Biology subjects

Patel, S.

Publications and source records attributed to Patel, S..

At least 73 records · Page 4Linked to original sources

bioRxiv2025

Replicating the Gold Standard: A Novel Female Chronic Social Defeat Stress Model (femCSDS) for Studying Sex Differences in Depression

Depression shows significant sex differences in prevalence and neurobiological underpinnings, yet preclinical research investigating the pathophysiology of depression and the efficacy of antidepressants has predominantly relied on male models. Here, we establish a novel female chronic social defeat stress paradigm by leveraging the natural aggression of parous CD1 females, co-housed with castrated males to induce aggression while eliminating confounding sexual behaviors and without hormonal or surgical manipulations. Selected aggressive females reliably displayed offensive behaviors toward C57BL/6NCrl intruders across repeated encounters. Defeated female mice exhibited pronounced depression-like behaviors, including social withdrawal, anhedonia, behavioral despair, and elevated anxiety-like responses. Biochemical analysis revealed elevated glutamate levels in Nucleus Accumbens (NAc) and caudate putamen (CPu). Alterations in EAAT1, GRIN2B, and Neurabin expression were observed in CPu, indicating excitotoxic stress and compromised synaptic integrity. Label free Quantitative MS-MS analysis of NAc revealed 1194 significantly dysregulated proteins. Ingenuity Pathway Analysis highlighted canonical pathway disruptions in synaptogenesis signaling pathway and glutamate signaling pathway. Disease and function analysis revealed enrichment in neuroinflammation, synaptic dysfunction, and mitochondrial dysfunction. Given the extensive literature on male CSDS and its established pathophysiology, we aimed and successfully developed female-specific replica model of traditional male CSDS, enabling direct comparison and elucidation of sex differences in depression pathophysiology.

neuroscience↗

bioRxiv2025

Dual-feature selectivity enables bidirectional coding in visual cortical neurons

Sensory neurons are traditionally viewed as feature detectors that respond with an increase in firing rate to preferred stimuli while remaining unresponsive to others. Here, we identify a dual-feature encoding strategy in macaque visual cortex, wherein many neurons in areas V1 and V4 are selectively tuned to two distinct visual features--one that enhances and one that suppresses activity--around an elevated baseline firing rate. By combining neuronal recordings with functional digital twin models--deep learning-based predictive models of biological neurons--we were able to systematically identify each neurons preferred and non-preferred features. These feature pairs served as anchors for a continuous, low-dimensional axis in natural image similarity space, along which neuronal activity varied approximately linearly. Within a single visual area, visual features that strongly or weakly activated individual neurons also had a high probability of modulating the activity of other neurons, suggesting a shared feature selectivity across the population that structures stimulus encoding. We show that this encoding strategy is conserved across species, present in both primary and lateral visual areas of mouse cortex. Dual-feature selectivity is consistent with recent anatomical evidence for feature-specific inhibitory connectivity, complementing the feature-detector principle through circuit mechanisms in which selective excitation and inhibition may together enhance the representational capacity of the neuronal population.

neuroscience↗

bioRxiv2025

Replication of SARS-CoV-2 Omicron lineages is defined by TMPRSS2 use in environments where ACE2 is complexed with solute carriers SLC6A19 and SLC6A20.

