Biology subjects

Zhao, Y.

Publications and source records attributed to Zhao, Y..

At least 343 records · Page 19Linked to original sources

bioRxiv2025

Nitrogen deposition reshapes plant nutrient acquisition strategies: a meta-analysis within the root economics space

Global nitrogen (N) deposition has fundamentally reshaped plant nutrient acquisition by altering mechanistic trade-offs between root exploration-exploitation strategies and mycorrhizal symbiosis. Through a meta-analysis of 135 studies spanning 153 sites, we demonstrate that N deposition stimulates root exploration (+84.6% root length density) and exploitation (+13.2% nitrogen content and +8.5% root biomass) while suppressing mycorrhizal dependence (-32.8% biomass, -18.73% colonized root length, -7.55% hyphal length), indicating a systemic shift toward root-autonomous nutrient acquisition. Divergent responses emerged between plant lifeforms: woody plants prioritize exploitation (+13.2% nitrogen content) over exploration capacity (-7.9% root length density), whereas herbaceous species exhibit synergistic enhancement of both strategies (+88% root length density, +13.2% nitrogen content and +71.3% root biomass). In contrast to N-only effects, Combined nitrogen and phosphorus additions reversed mycorrhizal suppression (+41.9% biomass), highlighting their persistent role in phosphorus acquisition under elevated nutrient conditions. The root economics space framework demonstrates predictive power, with plants possessing lower specific root length (SRL) and higher nitrogen content exhibiting stronger positive responses to N deposition, while those possessing higher SRL and lower nitrogen content showed reduced investment in root systems. These trait-mediated thresholds in carbon-nutrient tradeoffs refine our capacity to model belowground ecological responses to anthropogenic nitrogen perturbation, establishing a mechanistic basis for projecting ecosystem trajectories under global change.

plant biology↗

bioRxiv2025

Restoration of excitation/inhibition balance enhances neuronal signal-to-noise ratio and rescues social deficits in autism-associated Scn2a-deficiency

Social behavior is critical for survival and adaptation, which is profoundly disrupted in autism spectrum disorders (ASD). Social withdrawal due to information overload was often described in ASD, and it was suspected that increased basal noise, i.e., excessive background neuronal activities in the brain could be a disease mechanism. However, experimental test of this hypothesis is limited. Loss-of-function mutations (deficiency) in SCN2A, which encodes the voltage-gated sodium channel NaV1.2, have been revealed as a leading monogenic cause of profound ASD. Here, we revealed that Scn2a deficiency results in robust and multifaceted social impairments in mice. Scn2a-deficient neurons displayed an increased excitation-inhibition (E/I) ratio, contributing to elevated basal neuronal noise and diminished signal-to-noise ratio (SNR) during social interactions. Notably, the restoration of Scn2a expression in adulthood is able to rescue both SNR and social deficits. By balancing the E/I ratio and reducing basal neuronal firing, an FDA-approved GABAA receptor-positive allosteric modulator improves sociability in Scn2a-deficient mice and normalizes neuronal activities in translationally relevant human brain organoids carrying autism-associated SCN2A nonsense mutation. Collectively, our findings revealed a critical role of the NaV1.2 channel in the regulation of social behaviors, and identified molecular, cellular, and circuitry mechanisms underlying SCN2A-associated disorders. HIGHLIGHTSO_LINaV1.2 deficiency leads to pronounced social deficits in mice. C_LIO_LINaV1.2 deficiency results in an overall enhanced E/I ratio, elevated basal neuronal activity, and impaired signal-to-noise ratio. C_LIO_LIBoth the enhanced E/I ratio and impaired sociability are reversible through the restoration of NaV1.2 expression in adulthood. C_LIO_LITargeted restoration of NaV1.2 in striatum-projecting neurons rescues social impairments. C_LIO_LIGABA transmission is reduced in both mouse and human organoid models of SCN2A deficiency, and acute systemic administration of GABAA receptor-positive allosteric modulators restores sociability. C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=193 HEIGHT=200 SRC="FIGDIR/small/641498v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@154a323org.highwire.dtl.DTLVardef@1a27fbaorg.highwire.dtl.DTLVardef@16fa3beorg.highwire.dtl.DTLVardef@6712c2_HPS_FORMAT_FIGEXP M_FIG C_FIG Graphical abstract: Severe Scn2a deficiency leads to a predominate decrease in GABA transmission with an overall enhanced E/I ratio, elevated basal neuronal activity, impaired SNR, and social deficits in adult NaV1.2-deficient mice.

