Biology subjects

Brown, C.

Publications and source records attributed to Brown, C..

At least 37 records · Page 2Linked to original sources

bioRxiv2026

Direct-to-Biology Enables Rapid Identification of Potent FBXO22 Degraders

Proteolysis Targeting Chimeras (PROTACs) are heterobifunctional molecules that bring a ubiquitin E3 ligase into proximity of a target protein to polyubiquitinate and degrade the target. PROTACs act catalytically, offering distinct advantages over conventional inhibitors and are the subject of intense study. The development of PROTACs involves extensive optimization of the chemical moiety linking two different protein-binding chemotypes, often requiring the synthesis, purification and testing of hundreds of PROTAC candidates. We used this approach to rapidly explore the landscape of targeted degradation of four different targets in parallel, combining and comparing a recently reported FBXO22-recruiting chemical warhead with warheads for the commonly used CRBN and VHL E3 ligases. Using a limited number of compounds (175 compounds in total) we observed no FBXO22-dependent degradation of these four targets. However, our libraries generated potent FBXO22 homo-PROTACs inducing self-degradation, as well as CRBN- and VHL-mediated degraders of FBXO22.

cell biology↗

bioRxiv2026

Benchmarking cell type deconvolution in spatial transcriptomics and application to cancer immunotherapy

Cell type deconvolution is essential for resolving multicellular composition in spatial transcriptomics data, yet the accuracy and consistency of current methods across biological contexts remain uncertain. We introduce a benchmarking framework using realistic simulations that incorporate spatial and transcriptional complexity. We find that a simple marker gene signature scoring approach performs competitively, often outperforming more complex models, particularly for rare cell types. We further apply this method to a new spatial transcriptomics dataset profiling the early response to a single dose of anti-PD1 checkpoint blockade immunotherapy in a mouse cancer model, revealing pronounced spatially localized immune and microenvironmental changes in both tumors and draining lymph nodes. Our findings have implications for the interpretation of many published spatial transcriptomics studies that rely on cell type deconvolution, and they provide a robust, interpretable strategy for future analyses, even in the absence of matched single-cell references. Highlights- Framework for realistic benchmarking of spatial deconvolution methods - Marker gene scoring performs robustly for rare cell types across diverse datasets - Single-dose anti-PD1 cancer immunotherapy triggers profound localized response

genomics↗

bioRxiv2025

Reverberation exacerbates effects of interruption on auditory spatial selective attention

Everyday listening requires focusing on one talker while ignoring competing sounds, a process challenged by reverberation and unexpected distractions. Here, we asked whether reverberation decreases effects of distractions by reducing the salience of new onsets, or compounds disruption by increasing task difficulty. Across five online experiments, participants recalled spatialized syllable streams presented with or without interrupters under pseudo-anechoic and reverberant conditions. Interrupters consistently impaired recall, especially the syllables following the interrupter. For the syllable immediately after the interruption, this effect was larger in reverberation than in anechoic conditions. These results demonstrate that distractions are especially disruptive in reverberant settings.

neuroscience↗

bioRxiv2025

New genomic resources for an invasive mite pest help guide the development of KASP markers for pesticide resistance screening

