Biology subjects

Qin, X.

Publications and source records attributed to Qin, X..

At least 19 recordsLinked to original sources

bioRxiv2026

DYNAMIC CONFORMATIONS IN THE ACTIVATION OF PINK1 KINASE

Under mitochondrial stress, the mitochondrial kinase PINK1 is activated to phosphorylate ubiquitin, which in turn recruits the E3 ligase Parkin, thereby initiating clearance of damaged mitochondria. Dysregulation of the PINK1-Parkin signaling pathway is associated with early-onset autosomal recessive Parkinson's disease. However, the mechanisms governing PINK1 conformational dynamics and activation in response to mitochondrial stress remain poorly understood. Here, we report that Tribolium castaneum PINK1 (TcPINK1) forms specific symmetric dimers via interactions between the kinase N-lobe and the regulatory C-terminal domain. These dimers undergo dynamic conformational transitions across distinct states, including an autoinhibited state, an inactive intermediate state, a primed-activation state and an active state. Moreover, TcPINK1 undergoes lateral trans-phosphorylation through dimer-dimer interactions. Notably, symmetric dimer formation is essential for Parkin recruitment during mitophagy. These findings provide a framework for understanding the conformational plasticity and allosteric regulation of PINK1 in mitophagy.

biophysics↗

bioRxiv2026

MOLECULAR BASIS FOR PINK1 MATURATION

Phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (PINK1), a key regulator of mitophagy, has been linked to the pathogenesis of Parkinson's disease (PD). PINK1 recruits Parkin, an E3 ubiquitin ligase, triggering mitophagy in response to mitochondrial damage. During mitophagy, the quantity, stability, and activity of PINK1 must be strictly regulated; however, the mechanisms governing these parameters under cellular stress are still unclear. Herein, we determined the structural basis for PINK1 maturation mediated by heat shock protein 90/cell division cycle 37/FK506-binding protein 51 (HSP90/CDC37/FKBP51) chaperone complex. We identified PINK1-associated proteins using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and determined the structures of the complexes using Cryo-Electron Microscopy (Cryo-EM). Results showed that FKBP51 potentially interacts with a conserved leucine-proline-phenylalanine (LPF) motif on the activation loop of PINK1 and negatively regulates PINK1 functions in mitophagy. A PINK1 mutation located at the FKBP51 recognition site is linked to mitophagy deficiency, which can be partially rescued by specific inhibition of FKBP51. These findings reveal a general mechanism for PINK1 recognition by the HSP90/CDC37/FKBP51 chaperone complex and suggest a potential approach for upregulating PINK1 activity, which is impaired in PD.

biophysics↗

bioRxiv2026

Pharmacological up-regulation of CALM1 restore endothelial cells in intracranial aneurysm pathogenesis

Subarachnoid hemorrhage (SAH) resulted from intracranial aneurysm (IA) rupture is an especially severe form of stroke. Endothelial dysfunction represents the initiating event of IA pathogenesis. Understanding the role of endothelial cells (ECs) underlying formation of IAs is helpful to seek for pharmaceutical treatment strategy. Based on single-cell RNA sequencing, proteomics, and metabolic analysis, we discovered a change in cell population in IA samples, majorly in ECs and macrophages (MPs). Abnormal ECs exhibit senescence and death in IA samples, which is absent in the control arterial samples. Cross-analysis of multi-omics revealed that CALM1, a calcium detector involved in mechanotransduction, is downregulated in the abnormal ECs. CALM1 knockdown leads to senescence and inhibits the proliferation and maturation of ECs under turbulent flow. Through high-throughput virtual screening, this work identified compound ZC04329651 as a potent CALM1 activator in maintaining the stability of endothelial cell junctions and attenuating cellular senescence. Thus, our findings showed compound ZC04329651 up-regulate the expression of CALM1 to restore ECs, which maybe a promising pharmacological treatment strategy for IAs.

cell biology↗

bioRxiv2026

Transfer learning reveals species-specific olfactory preferences in Diptera and informs pest management strategies

