Biology subjects

Xu, L.

Publications and source records attributed to Xu, L..

At least 145 records · Page 8Linked to original sources

bioRxiv2025

Demethylating agents drive PARP inhibitor resistance in ovarian carcinomas with BRCA1 gene silencing

DNA methyltransferase 1 inhibitor (DNMT1i) therapy is a promising option for increasing immune response as part of combination cancer therapy. High-grade serous ovarian carcinoma (HGSOC) is a highly aggressive cancer with poor survival outcomes, where DNMT1i therapy is being increasingly explored. HGSOC with epigenetically silenced BRCA1 has been shown to respond to PARP inhibitor (PARPi) treatment - a core targeted therapy for HGSOC. However, loss of silencing of even a single BRCA1 allele causes PARPi and platinum chemotherapy resistance. We tested whether BRCA1 silencing was robust to DNMT1i therapy, or would be reversed, thus driving PARPi resistance. We previously generated two homozygously silenced BRCA1 HGSOC cell lines: WEHI-CS62 and an OVCAR8 derivative. DNMT1i treatment caused sustained BRCA1 promoter methylation loss, gene re-expression and PARPi resistance in both of these silenced BRCA1 lines, but not in mutated BRCA1/2 or RAD51C lines. Methylation arrays confirmed transient global CpG methylation losses following DNMT1i. CRISPR deletion of the re-expressed BRCA1 copy in WEHI-CS62 restored silencing and PARPi sensitivity. Furthermore, DNMT1i treatment of a silenced BRCA1 PDX caused heterogeneous BRCA1 promoter methylation loss. In summary, DNMT1 inhibitors caused sustained reduction of BRCA1 promoter methylation in HGSOC cells. This resulted in BRCA1 re-expression and PARP inhibitor resistance, presenting a significant risk for up to 17% of HGSOC patients with BRCA1 gene silencing who could benefit from PARP inhibitor therapy. We conclude that DNA demethylation therapy should be avoided for HGSOC patients with epigenetically silenced BRCA1.

cancer biology↗

bioRxiv2025

A transcriptional biosensor reveals mechanisms of α-ketoglutarate signaling to chromatin

Alpha-ketoglutarate (KG) is required for chromatin demethylation but mechanisms controlling KG abundance in the nucleus are poorly defined. Therefore, we designed a biosensor system to monitor this metabolite pool in human cells using an KG-responsive cyanobacterial transcription factor, NtcA. We then coupled this system with a genetic screen to identify genes that regulate KG in the nucleus, defining an inter-organelle pathway in which sequential mitochondrial activities of the GPT2 transaminase and SLC25A11 transporter supply nuclear KG. Using a mouse model of GPT2 deficiency, a human inborn error of metabolism, we found that this pathway controls chromatin methylation in the developing brain. Our work provides a tool to assess KG signaling to chromatin and a framework for leveraging forward genetics to study nuclear metabolite pools.

synthetic biology↗

bioRxiv2025

Super-Resolution Diffusivity Mapping Reveals Spatial Correlations Between Lateral Mobility and Structural Heterogeneities on Cellular Membranes

Cellular membranes orchestrate critical processes such as molecular transport and signal transduction, both regulated by the lateral mobility of lipids and proteins. However, resolving nanoscale diffusional heterogeneities and elucidating their underlying mechanisms remains a formidable challenge due to the membranes intricate architecture and compositional diversity. Here, we present point-cloud single-molecule diffusivity mapping (pc-SMdM), a cutting-edge super-resolution technique that offers a point-cloud data format with enhanced spatial resolution for diffusivity mapping. Using pc-SMdM, we visualize nanoscale diffusion slowdown clusters with [~]50 nm in diameter on plasma membranes. These clusters are predominantly governed by cholesterol content, alongside contributions from membrane protein assemblies and topographical features. Leveraging two-color pc-SMdM, we concurrently imaged multiple lipids and membrane probes, revealing distinct "fingerprint" diffusivity maps shaped by their interactions within lipid bilayers. Our findings position pc-SMdM as a transformative tool for spatially resolving molecular interactions and membrane dynamics in live cells, offering new insights into the underlying mechanisms that govern membrane mobility at the nanoscale.