neuroscience↗

bioRxiv2025

A comprehensive map of missense trafficking variants in rhodopsin and their response to pharmacologic correction

Rhodopsin (RHO) missense variants are a leading cause of autosomal dominant retinitis pigmentosa (adRP), a progressive retinal degeneration with no currently approved therapies. Interpreting the pathogenicity of the growing number of identified RHO variants is a major clinical challenge, and understanding their disease mechanisms is essential for developing effective therapies. Here, we present a high-resolution map of RHO missense variant trafficking using two complementary deep mutational scanning (DMS) approaches based on a surface abundance immunoassay and a membrane proximity assay. We generated a comprehensive dataset encompassing all 6,612 possible single-residue missense variants, revealing a strong correlation between the two methods. Over 700 variants were identified with pathogenic trafficking scores, significantly expanding the number of RHO variants with functional evidence supporting pathogenicity. We demonstrate a high concordance between the trafficking scores and ClinVar pathogenicity classifications, highlighting this approachs utility in resolving variants of uncertain significance (VUS). The data also identified structurally clustered trafficking-deficient variants, predominantly within the N-terminal region and second extracellular loop, in and above the extracellular/intradiscal beta-plug region. Furthermore, we evaluated the efficacy of the non-retinoid pharmacological chaperone YC-001, observing significant rescue of trafficking defects in a majority of mistrafficking variants. This comprehensive functional map of RHO missense variants provides a valuable resource for pathogenicity assessment, genotype-phenotype correlations, and the development of targeted therapeutic strategies for RHO-adRP, paving the way for improved diagnosis and treatment for patients.

genomics↗

bioRxiv2025

Persistent representation of a prior schema in the orbitofrontal cortex facilitates learning of a conflicting schema

Schemas allow efficient behavior in new situations, but reliance on them can impair flexibility when new demands conflict, culminating in psychopathology. Evidence implicates the orbitofrontal cortex (OFC) in deploying schemas in new situations congruent with previously acquired knowledge. But how does this role affect learning of a conflicting behavioral schema? Here we addressed this question by recording single-unit activity in the OFC of rats learning odor problems with identical external information but orthogonal rules governing reward. Consistent with schema formation, OFC representations adapted to track the underlying rules, and both performance and encoding was faster on subsequent than initial problems. Surprisingly however, when the rule governing reward changed, persistent representation of the prior schema was correlated with acquisition of the new. Thus, OFC was not a source of interference and instead supported new learning by accurately and independently representing the old schema as the new was acquired.

neuroscience↗

bioRxiv2025

Virus-host interactome reveals host cellular pathways perturbed by tick-borne encephalitis virus infection

Tick-borne encephalitis virus (TBEV) poses an increasingly significant threat to public health. Here, we delved into the virus-host interactome and discovered that TBEV has a profound impact on multiple host cellular pathways. Viral pre-membrane protein (prM), non-structural proteins NS1, NS2B3, NS4A, and NS5 remodel the hosts innate immune responses. PrM and NS2B3 are involved in autophagy, NS4A is associated with neurodegenerative diseases, and NS5 participates in spliceosome dynamics, ribosomal biogenesis, cell cycle regulation, and DNA damage response. Notably, TBEV infection causes G0/G1 cell cycle arrest in host cells. NS5 interacts with histone acetyltransferase P300 to upregulate P16 expression, suppressing CDK4 and resulting in cell cycle arrest. Elevated P16 and reduced CDK4 levels were observed in TBEV-infected brain organoids. The P300 inhibitors and CDK4 agonist can reverse virus-induced cell cycle arrest and inhibit viral replication. Further analysis uncovered potential antiviral targets like KAT6A, XIAP, and RIOK1/BRD9. These findings provide valuable insights into TBEV pathogenicity and hold promise for antiviral drug development.

