain tumors; however, it is often associated with delayed adverse effects, including cognitive decline and depression-like behavior. These effects are thought to arise, in part, from suppressed hippocampal neurogenesis, altered neuronal architecture, and microglial dysfunction. Despite this, the precise mechanisms underlying irradiation-induced cognitive deficits, as well as effective therapeutic interventions, remain poorly understood. In the present study, six-month-old male mice were subjected to a single 9 Gy dose of cranial irradiation, followed by behavioral assessments several weeks post-exposure. We observed that cranial irradiation significantly impaired hippocampal-, prefrontal cortex-, and cortical-dependent memory functions. Notably, treatment with dehydrozingerone (DH), a curcumin analog (50 mg/kg, oral administration for two weeks), markedly prevented these cognitive deficits. At the molecular level, irradiation disrupted the activity of key enzymes involved in the tricarboxylic acid (TCA) cycle and the glutamate-glutamine/GABA cycle, both of which were restored following DH treatment. Furthermore, irradiation induced dysregulation of genes and proteins associated with glycolysis (Atp2b1, mt-Nd2, mt-Atp6), mitochondrial energetics (mt-Atp8, mt-Cytb), glucose transport (Slc4a5), insulin resistance (Etnppl), lipid metabolism (Pla2g3, Plin4), and inflammation (Ighg2c), all of which were significantly normalized by DH. Importantly, DH also prevented irradiation-induced loss of cell-type-specific glucose transporter expression, including GLUT3 in neurons and GLUT5 in microglia. In conclusion, our findings suggest that DH is a promising therapeutic candidate for mitigating irradiation-induced energy deficits and cognitive impairments, likely through modulation of metabolic and mitochondrial pathways.

Immune checkpoint inhibitors (ICI) have revolutionized cancer treatment, but their efficacy has now reached a plateau. ICIs are the first class of treatment targeting the crosstalk between immune and tumor cells, making it crucial to understand the complex interactions within the tumor microenvironment (TME) to enhance therapeutic responses. The elevated consumption of resources by cancer cells, coupled with limited vascularization, often results in a TME that is deficient in nutrients, leading to competition for resources between cancer and stromal cells. Consequently, targeting tumor metabolism has emerged as a promising strategy to improve the efficacy of ICIs. Through metabolomic analysis, we have identified metabolic alterations in melanoma cells that are resistant to ICIs, specifically an increase in arginine synthesis and upregulation of ASS1, the rate-limiting enzyme in this pathway. By using gain and loss of function models, as well as a pharmacological inhibitor specific for ASS1, we demonstrated that modulations in the expression or activity of ASS1 is associated with translational reprogramming, characterized by an inhibition of the cap-dependent mRNA translation mediated through mTORC1/4EBP1 axis. We also demonstrated that targeting ASS1 in vivo, resensitize tumors initially resistant to ICI. Taken together, our results highlight the interaction between modulations of arginine synthesis pathway, mRNA translation reprogramming, antitumor immunity, and restauration of sensitivity to anti-PD-1. Our work also demonstrates the therapeutic potential of targeting arginine synthesis pathway, and especially ASS1, to offer new treatments to patients suffering from cutaneous melanoma resistant to ICIs.

Astrocytes are critical regulators of brain homeostasis and circuit function, and sulforhodamine 101 (SR101) are widely used for astrocyte labeling. However, SR101 also labels a significant portion of oligodendrocytes. Here, we identified Atto 643 carboxy, a far-red fluorescent dye, as a highly specific marker for astrocytes with minimal oligodendrocyte labeling. Furthermore, Atto 643 carboxy demonstrates minimal nonspecific uptake in myelin sheaths, and hippocampal pyramidal neuron as reported for SR101. We demonstrate robust imaging of astrocyte morphology and distribution in vivo and in acute brain slices. Our findings establish Atto 643 carboxy as a highly specific small-molecule probe for astrocyte-selective imaging, offering a powerful tool for dissecting astrocyte structure, dynamics, and function in intact brain tissue.