The Omicron variant of SARS-CoV-2 emerged in late 2021 and since then Omicron subvariants have continued to evolve and dominate globally. The viral S protein evolved towards highly efficient antibody evasion and replicative capacity in the upper respiratory tract resulting in high transmissibility. At the same time, the mutations acquired in the S protein diminish infection of the lung epithelium and pathogenic potential. The changing entry requirements for Omicron sub-lineages that lead to this shift in tropism remain poorly understood. We resolve the changing replication requirements of SARS-CoV-2 to be related to two distinct pools of ACE2. The first pool relates to ACE2s role in the renin angiotensin system (RAS) and this pool can complex with TMPRSS2 (RAS-ACE2). The second pool relates to ACE2s role as a protein solute carrier chaperone than cannot complex with TMPRSS2 (Chaperone ACE2). Here, we demonstrate that pre-Omicron lineages replicate in a TMPRSS2 dependent manner across both ACE2 pools, whilst Omicron lineages can only spread and replicate using chaperone ACE2. This provides a mechanistic basis for the evolving infectivity requirements of SARS-CoV-2 and furthermore provides approaches to track and monitor ACE2 utilizing coronaviruses. Graphical AbstractMechanistic basis for shift in SARS-CoV-2 tropism with the arrival of Omicron. A. Chaperone ACE2 is defined structurally as a heterodimer of dimers with a solute carrier protein-SLC6A19 or SLC6A20. Here this ACE2 structure can exist uncomplexed from TMPRSS2 and enables TMPRSS2 use by both pre-Omicron and Omicron lineages. B. Renin Angiotensin ACE2 is defined by ACE2 with an exposed collectrin-like domain (CLD), which enables binding of TMPRSS2 or ADAM-17. Here ACE2 can form a complex with TMPRSS2 in a manner that allows pre-Omicron but not Omicron lineages to utilize TMPRSS2 to facilitate infection. Here Omicron lineages are heavily attenuated as they cannot use TMPRSS2 to spread. C. to E. Based on single cell profiles, ACE2 can exist as a chaperone with SLC6A20 in C. the Nasal Cavity or D. primarily as RAS-ACE2 in the lung to respond to acute lung injury. E. The largest pool of ACE2 in our body resides within the small intestine on enterocytes and this further facilitates replication in this tissue by pre-Omicron and Omicron lineages. O_FIG O_LINKSMALLFIG WIDTH=198 HEIGHT=200 SRC="FIGDIR/small/663433v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@172aa47org.highwire.dtl.DTLVardef@109285eorg.highwire.dtl.DTLVardef@394f8corg.highwire.dtl.DTLVardef@958e31_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2025

Multimodal profiling of proinflammatory protease activity identifies caspase-1 as a target for lung cancer interception

The systemic inhibition of IL-1b, a key mediator of pulmonary inflammation, has been shown to reduce the incidence of lung cancer in patients in years following treatment, but knowledge gaps surrounding its activation and role in the tumor microenvironment are hindering approaches for cancer interception. We developed a suite of activity-based technologies to probe inflammation in early lung cancer and identified a translational target candidate. We designed probes sensitive to various IL-1b-activating proteases and applied them to a murine model of inflammatory lung cancer, Kras/Trp53-mutant with SIINFEKL expression (KPS). Our nanosensors revealed reduced cleavage of a caspase-1 reporter in the lungs of KPS mice treated with IL-1b antibody, as well as elevated caspase-1 expression and activity in naive tumor tissue sections, highlighting the importance of caspase-1 processing of IL-1b during cancer development. We conducted a pre-clinical trial of a novel combination intervention by administering both IL-1b blockade and caspase-1 inhibition shortly after tumor induction. Following treatment, we observed significant reduction in lung cancer formation, including complete ablation of tumor incidence in nearly 20% of KPS mice. Our approach to understand the interplay of protease activity and cytokine activation supports development of new strategies to mitigate inflammation and intercept lung cancer progression.

bioengineering↗

bioRxiv2025

Dopamine receptor 1 specific CRISPRa mice exhibit disrupted behaviors and striatal baseline cellular activity