The interaction between volatile organic compounds (VOCs) and odorant receptors (ORs), termed VOI, constitutes the molecular basis for species to recognize chemical information in the environment. Characterization of subtle differences in VOI relationships among closely related species is crucial for understanding the rapid evolution of ORs and for identifying species-specific chemical cues relevant to pest management. However, progress in this field has been constrained by both the scarcity of experimental data and limitations in computational prediction accuracy. To address this, we developed a virtual screening-enhanced transfer learning strategy that integrates large-scale molecular docking data with sparse functional experimental data. The resulting VOI prediction model was validated through functional experiments on the pest species Bactrocera dorsalis, demonstrating its cross-species predictive capacity. Using this model, we investigated the relationship between ecological niches and olfactory sensitivity in Diptera insects from multiple perspectives, with an emphasis on patterns that may inform pest behavioral research. The models predictive reliability was further supported by its consistency with known olfactory trends: it recapitulated the preferential responses of fruit flies and mosquitoes to esters and aromatics, respectively, and reproduced the previously reported negative correlation between olfactory and visual capacities in Drosophila. As a proof-of-concept application, we compared hematophagous and non-hematophagous mosquitoes, revealing overlapping chemical spaces but distinct patterns in their specifically recognized compounds. This study provides a reliable VOI prediction framework that reveals species-level olfactory preference patterns in Diptera, offering a computational pipeline to accelerate the discovery of behaviorally active compounds for species-specific pest monitoring and control.

bioinformatics↗

bioRxiv2026

Multi-Omics Characterization of Human Molecular Responses to Spaceflight Across Two Independent Missions

Human spaceflight has historically been led by government agencies, but the emergence of commercial organizations is enabling broader participation and new research opportunities. In this study, we present a comprehensive molecular characterization of early human responses to spaceflight, leveraging multi-omics data across the first three weeks of two commercial missions. Biospecimens from six individuals, four from the Axiom 2 mission (10 days) and two from Axiom 3 (21 days), were analyzed using single-cell and bulk RNA sequencing, alongside proteomic profiling. Individual and integrative analyses of these datasets reveal systemic changes in cell types, transcripts, and proteins related to immune regulation, osteoclast differentiation, NF-{kappa}B signaling, and blood homeostasis pathways. Importantly, several of the detected pathways align with physiological patterns observed in longer-duration missions. This work establishes a foundational resource for understanding early adaptation to spaceflight at the cellular and molecular levels, providing insights to reduce future space-travel health risks. HighlightsO_LIFirst integrated multi-omics analysis using single-cell, bulk RNA sequencing and proteomic profiling of early human spaceflight responses across two commercial missions (Ax-2 and Ax-3). C_LIO_LIDistinct PBMC clustering was observed across Ax-2 and Ax-3, and post-flight samples. It showed fewer monocytes, dendritic cells, and megakaryocytes with increased naive CD4 and cytotoxic T cells. C_LIO_LIMulti-omics integration identifies shared biological signatures, including osteoclast differentiation, metabolic stress, and coagulation changes. C_LIO_LIThese findings lay the foundation for developing countermeasures to protect immune, skeletal, and vascular health during spaceflight. C_LI

molecular biology↗

bioRxiv2026

IRF7 deficiency increases disease severity independently of TLR7 recognition in Influenza A infection in mice

