Biology subjects

Frey, N.

Publications and source records attributed to Frey, N..

At least 19 recordsLinked to original sources

bioRxiv2026

Tricuspid valve regurgitation accelerates heart failure via a cardio-intestinal innate immune circuit

Activation of the immune system impacts the progression of heart failure (HF), but the underlying mechanisms remain incompletely understood. Here, we identify a cardio-intestinal innate immune axis that links systemic venous congestion to myocardial inflammation, fibrosis, and functional decline. Using single-cell and single-nucleus transcriptomic profiling in patients and mice with tricuspid regurgitation (TR), we demonstrate that TR disrupts intestinal barrier integrity and elicits expansion of circulating monocytes which in turn orchestrate pathological crosstalk between the right and left heart. Monocyte-derived Interleukin-6 (IL-6) emerged as a key mediator of TR-driven myocardial fibrosis and dysfunction. Blockade of IL-6 attenuated cardiac fibrosis and improved cardiac function. In patients, catheter-based repair of TR resulted in reduced IL-6 levels. Together, these findings establish cardio-intestinal innate immunity as a mechanism linking altered hemodynamics to left ventricular remodeling and nominate TR patients as a selective target population for IL-6-directed therapy in HF. One Sentence SummaryThis work mechanistically resolves the heart-gut axis in tricuspid valve regurgitation, and its impact on heart failure progression as mediated by Interleukin-6.

immunology↗

bioRxiv2026

CD47 Blockade Reprograms the Monocyte-Macrophage Axis to Promote Inflammation Resolution in Atherosclerosis

Atherosclerosis is a chronic inflammatory disease and a leading cause of cardiovascular mortality worldwide. Disease progression is closely linked to defective efferocytosis, the impaired clearance of apoptotic cells, which drives necrotic core expansion and perpetuates arterial inflammation. Targeting the CD47-SIRP innate immune checkpoint, a dominant "dont-eat-me" signal, limits atherosclerosis in preclinical models and retrospective human studies. However, how pro-efferocytic intervention reshapes the immune landscape of established atherosclerotic lesions remains incompletely understood. Here, using single-cell transcriptomics, monocyte fate mapping, and functional analyses across complementary preventive and interventive murine atherosclerosis models, we demonstrate that CD47 blockade fundamentally reprograms the myeloid landscape of established lesions. Anti-CD47 therapy selectively suppresses inflammatory Ly6Chi monocyte recruitment and reduces local macrophage proliferation without altering overall plaque macrophage burden, indicating a qualitative rather than quantitative remodeling of the infiltrate. Concurrently, therapy enriches macrophage subsets bearing pro-efferocytic and macrophage survival-associated transcriptional programs, restoring defective apoptotic cell clearance in situ. Cross-species integration with an independent human coronary artery single-cell dataset identifies a conserved TREM2hi macrophage population that natively harbors the efferocytosis machinery reactivated by therapy in mice. Together, these findings demonstrate that innate immune checkpoint inhibition by CD47 blockade drives a coordinated reprogramming of monocyte-macrophage dynamics, simultaneously suppressing inflammatory influx and enriching efferocytic capacity. This dual mechanism advances our understanding of how pro-efferocytic therapies resolve vascular inflammation in atherosclerosis.

cell biology↗

bioRxiv2026

Cardiomyocyte autophagy promotes a pro-regenerative immune response during cardiac regeneration

Adult zebrafish possess a remarkable ability to regenerate their heart following cardiac injury. Over the past decades, our understanding of the diverse cell types involved in zebrafish cardiac regeneration has greatly advanced. However, the mechanisms governing their interaction and how heterocellular crosstalk drives regeneration remain poorly understood. Here, we identify cardiomyocyte autophagy as a key link between the cardiomyocyte injury response and heterocellular crosstalk between cardiomyocytes and macrophages. We find that cardiomyocyte autophagy is downstream of AP-1 transcription factors. Using newly generated genetic tools, we find that cardiomyocyte autophagy is an important regulator of cardiomyocyte protrusion into the fibrotic injured tissue and its disruption leads to defects in scar resolution. Notably, we find that blocking cardiomyocyte autophagy has a marked effect on the transcriptomic signatures in cardiac macrophages, shifting gene expression from phagocytic/pro-inflammatory/pro-reparative towards pro-angiogenic and pro-fibrotic states. Altogether, our results uncover autophagy as a mechanism linking cardiomyocyte injury responses to macrophage phenotype and coordinated tissue remodeling during heart regeneration.