biophysics↗

bioRxiv2025

Screening for pro-adhesive compounds and their relevance as therapeutic approach in Arrhythmogenic Cardiomyopathy

BackgroundArrhythmogenic Cardiomyopathy (ACM) is one of the major causes of sudden cardiac death in young adults. With the underlying patho-mechanisms not well understood, current therapeutic approaches for this genetic disease are solely symptomatic. A recent study demonstrates that loss of cell-cell adhesion is an important initial step leading to ACM. Because loss of cell-cell adhesion is considered a key initial step, we aim to identify new compounds from a drug library, which can restore intercellular adhesion and potentially serve as therapeutics for ACM. MethodsWe established a 2D cell adhesion-based high-throughput platform and screened an FDA-approved drug library. To model loss of intercellular adhesion, human cell lines deficient for the desmosomal adhesion molecule desmoglein-2 (DSG2) were employed and the revealed top candidates were validated in cells expressing different ACM patient mutations. The therapeutic potential of the top hit dexamethasone was evaluated in an inducible ACM disease mouse model using ECG, echocardiography and histology. Phospho-proteomic analysis was applied to investigate protective drug mechanisms. ResultsWe established a set-up to sensitively detect changes in cell-cell adhesion via a high-throughput platform and applied this approach to identify a large pool of adhesion-strengthening compounds in DSG2 deficient cells. The pro-adhesive effect of selected candidates was validated for different ACM patient mutations inducing defective cell cohesion. Importantly, in vivo administration of the selected top pro-adhesive drug dexamethasone rescued impaired right ventricular function in an ACM mouse model. Phospho-proteome analysis suggests modulation of AKT1/AMPK1 signaling and changes in cardiac contractility and junctional components as factors contributing to this protective effect. ConclusionsWe developed an adhesion-based high-throughput screening platform capable of identifying adhesion modulators. We revealed and validated several pro-adhesive compounds and showed a protective effect for the top candidate dexamethasone in an ACM mouse model. This indicates the potential of adhesion-strengthening compounds as therapeutic strategy in ACM and lays the basis for detailed follow-up studies. Moreover, these results and methods can be translated to other diseases with defective desmosomal adhesion. Clinical PerspectiveO_ST_ABSWhat is new?C_ST_ABSO_LIWe developed a novel high-throughput in vitro screening platform to identify compounds that strengthen cell-cell adhesion under conditions of disrupted adhesion, which is a central pathogenetic step in Arrhythmogenic Cardiomyopathy (ACM). C_LIO_LIWith aid of this screen, we identified and validated different compounds from an FDA-approved drug library, which restore disrupted cell-cell adhesion induced by ACM patient mutations including glucocorticoids such as dexamethasone. C_LIO_LIDexamethasone rescued impaired right ventricular function in a novel inducible ACM mouse model and reverted altered AKT1/AMPK1 signaling as well as changes in the phosphorylation state of components of the contractile and junctional apparatus. C_LI What are clinical implications?O_LIWe established a high-throughput cell-cell adhesion screening method as a viable tool for identifying drugs to combat ACM and other diseases featuring impaired desmosomal adhesion. C_LIO_LIWe identified the glucocorticoid dexamethasone as a promising compound to therapeutically approach ACM. C_LIO_LIThis study highlights strengthening of disrupted cell-cell adhesion as a novel therapeutic strategy to treat ACM. C_LI

pathology↗

bioRxiv2025

NeoAtlas and NeoBert: A Database and A Predictive Model for Canonical and Noncanonical Tumor Neoantigens

