Biology subjects

Fox, E.

Publications and source records attributed to Fox, E..

10 recordsLinked to original sources

bioRxiv2026

CMS4-focused multi-omic integration enhances antigen target identification in colorectal cancer

Colorectal cancer (CRC) remains a major cause of cancer mortality, with limited options for poor-prognosis subtypes such as CMS4. Antigen-targeted therapies show promise but tend to fail due to inadequate target selection and insufficient patient stratification. Effective prioritization requires large harmonized data capturing CRC heterogeneity - a resource that is currently lacking. To address this need, we built a harmonized multi-omic CRC knowledge base and applied a scalable discovery pipeline to identify antigen targets specifically associated with CMS4 biology and with strong translational potential. We constructed a harmonized CRC atlas by integrating 79 transcriptomics datasets (5,033 tumors, 161 normal samples) using proprietary AI-powered data scouting, integration, and curation technologies. Consensus Molecular Subtypes (CMS) were inferred to capture CMS4-specific expression patterns and this atlas was then combined with 3 bulk RNA-seq reference datasets, 2 single-cell atlases, and 8 protein annotation databases to form a unified multi-omic CRC knowledge base of unmatched scale. From this integrated system, we identified genes differentially expressed in CMS4 patients encoding druggable cell-surface proteins, which we then prioritized using a weighted efficacy- and safety-based scoring model. We identified 236 CMS4-enriched candidates, including 124 not detectable at the CRC-wide level, demonstrating the added resolution gained through subtype stratification. Recovery of known investigational CRC (LGR5, MET, TACSTD2) and CMS4-associated targets of clinical emerging interest (PDGFRB, ALK5/TGFBR1, FAP) support the biological and methodological validity of our approach. Benchmarking against thresholds from FDA-approved pan-cancer targets and terminated trials identified 32 candidates with comparable or superior therapeutic profiles. Among these, 11 were enriched for CMS4-defining pathways, including epithelial-mesenchymal transition, angiogenesis, and stromal invasion, and 5 showed strong profile similarity to established CRC and CMS4 benchmarks. After extensive data exploration, particularly promising candidates were shortlisted for further validation. This work shows that CMS4-focused molecular stratification, when combined with an unprecedentedly large harmonized multi-omic knowledge base, yields a refined set of antigen candidates with enhanced specificity, safety, and biological relevance. The prioritized targets illustrate the power of subtype-resolved discovery to uncover clinically actionable insights. Our pipelines modular design can extend to other tumor contexts, offering a robust foundation for accelerating targeted therapy development.

cancer biology↗

bioRxiv2025

The renin angiotensin system in cognitive flexibility: Effects of single-dose losartan on task-switching in healthy volunteers

IntroductionCognitive flexibility, defined as the ability to shift mental strategies when environmental demands change, may influence psychological well-being and psychiatric treatment outcomes. Emerging research increasingly implicates the renin angiotensin system (RAS) in cognition, yet its role in cognitive flexibility is inadequately understood. MethodsWe conducted a double-masked trial with N=60 healthy adults aged 18-50. Participants were randomly assigned to receive a single 50 mg dose of losartan, an angiotensin II receptor blocker, or placebo. Following a waiting period, participants completed a task-switching paradigm as a measure of cognitive flexibility, which required dynamically categorizing arrows based on location or direction. Statistical analyses were conducted to determine group (losartan vs. placebo), task (location vs. direction), and switch (switching vs. repeating a task) effects on reaction time (RT) and accuracy independently. A composite bin score was also computed, integrating both RT and accuracy data. ResultsAll participants displayed the expected switch costs. While participants on losartan exhibited numerically superior RTs and accuracy, losartan did not significantly improve task-switching performance relative to placebo. This was confirmed by the bin score analysis, which indicated no benefit of losartan when RT and accuracy were considered together. An exploratory probe suggested that losartan improved accuracy on direction switch trials, indicating a potential drug effect under specific cognitive conditions. ConclusionOverall, we found no significant benefits of losartan on task-switching in healthy adults. Our results may be explained by ceiling effects in our sample, which could have hindered the detection of group differences, or possibly reflect a negligible role for the RAS in cognitive flexibility.