cell biology↗

bioRxiv2026

FYCO1 improves postischemic cardiac remodeling via enhanced autophagic flux and attenuation of proinflammatory signaling

Acute myocardial infarction (MI) is associated with severe metabolic and oxidative stress that triggers cardiomyocyte death, pro-inflammatory signaling and progressive structural remodeling frequently culminating in heart failure. Although significant advances in reperfusion therapy improved acute survival in patients, therapeutic strategies that directly target intracellular processes in response to injury remain limited. One key response mechanism, autophagy, is rapidly activated to ameliorate ischemic stress. Yet, defective autophagic flux may exacerbate cardiomyocyte injury and maladaptive tissue remodeling. Here we identify FYCO1 as a cardiomyocyte-enriched key regulator of autophagy that enhances autophagic flux and promotes myocardial resilience following ischemic injury. Using cardiomyocyte-specific FYCO1 transgenic mice subjected to permanent coronary ligation, we demonstrate that FYCO1 overexpression limits infarct expansion, reduces cardiomyocyte injury, and preserves cardiac function during remodeling. In vivo RFP-EGFP-LC3 autophagy reporter analyses reveal that FYCO1 promotes a sustained increase of autophagic flux by coordinating autophagosome formation and efficient autolysosomal clearance. Transcriptomic profiling identifies a cardioprotective gene program in FYCO1-Tg animals subjected to MI, with suppression of proinflammatory, proapoptotic and stress-response pathways. Systemic serum cytokine and chemokine profiling as well as transcriptomic analyses of myocardium confirm reduced inflammatory signaling and subsequent reduction in macrophage recruitment into the infarct border zone. Together these findings position FYCO1 as a key regulator of cardiomyocyte autophagy and reveal a previously unrecognized link between autophagy and inflammation in shaping cardiac remodeling following myocardial infarction. FYCO1-mediated autophagy promotes myocardial preservation and functional recovery, highlighting autophagic flux as a promising target for cardioprotective interventions.

molecular biology↗

bioRxiv2026

Cardiomyocytes execute pro- and anti-inflammatory signaling of IFNγ-induced GBP5 by differential regulation of the inflammasome

Infiltration of conventional immune cells has been ascribed as the fundamental drivers of innate immune signaling in the damaged myocardium. However, the emerging intrinsic immunoregulatory potential of cardiomyocytes still remains poorly understood. Interferon gamma (IFN{gamma}) is a pleiotropic cytokine with context-dependent detrimental as well protective role in regulating cardiac inflammatory circuits. The prevailing view of IFN{gamma} as a prime pro-inflammatory cytokine has been challenged due to its paradoxical actions both as an inducer as well as negative regulator of inflammation, but the players involved in these converse processes remains enigmatic. Here we show that cardiomyocytes exhibit a cell-autonomous immunocompetent response upregulating innate inflammatory signaling upon type I and type II IFN stimulus. Notably, hiPSC-derived cardiomyocytes display a robust increase in guanylate binding protein 5 (GBP5), one of the major IFN{gamma}-induced GTPase involved in inflammasome signaling, followed by upregulation of AIM2/CASP1 pathway whereas NLRP3 levels remain unaltered by IFN{gamma} stimulation. GBP5 knockdown and overexpression studies in hiPSC-derived cardiomyocytes identify GBP5/TGF{beta} axis as a non-canonical anti-inflammatory feedback regulation on the IFN{gamma}-induced inflammatory cascade.