Neoantigens are classified into canonical and noncanonical types. Noncanonical neoantigens include those derived from noncoding regions, transposable elements (TE), and intron retention events, and they have recently gained considerable attention in cancer immunity. We curated 35,574 non-redundant neoantigen-HLA pairs from 14 immunopeptidomes studies, by analyzing unique features and differences across various sources of neoantigens. This knowledge enabled us to develop machine learning models for the prediction of different types of neoantigens. Our data and models are available at a public portal (https://ngdc.cncb.ac.cn/neoatlas) to facilitate broad access and future research. This resource offers advanced functionalities, including integration with epigenome browsers which allow easy navigation of epigenomic datasets to support and confirm the expression of neoantigens. We further demonstrate that combining our database with mass spectrometry analysis can identify noncanonical neoantigens. The resource we constructed holds significant value and promise for the development of neoantigen-based vaccines.

systems biology↗

bioRxiv2025

Leveraging cell-type specificity and similarity improves single-cell eQTL fine-mapping

Identifying cell-type-specific expression quantitative trait loci (eQTLs) is important to understanding the genetic regulation of gene expressions at the cell-type level and its relevance to complex traits. However, existing eQTL fine-mapping methods are limited in power and accuracy when cell types are analyzed separately. To improve eQTL mapping, we present CASE, a Bayesian framework to perform Cell-type-specific And Shared EQTL fine-mapping that simultaneously analyzes multiple cell types. CASE can effectively capture effect-sharing patterns across cell types while disentangling the confounding effects of linkage disequilibrium (LD). We demonstrate that CASE outperforms the existing single-trait (SuSiE) and multi-trait (mvSuSiE) eQTL methods through comprehensive simulations. When applied to the OneK1K data, CASE identified more genetic regulations of gene expressions, better capturing cell type specificity and functionally enriched and disease-associated eQTLs. The CASE framework for single-cell eQTL fine-mapping can be broadly applied to multi-tissue and multi-trait genetic studies.

genetics↗

bioRxiv2025

DNA polymerase actively and sequentially displaces single-stranded DNA-binding proteins

Single-stranded DNA-binding proteins (SSBs) play a crucial role in stabilizing and protecting transiently exposed single-stranded DNA (ssDNA), yet the mechanisms governing their displacement by DNA polymerase (DNAp) during replication remain largely unexplored. Using bacteriophage T7 DNAp and its SSB, T7 gp2.5, we investigated the molecular mechanisms and visualized the dynamic process underlying SSB displacement. Our single-molecule force spectroscopy demonstrates that T7 SSB modulates DNA replication in an ssDNA conformation-dependent manner, regulated by tension applied to the DNA template. By integrating dual-color single-molecule imaging, we observe that T7 SSB remains stationary as DNAp approaches, indicating that SSB molecules are sequentially displaced rather than pushed forwards. Molecular dynamics (MD) simulations revealed reduced energy barriers for SSB dissociation in the presence of DNAp. This finding, combined with the detected FRET signals when DNAp approaches an SSB-bound ssDNA region and observations of faster replication rates compared to relative slow intrinsic SSB dissociation, collectively support an active displacement mechanism. Using both ensemble and single-molecule analyses, we demonstrated that SSB saturation of ssDNA is critical for optimal replication efficiency, with each SSB molecule contributing positively to the process. Taken together, the uncovered spatial-temporal coordination between SSB and DNAp is necessary for resolving molecular collisions during DNA replication, and may represent a universal strategy employed by other DNA translocating motors to ensure genomic integrity.

biophysics↗

bioRxiv2025

A functional anatomical shift from the lateral frontal pole to dorsolateral prefrontal cortex in emotion action control underpins elevated levels of anxiety: partial replication and generalization of Bramson et al., 2023