microbiology↗

bioRxiv2025

Dynamic fingerprinting of the human functional connectome

Resting-state functional connectivity (FC) have distinct, personalized patterns that could serve as a unique fingerprint of each individuals brain. While previous brain fingerprinting methods have used functional connectivity maps over a scanning session (static method), it has been shown that the brain is a dynamic system that switches between several metastable states, each of which having a different FC map. Taking the dynamic nature of brain connectivity into account will likely lead to more subject-specific information and better individual identification. In this paper, we derived the state-specific FCs using sliding window correlation and clustering and evaluated their performance in individual identification and cognitive score prediction. The resultant dynamic fingerprints outperformed the static fingerprints in identification accuracy. Furthermore, some of the brain states were more accurate in predicting cognitive scores, indicating that connectivity in some brain states is informative of cognition abilities, possibly useful as biomarkers for brain disorders. Impact StatementOur findings suggest that state-specific functional connectivity patterns of the brain are unique for each individual. These brain states predicted participants cognitive performance, suggesting that they have the potential to be used as biomarkers for cognitive function or neurological disorders. Integrating state-based functional connectivity into clinical frameworks could potentially enhance early diagnosis and patient stratification, and lead to targeted interventions for neuropsychiatric and neurodegenerative conditions.

neuroscience↗

bioRxiv2025

Distinct prelimbic cortex ensembles encode response execution and inhibition

Learning when to initiate or withhold actions is essential for survival and requires integration of past experiences with new information to adapt to changing environments. While stable prelimbic cortex (PL) ensembles have been identified during reward learning, it remains unclear how they adapt when contingencies shift. Does the same ensemble adjust its activity to support behavioral suppression upon reward omission, or is a distinct ensemble recruited for this new learning? We used single-cell calcium imaging to longitudinally track PL neurons in rats across operant food reward Training, Extinction and Reinstatement, trained rat-specific decoders to predict trial-wise behavior, and implemented an in-silico deletion approach to characterize ensemble contributions to behavior. We show that operant training and extinction recruit distinct PL ensembles that encode response execution and inhibition, and that both ensembles are re-engaged and maintain their roles during Reinstatement. These findings highlight ensemble-based encoding of multiple learned associations within a region, with selective ensemble recruitment supporting behavioral flexibility under changing contingencies.

neuroscience↗

bioRxiv2025

Highly accurate prediction of broad-spectrum antiviral compounds with DeepAVC

Lethal viruses, such as HIV, pose a significant threat to human life, with each pandemic causing millions of fatalities globally. Small-molecule antiviral drugs provide an efficient and convenient approach to antiviral therapy by either inhibiting viral activity or activating the host immune system. However, conventional antiviral drug discovery is often labor-intensive and time-consuming due to the vast chemical space. Although some existing computational models mitigate this problem, there remains a lack of rapid and accurate method specifically designed for antiviral drug discovery. Here, we propose DeepAVC, a universal framework based on pre-trained large language models, for highly accurate broad-spectrum antiviral compounds discovery, including two models, DeepPAVC for phenotype-based prediction and DeepTAVC for target-based prediction. As a result, DeepAVC greatly outperforms other in silico methods. More importantly, in the top predictions, MNS and NVP-BVU972 were identified as novel compounds with promising broad-spectrum antiviral activities by in vitro experiments. Finally, DeepAVC demonstrates high interpretability, one of the bottlenecks of current AI methods, due to its ability of analyzing key functional groups of antiviral compounds and important binding sites on targets.

bioinformatics↗

bioRxiv2025

Structural and Functional Insights into GGCX-FIX Interaction: Implications for Vitamin K-Dependent Bleeding Disorders