cell biology↗

bioRxiv2026

Muscle LIM Protein is a Non-Canonical RNA-Binding Protein that Engages in RNA-Dependent Cytoskeletal Assemblies

Muscle LIM Protein (MLP) is a key component of the cardiomyocyte Z disc that is essential for sarcomere integrity and cardiac homeostasis. Accordingly, loss of MLP function results in (cardio)myopathy phenotypes in animal models and human patients. In this study, we present the first RNA-binding proteome (RBPome) of human cardiomyocytes using enhanced RNA interactome capture (eRIC). Data integration with existing RBPome datasets identified MLP as an evolutionary conserved and previously unrecognized non-canonical RNA-binding protein (RBP) in cardiomyocytes. Using complementary biochemical approaches, we confirmed direct RNA association of MLP. Moreover, RNA integrity was required for MLP interactions with cytoskeletal proteins and for the formation of higher-order MLP-containing assemblies, revealed by co-immunoprecipitation and RNA-dependent sedimentation profiling. Subcellular fractionation and imaging analyses indicated that MLPs association with cytoskeletal complexes may be weaker and more dynamic compared to core sarcomere proteins like -actinin. Domain mapping identified two glycine-rich regions within MLP as RNA contact sites, and RNA-binding-deficient MLP mutants showed impaired association with cytoskeletal complexes. Together, our findings suggest RNA binding as a regulated property of MLP and uncover an RNA-dependent layer of cytoskeletal organization in cardiomyocytes.

molecular biology↗

bioRxiv2026

Transcriptional regulation of the TASK-1 potassium channel by ETV1 -Implications for atrial excitability

BackgroundAtrial fibrillation (AF), the most common sustained arrhythmia, is driven by electrical and structural remodelling, including altered ion channel expression. The atrial-specific potassium channel TASK-1 regulates action potential duration (APD) and is differentially expressed in AF and left ventricular dysfunction, but the mechanisms controlling its expression are not well understood. ObjectiveThis study examines whether the transcription factor ETV1 regulates TASK-1 and contributes to atrial electrical remodelling. MethodsAtrial tissue from patients with and without AF was analysed to assess the relationship between ETV1 and TASK-1 (KCNK3) expression. In HL-1 cardiomyocyte-like cells and native fibroblasts, ETV1 activity was reduced using pharmacological inhibition or siRNA-mediated knockdown. TASK-1 expression, TASK-1 current, and APD at 90% repolarization were measured. Pacing experiments tested activity-dependent TASK-1 regulation. Direct transcriptional regulation was evaluated using ChIP-qPCR and ChIP-seq to detect ETV1 binding at the KCNK3 promoter. ResultsETV1 and TASK-1 levels were positively correlated in human atrial tissue. In HL-1 cells and fibroblasts, ETV1 inhibition or knockdown decreased TASK-1 expression and current and selectively prolonged APD90. Pacing-induced upregulation of TASK-1 was prevented by ETV1 inhibition, indicating a protective effect against pro-arrhythmic remodelling. ChIP-qPCR and ChIP-seq confirmed direct ETV1 binding to the KCNK3 promoter. ConclusionETV1 directly regulates TASK-1 expression and contributes to atrial electrical remodelling, identifying ETV1 as a potential upstream therapeutic target in AF. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=89 SRC="FIGDIR/small/711402v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@3e5b25org.highwire.dtl.DTLVardef@1d1f909org.highwire.dtl.DTLVardef@34cf37org.highwire.dtl.DTLVardef@1105539_HPS_FORMAT_FIGEXP M_FIG C_FIG Translational perspectiveAtrial fibrillation is sustained by maladaptive electrical remodelling that remains insufficiently addressed by current rhythm-control therapies. Direct inhibition of individual ion channels has shown efficacy but is limited by phenotype dependence and proarrhythmic risk. The present data identify ETV1 as an upstream transcriptional regulator of the atrial-specific potassium channel TASK-1. Modulation of ETV1 reduced TASK-1 expression, prolonged atrial repolarisation, and prevented tachycardia-induced electrical remodelling in vitro. Targeting ETV1 may therefore represent a disease-modifying strategy that intervenes earlier in the remodelling cascade than conventional antiarrhythmic drugs. This approach could enable phenotype-guided therapy in atrial cardiomyopathy, particularly in patients with preserved ventricular function, and warrants validation in translational large-animal and clinical studies.