Flexible control over emotional behavior represents a promising target for novel interventions for mental disorders. Accumulating evidence has indicated a key role of the lateral frontal pole (FPl) and its connections with other cortical and subcortical systems in emotional action regulation. A recent study from Bramson et al., (2023) employed a multi-modal neuroimaging approach to demonstrate a functional-anatomical shift from FPI to dorsolateral prefrontal cortex (DLPFC) in a sample of anxious individuals during emotional action control. While these findings might represent a venue for interventions in anxiety disorders, conventional neuroimaging strategies are often limited with respect to generalizability and reproducibility. Against this background we capitalized previous large-scale fMRI data in n = 250 participants using an affective linguistic Go/NoGo paradigm to examine the robustness of the reported associations with trait social anxiety across samples, cultures and paradigms. Additionally, context-dependent functional connectivity patterns were explored to examine action control in different emotional contexts. In line with previous study, we found no difference between high- and non-anxious group on the behavioral congruency-effect. The neural results showed that non-social anxious group engaged the left FPl while the high-social anxious group specifically recruited the DLPFC, however in the absence of significant between-group differences. Importantly, the level of trait social anxiety was significantly positively related with DLPFC activity and negatively with left FPl activation across groups. Furthermore, context-dependent functional connectivity analyses revealed a negative context-specific neural shift from the sgACC-FPl to sgACC-DLPFC specifically in the high anxiety group. Together, the present study employed a different task paradigm, population and analytic methods, partially replicated the findings described by Bramson et al., (2023) and additionally determined context-specific changes in the communication with the sgACC in high anxiety. The findings provide further evidence for target-based interventions of persistent emotional control deficits in anxiety disorders.

neuroscience↗

bioRxiv2025

Evaluation of Aggregate Oral Fluid Sampling for Early Detection of African Swine Fever Virus Infection

African swine fever (ASF) is a highly infectious viral disease that poses significant threat to the United States and global pig industries. Given the lack of effective vaccines, control and prevention of the spread of African swine fever virus (ASFV) is dependent on enhanced surveillance and early disease detection. Commercial swine operations in the US are characterized by comparatively large number of pigs, and sampling individual pigs, which represents the main strategy for current ASF surveillance, is both costly and labor intensive. The major objective of this study was to estimate the diagnostic sensitivity of pen-based aggregate oral fluid testing for ASFV in infected pigs in a pen of 30 animals and evaluate its utility as a tool to support surveillance of ASF in the United States. The study was performed in three phases: (i) Virus (Ghana ASFV24) amplification in a target host species to generate the challenge inoculum, (ii) Titration of the inoculum (10% spleen homogenate) in target host species to determine the minimum dose inducing acute ASF in pigs with survival up to 5 - 6 days post-inoculation (dpi), and (iii) The main study involving 186 pigs consisting of 6 replicates of 30 pigs per pen and one seeder pig inoculated with the Ghana ASFV24 per pen. Daily sampling of aggregate oral fluids, uncoagulated blood, oropharyngeal swabs, fecal and water nipple swabs, and recording of rectal temperatures and clinical observations, was carried out. The seeder pigs were each inoculated intramuscularly with 0.5 ml of the 10% spleen homogenate which induced the desired clinical course of ASF in the pigs with survival of up to 6 dpi. ASFV DNA could be detected in the seeder pigs as early as 1 dpi and 2 dpi in the blood and oropharyngeal swabs, respectively. Transmission of ASFV from the seeder pigs to the contact pig population was detected via positive amplification of ASFV DNA in aggregate oral fluid samples at 3 days post-contact (dpc) in 4 out of 6 pens, and in all 6 pens at 4 dpc. Testing of oropharyngeal swabs and blood samples from individual pigs revealed variable number of ASFV positive pigs between 3 and 5 dpc, with detection of 100% positivity between 6 and 18 dpc, the study endpoint. These findings demonstrate the potential utility of aggregate oral fluid sampling for sensitive and early detection of ASFV incursion into naive swine herds. It also demonstrates that testing of environmental samples from the premises could further enhance overall ASF early detection and surveillance strategy. Author summaryEarly detection of ASFV in swine farms requires robust passive surveillance using sample types and sampling methods that allow sensitive and timely detection. Commercial swine operations in the US or North America are characterized by comparatively large number of pigs, and sampling individual pigs, which represents the current strategy for ASF surveillance, is both costly and labor-intensive. Oral fluid has been shown to be an acceptable sample type for detection of ASFV in individual infected pigs. For the first time on such a scale, we conducted a study enrolling 186 pigs, with daily sampling, in six experimental replicates at 3.2% pen prevalence using a highly virulent ASFV (Ghana ASFV24) to evaluate the utility of aggregate oral fluids for early detection of ASFV. Whole genome sequencing and characterization confirmed grouping of the virus with those in the p72 genotype II cluster. We demonstrate that intramuscular inoculation of the seeder pigs induces acute ASF and transmission to the contact pigs which is detectable in aggregate oral fluids as early as 3 - 4 dpc. We have shown that ASFV DNA detections in aggregate oral fluids correlate with the oropharyngeal swabs of individual pigs. In comparison, ASFV DNA in individual blood samples is detected 1-3 days later. The study demonstrates the potential utility of aggregate oral fluid sampling for enhanced surveillance of ASFV in large commercial swine operations.