Gamma-carboxylation, catalyzed by {gamma}-glutamyl carboxylase (GGCX), is a critical post-translational modification essential for the biological activity of vitamin K-dependent proteins (VKDPs). Mutations in GGCX, depending on their specific location, result in vitamin K-dependent coagulation factor deficiency type 1 (VKCFD1), which encompasses a broad spectrum of clinical manifestations ranging from mild to severe, including bleeding disorders, osteoporosis, and vascular calcification. The limited knowledge of GGCXs structure and functional regions hinders our understanding of the consequences of GGCX mutations and the treatment for VKCFD1. This study aimed to identify key functional regions of GGCX and their interactions with VKDPs to better elucidate the molecular mechanisms underlying these diverse clinical symptoms. Using AlphaFold 3 and molecular dynamics simulations, we developed a complex binding model of GGCX, FIX, and reduced vitamin K, which revealed critical regions and residues involved in their interaction. Site-directed mutagenesis and cell-based assays further validated the model, confirming that multisite and regional cooperative binding of FIX to GGCX plays a key role in modulating {gamma}-carboxylation efficiency. Additionally, novel residues (I296, M303, M401, M402) were identified as essential for GGCXs dual enzymatic activities: carboxylation and vitamin K epoxidation. We further demonstrated that the spatial proximity of these active sites supports the hypothesis that GGCXs carboxylation and vitamin K epoxidation centers are interconnected, ensuring the efficient coupling of these processes. Our GGCX-FIX binding and carboxylation model aligns with known pathogenic GGCX mutations, providing valuable insights into the molecular basis of coagulation disorders caused by GGCX mutants.

pathology↗

bioRxiv2025

Key residue Unlocks Dual PIP2-Dependent and Independent Gating in G Protein-Gated Inwardly Rectifying Potassium Channels

G protein-gated inwardly rectifying potassium (GIRK) channels mediate membrane hyperpolarization in response to G protein-coupled receptor activation and are critical for regulating neuronal excitability. The membrane phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2) is essential for regulating a large family of ion channels, and disruptions in PIP2 interactions contribute to some neurological diseases. Structural analyses have identified key residues in PIP2-mediated gating of the GIRK2 channel. Notably, Arginine-92 (R92), a highly conserved basic residue at the membrane interface in GIRK2, interacts with PIP2 as well as cholesteryl hemisuccinate (CHS), a potentiator of GIRK2. Here, we used a combination of electrophysiological assays, fluorescent K+ flux measurements, cryo-electron microscopy, and molecular dynamics simulations, and find that mutations at R92 (Y, F, and Q) not only alter PIP2 sensitivity but can also reveal a novel gating mechanism that is independent of PIP2. These findings indicate that R92 plays a crucial role in modulating GIRK2 channel gating, offering new insights into developing potential therapeutic targets for treating neurological disorders linked to GIRK channel dysfunction.

biophysics↗

bioRxiv2025

Generalization but not strengthening of negative memories drives the development of depression-like behaviors

Individuals with depression exhibit intensified and generalized negative memories. However, how prior memories actually influence onset and/or progression of depression, particularly whether the strengthening or the generalization of prior memories play a key role, is poorly understood. Here, we introduced behavioral paradigms to differentiate between memory strengthening and generalization, and found that negative experiences that produced memory overgeneralization, but not strengthening, induced depression-like behaviors. Furthermore, we identified that the medial prefrontal cortex (mPFC) to the bed nucleus of the stria terminals (BNST) projection functionally linked memory generalization to depression, together with single-cell calcium imaging which revealed a significant overlap in the mPFCBNST neuronal ensemble encoding depression-like behaviors with that encoding generalized memory, but not memory strength. Circuit-based transcriptomic analysis and chromophore-assisted light inactivation demonstrated that triggering actin remodeling in the mPFCBNST neurons, a memory consolidation mechanism enhanced generalization while reducing memory strength, also induced depression-like behaviors. Collectively, these findings suggest generalization of negative memories as a primary factor that drives depression-like behaviors, highlighting the importance of early identification of individuals at risk for depression who exhibit overgeneralized negative memories, and targeting overgeneralized negative memories in the treatment of depression.