molecular biology↗

bioRxiv2026

HDAC inhibition via suberoylanilide hydroxamic acid (SAHA) ameliorates Doxorubicin-induced cardiotoxicity

BackgroundAnthracycline-induced cardiotoxicity remains a major limitation of cancer therapy, and effective preventive strategies are lacking. Topoisomerase II{beta} (Topo IIb) has been implicated as a central driver of this toxicity, suggesting that epigenetic regulators may interfere with pathological cardiac response. Methods and ResultsHere, we show that doxorubicin promotes Topo IIb accumulation at cardiomyocyte gene promoters (e.g., Actc1, Myl2, and Myh7) overlapping myocyte enhancer factor 2 (MEF2) binding sites and enhances MEF2-dependent transcription. This response is attenuated by the pan-histone deacetylase (HDAC) inhibitor suberoylanilide hydroxamic acid (SAHA). SAHA-mediated cardioprotection requires class IIa HDACs, as genetic loss of HDAC4 abolishes its effect. Mechanistically, SAHA induces acetylation of the chaperone 14-3-3, disrupting its interaction with HDAC4/5, promoting their nuclear accumulation, and repressing MEF2-driven transcription. In vivo, SAHA mitigates doxorubicin-induced cardiotoxicity. ConclusionThese findings identify HDAC inhibition as a cardioprotective repurposing strategy and reveal a mechanistic link between epigenetic regulation and anthracycline-associated cardiotoxicity.

molecular biology↗

bioRxiv2026

Multicellular signaling and partial recovery define reverse cardiac remodeling

Heart failure results from maladaptive multicellular remodeling triggered by sustained biomechanical stress. Although mechanical unloading can promote reverse remodeling, recovery is frequently incomplete and its mechanistic basis remains unclear. Using a reversible murine pressure-overload model combined with bulk and single-nucleus transcriptomics, we demonstrate that reverse remodeling represents an actively maintained yet constrained multicellular state. Unloading improved cardiac function and partially restored extracellular matrix and metabolic programs, whereas inflammatory and mitochondrial dysfunction signatures persisted. Cardiomyocytes and endothelial cells largely re-established homeostatic transcriptional states, while fibroblasts retained activated programs that dominated residual pathology. Multicellular factor analysis delineated a reversible metabolic stress program and a persistent inflammatory-mitochondrial program coordinated across cardiac cell types. Cell-cell communication analysis identified lymphatic endothelial cells as key instructive regulators of recovery. Notably, lymphatic-derived Reelin directly suppressed pathological fetal gene activation in murine and human cardiomyocytes, uncovering a previously unrecognized lymphoangiocrine mechanism that constrains myocardial recovery in chronic heart failure.

molecular biology↗

bioRxiv2025

TRIM24 preserves cardiomyocyte immune quiescence by repressing interferon/STAT signaling