The two main cell types in the striatum, dopamine receptor 1 and adenosine receptor 2a spiny projection neurons (D1-SPNs and A2A-SPNs), have distinct roles in regulating motor- and reward-related behaviors. Cre-selective CRISPR-dCas9 systems allow for cell-type specific, epigenomic-based manipulation of gene expression with gene-specific single guide RNAs (sgRNAs) and have potential to elucidate molecular mechanisms underlying striatal subtype mediated behaviors. Conditional transgenic Rosa26:LSL-dCas9-p300 mice were recently generated to allow for robust expression of dCas9-p300 expression with Cre-driven cell-type specificity. This system utilizes p300, a histone acetyltransferase which regulates gene expression by unwinding chromatin and making that region of the genome more accessible for transcription. Rosa26-LSL-dCas9-p300 mice were paired with Drd1-Cre and Ador2a-Cre mice to generate Drd1-Cre:dCas9-p300 and Ador2a-Cre:dCas9-p300 mouse lines and underwent behavioral phenotyping when sgRNAs were not present. Both Drd1-Cre:dCas9-p300 and Ador2a-Cre:dCas9-p300 have cell-type specific expression of spCas9 mRNA. Baseline behavioral assessments revealed that, under a sgRNA absent nontargeted state, Drd1-Cre:dCas9-p300 mice display repetitive spinning behavior, hyperlocomotion and enhanced acquisition of reward learning in comparison to all genotypic littermates. In contrast, Ador2a-Cre:dCas9-p300 do not exhibit any changes in behavior in comparison to their littermates. Electrophysiological recordings of dorsal striatum D1-SPNs revealed that Drd1-Cre:dCas9-p300 mice have increased input resistance and increased spontaneous excitatory postsynaptic current amplitude, together suggesting greater excitatory drive of D1-SPNs. Overall, these data demonstrate the necessity to validate CRISPR-dCas9 lines for research investigations. Additionally, the Drd1-Cre:dCas9-p300 line has the potential to be used to study underlying mechanisms of stereotypy and reward-learning. Significance StatementUsing CRISPR-based tools to identify cell-type specific epigenomic and transcriptional mechanisms in disease and behavior has high utility for the neuroscience field. Previous limitations related to implementation of CRISPR-editing systems in mice were thought to be overcome by the generation of transgenic mouse lines, including a novel Cre-dependent dCas9-p300 mouse line. Our data shows however that Drd1-Cre:dCas9-p300 mice, generated from breeding Drd1-Cre mice with the dCas9-p300 mice, have cellular and behavioral disruptions under a nontargeted sgRNA absent state. Overall, these data suggest caution in employing CRISPR-dCas9 systems, particularly transgenic mouse lines, for research investigations.

neuroscience↗

bioRxiv2025

Harnessing cell-encapsulated hydrogels to study astrocyte mechanoresponse in 4D

In glaucoma, the optic nerve head (ONH) is exposed to increased biomechanical strain, impacting the resident astrocytes that maintain neural homeostasis. During disease progression astrocytes exhibit morphologic and metabolic shifts; however, the specific impact of glaucoma-related biomechanical strains on astrocyte behavior remains poorly understood. To address this, we used our previously established 3D cell-encapsulated extracellular matrix (ECM) hydrogel to investigate ONH astrocyte cellular and transcriptomic responses to varying biomechanical strain levels over time. Murine ONH astrocyte were encapsulated within an ECM hydrogel made from photocrosslinkable collagen type I and hyaluronic acid, and subjected to 0, 3, or 10% cyclic compression for 4h and 24h. We found significant restructuring of cytoskeletal morphology, metabolic dysregulation, and astrocyte-mediated ECM modulation that were strain-, duration- and hydrogel subregion-dependent. These phenotypic alterations were associated with diverse transcriptional changes in genes related to cell cycle and morphology, inflammation, metabolism and matrix remodeling that were driven by compressive strain intensity and duration. Our work reveals the direct role of compressive strain in eliciting a complex astrocyte response, supports targeting mechanosensation to prevent these pathologic astrocyte responses, and establishes ECM-based hydrogels as a platform to test mechanisms driving astrocyte mechanodysfunction. Altogether, our study offers new insights into astrocyte responses to biomechanical insult and demonstrates the use of a tunable 3D ECM hydrogel for future mechanistic studies of neurodegeneration. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/660800v2_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@f355fborg.highwire.dtl.DTLVardef@191e096org.highwire.dtl.DTLVardef@2fae32org.highwire.dtl.DTLVardef@55913c_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

The Spatial Atlas of Human Anatomy (SAHA): A Multimodal Subcellular-Resolution Reference Across Human Organs