The redlegged earth mite, Halotydeus destructor, is an invasive pest of grain and pasture crops in Australia. Australian populations are increasingly developing resistance to broad-spectrum pesticides. Here, we assembled a chromosome-level genome for H. destructor, comprising nine chromosomes and totalling approximately 46 Mb. We also generated a genomic dataset from 190 populations collected across thousands of kilometres of its invasive Australian range and across four years (2018, 2021, 2022, and 2023), and used this dataset to develop KASP (Kompetitive allele-specific PCR) markers for identifying resistance mutations. We combined pooled whole-genome sequencing with resistance testing for organophosphate pesticides, chlorpyrifos and omethoate, and the pyrethroid pesticide bifenthrin. Target-site resistance to chlorpyrifos was strongly associated with the common ace G119S mutation. A second mutation, ace A201S, was also common, although its association with chlorpyrifos or omethoate resistance was less clear. Target-site resistance to bifenthrin was strongly associated with the common para L1024F mutation. We also detected several rare target-site mutations that likely contribute to resistance in specific populations, including F331Y in ace, and M918T, M918V, L925I, F1020S, and F1538I in para. Copy number variation in ace showed mixed associations with resistance, potentially reflecting the co-occurrence of variable gene copies and target-site mutations segregating within and among populations. Finally, we observed putative compositional and structural differences in genomic regions containing carboxylesterases with high ace-like homology, a group of genes that might be involved in organophosphate metabolism. This study advances the genomic and molecular resources available for H. destructor, and represents the most comprehensive investigation to date of resistance mechanisms and their spatiotemporal distribution in this invasive pest. Our data showed that pesticide resistance in H. destructor likely involves many genes. Nevertheless, target-site mutations are likely to remain key markers for rapid resistance diagnostics, for which our KASP markers will provide valuable tools.

genomics↗

bioRxiv2025

Streptococcus pneumoniae accessory capsular genes modulate fitness, pathogenicity and immune evasion

Globally, Streptococcus pneumoniae disproportionally affects children in resource-poor settings, older adults and people living with HIV. Frequently found as an asymptomatic colonizer of the nasopharynx, this versatile pathogen is a prominent cause of pneumonia, meningitis, bacteraemia and otitis media. Recently, a serotype 3 capsule variant (GPSC10-ST700) has expanded in Malawi with enhanced vaccine escape potential. Here, using a mutational and complementation approach, we show that loss of accessory capsular genes in GPSC10-ST700 contribute to increased opsonophagocytic resistance in this lineage. Although originally thought to be nonfunctional pseudogenes, we show that these genes modulate fitness and the global phosphoproteome in serotype 3 strains. These findings highlight that vaccine escape may be mediated through variations in the pneumococcal capsular locus that enhance fitness, pathogenicity and immune evasion, without capsule switching. IMPORTANCEPneumococcal polysaccharide-conjugate vaccines (PCV) target the polysaccharide capsule (CPS), which is a dominant virulence factor. However, current PCVs induce suboptimal protection against serotype 3 strains, which produce a thicker capsule that when released from the bacterial surface, interferes with antibody-mediated bacterial killing and protection. We recently described the clonal expansion of a sequence type (ST) 700-GPSC10 serotype 3 lineage in Malawi post-PCV13 introduction. This lineage is characterized by the absence of at least 6 genes in its cps locus and a distinct antimicrobial resistance (AMR) profile compared to other serotype 3 strains. Here we uncovered a functional role for the accessory capsular genes (acl) in serotype 3, previously considered to be pseudogenes, which modulate capsule production, shedding, serum tolerance, and bacterial fitness. By linking genotype to phenotype, our work provides new insights into the molecular basis of serotype 3 immune evasion, informing the design of more effective pneumococcal vaccines.