Innate immune activation in cardiomyocytes is a key driver of inflammatory heart disease and heart failure progression, yet the mechanisms by which cardiomyocytes maintain transcriptional immune quiescence remain poorly understood. TRIM24, a multidomain chromatin reader implicated in cancer and inflammation, has not been previously studied in the heart. Here, we investigated whether TRIM24 regulates interferon/STAT-driven inflammatory programs and paracrine signaling in cardiomyocytes. Single-nucleus RNA sequencing revealed a pronounced downregulation of TRIM24 specifically in cardiomyocytes from human ischemic cardiomyopathy, a change not detected by bulk RNA-sequencing in either human or mouse myocardial infarction samples, likely due to masking by non-cardiomyocyte cell populations. Fractionated protein analysis further demonstrated that TRIM24 is enriched in cardiomyocyte nuclei. Functional studies in neonatal rat ventricular cardiomyocytes showed that TRIM24 represses interferon-stimulated and STAT-dependent genes, including Mx1, Irf7, and Ifit3. ChIP-Seq revealed TRIM24 occupancy at STAT-bound regulatory regions, suggesting cooperative suppression of inflammatory gene networks. Mechanistically, TRIM24 decreased Stat1a/b and Stat3 transcription, reduced protein abundance, and inhibited phosphorylation independently of proteasomal degradation; these effects were partially reversible by bromodomain inhibition. Functionally, TRIM24 overexpression dampened paracrine macrophage recruitment, whereas TRIM24 depletion amplified it. Collectively, these findings identify TRIM24 as a cardiomyocyte-enriched chromatin regulator that restrains STAT1/3 signaling and suppresses paracrine inflammatory activation. TRIM24 acts as a transcriptional safeguard of immune quiescence in the heart and represents a potential therapeutic target for limiting cardiac inflammation.

molecular biology↗

bioRxiv2025

ZEB1 Is a Key Regulator of Cardiomyocyte Structure and Function

BackgroundOur previous work identified the transcription factor "Zinc Finger E-Box Binding Homeobox 1" (ZEB1) as a downstream effector of Cytoplasmic Polyadenylation Element Binding Protein 4 (CPEB4), an RNA-binding protein responsive to cardiac stress. While ZEB1 is known for its role in cancer metastasis and epithelial-to-mesenchymal transition (EMT), its function in cardiomyocytes is not well understood. Based on previous findings, we hypothesize that ZEB1 is essential for maintaining the structural integrity and mitochondrial function of cardiomyocytes. MethodsAAV9-Zeb1 was used for the overexpression of Zeb1. Using a myosin heavy chain alpha (MHC) Cre system, we created a Zeb1 conditional knockout mouse. To evaluate cardiac function and structure, we used echocardiography, electron microscopy, immunohistochemistry. We identified differentially expressed genes following Zeb1 deletion using RNA-seq and determined direct Zeb1 target genes by integrating this transcriptome data with a Zeb1 ChIP-seq dataset. ResultsZEB1 deletion leads to sarcomere damage, mitochondrial dysfunction, and dedifferentiation, with more pronounced effects in females. Overexpression promotes hypertrophic remodeling. Echocardiographic analysis showed progressive systolic dysfunction, and histology revealed sarcomeric disarray, again especially in females. A tamoxifen-inducible ZEB1 knockout mouse model only confirmed ZEB1s crucial role in fully differentiated cardiomyocytes in female mice. Integrated analysis of RNA-seq and ChIP-seq revealed that Zeb1 directly regulated mitochondrial genes, thereby playing a critical role cardiomyocyte energy supply and having secondary effects on cardiac structure and function. ConclusionsZEB1 is critical for cardiomyocyte homeostasis, and maintaining its function is necessary for normal cardiac performance and structure.

molecular biology↗

bioRxiv2025

Deep Evolutionary Fitness Inference for Variant Nomination from Directed Evolution

Iterative screening techniques, such as directed evolution, enable high-throughput affinity maturation to optimize binders to molecular interfaces. However, the decision problem of selecting variants from rich, evolved populations to enter low-throughput follow-up methods remains a significant bottleneck. Here, we present evolutionary fitness inference (EVFI) and DeepEVFI, two machine learning methods that model directed evolution from time-series sequencing data, and infer fitness, a variants ability to enrich under selection pressure. Our methods flexibly handle mutation mechanisms and starting populations that may be partially unknown - settings relevant to drug discovery - and achieve strong performance on a diverse set of experimental data. We conducted two experimental directed evolution campaigns, using antibodies and macrocyclic peptides libraries to identify and optimize binders to therapeutically relevant targets. EVFI and DeepEVFI identified tighter binders that were missed by human experts using conventional frequency-based approaches, including "rising stars" with low frequency. Beyond initial hit discovery, EVFI and Deep-EVFI enables labeling large-scale sequence-fitness datasets and identifying variants of initial binders with diverse properties.