The Spatial Atlas of Human Anatomy (SAHA) represents the first multimodal, subcellular- resolution reference of healthy adult human tissues across multiple organ systems. Integrating spatial transcriptomics, proteomics, and histological features across over 15 million cells from more than 100 donors, SAHA maps conserved and organ-specific cellular niches in gastrointestinal and immune tissues. High-resolution profiling using CosMx SMI, 10x Xenium, RNAscope, GeoMx DSP, and single-nucleus RNA-seq reveals spatially organized cell states, rare adaptive immune populations, and tissue-specific cell-cell interactions and ligand-receptor pairs. Comparative analyses with colorectal cancer and inflammatory bowel disease demonstrate the power of SAHA to detect disease-associated spatial disruptions, including crypt dedifferentiation, perineural invasion, and therapy-resistant immune remodeling. All data are openly accessible through a FAIR-compliant interactive portal to support exploration, benchmarking, and machine learning model training. Through SAHA, we provide a foundational framework for spatial diagnostics and next-generation precision medicine grounded in a comprehensive human tissue atlas, enabling the development of context-aware models that simulate tissue behavior, decode complex pathologies, and accelerate therapeutic innovation at unprecedented scale.

systems biology↗

bioRxiv2025

Differential Sensitivity of Impedance Plethysmography andPhotoplethysmography Sensors to Temperature-Induced PeripheralVasoconstriction

Impedance plethysmography (IPG) and photoplethysmography (PPG) are non-invasive techniques for measuring blood volume changes. This study investigated the differential responses of IPG and PPG to temperature-mediated vasoconstriction induced by localized cooling. Twenty-one participants underwent control and treatment conditions, with real or fake ice cubes applied to the forearm. PPG signal amplitude significantly decreased with cooling (p < 0.001), indicating sensitivity to capillary blood flow changes. In contrast, IPG signal amplitude remained stable, suggesting it primarily reflects blood flow in larger/deeper vessels. Blood pressure remained stable, while heart rate decreased. These findings suggest IPG is less sensitive to capillary-level changes than PPG and may be more suitable for monitoring deeper blood flow. This study provides insights into the distinct sensitivities of IPG and PPG, with implications for wearable device development and cardiovascular monitoring.

physiology↗

bioRxiv2025

A Multimodal Spatial and Epigenomic Atlas of Human Adult Lung Topography

Developing high-resolution reference maps of disease-susceptible spatial niches is a critical step to mitigating the profound effects of lung disease. Here, we present an integrated multimodal single-nucleus human lung atlas (snHLA) profiling 746,047 nuclei from 49 mapped lung blocks spanning clinically relevant distal airways, alveoli, and interstitium across 11 healthy adults. Integrating snRNA-seq and SNARE-seq2, which co-assays chromatin accessibility and gene expression from the same nucleus, we resolved 70 molecularly distinct populations and captured 332,846 accessible chromatin regions, nominating new transcriptional regulators of human lung cell diversity. Spatial transcriptomics using MERFISH mapped 25 cell populations across 7 structural neighborhoods and multiplexed immunofluorescence localized cell subtypes and distal airway-defining protein markers, expanding and validating distinct lung structure-specific cell populations. This open access snHLA and companion Cell Type and Marker Gene Dictionary with anatomically aligned nomenclature delivers a foundational resource at an unprecedented resolution to interrogate the origins of lung pathophysiology.

cell biology↗

bioRxiv2025

Temporal coding carries more stable cortical visual representations than firing rate over time

The brains ability to stably represent recurring visual scenes is crucial for behavior. Previous studies have used slow dynamic (1-5 seconds) rate code measurements to study visual tuning, revealing varying degrees of gradual activity changes over time or "representational drifts." However, it remains unclear if there is an underlying neural code that maintains the encoding of information stable over time. In this study, we extracted structures in fast (tens of milliseconds) temporal responses and explored the role of such temporal codes in supporting the stability of visual representations. We tracked the spiking activity of the same visual cortical populations in male mice for 15 consecutive days using custom-developed, large-scale, ultraflexible electrode arrays. Across various types of stimuli, we found that neurons exhibited varying degrees of day-to-day stability in their firing rate-based tuning. The across day stability correlated with tuning reliability. Notably, accounting for spiking temporal dynamics increased single neuron tuning stability, especially for less reliable neurons. Temporal coding further improved population representation discriminability and decoding accuracy. The stability of temporal codes was more correlated with network functional connectivity than rate coding. These results show that temporal coding is crucial for stably encoding sensory stimuli, suggesting its significant role in ensuring consistent sensory experiences.

neuroscience↗

bioRxiv2025

DNA Methylation Dynamics of Dose-dependent Acute Exercise, Training Adaptation, and Detraining