microbiology↗

bioRxiv2025

SpaFun: Discovering Domain-specific Spatial Expression Patterns and New Disease-Relevant Genes using Functional Principal Component Analysis

SpaFun is a novel, non-model-based method developed to address limitations in existing spatially variable gene (SVG) detection techniques, particularly for large-scale spatially resolved transcriptomics (SRT) datasets. These limitations include computational inefficiency, limited statistical power with increasing data size, and the inability to capture spatial heterogeneity and co-expression patterns among genes. Built on functional principal component analysis (fPCA), SpaFun identifies domain-representative genes (DRGs) with significantly better computational efficiency and greater statistical power while accounting for spatial heterogeneity and co-expression patterns among genes. We applied SpaFun to three SRT datasets and demonstrated that SpaFun outperformed state-of-the-art algorithms for identifying representative genes for tumor regions (e.g., DESeq, edgeR, and limma), as well as recently developed novel algorithms designed for spatial omics to identify the representative genes (e.g., SPARK and CSIDE). This highlights SpaFuns ability to accurately identify genes most representative of each spatial domain (e.g., tumor, immune, or stroma regions). By uncovering novel disease-relevant genes overlooked by existing algorithms, SpaFun could provide insights into new molecular mechanisms and propose innovative therapeutic strategies to improve patient outcomes. Key PointsO_LISpaFun is the first method dedicated to identifying DRGs, capturing spatially representative expression patterns within annotated tissue regions, setting it apart from all traditional SVG detection methods. C_LIO_LIIt leverages fPCA to model gene expression as a function of spatial location, avoiding reliance on predefined spatial distribution assumptions. C_LIO_LIThe non-model-based framework ensures compatibility with different SRT platforms and experimental designs, making it a scalable and widely applicable tool for SRT research. C_LI

genomics↗

bioRxiv2025

Predicting Mutation-Induced Relative Protein-Ligand Binding Affinity Changes via Conformational Sampling and Diversity Integration with Subsampled Alphafold2 in Few-Shot Learning

Predicting the impact of mutations on protein-ligand binding affinity is crucial in drug discovery, particularly in addressing drug resistance and repurposing existing drugs. Current structure-based methods are constrained by their dependence on known protein-ligand complex structures. The absence of structural data for mutated proteins, along with the potential of mutations to modify protein conformational states, brings challenges for reliable predictions. The downstream application of conformational sampling of Alphafold, such as the prediction of binding free energy changes induced by mutation, has not been fully explored. To tackle the challenge, we propose a few-shot learning approach that integrates AlphaFold2 subsampling, ensemble docking, and Siamese learning to predict the relative binding affinity changes induced by mutations. We validate our approach for predicting binding affinity changes in 31 variants of the mutated Abelson tyrosine kinase (ABL). The conformation ensembles for each ABL variant are generated using AlphaFold2 subsampling. The most likely conformational states are selected by Gaussian fitting to construct the pairwise structural inputs and compute relative binding energies. Then the relative binding affinities are derived by averaging the values predicted by the Siamese learning model across the constructed conformational pairs. By benchmarking on the Tyrosine kinase inhibitors (TKI) dataset and the direct binding affinity using the refined set of PDBbind database, our proposed approach achieves an increased Spearman correlation coefficient for five out of six TKI molecules across 31 mutants when estimating relative binding affinities. Although the proposed method is validated on the ABL, it can be transferred and applied to other drug-target interaction predictions when the conformational flexibility of proteins needs to be considered. We anticipate the approach will be valuable for predicting binding energy changes induced by mutations in proteins or ligands, facilitating drug discovery efforts.