animal behavior and cognition↗

bioRxiv2025

MethylProphet: A Generalized Gene-Contextual Model for Inferring Whole-Genome DNA Methylation Landscape

AO_SCPLOWBSTRACTC_SCPLOWDNA methylation (DNAm) is a key epigenetic modification that regulates gene expression and is pivotal in development and disease. However, profiling DNAm at genome scale is challenging: of [~]28 million CpG sites in the human genome, only about 1-3% are typically assayed in common datasets due to technological limitations and cost. Recent deep learning approaches, including masking-based generative Transformer models, have shown promise in capturing DNAm-gene expression relationships, but they rely on partially observed DNAm values for unmeasured CpGs and cannot be applied to completely unmeasured samples. To overcome this barrier, we introduce MethylProphet, a gene-guided, context-aware Transformer model for whole-genome DNAm inference without any measured DNAm input. MethylProphet compresses comprehensive gene expression profiles ([~]25K genes) through an efficient bottleneck multilayer perceptron, and encodes local CpG sequence context with a specialized DNA tokenizer. These representations are integrated by a Transformer encoder to predict site-specific methylation levels. Trained on large-scale pan-tissue whole-genome bisulfite sequencing data from ENCODE (1.6 billion CpG-sample pairs, [~]322 billion tokens), MethylProphet demonstrates strong performance in hold-out evaluations, accurately inferring DNAm at unmeasured CpGs and generalizing to unseen samples. Furthermore, application to TCGA pan-cancer data (chromosome 1, 9,194 samples; [~]450 million training pairs, 91 billion tokens) highlights its potential for pan-cancer whole-genome methylome imputation. MethylProphet offers a powerful and scalable foundation model for epigenetics, providing high-resolution methylation landscape reconstruction and advancing both biological research and precision medicine.

bioinformatics↗

bioRxiv2025

Dietary quorum quenching AHL lactonase impairs the adaptation of commensal bacterium to host innate immunity

Quorum sensing (QS) is the communication system of bacteria that depends on QS signals. Quorum quenching (QQ) enzymes degrade QS signals and are promising alternatives of antibiotics to treat bacterial infections. Here, we found that dietary QQ N-acyl homoserine (AHL) lactonase led to microbiota dysbiosis in zebrafish, with reduction of Aeromonas and enrichment of Plesiomonas. Through gnotobiotic zebrafish colonized with a minimal microbiota, we found that QQ-mediated microbial alteration relies on host Myd88 signaling and neutrophil elastase. Mechanistically, quorum quenching increased the susceptibility of commensal Aeromonas to host neutrophil elastase by impairing bacterial lateral flagellar system, leading to reduced colonization of Aeromonas and subsequent enrichment of Plesiomonas due to ecological competition of the two species. Together, we found that dietary QQ lactonase led to microbiota alteration by impairing the adaptation of commensal Aeromonas to host innate immunity, which provided novel insight in the role of quorum sensing in host-microbiota interaction.

microbiology↗

bioRxiv2025

Differences between Scaly-sided Merganser (Mergus squamatus) and Common Merganser (M. merganser) feather microstructure

The microstructures of breast feathers, flank feathers, rectrices, and down feathers of both Scaly-sided Merganser (Mergus squamatus) and Common Merganser (M. merganser) were meticulously examined using scanning electron microscope. The observed data were subjected to the Mann-Whitney U test, followed by hierarchical clustering and Fishers discriminant analysis of indicators with significant differences in morphological structure. The findings revealed statistically significant differences between Scaly-sided Merganser and Common Merganser in terms of the length of the base handle from distal barbules in the breast feathers, the length of the base handle, the number of hooked barbules, the number of cilia from distal barbules, and the number of ventral teeth from proximal barbules in rectrices. Additionally, variations in the distance between nodules, the nodular diameter, and the length of the base nodular of the down feathers were noted. Hierarchical clustering successfully categorized Scaly-sided Merganser and Common Merganser into distinct groups, achieving a 100% accuracy rate. Moreover, Fishers discriminant analysis demonstrated a 100% correctness rate when the back-substitution method was used to test the discriminant equation. Consequently, the length of the base handle of distal barbules in breast feathers, along with all selected indicators from rectrices and down feathers, may provide references to identify Scaly-sided Merganser and Common Merganser.