microbiology↗

bioRxiv2025

Runx/Cbfβ regulates the development of tolerogenic Thetis cells

Establishing immune tolerance to gut microbiota and food antigens upon first exposures during early life is essential to prevent inflammatory bowel diseases and food allergy and depends on induction of peripherally induced Ror{gamma}t expressing regulatory T (Ror{gamma}t+ pTreg) cells1, 2, 3, 4, 5, 6. Recent studies have identified a critical role for Ror{gamma}t expressing antigen-presenting cells (APC), Thetis cells (TCs), in peripheral regulatory T (pTreg) cell differentiation and tolerance to food and commensal microbes7, 8, 9, 10, 11. TCs encompass four distinct subsets, and a subset of TCs, TC IV induces pTreg differentiation, but the transcription factors that control their differentiation are not fully known. Here, using orthogonal genetic approaches to impair Runx/Cbf{beta} activity, we show that development of specific TCs subsets is regulated by Runx/Cbf{beta} transcriptional factor complexes. While attenuated Runx3 by germline mutations resulted in a severe reduction of all Ror{gamma}t+ APCs, mice lacking one of two Cbf{beta} splicing variants, Cbf{beta}2, exhibited a loss of TC II, III and TC IV subsets with associated loss of Ror{gamma}t+ pTreg cells. Conditional inactivation of Runx1 and Runx3 genes by CD11c-Cre led to a specific loss of TC III and TC IV subsets. Strikingly, CD11c-Cre driven transgenic Runx expression, particularly Runx1, led to enhanced TC IV differentiation and thus Ror{gamma}t+ pTreg cells. Furthermore, transgenic Cbf{beta}2 by CD11c-Cre recovered only TC IV subset with restoration of Ror{gamma}t+ pTreg in Cbf{beta}2-deficient mice. Collectively, our findings establish a critical pathway for TC IV differentiation and provide new insights into therapeutic interventions to promote Ror{gamma}t+ pTreg induction in autoimmune and inflammatory diseases.

immunology↗

bioRxiv2025

Phase separation of mycobacterial Rho factor is associated with acid stress

Rho is a bacterial transcription termination factor that contains additional regions in some species, including many mycobacteria. These additional regions are compositionally biased and are predicted to be intrinsically disordered and associated with Liquid-Liquid Phase Separation (LLPS). However, LLPS has only been experimentally demonstrated for Rho from Bacteroides thetaiotaomicron. Here, we extend a prior bioinformatic analysis of Rho from mycobacterial species, where Rho has additional regions that differ from Rho in B. thetaiotaomicron. We show that Mycolicibacterium smegmatis Rho forms in vitro and in vivo condensates by LLPS, and that LLPS depends upon the additional regions. In vitro, droplet formation for Rho from four mycobacterial species was modulated by the presence of RNA and was observed for the intrinsically disordered regions alone, suggesting that these regions are sufficient for droplet formation. Moreover, Rho phase separation in living M. smegmatis cells occurs during acid stress and may impact cell fitness. This phenotype is due to Rhos additional regions, since engineered mycobacterial Rho lacking the additional regions does not undergo LLPS. In summary, our results indicate that LLPS is a widespread mechanism in which mycobacterial Rho assembles into condensates, possibly terminating the transcription of different genes, which allows tolerance to stress conditions.

microbiology↗

bioRxiv2025

REM Sleep Misfires: Intruding Delta Waves Forecast Tau, Amyloid, and Forgetting in Aging

Rapid Eye Movement (REM) sleep degrades with age, and more severely in Alzheimers disease (AD). REM sleep comprises about twenty percent of adult sleep, alternates between phasic and tonic periods, and includes delta waves (1-4Hz) in two forms: fast sawtooth waves and slower, NREM-like waves, whose expression dynamically varies across REM periods. Yet, the functional relevance of these REM sleep delta waves remains unknown. Here, using two independent cohorts, we show that aging is associated with a shift from fast sawtooth to slow NREM-like delta waves, particularly during phasic REM sleep--a period typically marked by high cortical activation. Beyond chronological age, this shift is associated with amyloid-beta and tau burden, suggesting that AD pathology disrupts REM-specific oscillatory patterns. Furthermore, this shift in REM oscillations is linked to impaired overnight memory consolidation, independent of NREM sleep quality. Moreover, variation in ApoE alleles, a major genetic risk factor for AD, was independently associated with a reduction in fast sawtooth wave density, thereby linking a genetic predisposition for AD to these specific REM microstructural changes. These findings identify a novel signature of memory decline in aging and implicate REM sleep as a distinct vulnerable substrate through which AD pathology may impair brain function.

neuroscience↗

bioRxiv2025

Decoding the interconnected splicing patterns of hepatitis B virus and host using large language and deep learning models