Exercise and diet are direct physical contributors to human health, wellness, resilience, and performance1-5. Endurance and resistance training are known to improve healthspan through various biological processes such as mitochondrial function6-8, telomere maintenance9, and inflammaging10. Although several training prescriptions have been defined with specific merits 1,10-20, the long-term effects of these in terms of their molecular alterations have not yet been well explored. In this study, we focus on two combined endurance and resistance training programs: (1) traditional moderate-intensity continuous endurance and resistance exercise (TRAD) and (2) a variation of high-intensity interval training (HIIT) we refer to as high intensity tactical training (HITT), to assess the dynamics of DNA methylation (DNAm) in blood and muscle derived from males (N=23) and females (N=31), over a 12-week period of training followed by a 4-week period of detraining, sampled at pre-exercise and acute time points, totaling 528 samples. Due to its rapid responsiveness to stimuli and its stability, DNAm has been known to facilitate regulatory cascades that significantly affect various physiological processes and pathways. We find that several thousand differentially methylated regions (DMRs) associated with acute exercise in blood, many of which are shared across males and females. This trend is reversed when comparing the baseline (pre-exercise) time points or post-exercise timepoints at the untrained state with those at the post-conditioned state. Here, muscle shows majority of DNAm changes, with most of those being unique. We also find several hundred "memory" DMRs in muscle that maintain the gain or loss of methylation after four weeks of inactivity. Comparing phenotypic measurements, we find specific DMRs that correlate significantly with mitochondrial function and myofiber switching. Using machine learning, we select a subset of DMRs that are most characteristic of training modalities, sex and timepoint. Most of the DMRs are enriched in pathways associated with immune function, cell differentiation, and exercise adaptation. These findings reveal mechanisms by which exercise- and training-induced epigenetic changes alter immune surveillance, mitochondrial function, and inflammatory response, and underscore the relevance of epigenetic plasticity to health monitoring and wellness.

genomics↗

bioRxiv2025

Inhibition Of One-Carbon Metabolism In Ewing Sarcoma Results In Profound And Prolonged Growth Suppression Associated With Purine Depletion

Ewing sarcoma (EWS) is the second most common primary bone malignancy in adolescents and young adults. Patients who present with localized disease have experienced a steadily improving survival rate over the years, whereas those who present with metastatic disease have the same dismal prognosis as 30 years ago, with long term survival rates less than 20%, despite maximal intensification of chemotherapy. Thus, novel treatment approaches are a significant unmet clinical need. Targeting metabolic differences between EWS and normal cells offers a promising approach to improve outcomes for these patients. One-carbon metabolism utilizes serine and folate to generate glycine and tetrahydrofolate (THF)-bound one-carbon units required for de novo nucleotide biosynthesis. Elevated expression of several one-carbon metabolism genes is significantly associated with reduced survival in EWS patients. We show that both genetic and pharmacological inhibition of a key enzyme of the mitochondrial arm of the one-carbon metabolic pathway, serine hydroxymethyltransferase 2 (SHMT2), leads to substantial inhibition of EWS cell proliferation and colony-forming ability, and that this effect is primarily caused by depletion of glycine and one-carbon units required for synthesis of purine nucleotides. Inhibition of one-carbon metabolism at a different node, using the clinically relevant dihydrofolate reductase inhibitor Pralatrexate, similarly yields a profound growth inhibition, with depletion of thymidylate and purine nucleotides. Genetic depletion of SHMT2 dramatically impairs tumor growth in a xenograft model of EWS. Together, these data establish the upregulation of the one-carbon metabolism as a novel and targetable vulnerability of EWS cells, which can be exploited for therapy. Statement of SignificanceUsing both genetic and pharmacologic approaches, this study identifies Ewing sarcomas dependence on the mitochondrial arm, but not the cytoplasmic arm, of one-carbon metabolism as a targetable vulnerability that can be effectively harnessed for therapy.

cancer biology↗

bioRxiv2025

Mechanistic basis for protection against fatty liver disease by CIDEB loss-of-function mutations