Influenza A virus (IAV) remains a major cause of respiratory morbidity and mortality, yet the role of Toll-like receptor 7 (TLR7), an RNA sensor, and its downstream signaling events, such as interferon regulatory factor 7 (IRF7), in IAV infection remain unclear. To address this question, we used single-cell RNA sequencing, genetic mouse models, and immunological analysis. Single -cell transcriptomic profiling of the infected lungs revealed robust upregulation of Tlr7 and genes associated with interferon pathways in dendritic cells and B cells, alongside widespread induction of Irf7 across immune and non-immune compartments. Tlr7-deficient mice exhibited normal viral control, lung pathology, and survival following IAV challenge. In contrast, Irf7 deficiency resulted in significantly increased disease severity, impaired early interferon responses, exacerbated bronchial epithelial hyperplasia, and defective early humoral priming. In assessing adaptive immunity, both Irf7-deficient andTlr7-deficient mice had reduced antihemagglutinin antibody production. Mechanistically, IRF7 protein expression and downstream signaling were largely preserved in TLR7-deficient mice, indicating that IRF7 activation during IAV infection occurs independently of TLR7. Collectively, these findings identify IRF7 as a non-redundant determinant of innate immunity and disease outcomes during IAV infection, while positioning TLR7 as a modulatory factor primarily influencing adaptive immune maturation. Our study refines current models of antiviral sensing by uncoupling receptor induction from functional necessity and highlights IRF7 as a critical downstream regulator dictating host defense against acute influenza A infection. ImportanceInfluenza A virus is a respiratory pathogen that remains a major threat to global health as a seasonal disease and a source of periodic pandemics. The outcomes of the infection can range from mild illness to severe pneumonia and death, particularly in vulnerable populations, yet the reasons why some individuals develop more severe disease are not fully understood. Early immune defenses in the lungs are critical for controlling the virus, but they can also contribute to harmful inflammation if not properly regulated. In particular, key sensors that detect viral genetic material and the signaling pathways that activate antiviral responses play an essential role in shaping these outcomes. The significance of our study lies in defining how these early immune mechanisms influence the course of influenza A infection, providing insight that may guide the development of improved therapies for influenza and related respiratory viruses.

immunology↗

bioRxiv2026

The Vagus Nerve conducts viable translocation of gut flora to the lungs that impacts interstitial lung disease severity in mice

Communication between gut microbiota and extraintestinal organs is increasingly recognized, yet elucidation of relevant translocation mechanism(s) remains enigmatic. Vagus neuroanatomy and reports of vagal protein transfer to extraintestinal organs suggest that this "superhighway" could translocate bacteria. Here we explore whether the vagus superhighway can translocate bacteria to extraintestinal organs. Gavage of green fluorescent protein-expressing Escherichia coli (GFP-E. coli) into germ-free (GF) or specific-pathogen free (SPF) C57BL/6 mice yielded high bacillary loads in the stomach and lungs, followed by the heart, stool and peripheral muscles, despite negative blood cultures. Notably, confocal microscopy and culture revealed GFP-E. coli within the vagus nerve within five minutes of gavage suggesting rapid translocation. Metagenomic analysis of stool, lung, heart, vagus nerve, and muscle from non-gavaged SPF mice demonstrated significant microbial overlap, supporting that bacterial translocation occurs despite the presence of endogenous microflora. Remarkably, subdiaphragmatic vagotomy performed prior to GFP-E. coli gavage resulted in marked reductions of bacterial transduction in the lungs and other extraintestinal organs, except muscle. Furthermore, vagotomy significantly reduced lung fibrosis in SPF mice following intranasal bleomycin administration. In lung cancer patients undergoing lobectomy, vagotomy inhibited postsurgical reductions in forced vital capacity. These findings identify the vagus nerve as a literal gut-lung axis, facilitating viable bacterial translocation and influencing lung severity.

microbiology↗

bioRxiv2026

Postnatal Reprogramming Shapes Human Intestinal Epithelial Immune Competency

At birth, the intestine must rapidly adapt to enable nutritional function and immune microbial tolerance. Here, integrating single-cell multi-omics and spatial transcriptomics we define the circuits underpinning this process. We identify asynchronous developmental trajectories with postnatal epithelial reprogramming characterised by coordinated changes in metabolism, junctional structure and innate defence. At birth epithelial stem cells demonstrate dynamic enhancer remodelling, with accessibility often preceding transcription. Fetal stemness elements remain accessible despite reduced transcription across epithelial lineages, retaining plasticity potential. Post-natal epithelia experience sequential homing of myeloid cells followed by innate T cells with peri-epithelial B cells localising later in infancy. Using developmentally staged organoids, we show that epithelial responses to inflammatory stimuli are age-dependent and constrained in early life. We identify BHLHE40 as an early-life regulator that attenuates the impact of interferon- and NF-{kappa}B-driven signalling. Altogether we define the events driving epithelial licensing and barrier adaptation at birth and through infancy.