bioinformatics↗

bioRxiv2025

Sarcomeric Remodelling in Human Heart Failure unraveled by single molecule long read sequencing

Dysregulation of alternative splicing - mediated by factors such as RBM20 or SLM2 - can affect proper gene isoform control disrupts gene isoform homeostasis and underpins severe cardiomyopathy in both animal models and patients. Although innovative therapies target various sarcomeric components, the impact of isoform switching in cardiac disease remains poorly understood. Here, we applied nanopore long-read sequencing to map the full-length transcriptome of left ventricular tissue from thirteen nonfailing controls, ten patients with dilated cardiomyopathy (DCM), and ten with ischemic cardiomyopathy (ICM). Our analysis identified 78,520 transcripts, 31% of which represent novel isoforms of known genes. Notably, the transcriptomes of DCM and ICM were largely indistinguishable, indicating that end-stage heart failure is characterized by a convergent isoform landscape, irrespective of disease etiology. Among 11 prototypical sarcomere genes, 10 displayed highly significant isoform shifts (p= 5.23x10-45 - 2.89x10-200). Focusing on tropomyosin, we observed that while the predominant cardiac gene TPM1 showed moderate up-regulation of its transcript isoforms, transcripts derived from TPM3--typically expressed at lower levels in the healthy heart--were markedly increased in heart failure.

genetics↗

bioRxiv2025

A cross-species multimodal database of gene expression in cardiac hypertrophy

Changes in gene expression determine the pathophysiological response of the heart during increased workload and biomechanical stress.1,2 Understanding the complex gene expression regulation during physiological and pathological stress provides insight into mechanisms that mediate the transition from cardiac hypertrophy to heart failure. We curated and generated a multi-omics database of cardiac hypertrophy and heart failure and exemplified its usage by evaluating the regulation of UCK2, a gene that belongs to a conserved cardiomyocyte hypertrophy gene expression signature with unknown function in heart disease. To provide the community with simple access to the database, we developed CHEERIO (Cardiac HypErtrophy gEne expRessIOn Database), a free and easy-to-use interactive web application to explore rodent and human gene expression during early, late, physiological and pathological cardiac hypertrophy, as well as heart failure at transcriptome, translatome, proteome, bulk and single-cell resolution. CHEERIO is freely available at https://voelkerslab.shinyapps.io/CHEERIO/.

physiology↗

bioRxiv2024

Generating All-Atom Protein Structure from Sequence-Only Training Data

AbstractO_ST_ABSPurposeC_ST_ABSDesigning proteins with atomic-level functional control remains a central challenge in de novo design, exacerbated by limitations in structural data availability. MethodsWe introduce PLAID (Protein Latent Induced Diffusion), a generative model that efficiently co-generates discrete sequence and all-atom structure by sampling directly in the shared sequence-structure latent of a pretrained sequence-to-structure predictor. Unlike existing de novo generative models, PLAID trains its diffusion model exclusively on sequences, expanding effective training corpora by 2-4 orders of magnitude relative to structural databases. Using classifier-free guidance, PLAID supports controllable generation based on function (Gene Ontology) and organism keywords. ResultsIn silico, PLAID can unconditionally generate all-atom structures without explicit structural supervision during diffusion model training. Function conditioned proteins can recapitulates catalytic side-chain positions for residues at non-adjacent positions, and transmembrane proteins with expected hydrophobicity patterns and predicted topologies. We further experimentally validate that PLAID can be prompted to generate heme binding proteins with high sequence novelty. ConclusionOverall, PLAID unifies sequence-scale training with atomic-level generation, enabling more precise functional control in protein design. Code and weights: github.com/amyxlu/plaid.