Hepatitis B virus (HBV) infection causes approximately one million deaths annually and remains a major driver of hepatocellular carcinoma. Despite its compact 3.2-kb genome, HBV exhibits extensive alternative splicing. Functionally, HBV splice variants contribute to immune evasion and reduce the likelihood of achieving a functional cure. Here, we show that HBV splicing efficiency--quantified from 279 RNA-seq libraries of HBV-associated liver biopsies and cultured cells--correlates more strongly with disease progression than the overall proportion of spliced HBV RNAs, an emerging biomarker. All HBV splice sites are embedded within protein-coding regions, forming a genetic architecture distinct from typical host splice sites. To decode the sequence determinants of HBV splicing, we apply transformer-based and deep learning models to 4,707 HBV genomes. These models reveal that more highly used HBV splice sites are more conserved and share features with host splice sites that are less frequently used but still functional. This similarity likely reflects constraints imposed by HBVs compact genome, which must accommodate overlapping protein-coding regions. HBV may have evolved to exploit suboptimal but spliceable host-like motifs without disrupting its genetic architecture. Further analysis of splicing propensity across HBV genomes reveals genotype-specific patterns, indicating regulation by sequence context in a site- and genotype-dependent manner. HBV genotypes may have coevolved with their human hosts to fine-tune splicing through host-like features, supporting mechanisms of viral persistence and immune evasion. This study demonstrates the utility of AI in decoding viral splicing architectures and provides a framework for investigating co-transcriptional processes in other clinically important viruses. Impact StatementHepatitis B virus (HBV) is a major global health concern, causing 1.1 million deaths in 2022 and 1.2 million new infections each year. It is a leading cause of serious liver conditions, including cirrhosis and liver cancer. Although vaccines can prevent HBV infection, there is currently no cure. HBV produces different types of genetic messages (RNAs), including spliced versions that are processed by the host cells machinery. These spliced RNAs help the virus evade the immune system and make it harder for treatments to fully clear the infection. In this study, we analysed 279 HBV samples from liver tissues and lab-grown cells and found that the efficiency of host-mediated splicing of viral RNA reflects the severity of disease. Using advanced artificial intelligence tools, we mapped the splicing patterns in both the virus and human host, and investigated over 4,700 HBV genomes. We discovered that HBV splice sites resemble host splice sites that are used less frequently but remain functional, suggesting the virus has evolved RNA sequences that are compatible with the host cells splicing machinery while accommodating its compact genome. Insights into this viral adaptation may help researchers identify new biomarkers for disease severity and develop therapeutic strategies that disrupt the viruss ability to exploit the host cells machinery. Data SummaryThe raw RNA-sequencing (RNA-seq) libraries analysed in this study were previously published and are available in the Gene Expression Omnibus under accession number GSE155983. These libraries form part of a curated collection of 279 RNA-seq datasets derived from HBV-associated liver biopsy tissues and cultured cells. Further details and metadata are available in the associated GitHub repository: https://github.com/lcscs12345/HBV_splicing_paper_2025.

genomics↗

bioRxiv2025

A periplasmic protein complex mediates cell envelope integrity and intrinsic multidrug resistance in Mycobacterium tuberculosis

The intrinsic drug resistance of Mycobacterium tuberculosis (Mtb) is a major barrier to effective tuberculosis (TB) treatment, largely due to its complex, impermeable cell envelope. We identified a periplasmic protein complex comprising FecB and Rv3035 that is essential for maintaining envelope integrity and mediating intrinsic multidrug resistance in Mtb. FecB interacts with Rv3035, forming a stable heterodimer that associates with the cell envelope biosynthesis protein AftB. We report the structures of Rv3035 alone and in complex with FecB and identify critical residues for complex formation and function. Co-essentiality and genetic interaction analyses support a functional link between FecB, Rv3035 and AftB, an arabinofuranosyltransferase which synthesizes arabinogalactan and lipoarabinomannan. Loss of FecB or Rv3035 disrupted AftB-mediated arabinan synthesis, suggesting that these proteins support AftBs enzymatic activity. Importantly, FecB is required for Mtb virulence in mice, underscoring its physiological relevance. These findings highlight FecB, Rv3035 and AftB as promising therapeutic targets.