cell biology↗

bioRxiv2026

Comprehensive bioinformatic analysis reveals novel potential diagnostic biomarkers associated with monocytes in osteoporosis

Osteoporosis affects millions of women globally. In this study, we applied bioinformatics methods to screen for novel diagnostic biomarkers of osteoporosis in women using the GSE62402 and GSE56814 datasets. PCSK5, ZNF225, and H1FX were used to construct a diagnostic model. ROC, calibration, and decision curve analyses were performed to assess the diagnostic performance on the training (GSE56814) and external (GSE56815) datasets. The expression level of model genes was validated in GEO datasets. Furthermore, five transcription factors (ETS1, NOTCH1, MAZ, ERG, and FLI1) were identified as common upstream regulators of model genes. PCSK5, ZNF225, and H1FX serve as novel diagnostic biomarkers, providing new insights into the pathogenesis of and treatment strategies for osteoporosis in women.

genetics↗

bioRxiv2026

Rapid Podocyte ablation Causes Acute Renal Tubule Cell Necrosis and Interstitial Fibrosis

It remains unclear whether podocyte loss directly causes acute renal tubular cell (RTC) damage and interstitial fibrosis, thereby leading to renal failure. Here, we applied intermedilysin (ILY)-mediated human CD59 (hCD59) cell ablation to generate an acute, specific podocyte-ablation mouse model. Cre-induced hCD59 transgenics (ihCD59) were crossed with Nphs2Cre to generate ihCD59+/-/Nphs2Cre+/- mice. The specific and rapid podocyte-ablation mediated by ILY injection directly caused RTC necrosis, leading to renal failure and even death within 2-3 days in a dose-dependent manner. Treating mice that received an ILY lethal dose with peritoneal dialysis or administering a non-lethal dose, we extended their survival beyond six weeks and found that mice developed interstitial fibrosis and glomerulosclerosis with persistent proteinuria and tubule damage. Podocyte-ablation caused massive disruption of glomerular function at week 1, and then partial recovery by week 2. Genes and pathways of TLRs and apoptosis, and mitochondrial functions were respectively upregulated and downregulated in both ablated-podocyte mouse and biopsied-glomerulonephritis patient kidney samples. Together, this rapid podocyte-ablation causes acute RTC necrosis that progresses to interstitial fibrosis in this mouse model, which is applicable for dissecting mechanisms underlying podocyte injury-mediated tubular damage and glomerular repair, with the potential to reveal novel therapeutic targets for kidney diseases.

pathology↗

bioRxiv2026

Early demyelination by off-target complement injury in a mouse model of neuromyelitis optica

Neuromyelitis optica spectrum disorder (NMOSD) is an autoimmune CNS disease characterized by serum antibodies targeting astrocytes for complement-mediated lysis. NMOSD lesions show not only astrocyte loss but also early demyelination and oligodendrocyte injury - histological hallmarks that converge with those of other primary demyelinating conditions. How pathology spreads to cause demyelination, particularly in early lesions, remains unclear. Using spinal cord imaging in an acute mouse NMOSD model, we directly observe the spread of pathology from astrocytes to oligodendrocytes in evolving lesions. This spread is characterized by initial calcium dyshomeostasis in oligodendrocytes followed by delayed, non-lytic cell death. Oligodendrocyte death is driven not by astrocyte loss per se but by spill-over of soluble complement proteins, as oligodendrocytes can be cell-autonomously preserved by the cell-type-specific expression of the complement inhibitor CD59. Our findings explain the convergence of glial pathology in antibody-mediated CNS autoimmunity and point towards new approaches to prevent secondary glial injury.