Angiogenesis, the process by which new blood vessels form from existing vasculature, is fundamental to tissue repair and regeneration but also underlies pathological conditions such as cancer progression. Targeting angiogenesis has thus become a promising approach for developing novel cancer therapeutics. While various phytochemicals have demonstrated anti-angiogenic effects, the role of 2-5(H)-Furanone, a naturally occurring lactone found in various plants and marine sources with diverse biological activities, remains insufficiently explored. In this study, we systematically evaluate the anti-angiogenic potential of 2-5(H)-Furanone using Human Umbilical Vein Endothelial Cells (HUVECs) as an in vitro model and zebrafish embryos as an in vivo model. Experimental findings demonstrated that treatment of HUVECs with increasing concentrations of 2-5(H)-Furanone led to significant, dose-dependent reductions in proliferation, invasion, migration, and tube formation. Analyses of gene expression revealed marked downregulation of key pro-angiogenic mediators, VEGF, and HIF-1. Complementing these in vitro results, in vivo studies in zebrafish embryos showed robust, dose-dependent inhibition of intersegmental vessel (ISV) formation, accompanied by suppression of critical angiogenesis-related genes. Molecular docking further supported these observations by indicating stable binding of 2-5(H)-Furanone to major angiogenic targets, including VEGFR2, MMP2, HIF-1, and PIK3CA. Collectively, our data demonstrate that 2-5(H)-Furanone potently inhibits angiogenesis, as evidenced in both HUVEC and zebrafish models, through functional and molecular mechanisms. These findings support the further development of 2-5(H)-Furanone as a promising anti-angiogenic therapy candidate.

Colony stimulating factor 1 receptor (CSF1R) is a tyrosine kinase receptor that is expressed exclusively in microglia within the CNS. Its endogenous ligands, colony stimulating factor-1 (CSF1) and interleukin-34 (IL-34), are released from neurons, positioning CSF1R as a key mediator receptor of neuron-glia communication. CSF1R is considered not only a potential drug target, but also a biomarker of neuroinflammation. From that perspective, selective radioligands for neuroimaging are of great interest for imaging neuroinflammation and determining drug occupancy. In this study, we have validated the binding characteristics of a CSF1R inhibitor, 4-((5-MethOxy-6-((5-methoxypyridin-2-yl)methoxy)pyridin-3-yl)methyl)-2-(1-methyl-1H-pyrazol-4-yl)pyrimidine (5-MOP) as a novel CSF1R radioligand, by performing in vitro saturation binding experiments in human and murine tissues. 5-MOP was found to be selective for CSF1R among a broad range of kinases. Autoradiography revealed that [3H]5-MOP binds with high affinity (KD = 9.8 nM) to a single saturable binding site in human meningioma tissues, and this binding was displaced with known CSF1R inhibitors, including CPPC, sCSF1inh and GW-2580. In contrast, CPPC, which has been extensively used as a CSF1R radioligand showed substantial cross-reactivity to other brain kinases, including Trk A/B/C, and [3H]CPPC could only be displaced with CPPC itself, not by other ligands, including 5-MOP. These results identify [3H]5-MOP as the most selective radioligand currently available, enabling accurate detection of drug occupancy and activated microglia. Significance of the studyThis study identifies and validates a novel selective radioligand that binds CSF1R with high selectivity and low nanomolar affinity. Because CSF1R is selectively expressed in activated microglia, this radioligand could be useful for detecting neuroinflammatory activity.

Resistance to targeted therapy in HER2-positive breast cancer remains a clinical challenge, especially for patients with relapsed or metastatic disease. Particularly, persistent activation of hypoxia-inducible factor 1 (HIF-1) signalling is well documented in the context of trastuzumab and trastuzumab emtansine resistance. To achieve a deeper understanding of how HIF-1 activity modulates the response to anti-HER2 treatment, we functionally characterized a cellular model of hypoxia-induced drug resistance for HER2-positive breast cancer using shotgun proteomics. By global phosphoproteomics profiling, the Rac1 pathway was identified as one of the most enriched signalling networks under hypoxia. Furthermore, the selective Rac1 blockade with the 1A-116 small-molecule inhibitor sensitised HER2-positive cells to trastuzumab in both 2D and 3D culture systems. Altogether, our findings demonstrate that hypoxic conditions induce the resistance of HER2-positive breast cancer cells to targeted therapy and suggest the therapeutic potential of Rac1 inhibition to enhance trastuzumab efficacy. HighlightsO_LIHypoxic conditions induce trastuzumab resistance in HER2-positive breast cancer. C_LIO_LIRac1 signalling was mapped under hypoxia by phosphoproteomics profiling. C_LIO_LIRac1 inhibition sensitises HER2-positive cells to trastuzumab. C_LI