microbiology↗

bioRxiv2025

Study of combination CAR T-cell treatment for glioblastoma using mathematical modeling

Glioblastoma is a highly aggressive and difficult-to-treat brain cancer that resists conventional therapies. Recent advances in chimeric antigen receptor (CAR) T-cell therapy have shown promising potential for treating glioblastoma; however, achieving optimal efficacy remains challenging due to tumor antigen heterogeneity, the tumor microenvironment, and T-cell exhaustion. In this study, we developed a mathematical model of CAR T-cell therapy for glioblastoma to explore combinations of CAR T-cell treatments that take into account the spatial heterogeneity of antigen expression. Our hybrid model, created using the multicellular modeling platform PhysiCell, couples partial differential equations that describe the tumor microenvironment with agent-based models for glioblastoma and CAR T-cells. The model captures cell-to-cell interactions between the glioblastoma cells and CAR T-cells throughout treatment, focusing on three target antigens: IL-13R2, HER2, and EGFR. We analyze tumor antigen expression heterogeneity informed by expression patterns identified from human tissues and investigate patient-specific combination CAR T-cell treatment strategies. Our model demonstrates that an early intervention is the most effective approach, especially in glioblastoma tumors characterized by mixed antigen expression. However, in tissues with clustered antigen patterns, we find that sequential administration with specific CAR T-cell types can achieve efficacy comparable to simultaneous administration. In addition, spatially targeted delivery of CAR T-cells to specific tumor regions with matching antigen is an effective strategy as well. Our model provides a valuable platform for developing patient-specific CAR T-cell treatment plans with the potential to optimize scheduling and locations of CAR T-cell injections based on individual antigen expression profiles.

systems biology↗

bioRxiv2025

Altered Hepatic Metabolism in Down Syndrome

Trisomy 21 (T21) gives rise to Down syndrome (DS), the most commonly occurring chromosomal abnormality in humans. T21 affects nearly every organ and tissue system in the body, predisposing individuals with DS to congenital heart defects, autoimmunity, and Alzheimers disease, among other co-occurring conditions. Here, using multi-omic analysis of plasma from more than 400 people, we report broad metabolic changes in the population with DS typified by increased bile acid levels and protein signatures of liver dysfunction. In a mouse model of DS, we demonstrate conservation of perturbed bile acid metabolism accompanied by liver pathology. Bulk RNA-sequencing revealed widespread impacts of the Dp16 model on hepatic metabolism and inflammation, while single-cell transcriptomics highlighted cell types associated with these observations. Modulation of dietary fat profoundly impacted gene expression, bile acids, and liver pathology. Overall, these data represent evidence for altered hepatic metabolism in DS that could be modulated by diet.

genetics↗

bioRxiv2025

The effects of predator recovery and climate change on the long-term demography of a flagship herbivore