bioinformatics↗

bioRxiv2024

Tokenized and Continuous Embedding Compressions of Protein Sequence and Structure

Existing protein machine learning representations typically model either the sequence or structure distribution, with the other modality implicit. The latent space of sequence-to-structure prediction models such as ESMFold represents the joint distribution of sequence and structure; however, we find these embeddings to exhibit massive activations, whereby some channels have values 3000x higher than others, regardless of the input. Further, on continuous compression schemes, ESMFold embeddings can be reduced by a factor of 128x along the channel and 8x along the length, while retaining structure information at <2[A] scale accuracy, and performing competitively on protein function and localization benchmarks. On discrete compression schemes, we construct a tokenized all-atom structure vocabulary that retains high reconstruction accuracy, thus introducing a tokenized representation of all-atom structure that can be obtained from sequence alone. We term this series of embeddings as CHEAP (Compressed Hourglass Embedding Adaptations of Proteins) embeddings, obtained via the HPCT (Hourglass Protein Compression Transformer) architecture. CHEAP is a compact representation of both protein structure and sequence, sheds light on information content asymmetries between sequence and structure, democratizes representations captured by large models, and is designed to have flexible downstream applications such as generation, search, and prediction.

bioinformatics↗

bioRxiv2024

Spatial omics of acute myocardial infarction reveals a novel mode of immune cell infiltration

Myocardial infarction (MI) continues to be a leading cause of death worldwide. Even though it is well-established that the complex interplay between different cell types determines the overall healing response after MI, the precise changes in the tissue architecture are still poorly understood. Here we generated an integrative cellular map of the acute phase after murine MI using a combination of imaging-based transcriptomics (Molecular Cartography) and antibody-based highly multiplexed imaging (Sequential Immunofluorescence), which enabled us to evaluate cell-type compositions and changes at subcellular resolution over time. One striking finding of these analyses was the identification of a novel mode of leukocyte accumulation to the infarcted heart via the endocardium - the inner layer of the heart. To investigate the underlying mechanisms driving this previously unknown infiltration route, we performed unbiased spatial proteomic analysis using Deep Visual Proteomics (DVP). When comparing endocardial cells of homeostatic hearts and infarcted hearts, DVP identified von Willebrand Factor (vWF) as an upregulated mediator of inflammation 24 hours post-MI. To further explore the immune mediating capabilities of vWF and its effect on tissue repair, we performed functional blocking of vWF during acute murine MI. This resulted in a reduced amount of infiltration by CCR2+ monocytes and worse cardiac function post-MI. Our study provides the first spatial map of acute murine MI with subcellular resolution and subsequently discovers a novel route of immune infiltration. Furthermore, we identified vWF as a critical immune mediating agent for endocardial immune cell infiltration.

systems biology↗

bioRxiv2024

Directed evolution-based discovery of ligands for in vivo restimulation of CAR-T cells

Chimeric antigen receptor (CAR) T cell therapy targeting CD19 elicits remarkable clinical efficacy in B-cell malignancies, but many patients relapse due to failed expansion and/or progressive loss of CAR-T cells. We recently reported a strategy to potently restimulate CAR-T cells in vivo, enhancing their functionality by administration of a vaccine-like stimulus comprised of surrogate peptide ligands for a CAR linked to a lymph node-targeting amphiphilic PEG-lipid (termed CAR-T-vax). Here, we demonstrate a general strategy to generate and optimize peptide mimotopes enabling CAR-T-vax generation for any CAR. Using the clinical CD19 CAR (FMC63) as a test case, we employed yeast surface display to identify peptide binders to soluble IgG versions of FMC63, which were subsequently affinity matured by directed evolution. CAR-T vaccines using these optimized mimotopes triggered marked expansion and memory development of CD19 CAR-T cells in both syngeneic and humanized mouse models of B-ALL/Lymphoma, and enhanced control of disease progression. This approach thus enables vaccine boosting to be applied to any clinically relevant CAR-T cell product.

bioengineering↗