Background & AimsSomatic and germline CIDEB mutations are associated with protection from chronic liver diseases. The mechanistic basis and whether CIDEB suppression would be an effective therapy against fatty liver disease remain unclear. Methods21 CIDEB somatic mutations were introduced into cells to assess functionality. In vivo screening was used to trace Cideb mutant clones in mice fed normal chow, western (WD), and choline-deficient, L-amino acid-defined, high-fat (CDA-HFD) diets. Constitutive and conditional Cideb knockout mice were generated to study Cideb in liver disease. Isotope tracing was used to evaluate fatty acid oxidation and de novo lipogenesis. Transcriptomics, lipidomics, and metabolic analyses were utilized to explore molecular mechanisms. Double knockout models (Cideb/Atgl and Cideb/Ppara) tested mechanisms underlying Cideb loss. ResultsMost CIDEB mutations showed that they impair function, and lineage-tracing showed that loss-of-function clones were positively selected with some, but not all fatty liver inducing diets. Cideb KO mice were protected from WD, CDA-HFD, and alcohol diets, but had the greatest impact on CDA-HFD induced liver disease. Hepatocyte-specific Cideb deletion could ameliorate disease after MASLD establishment, modeling the impact of therapeutic siRNAs. Cideb loss protected livers via increased {beta}-oxidation, specifically through ATGL and PPARa activation. ConclusionsCideb deletion is more protective in some types of fatty liver disease. {beta}-oxidation is an important component of the Cideb protective mechanism. CIDEB inhibition represents a promising approach, and somatic mutations in CIDEB might predict the patient populations that might benefit the most.

physiology↗

bioRxiv2025

Cumulative pregnancy and postnatal environmental exposures impact social behaviour in male mice associated with epigenetic, ribosomal, and immune dysregulation

Environmental exposures across critical developmental windows can significantly influence brain development and contribute to the pathogenesis of neurodevelopmental disorders (NDDs). Emerging clinical evidence suggests that cumulative environmental factors during early development result in more pronounced disease phenotypes in offspring. Here, we developed a triple-hit model using C57Bl/6JAusb mice to examine the cumulative effects of antenatal social stress, antenatal chronic high-fat diet consumption, and postnatal poly(I:C) exposure on neurodevelopmental outcomes in offspring. Male triple-hit offspring displayed autism-like social deficits and an overall increased susceptibility to neurodevelopmental behavioural alterations in adulthood compared to male non-stressed controls. Conversely, these behavioural changes were not observed amongst female triple-hit offspring. Single-cell RNA (scRNA) transcriptomic and bulk proteomic sequencing were performed in male triple-hit offspring across peripheral blood immune cells and brain tissue. scRNA sequencing in microglia, astrocytes, and oligodendrocytes revealed dysregulation in critical glial cell processes, ribosomal functions, and chromatin remodelling. Similar functional themes were observed across peripheral blood macrophages, neutrophils, and naive B cells - displaying immune and ribosomal dysregulation at the transcriptional level. Proteomics pathway enrichment analyses revealed significantly reduced protein abundance in ribosomal biogenesis, translation, and chromatin remodelling functions in both peripheral blood immune cells and brain tissue. We also observed synaptic dysfunction in the blood and brain proteome. Overall, using our unique triple-hit model, we demonstrate that multiple early life environmental exposures drive NDD-associated behaviours in a sex-specific manner; and is associated with overlapping central and peripheral molecular mechanisms that have clinical relevance to NDD pathogenesis.

developmental biology↗

bioRxiv2025

Balanced Permeability Index is a Strong Predictor of Intestinal Absorption and Oral Bioavailability for Heterobifunctional Ligand-Directed Degraders

Ligand-directed degraders (LDDs) are heterobifunctional molecules that degrade proteins by engaging the ubiquitin-protein ligase (E3) system. LDDs consist of a target-engaging moiety, an E3 ligase-binding moiety and a bridging linker. Due to their size and physicochemical complexity these molecules do not adhere to well-established rules of lead optimization. The optimization of passive permeability remains a key challenge to develop orally bioavailable LDDs. To overcome this challenge, in this study we demonstrate that the Balanced Permeability Index (BPI)--a new metric that combines size, polarity and lipophilicity--is highly predictive of oral bioavailability for LDDs. Here, we introduce an additional parameter--called smallest maximum intramo-lecular distance (SMID)--to the original BPI index to account for cross sectional area of LDDs, termed BPILDD. With this new parameter, BPILDD can differentiate oral bioavailability of LDDs in our dataset more effectively than polarity, lipophilicity, or size separately. In addition, BPILDD is also more effective at identifying orally bioavailable LDDs than some in vitro measurements of cell permeability that traditionally inform bioavailability. This finding opens the possibility of employing BPILDD for the design and optimization of orally bioavailable LDDs to improve their drug metabolism and pharmacokinetics properties.