neuroscience↗

bioRxiv2026

Quantitative dissection of the metastatic cascade at single colony resolution

Metastasis is the leading cause of cancer-related deaths. However, the core determinants and mechanistic principles underlying the metastatic cascade remain elusive. Small cell lung cancer (SCLC) is a highly aggressive malignancy with exceptional metastatic potential and limited therapeutic options. Here, we present Metastasis Originated Barcode Sequencing (MOBA-seq), a high-throughput in vivo platform that systematically maps genetic regulators across the metastatic cascade at single-colony resolution. MOBA-seq integrates scalable barcode-based lineage tracing with a computational pipeline that quantitatively deconvolutes genotype-specific effects on metastatic seeding, dormancy, and clonal expansion across hundreds of thousands of metastatic events. Applying this approach to more than 400 candidate regulators of SCLC, we uncovered tissue-specific metastatic suppressors and universal metastatic essential genes. We identified metastatic seeding as the predominant determinant of metastasis. Comparative analysis across recipient mice of distinct genetic backgrounds further revealed that innate immune surveillance constrains metastatic progression by reducing metastatic seeding and enforcing dormancy, with additional modulation by sex and tissue context. We validated the frequently mutated gene CREBBP as a key metastasis suppressor whose loss enhances SCLC metastasis through both tumor-intrinsic and immune-modulatory mechanisms. This work establishes a scalable and quantitative platform for mapping the metastatic fitness landscape at single-colony resolution across hundreds of thousands of in vivo data points. Our approach offers a broadly applicable framework for dissecting the interactions between cancer-intrinsic and microenvironmental factors governing tumor initiation, progression, and therapeutic response.

cancer biology↗

bioRxiv2026

Molecular architecture of the tumor microenvironment caused by BRCA1 and BRCA2 somatic mutations in lung adenocarcinoma

ObjectivesHomologous recombination repair (HRR) deficiency is associated with improved immunotherapy responses in non-small cell lung cancer (NSCLC) patients. The HRR genes BRCA1/2 are key regulators of DNA repair, yet their impact on the tumor microenvironment (TME) in lung adenocarcinoma (LUAD) remains unclear. MethodsUsing single-cell sequencing and multi-omics data, we characterized BRCA1/2 mutation-associated transcriptional programs, immune cell composition, and functional alterations in T cells, investigating the molecular and immune architecture of BRCA-mutant LUAD patients. ResultsBRCA1/2 mutations were associated with increased genomic instability and poor prognosis in LUAD patients, but predicted better clinical outcomes following immune checkpoint blockade (ICB) treatment. BRCA1 mutations correlated with an upregulated type I IFN/IFN-{gamma} signature and CD8+ T cell activation. BRCA2 mutations were associated with alveolar/stress/inflammatory responses and enhanced MHC-II antigen presentation, linked to CD4+ T cell differentiation. Both alterations coincided with reduced CD28 co-stimulation and CTL activity, hinting of immune evasion. We identified two tissue-resident memory T cell (Trm) subsets as predictors of clinical outcomes and ICB response. BRCA1 mutations were associated with CD8+ Trm expansion, whereas BRCA2 mutations linked to tumor CD4+ Trm expansion and peripheral T/NK cell cytotoxicity. Furthermore, a cancer-promoting program activated by BRCA1 mutation was vulnerable to histone deacetylase inhibitors, which inhibited LUAD tumor growth. ConclusionsThis study provides a preliminary characterization of the BRCA-mutant TME in LUAD patients, revealing distinct transcriptional and immune patterns that highlight differences in BRCA1/2-associated molecular architecture and offer a framework for improving therapy efficacy in LUAD.

cancer biology↗

bioRxiv2026

Extracellular vesicles facilitate the horizontal transfer of drug resistance and stem-like properties between ovarian tumor cells