neuroscience↗

bioRxiv2025

T cell protrusions enable fast, localised initiation of CAR signalling

Actin-rich protrusions densely cover the surface of T cells and are well characterised for their role in cell migration. However, recent studies have uncovered their role in antigen surveillance and immune signalling initiation. To investigate how membrane protrusions initiate and contribute to signalling, from the first cell-cell contact to immunological synapse formation, we performed dynamic imaging experiments of endogenously tagged signalling proteins in T cells. To quantitatively capture the early dynamics of cell-cell interactions, we employed HER2-CAR-expressing T cells targeting HER2+breast cancer cells. By harnessing live-cell imaging and super-resolution stimulated emission depletion (STED) microscopy we were able to capture topological membrane changes and their correlation with mesoscale protein rearrangements over time. Our findings indicate that, prior to activation, key molecular players in T cell activation, including the kinase Lck, the phosphatase CD45 and the adaptor LAT, as well as the exogenously expressed CAR, lack any enrichment in actin-rich protrusions. However, upon initial contact of the T cell with the target cell, a dynamic and fast rearrangement of the surface receptors, phosphatases, and kinases occurs within the protrusions, ensuring a rapid and effective initiation of the immune signalling cascade. The rapid clustering of the HER2-CAR occurs preferentially within protrusions rather than flat membrane regions and is accompanied by enhanced recruitment of the kinase ZAP-70 and LAT. While the localisation of the kinase Lck remained unchanged, protrusion-cell contacts trigger a pronounced exclusion of the phosphatase CD45, an effect observed both with and without the cytosolic signalling domain of the CAR. Overall, the signalling machinery rearranged more rapidly and efficiently at contacts mediated by protrusive structures compared to non-protrusive regions. Together, our data provide a quantitative framework illustrating how signalling proteins are dynamically reorganised to facilitate CAR-mediated activation within these specialised structures.

cell biology↗

bioRxiv2024

Extending the Coding-variant Allelic Series Test to Summary Statistics

Genes for which a dose-response relationship exists between mutational severity and phenotypic impact make for logical therapeutic targets, as the effects of pharmacological modulation can be anticipated from the natural variation present in a population. We refer to genes where such a dose-response relationship exists as harboring allelic series, and have introduced the rare coding-variant allelic series test (COAST) for their identification. The original COAST required access to individual-level data. However, such data are often unavailable due to privacy concerns or logistical challenges. Meanwhile, single-variant summary statistics of the type produced by genome-wide association studies are plentiful. Here we introduce COAST-SS, an extension of COAST that accepts summary statistics as input, namely the per-variant effect sizes and standard errors, along with estimates of the minor allele frequency and local linkage disequilibrium (LD). As a running example, we consider identifying allelic series for circulating lipid traits, drawing on data from the UK Biobank, Million Veterans Program, and Trans-Omics of Precision Medicine Program. Through extensive analyses of real and simulated data, we demonstrate that COAST-SS provides p-values effectively equivalent to those from the original COAST. Interestingly, we find that when LD is low, as is expected among rare variants, COAST-SS is robust to misspecification of the LD matrix, providing valid inference even when the LD matrix is set to the identity matrix. We explore several strategies for annotating the pathogenicity of variants supplied to COAST-SS, finding that they often yield similar power for detecting candidate allelic series. Lastly, we employ COAST-SS to screen for lipid trait allelic series in a meta-analyzed cohort of up to 840K subjects. COAST-SS has been incorporated into the publicly available AllelicSeries R package.

genetics↗

bioRxiv2023

Multi-functional ARF1 compartments serve as a hub for short-range cargo transfer to endosomes

Cellular membrane homeostasis is maintained via a tightly regulated membrane and cargo flow between organelles of the endocytic and secretory pathways. Adaptor protein complexes (APs), which are recruited to membranes by the small GTPase ARF1, facilitate cargo selection and incorporation into trafficking intermediates. According to the classical model, small vesicles would facilitate bi-directional long-range transport between the Golgi, endosomes and plasma membrane. Here we revisit the intracellular organization of the vesicular transport machinery using a combination of CRISPR-Cas9 gene editing, live-cell high temporal (fast-confocal) or spatial (stimulated emission depletion (STED)) microscopy as well as correlative light and electron microscopy. We characterize novel tubulo-vesicular ARF1 compartments that harbor clathrin and different APs. Our findings reveal two functionally different classes of ARF1 compartments, each decorated by a different combination of APs. Perinuclear ARF1 compartments facilitate Golgi export of secretory cargo, while peripheral ARF1 compartments are involved in endocytic recycling downstream of early endosomes. Contrary to the classical model of long-range vesicle shuttling, we observe that ARF1 compartments shed ARF1 and mature into recycling endosomes. This maturation process is impaired in the absence of AP-1 and results in trafficking defects. Collectively, these data highlight a crucial role for ARF1 compartments in post-Golgi sorting.

cell biology↗

bioRxiv2023

CRISPR/Cas9 gene editing to generate Drosophila LexA lines in secondary school classes

Genome editing in vivo with CRISPR/Cas9 generates powerful tools to study gene regulation and function. We developed CRISPR-based methods that permitted secondary school student scientists to convert Drosophila GAL4 lines to LexA lines. Our novel curricula implement a new donor strain optimizing Homology-assisted CRISPR knock-in (HACK) that simplifies screening using light microscopy. Successful curricula adoption by a consortium of schools led to the generation and characterization of 16 novel LexA lines. This includes extensive comparative tissue expression analysis between the parental Gal4 and derived LexA lines. From this collaboration, we established a workflow to systematically generate LexA lines from frequently-used GAL4 lines. Modular courses developed from this effort can be tailored to specific secondary school scheduling needs, and serve as a template for science educators to innovate courses and instructional goals. Our unique collaborations highlight that resources and expertise harnessed by university-based research laboratories can transform experiential science instruction in secondary schools while addressing research needs for the community of science.

genetics↗

bioRxiv2023

Efficient methods for target gene manipulation in haematopoietic stem cell derived human neutrophils.