bioinformatics↗

bioRxiv2025

CRISPR targeting of H3K4me3 activates gene expression and unlocks centromeric crossover recombination in Arabidopsis

H3K4me3 is a fundamental and highly conserved chromatin mark across eukaryotes, playing a central role in many genome-related processes, including transcription, maintenance of cell identity, DNA damage repair, and meiotic recombination. However, identifying the causal function of H3K4me3 in these diverse pathways remains a challenge, and we lack the tools to manipulate it for agricultural benefit. Here we use the CRISPR-based SunTag system to direct H3K4me3 methyltransferases in the model plant, Arabidopsis thaliana. Targeting of SunTag-SDG2 activates the expression of the endogenous reporter gene, FWA. We show that SunTag-SDG2 can be employed to increase pathogen resistance by targeting the H3K4me3-dependent disease resistance gene, SNC1. Meiotic crossover recombination rates impose a limit on the speed with which new traits can be transferred to elite crop varieties. We demonstrate that targeting of SunTag-SDG2 to low recombining centromeric regions can significantly stimulate proximal crossover formation. Finally, we reveal that the effect is not specific to SDG2 and is likely dependent on the H3K4me3 mark itself, as the orthogonal mammalian-derived H3K4me3 methyltransferase, PRDM9, produces a similar effect on gene expression with reduced off-target potential. Overall, our study supports an instructive role for H3K4me3 in transcription and meiotic recombination and opens the door to precise modulation of important agricultural traits.

plant biology↗

bioRxiv2025

ST-FFPE-mIF: Integrating Spatial Transcriptomics and Multiplex Immunofluorescence in Formalin-Fixed Paraffin-Embedded Tissues Using Stereo-seq

Stereo-seq is the state-of-the-art technique, which allows capturing of spatial transcriptomic data at sub-cellular resolution and in large scale of view. Stereo-seq FFPE technique is recently launched with the capability to obtain spatially resolved gene expression in FFPE tissue sections. However, the omic information is limited to transcriptome. To delineate spatial molecular hierarchy from transcriptome to phenome, we present the protocol that implements nine-plex immunofluorescence staining with detection of spatial transcriptomics in one FFPE tissue section (short for ST-FFPE-mIF). This protocol details the modified biochemistry procedure and also provides analytical framework to achieve single-cell resolution through nuclear or membrane staining-based cell segmentation. We demonstrated the application of ST-FFPE-mIF in colorectal cancer sample to illustrate how ST-FFPE-mIF facilitates a robust analysis of spatially resolved multi-omic data at the single-cell level.

biochemistry↗

bioRxiv2025

Benchmarking and optimizing Perturb-seq in differentiating human pluripotent stem cells

Perturb-seq is a powerful approach to systematically assess how genes and enhancers impact the molecular and cellular pathways of development and disease. However, technical challenges have limited its application in stem cell-based systems. Here, we benchmarked Perturb-seq across multiple CRISPRi modalities, on diverse genomic targets, in multiple human pluripotent stem cells, during directed differentiation to multiple lineages, and across multiple sgRNA delivery systems. To ensure cost-effective production of large-scale Perturb-seq datasets as part of the Impact of Genomic Variants on Function (IGVF) consortium, our optimized protocol dynamically assesses experiment quality across the weeks-long procedure. Our analysis of 1,996,260 sequenced cells across benchmarking datasets reveals shared regulatory networks linking disease-associated enhancers and genes with downstream targets during cardiomyocyte differentiation. This study establishes open tools and resources for interrogating genome function during stem cell differentiation.

genomics↗

bioRxiv2025

Actomyosin and the Arp2/3 Complex Are Involved in the Internalization of Cellulose Synthase Complexes