Ovarian cancer stem cells (CSCs) can seed recurrent drug-resistant disease. Likewise, non-CSCs can acquire CSC phenotypic properties. How this process is orchestrated is of interest to inform how it might be prevented. We tested the hypothesis that ovarian CSC and/or drug-resistant tumor cells confer stem-like properties via extracellular vesicles (EVs). We focused our investigation on how EVs might mediate EZH2 signaling to promote a phenotypic change in drug-sensitive, non-CSCs. To accomplish this, we utilized paired PARP inhibitor-sensitive and - resistant ovarian cancer (OvCa) cell lines, EZH2 knockdown lines, and patient-derived organoids (PDOs) originating from recurrent high-grade serous OvCa. Small EVs isolated from drug-sensitive, CSC and/or drug-resistant enriched cultures, PARP inhibitor (olaparib) resistant lines, or drug-treated (olaparib or carboplatin) lines were cultured with treatment naive or sensitive lines for defined time points. The impact of small EV exposure was determined by assessing cell number, metabolic activity, viability, sphere and colony-forming capacity, ALDH activity, DNA damage, and changes in associated signaling pathways. We found that EVs from CSC or drug-resistant enriched cell fractions communicate CSC-like phenotypes to the more sensitive tumor cells via EZH2 canonical and non-canonical signaling pathways, promoting stemness. We conclude that EV-mediated activation of EZH2 signaling represents a targetable mechanism contributing to stemness-associated drug resistance in OvCa. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=160 SRC="FIGDIR/small/699925v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@6b4ffborg.highwire.dtl.DTLVardef@1501931org.highwire.dtl.DTLVardef@1a5f31aorg.highwire.dtl.DTLVardef@1fb475f_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2026

High-Affinity Protein Binder Design via Flow Matching and In Silico Maturation

The de novo design of high-affinity protein binders remains a central challenge in protein engineering and therapeutic discovery. While deep generative models have advanced backbone generation and interface design, achieving picomolar or nanomolar binding affinities typically demands extensive experimental screening or iterative in vitro maturation. A general computational approach for the direct production of high-affinity binders has remained elusive. Here, we introduce an integrated framework that synergizes PPIFlow, a flow-matching-based generative model, with a novel in silico maturation strategy. PPIFlow employs a pairformer architecture to explicitly reason over pair-wise geometric and chemical interactions, modeling protein backbone rigid-body transformations as continuous flows. To bridge the affinity gap, we implement a dedicated in silico affinity maturation stage that combines interface rotamer enrichment with partial flow refinement to optimize energetic packing. This pipeline is further accelerated by AF3Score, a score-only adaptation of AlphaFold3 that enables high-fidelity and computationally efficient candidate prioritization. Across a diverse set of therapeutic targets, this synergistic approach consistently produces picomolar and nanomolar affinity binders without experimental affinity maturation. Notably, the framework proves highly effective for the de novo design of single-domain antibodies (VHHs), producing sub-nanomolar binders across multiple targets. These results establish that coupling robust backbone generation with focused in silico maturation renders the purely computational design of high-affinity binders feasible.

biochemistry↗

bioRxiv2025

MECP2 Duplication Uncouples Mitochondrial and Purine Metabolism During neuronal maturation

Mitochondria and nucleotide metabolism are critical for cellular and developmental homeostasis, yet their potential interdependence and role in neurodevelopmental disease remain unclear. In MECP2 Duplication Syndrome (MDS), we identify a conserved correlation between mitochondrial function and purine metabolism that is disrupted across human, organoid, and mouse models. Multiomics integration reveals Complex III as the focal point of mitochondrial collapse, leading to redox stress, DNA damage, and hyperactivation of the de novo purine biosynthesis via purinosome assembly. The breakdown of mitochondria-purinosome coupling compromises genome stability, impairs radial glia proliferation, and delays neuronal maturation. By linking a defined genetic dosage imbalance to metabolic network failure, our study positions the mitochondria-purinosome coordination as a fundamental control axis for neurodevelopment and a therapeutic entry point across metabolic and neurodevelopmental disorders. Metabolic control is fundamental to cellular function, influencing energy production, signaling, epigenetic regulation, and tissue homeostasis1. Nowhere is this more critical than in the brain, where tightly regulated metabolic networks sustain high energetic demands and support neuronal development, synaptic plasticity, and circuit formation2. Disruptions in these networks are increasingly implicated across a spectrum of neurodevelopmental disorders3-5, yet their precise metabolic signatures and mechanistic contributions remain poorly understood.