zoology↗

bioRxiv2025

Esophageal adenocarcinoma relapse after chemoradiation is dominated by a basal-like subtype

Neoadjuvant chemoradiation therapy (RCT) is a frequently used treatment regimen for esophageal adenocarcinoma (EAC); however, the response varies dramatically, and resistance is a clinical challenge. We aimed to identify the molecular mechanisms underlying RCT resistance. We established a mouse xenograft RCT model with human EAC cell lines representing different response groups, and tested enhanced genomic instability as a potential evolutionary modulator by reducing BRCA2 function. Xenografts that relapsed after RCT displayed upregulation of stress response keratins, including KRT6 and KRT16 connected with a basal-like transcriptomic/ proteomic phenotype. We screened our cohort of 728 patients with EAC and found significantly shorter overall survival for patients with KRT6-high tumors, driven by patients receiving neoadjuvant treatment. Overall, we identified a basal-like cell state in EAC that reflects RCT relapse. The basal-like subtype is a marker of treatment failure, providing a new avenue for translational research to overcome RCT resistance.

cancer biology↗

bioRxiv2025

SNARE complex assembly and disassembly dynamics in response to Ca2+ current activation in live cells

A SNAP25 based FRET construct named SCORE (SNARE COmplex REporter) has revealed a transient FRET increase that specifically occurred at fusion sites preceding fusion events by tens of milliseconds and presumably reflects vesicle priming. The FRET increase lasts for a few seconds until it is reversed. In those experiments, the FRET increase was found to be localized to areas <0.5 {micro}m2 at sites of transmitter release as detected amperometrically using electrochemical detector arrays. Due to the localization to such small areas, it was unknown if the reversal of the FRET increase is due to local dispersion of high-FRET SCORE copies leaving the site after fusion and exchange with surrounding low-FRET copies, or if it reflects disassembly of the high-FRET complexes. To resolve this question, we performed whole-cell patch clamp pulse stimulation experiments, imaging the entire footprint of the cells in Total Internal Reflection Fluorescence (TIRF) excitation mode such that diffusional exchange between high-FRET and low-FRET copies does not produce a net FRET change. We show here that pulse stimulation of calcium currents results in FRET ratio transients with a time course very similar that related to fusion events. By comparing the kinetics of the FRET ratio decay with analytical and numerical diffusion simulation results, we show that the experimentally observed kinetics cannot be explained by diffusional exchange and conclude that the SCORE FRET ratio transients reflect incorporation of SCORE in SNARE complexes followed by SNARE complex disassembly. Experiments using Synaptobrevin 2/Cellubrevin double knock-out mouse embryonal chromaffin cells showed no pulse induced FRET change, indicating that the vSNARE is required for the incorporation of SCORE (or SNAP25 in wild type cells) in the SNARE complex during priming. Statement of SignificanceIn chromaffin cells, SNAP25-based FRET constructs (SCORE) revealed transient FRET increases within <0.5 {micro}m2 areas, preceding individual fusion events that reversed within seconds. It remained unknown whether this reversal stems from high-FRET complex disassembly or diffusion-mediated exchange with low-FRET complexes. Here, we performed whole-cell patch-clamp pulse stimulation with TIRF microscopy, imaging large [~]30 {micro}m2 areas of the cell footprint. Calcium currents induced FRET transients with the same decay time constant of [~]1.5 s, significantly shorter than the time scale of diffusion. The SCORE FRET ratio thus reports in real time the dynamics of SNRE complex assembly and disassembly in live cells. Using Synaptobrevin 2/Cellubrevin double knock-out mouse chromaffin cells we show that vSNAREs are required for SNAP25 incorporation into SNARE complexes during priming.