Neutrophils are the most abundant leukocyte in humans and the principal effectors of the innate immue response. Genetic modification of human neutrophils is challenging due to their short lifespan and tendency to activate in response to even minor perturbation. However, genetic manipulation of haematopoietic progenitor cells and subsequent directed differentation into neutrophils represents a potential avenue to study the contributions of individual genes and pathways to human neutrophil function. Here we present a method of directed granulocytic CD34+ progenitor differentiation into neutrophils capable of key functions such as priming and neutrophil extracellular trap (NET) formation. We further show that differentiating progenitors can be efficiently and stably modified by lentiviral gene delivery and Cas9-gRNP nucleofection to produce potent and activation-free gene knockdown in mature neutrophils, thereby providing new tools for understanding the contribution of neutrophils to health and disease. Using this model we have shown that, contrary to previous reports, CD11b is not required for phagocytosis of serum-opsonised bacterial particles.

cell biology↗

bioRxiv2022

Gene editing and super-resolution microscopy reveal multiple distinct roles for ARF GTPases in cellular membrane organization

ADP-ribosylation factor (ARF) GTPases are major regulators of cellular membrane homeostasis. High sequence similarity and multiple, possible redundant functions of the five human ARFs make investigating their function a challenging task. To shed light on the roles of the different Golgi-localized ARF members in membrane trafficking, we generated CRISPR-Cas9 knock ins (KIs) of type I (ARF1 and ARF3) and type II ARFs (ARF4 and ARF5) and mapped their nanoscale localization with stimulated emission depletion (STED) super-resolution microscopy. We find ARF1, ARF4 and ARF5 on segregated nano-domains on the cis-Golgi and ER-Golgi intermediate compartments (ERGIC), revealing distinct roles in COPI recruitment on early secretory membranes. Interestingly, ARF4 and ARF5 define Golgi-tethered ERGIC elements decorated by COPI and devoid of ARF1. Differential localization of ARF1 and ARF4 on distal ERGICs suggests the presence of functionally different classes of intermediate compartments that could regulate bi-directional transport between the ER and the Golgi. Furthermore, ARF1 and ARF3 localize to segregated nano-domains on the trans-Golgi network (TGN) and TGN-derived post-Golgi tubules, strengthening the idea of distinct roles in post-Golgi sorting. This work provides the first map of the nanoscale organization of human ARF GTPases on cellular membranes and sets the stage to dissect their numerous cellular roles.

cell biology↗

bioRxiv2021

Transgenic Drosophila lines for LexA-dependent gene and growth regulation

Conditional expression of short hairpin RNAs (shRNAs) with binary genetic systems is an indispensable tool for studying gene function. Addressing mechanisms underlying cell-cell communication in vivo benefits from simultaneous use of two independent gene expression systems. To complement the abundance of existing Gal4/UAS-based resources in Drosophila, we and others have developed LexA/LexAop-based genetic tools. Here, we describe experimental and pedagogical advances that promote the efficient conversion of Drosophila Gal4 lines to LexA lines, and the generation of LexAop-shRNA lines to suppress gene function. We developed a CRISPR/Cas9-based knock-in system to replace Gal4 coding sequences with LexA, and a LexAop-based shRNA expression vector to achieve shRNA-mediated gene silencing. We demonstrate the use of these approaches to achieve targeted genetic loss-of-function in multiple tissues. We also detail our development of secondary school curricula that enable students to create transgenic flies, thereby magnifying the production of well-characterized LexA/LexAop lines for the scientific community. The genetic tools and teaching methods presented here provide LexA/LexAop resources that complement existing resources to study intercellular communication coordinating metazoan physiology and development.

genetics↗

bioRxiv2018

Microcapsuled entomopathogenic fungi against fire ants

A new microencapsulation method of Metarhizium anisopliae based on gelatin (GE) and gum arabic (GA) is presented. Conditions to produce spheres 35-50 m relied on 1% wall material, GE:GA ratio 1:1-1:2, core:wall ratio 1:1, under pH 4.0, 35-50{degrees}C and 500-700 r/min agitation. Microcapsulation provided protection against UV and longer shelf life as compared to unencapsulated conidia. The obtained preparation proved active against the red imported fire ant Solenopsis invicta.

microbiology↗