As potent triggers of innate immunity, STING agonists hold promise as active immunotherapeutic agents for cancer treatment. Second-generation STING agonists, suitable for systemic delivery, are being investigated in preclinical research and have entered clinical trials. Here, the novel synthetic STING agonist, BI-1703880 (STINGa), which was designed for intravenous delivery, was investigated for anti-tumour and immunological effects. We show that STINGa activates the STING pathway and results in a transient and dose-dependent upregulation and secretion of interferons and proinflammatory cytokines in vitro and in vivo. We show that intravenous administration of repeated dosing with low-dose STINGa is well tolerated. We report that radiotherapy (RT) and STING agonism synergizes to generate innate immune cell and CD8+ T cell responses that control tumour growth. Anti-tumour activity induced by combined RT / STINGa was reduced in mice lacking a functional immune system. RT / STINGa combination treatment also initiated development of protective immune memory. RT / STINGa upregulated PD-L1, PD-1 and CTLA-4 in the tumour microenvironment. Our findings show that combining RT / STINGa with immune checkpoint inhibitors further increases therapeutic benefit. Our data confirm STING as a therapeutic target in cancer and support the clinical development of BI-1703880 STING agonist, thereby suggesting radiotherapy as a potential combination for enhancing anti-tumour efficacy.

Epigenetic enzymes, writers, readers and erasers regulate chromatin landscapes and participate in tumor heterogeneity. While therapeutic targeting of these enzymes has shown clinical promise, the comparative efficacy of mono-versus dual-inhibitor strategies remain unclear. Here, we introduce a multi-modal platform that uses NicE-viewSeq and integrates automated deep learning based spatially resolved chromatin accessibility profiling with high-throughput sequencing following epigenetic inhibitor application. Accessible chromatin landscapes were altered along with nucleosome positioning following inhibition of either LSD1 or HDACs alone, or both together. Coordinated modulation of histone marks and the CoREST complex on chromatin was observed across inhibitory conditions. Transcription factor binding analysis identified three predominant families, ETS, RUNT, and bZIP with enhanced chromatin association upon treatments. Mechanistically, a CoREST-RUNX regulatory axis was uncovered wherein JunB, a member of bZIP family displaces CoREST-RUNX at differentially accessible regions, triggering apoptotic pathways. Therefore, JunB-mediated mechanism reveals a convergent therapeutic vulnerability, offering new avenues for optimizing different combinatorial epigenetic therapy in cancer.

Meningeal lymphatic vessels (mLVs) are vital for brain waste clearance, making them a promising therapeutic target. However, effective modulation strategies for mLVs with translational potential remain underdeveloped. Here, we develop a low-intensity focused ultrasound (LIFU) strategy that precisely targets the vault cranial meninges to non-invasively facilitate mLVs drainage. Using models of Alzheimers disease (AD) and aging, we demonstrate that this approach promotes CSF drainage, prevents cognitive decline, and reduces pathological biomarkers. Mechanistically, RNA sequencing combined with calcium imaging in vitro reveals that LIFU activates the Piezo1 ion channel in lymphatic endothelial cells, whereas pharmacological inhibition of Piezo1 abolishes LIFUs therapeutic effects. Compliant with FDA safety guidelines, this LIFU protocol demonstrates strong clinical translatability. If its efficacy is clinically confirmed, LIFU offers a promising therapy for neurodegenerative diseases triggered by waste accumulation.

Pioneering research is adapting chimeric antigen receptors (CARs) from oncology to Alzheimers disease (AD) by targeting amyloid beta (A{beta}). Newer synthetic receptor systems can go beyond, transforming cells into targeted biological drug factories that can couple A{beta} detection to synthesis and secretion of genetically encoded therapeutics. Among candidate systems, T cells Redirected for Universal Cytokine Killing (TRUCK), synthetic Notch (synNotch), and Synthetic Intramembrane Proteolysis Receptors (SNIPR) have shown promise in oncology. Here, we adapt these platforms to AD using a shared A{beta}-targeting binding domain derived from Aducanumab (Aduhelm), coupled to inducible expression cassettes driving identical transgenes: secreted Metridia luciferase (MetLuc) and a Lecanemab (Leqembi)-based chimeric human-mouse antibody (chLecanemab). To validate these systems in vitro, Jurkat clones expressing each receptor were treated with oligomer-enriched A{beta} (A{beta}O) to model AD, and receptor output was quantified by media MetLuc levels and chLecanemab colocalization with A{beta} aggregates. For TRUCK systems, we show the A{beta}-targeting CAR successfully activated Jurkat cells by flow cytometry. We also show that six Nuclear Factor of Activated T-cells (NFAT) tandem repeat response elements (6xNFAT) paired with either minimal interleukin-2, synthetic TATA box, or minimal cytomegalovirus promoters resulted in functional regulatory regions. Despite this, all TRUCK variants failed to significantly upregulate MetLuc in response to A{beta}O. In contrast, both synNotch and SNIPR responded robustly to A{beta}O, with SNIPR outperforming synNotch in both MetLuc and chLecanemab production. These findings establish SNIPR and synNotch as promising platforms for future research on cell-based targeted therapeutic delivery in AD.