neuroscience↗

bioRxiv2025

Developing High-Yield and Safe Therapeutic EVs by Ablating Tissue Factor-Mediated Toxicity

The clinical translation of extracellular vesicles (EVs) as next-generation drug delivery vehicles is currently stalled by two interrelated challenges: the difficulty of manufacturing EVs at an industrial scale ("the yield problem") and the risks of toxicity associated with systemic administration of high doses ("the safety ceiling"). In this comprehensive study, we present a two-pronged genetic engineering strategy to overcome these limitations. First, we discovered that overexpression of glycosylphosphatidylinositol (GPI)-anchored proteins, such as CD55 and CD59, significantly enhances EV production in HEK293T and Expi293F cells. By engineering a truncated CD55 variant (TR3) that retains the GPI-anchor but lacks functional domains, we developed the ExoBoost cell line, which boosts EVs yield by approximately 50-fold without altering vesicle morphology. Second, we observed acute toxicity in mice following intravenous administration of high-dose ExoBoost EVs, with symptoms resembling venous thromboembolism. Through comparative analysis of EVs derived from other cell types, including keratinocyte stem cells (KSCs) and mesenchymal stem cells (MSCs), we identified Tissue Factor (TF/F3) as the critical causative agent of this lethal response. Moreover, we completely abolished this toxicity by generating a F3-knockout cell line (ExoBoost2.0) using CRISPR/Cas9, thereby raising the safe dose to as high as 1.5E12 particles per mouse, as quantified by nano flow-cytometry (nFCM). Finally, using a novel CD46-nanoluciferase reporter system, we demonstrated that high-dose administration of these safe EVs saturates hepatic and splenic clearance pathways, leading to a dramatic increase of systemic EVs accumulation, particularly in hard-to-target tissues such as the brain ([~]10,000-fold) and muscle ([~]1,000-fold). Additionally, repeated high-dose administration of ExoBoost2.0 EVs did not upregulate inflammatory cytokines or production of IgG and IgM. In summary, these findings establish a scalable, safe, and highly biocompatible EVs platform, potentially revolutionizing the drug delivery system in clinical application.

bioengineering↗

bioRxiv2025

BCL-XL Dependence is a Subtype Agnostic Actionable Feature of Difficult-to-Treat Kidney Cancers

The BCL-XL anti-apoptotic protein is a clear cell Renal Cell Carcinoma (ccRCC) dependency; however, the mechanism of this dependence and its relevance in other aggressive kidney cancer contexts, including metastatic and/or rare RCC subtypes [e.g., Fumarate Hydratase (FH)-deficient and sarcomatoid RCCs], is unknown. Computational predictions, using a machine learning model trained on the human RCC TCGA dataset, and cell-based validations, confirmed BCL-XL dependence in all RCC subtypes. Remarkably, cell state changes, anoikis programs, inflammatory state, and metabolic perturbations (e.g., fumarate production in FH-deficient RCCs) independently conferred increased BCL-XL dependence. Correlation studies revealed that increased AMPK isoform 2 (PRKAA2) expression is a kidney-specific biomarker of BCL-XL dependence. Indeed, pharmacological AMPK activation sensitized RCCs to BCL-XL blockade. Finally, using functional studies, we developed a multivariate model that accurately predicted BCL-XL dependence in RCC. Our studies offer biomarkers for patient stratification and credential BCL-XL as a subtype agnostic vulnerability in difficult-to-treat RCCs.

cancer biology↗