pharmacology and toxicology↗

bioRxiv2025

Tuning antibody stability and function by rational designs of framework mutations

Artificial intelligence and machine learning models have been developed to engineer antibodies for specific recognition of antigens, however these approaches often focus on the antibody complementarity determining region (CDR) whilst ignoring the immunoglobulin framework (FW) which provides structural rigidity and support for the flexible CDR loops. Here we present an integrated computational-experimental workflow, combining static structure analyses, molecular dynamics simulations and in vitro physicochemical and functional assays to generate rational designs of FW mutations for modulating antibody stability and activity. We first showed that recent antibody-specific language models lacked insights in FW mutagenesis, in comparison to approaches which utilised antibody structure information. Using the widely-used breast cancer therapeutic trastuzumab as a use case, we designed stabilising mutants which were distal to the CDR and preserved the antibodys functionality to engage its cognate antigen (HER2) and induce antibody-dependent cellular cytotoxicity (ADCC). Interestingly, guided by local backbone motions predicted using molecular dynamics simulations, we designed a FW mutation on the trastuzumab light chain which retained antigen-binding effects but lost Fab-mediated and Fc-mediated effector functions. This highlighted effects of FW on immunological functions engendered in distal areas of the antibody, and the importance to consider attributes other than binding affinity when assessing antibody function. Our approach incorporates interdomain dynamics and distal effects between FW and the Fc domains, expands the scope of antibody engineering beyond the CDR, and underscores the importance of a holistic perspective that considers the entire antibody structure as a whole in optimising antibody stability, developability and function.

immunology↗

bioRxiv2025

Dynamic subcellular proteomics identifies novel regulators of adipocyte insulin action

Insulin acts on adipocytes to suppress lipolysis and increase glucose uptake to control whole-body glucose and lipid metabolism. Regulation of these processes by insulin signalling depends on changes in protein localisation. However, the extent of insulin-stimulated changes to the adipocyte spatial proteome, and the importance of these in the cellular insulin response, is unknown. Here, we use subcellular proteomics approaches to map acute insulin-stimulated protein relocalisation in adipocytes on a cell-wide scale. These data revealed extensive insulin-regulated protein redistribution, with hundreds of novel insulin-responsive proteins. These included the uncharacterised protein C3ORF18, which redistributed to the PM in response to insulin. Studies in C3ORF18-depleted adipocytes suggest this protein is required for maximal insulin signalling. Overall, our data highlight the scale of protein relocalisation in the adipocyte insulin response, and provide an accessible resource to inform further studies into how changes in protein localisation contribute to cellular insulin responses.

cell biology↗

bioRxiv2025

EVOFLOW-RNA: GENERATING AND REPRESENTINGNON-CODING RNA WITH A LANGUAGE MODEL

RNA plays a critical role across numerous biological functions. Recent advances in language modeling show promise with representing RNA, but the possibility of large-scale RNA design and optimization has not been fully explored. We propose EvoFlow-RNA, a bidirectional non-coding RNA language model leveraging a masked discrete diffusion model (MDM) formulation for both generative modeling and representation learning. EvoFlow-RNA bridges the gap between RNA sequence representation and design. It outperforms leading RNA models on three BEACON tasks critical to understanding RNA function, spanning from structure prediction to gene editing. For unconditional generation, it synthesizes diverse RNA sequences with native-like structural and binding properties. Additionally, EvoFlow-RNA can globally redesign aptamer sequences around preserved binding recognition sites with enhanced functionality. Our results demonstrate the effectiveness of EvoFlow-RNA in RNA modeling, highlighting the capability and potential of masked discrete diffusion for both recapitulating and enhancing existing RNAs.

synthetic biology↗