biophysics↗

bioRxiv2025

Avoidance of fratricide and improved efficacy of CD33-MSLN CAR-iNK cells in treating acute myeloid leukemia

Acute myeloid leukemia (AML) patients are often older, which brings challenges of endurance and persistent efficacy of autologous CAR-T cell therapies. Allogenic CAR-NK cell therapies may offer reduced toxicities and enhanced anti-leukemic potential against AML. In this study, we designed a novel CD33-mesothelin loop CAR (Loop CAR) and evaluated its anti-tumor efficacy in human umbilical cord blood-derived NK (UCB-NK) cells and human pluripotent stem cell-derived NK (hPSC-iNK) cells. The Loop CAR exhibited superior cytotoxicity against dual-antigen-positive tumor cell lines and primary AML cells. To further avoid fratricide caused by endogenous CD33 expression in NK cells, we established a hPSC-derived cell line via knockout of CD33 gene (CD33KO) and engineered Loop CAR. Rather than enforced expression of exogenous CD16, we generated abundant mature CD33KO-Loop CAR-iNK cells highly expressing endogenous CD16 via an organoid induction approach. This innovative strategy effectively mitigated NK cell fratricide and significantly enhanced CD33 and mesothelin-mediated specific cytotoxicity. Moreover, the CD33KO-Loop CAR-iNK cells demonstrated superior tumor-killing activity in AML xenograft mice and significantly prolonged survival. hPSC-derived CD33KO-Loop CAR-iNK cells possess unique advantages and translational potential for treating AML.

cancer biology↗

bioRxiv2025

Engineered CRO-CD7 CAR-NK cells derived from pluripotent stem cells avoid fratricide and efficiently suppress human T-cell malignancies

BackgroundT cell malignancies are highly aggressive hematological tumors with limited effective treatment options. CAR-NK cell therapy targeting CD7 has emerged as a promising approach for treating T-cell malignancies. However, conventional CAR-NK cell therapy faces the challenges of cell fratricide due to CD7 expression on both malignant cells and normal NK cells. Additionally, engineering CARs into human tissue-derived NK cells demonstrates heterogeneity, low transduction efficiency, and high manufacturing costs. MethodsThe human pluripotent stem cells (hPSCs) were genetically modified by knocking out the CD7 gene and introducing the CD7 CAR expression cassette to generate CD7 KO-CD7 CAR-hPSCs. These modified hPSCs were subsequently differentiated into CD7 KO-CD7 CAR-iNK cells using an efficient organoid induction method. The cytotoxicity of CD7 KO-CD7 CAR-iNK cells against CD7+ tumor cells was evaluated. Furthermore, we overexpressed the CXCR4 gene in CD7 KO-CD7 CAR-hPSCs and derived CXCR4-expressing CD7 KO-CD7 CAR-iNK (CRO-CD7 CAR-iNK) cells. The dynamics of CRO-CD7 CAR-iNK cells in vivo were tracked, and their therapeutic efficacy was assessed using human T-cell acute lymphoblastic leukemia (T-ALL) xenograft models. ResultsThe CD7 KO-CD7 CAR-iNK cells derived from CD7 KO-CD7 CAR-hPSCs effectively avoided fratricide, demonstrated normal expansion, and exhibited potent and specific anti-tumor activity against CD7+ T-cell tumor cell lines and primary T-ALL cells. CXCR4 overexpression in CRO-CD7 CAR-iNK cells improved their homing capacity and extended their persistence in vivo. The CRO-CD7 CAR iNK cells significantly suppressed tumor growth and prolonged the survival of T-ALL tumor-bearing mice. ConclusionsOur study provides a reliable strategy for the large-scale generation of fratricide-resistant CD7 CAR-iNK cells with robust anti-tumor effects from hPSCs, offering a promising cell product to treat T-cell malignancy.

cancer biology↗