The coupling of exo- and endocytic trafficking of Cellulose Synthase Complexes (CSCs) has been proposed to be important for maintaining the population of active CSCs at the plasma membrane (PM) and thus appropriate levels of cell wall assembly. Although actin and myosin are known to participate in the late stages of exocytosis of CSCs, their exact role during CSC internalization events remains controversial. We constructed a functional, photoconvertible fluorescent mEOS2-CESA6 reporter and developed single-particle live-cell imaging approaches to visualize and quantify the dynamic behavior of CSCs at the PM during internalization. Using the small molecule inhibitor of clathrin, Endosidin 9-17 or ES9-17, we confirmed that clathrin-mediated endocytosis is a major pathway for CSC internalization. We also found that the actin cytoskeleton is involved in CSC internalization. Genetic or chemical inhibition of actin, myosin, or the Arp2/3 complex significantly reduced the frequency of CSC internalization events and prolonged the CSC pause time prior to internalization. Additionally, we found that the Arp2/3 complex contributes to the late stage of exocytosis of CSCs into the PM. These results reveal a role for actomyosin and the Arp2/3 complex in both CSC secretion as well as internalization that was previously undescribed in plant cells. One sentence summaryDirect visualization of individual CSC internalization events reveals that actomyosin participates in CSC internalization and the Arp2/3 complex contributes to both exocytosis and internalization of CSCs through regulating the dynamic homeostasis of the cortical actin cytoskeleton.

cell biology↗

bioRxiv2025

Modeling mucociliary mixing and transport at tissue scale

Mucociliary clearance is the primary defense mechanism in our respiratory system against aerosol pathogens and allergens. The rhythmic movement of cilia on airway-lining cells propels mucus flow, driving the movement of trapped particles. We want harmful particles, e.g., pathogens and allergens to be transported rapidly out of the airway and beneficial particles, e.g. vaccines and medicine, to stay and disperse. However, the impact of cilia density and distribution on mucociliary mixing and transport at the tissue scale remains poorly understood. We present three-dimensional (3D) simulations of mucociliary mixing and transport at the tissue scale. We investigate the influence of ciliary cluster spacing, metachronal wave, and ciliary density on mucus mixing and transport. Our findings reveal that: (i) cilia clusters produce swirls of mucus flow, their size scales with ciliary density; (ii) a single cilia cluster generates horizontal and upward transport with horizontal mixing; (iii) for three aligned cilia clusters, an optimal cluster spacing exists for achieving maximum horizontal transport; (iv) asynchronous beating among clusters enhances mixing but hinders transport; and (v) the diffusion of particles exhibits spatial inhomogeneity, leading to particle aggregation into discrete groups over time. These discoveries have implications for understanding particle distributions in the airway and contribute to drug delivery design considerations. Author summaryOur airways are exposed to various foreign particles, including allergens, pathogens, and aerosolized vaccines or medications. The mucus coating the airway surface traps these particles, and the beating cilia extending from the ciliated epithelial cells play a crucial role in propelling the flow of mucus to transport the particles out of the airway. This mucociliary clearance process is closely associated with the initiation and progression of airway tissue response, such as allergic responses, infections, and treatment effects. Changes in ciliated cell density or mucus production can significantly impact this process. Conversely, achieving a homogeneous distribution of aerosol drug particles within the airway is essential for optimizing treatment outcomes. We simulate cilia-mucus coupled flow to examine how the motion of cilia influences the transport and mixing of particles in the mucus. Our key findings suggest that an optimal spacing for ciliated cell clusters that enhances directional transport. Intriguingly, while the metachronal wave promotes mixing, it also tends to hinder the overall transport of particles.

systems biology↗

bioRxiv2024

Direct and abscopal anti-tumor responses elicited by peripheral nerve schwannoma irradiation synergize with anti-PD1 treatment in vestibular schwannoma models