Diffuse hemispheric gliomas (DHGs) are highly aggressive and infiltrative CNS tumors that are refringent to treatment, and with a 5-year overall survival of around 20%. A fraction of DHGs is driven by mutations in the histones H3.1 and H3.3. In this study, we demonstrate that the expression of histone H3.3 glycine 34 to arginine mutations (H3.3-G34R) result in the epigenetic and transcriptional activation of the NF-{kappa}B signaling pathway in DHG. To target this vulnerability, we designed high density lipoprotein (HDL) nanoparticles loaded with unmethylated CpG dinucleotides, which mimic the immune stimulatory activity of bacterial DNA. CpG are recognized by Toll-like receptor 9 (TLR9), activating the NF-{kappa}B signaling. The CpG-mediated NF-{kappa}B activation results in the release of immuno-stimulating cytokines that promote an antitumoral response. As we previously established that G34-mutant DHGs are characterized by DNA repair impairment, we combined CpG dinucleotides with a PARP (poly (ADP-ribose) polymerase) inhibitor, olaparib, in the HDL nanoparticles.

Clot resistance to pharmacological thrombolysis remains a critical challenge in ischemic stroke (IS) management. Thrombus heterogeneity, particularly the presence of thrombolysis-resistant domains composed of dense fibrin and non-fibrin components, including neutrophil extracellular traps (NETs), significantly limits the efficacy of recombinant tissue-type plasminogen activator (r-tPA) and its variant, Tenecteplase (TNK). Consequently, novel therapeutic strategies are urgently required. Emerging evidence suggests that co-administration of deoxyribonuclease I (DNase I) with r-tPA can degrade DNA fibers and enhance clot lysis. In this study, we optimized a previously developed theranostic agent--iron oxide microparticles coated with polydopamine--by dual-grafting both r-tPA and DNase to target resistant thrombi. Using functional ultrasound imaging (fUS) during the acute phase of IS, we demonstrated accelerated reperfusion with this dual-functionalized platform in a r-tPA resistant IS model. Furthermore, MRI analysis confirmed a significant reduction in lesion volume at 24 hours, correlating with improved functional recovery five days post-ischemia.

The synaptic vesicle protein SV2C, predominantly found in the basal ganglia, has been associated with Parkinsons disease through genetic studies. It plays a crucial role in regulating dopamine release and has been shown to be disrupted in PD animal models and brain tissues from PD patients. In the context of PD-related synaptopathy, SV2C may serve as a potential imaging target for monitoring disease progression and response to treatment. [18F]UCB-F is a radioligand binding to SV2C developed by UCB. Preliminary autoradiography and PET studies in rats showed that [18F]UCB-F displays a brain distribution consistent with the expression of SV2C in vitro but does not display any specific binding in vivo. This study was therefore designed to further investigate the affinity and selectivity of [18F]UCB-F for SV2C and to examine the in vitro and in vivo properties of the radioligand in non-human primates. In vitro binding studies were performed to measure the affinity of UCB-F to SV2A, SV2B, and SV2C. Insilico modeling was used to assess the binding mode and energy of UCB-F. Autoradiography studies on rat and non-human primate (NHP) brain tissues were performed to confirm that [18F]UCB-F showed similar distribution in rat and NHP tissue. Finally, PET studied in NHPs were performed to examine the in vivo pharmacokinetic properties of [18F]UCB-F. [18F]UCB-F was successfully synthesized from the corresponding precursor with high yield. Autoradiography on brain slices from rats and NHPs demonstrated specific binding of [18F]UCB-F in the pallidum, striatum, substantia nigra, and brainstem, consistent with the known brain expression of SV2C. In NHPs, [18F]UCB-F rapidly crossed the blood-brain barrier, reaching peak uptake values of 2.8 %ID in NHP1 and 2.1 %ID in NHP2 at 4 minutes post-injection. The tracer wasrapidly washed out from the brain, with no clear regional distribution. Radiometabolite analysis revealed the formation of only more polar radiometabolites, with approximately 15% of unchanged radioligand remaining in plasma at 15 minutes post-injection. In vitro and in-silico studies demonstrated that the affinity of [18F]UCB-F decreased by approximately one factor of magnitude with increase of temperature from 4{degrees} to 37{degrees} C. This temperature-related decrease of the affinity for SV2C together with rapid in vivo radiometabolism might explain the discrepancy between in vitro and in vivo performance of [18F]UCB-F. Overall, these findings suggest that [18F]UCB-F is not a suitable PET radioligand for imaging SV2C. Further research is needed to identify alternative candidates with improved in vivo stability and brain retention.