NF2-related schwannomatosis (NF2-SWN) is a progressive and disabling disease requiring effective treatments. The hallmark of NF2-SWN is bilateral vestibular schwannomas (VSs), which progressively enlarge, leading to permanent sensorineural hearing loss and severely impacting patients quality of life. Currently, there are no FDA-approved drugs for VS or the associated hearing loss. Immune checkpoint inhibitors (ICIs) have revolutionized cancer treatment, but have not yet been systematically investigated in non-malignant tumors such as VS. In our studies, we demonstrated that combining anti-PD1 (PD1) treatment with radiation therapy (RT) provides three significant therapeutic benefits: i) Enhanced PD1 efficacy and immune memory: RT induces immunogenic cell death and activates the STING pathway, enhancing PD1 efficacy and generating long-term immune memory, ii) Reduced RT dose and associated tissue injury: The combination strategy reduces the required RT dose necessary for effective tumor control, potentially minimizing RT injury to surrounding normal tissues, and iii) Elicited abscopal effects on cerebellopontine angle (CPA) schwannomas: RT to peripheral nerve tumor induces a systemic abscopal effect, which synergizes with PD-1 to effectively control intracranial schwannomas without direct irradiation, sparing the cochlea from radiation exposure and avoiding auditory radiation injury. Together, our findings provide a compelling rationale for deploying ICIs in combination with radiotherapy as a novel treatment approach for patients with VS and NF2-SWN.

cancer biology↗

bioRxiv2024

Enhanced Tumor Control and Hearing Loss Prevention Achieved with Combined Immune Checkpoint Inhibitor and Anti-VEGF Therapy in Vestibular Schwannoma Model

BackgroundNF2-related schwannomatosis (NF2-SWN) is a debilitating condition that calls for robust treatment options. The defining feature of NF2-SWN is the presence of bilateral vestibular schwannomas (VSs), which grow over time and can result in irreversible sensorineural hearing loss, significantly affecting the quality of life for those affected. At present, there are no FDA-approved medications specifically for treating VS or related hearing loss. VS management involves radiotherapy or surgical resection, while bevacizumab, an anti-vascular endothelial growth factor (VEGF) monoclonal antibody (VEGF) may be used off-label in NF2-SWN to shrink the tumor. However, not all patients respond, and the effect is not always durable. There is a critical need for effective medications that can stop the growth of VS and prevent hearing loss associated with these tumors. While immune checkpoint inhibitors have transformed cancer therapy, their potential has not been thoroughly explored in non-malignant tumors such as VS. MethodsWe characterize the effects of anti-PD1 (PD1) treatment on tumor growth and hearing function in two syngeneic, immune-competent VS models. ResultsWe demonstrated that combining VEGF treatment with PD1 significantly enhances the efficacy of each monotherapy. Specifically, i) VEGF enhances PD1 efficacy by normalizing the tumor vasculature to improve drug delivery and immune cell infiltration, and by activating T cell and NK cell anti-tumor cytotoxicity via NKG2D upregulation; and ii) combining PD1 with VEGF treatment effectively controls tumors that progressed despite VEGF treatment. ConclusionThese findings provide a strong foundation for the development of PD1 with VEGF combination therapies for patients with NF2-SWN. Key pointsWe filled a critical gap in NF2 research: 1) we characterized the effects of immunotherapy on tumor growth and hearing function in non-malignant vestibular schwannomas 2) We showed combined anti-VEGF and anti-PD1 enhances the efficacy of each monotherapy Importance of the studyTreatment options for patients with NF2-SWN are limited or are associated with significant co-morbidities. There are no approved medical treatments for NF2-related tumors. While immune checkpoint inhibitors have transformed cancer therapy, their potential has not been thoroughly explored in non-malignant tumors such as VS. Our work filled this critical gap in NF2-SWN research. For the first time, we systemically evaluated ICI efficacy on tumor growth and hearing function in non-malignant schwannomas. Furthermore, we demonstrated that combining VEGF treatment with PD1 significantly enhances the efficacy of each monotherapy. Specifically: i) VEGF enhances PD1 efficacy by normalizing the tumor vasculature to improve drug delivery and immune cell infiltration, and by activating T cell and NK cell anti-tumor cytotoxicity via NKG2D upregulation; and ii) combining PD1 with VEGF treatment effectively controls tumors that progress despite VEGF treatment. Our findings provide a strong foundation for the development of PD1 with VEGF combination therapies for patients with NF2-SWN.

cancer biology↗