In the western United States, ecosystems are being reshaped from both bottom-up and top-down processes. Widespread carnivore recolonizations after 20th century extirpations have returned top-down pressures to ecosystems, and climate change is reshaping bottom-up forces through shifts in the timing and length of growing seasons, resulting in reduced forage quality and quantity. Large herbivores, such as elk (Cervus canadensis), function as flagship species for conservation and have potential for both top-down and bottom-up forces to shape demography as ecosystems change. To test the roles of top-down (puma density (Puma concolor)) and bottom-up (drought severity and elk density) processes in shaping large herbivore demography, and to tie those effects to population trajectories, we built an integrated population model (IPM) using 36 years of elk data spanning the recolonization of pumas and long-term climate change in eastern Oregon, USA. We tested the effects of top-down and bottom-up forces on elk recruitment, calf survival, and population growth. We also tested effects of bottom-up forces on elk pregnancy rates. Puma recolonization corresponded with declines in mean recruitment from 0.44 to 0.32 calves per female and in calf survival from 0.92 to 0.69, corresponding with a 4% drop in population growth. Drought severity increased over our study period, explaining a decline in mean recruitment from 0.39 to 0.32 calves per female, which corresponds to a 3% reduction in population growth. Drought severity likely acted on recruitment through reduced pregnancy rates. Lactating females showed lower pregnancy rates in drought conditions, while drought from fall breeding periods to the following fall had no effect on recruitment. While both top-down and bottom-up forces explained variation in elk demography over the course of our study, bottom-up forces are likely to be more influential on large herbivores moving forward. Predator populations stabilized following recovery, limiting the top-down contribution to annual variation in vital rates. Climate change-induced drought patterns, however, are accelerating during summer and fall throughout large areas of the interior western U.S. Thus, our results indicate the potential for increasingly strong bottom-up negative effects on herbivores whose performance depends on a narrow period of summer nutrition. Open Research StatementData are not yet provided. Many of the data from Starkey Experimental Forest and Range are already publicly available, and we are currently working to post data for this manuscript on the Forest Service Research Data Archive (FSRDA) linked to the rest of the data from Starkey. Upon acceptance, all data and the relevant code will be archived the FSRDA. Currently, the code for the paper is publicly available on Github at https://github.com/keloonam/starkey_ipm

ecology↗

bioRxiv2025

CellTune: An integrative software for accurate cell classification in spatial proteomics

Spatial proteomics measures multiple proteins in situ, capturing tissue complexity. However, cell classification in densely packed tissues remains challenging due to the lack of efficient classification algorithms, annotation tools, and high-quality labeled datasets to benchmark computational methods. We introduce CellTune, an integrated software for analysis of large spatial proteomics datasets, which streamlines precise cell classification through an optimized human-in-the-loop active learning workflow. It advances core capabilities across within a unified, intuitive, and code-free interface. To evaluate CellTune, we created CellTuneDepot, a resource of 40k manually-annotated cells and 3.5 million high-quality labeled cells across 60 cell types. CellTune outperforms alternative methods, achieving accuracy comparable to human performance while enabling increased classification resolution and discovery of novel cell types. Together, CellTune and CellTuneDepot provide researchers with a tool for state-of-the-art classification accuracy and resolution at scale to drive biological insights.

bioinformatics↗

bioRxiv2025

Visual lateralisation in female guppies demonstrates social conformity but is reduced when observing a live predator

Living in groups offers individuals a way of reducing their risk of predation. Visual lateralisation, characterised as an asymmetry in eye-use, may offer an additional advantage to group-living animals by enabling them to manage two concurrent visual tasks simultaneously. This could enhance multi-tasking efficiency by facilitating cohesion with group mates while monitoring for threats. In our study, we examined visual lateralisation of Trinidadian guppies (Poecilia reticulata) tested either alone or in groups, in either the presence or absence of a live predator, the blue acara (Andinoacara pulcher). We consistently observed low levels of visual lateralisation across all treatments. Contrary to our expectations, however, guppies exhibited significantly higher absolute lateralisation when tested alone in the absence of the predator compared to the other treatments. Moreover, a significant left-eye bias was observed when the predator was present, and the fish showed a right-eye bias when the predator was absent. Use of a repeated measures design and assessing individual and group ID as random effects provided evidence that both relative and absolute laterality were repeatable at the group level, but there was limited evidence for repeatability at the level of the individuals. This repeatability in lateralisation when tested as a group, but not when individual fish composing these groups were tested alone, suggests social conformity in lateralisation. Our results suggest that social processes may have a significant impact on within-population variation in lateralisation.

animal behavior and cognition↗

bioRxiv2025

Familial Alzheimers disease mutations impair neutrophil phagocytosis and release of immune modulators

Recent reports show that neutrophil activity plays a role in the cognitive decline associated with Alzheimers Disease (AD). There is evidence of altered functions in neutrophils isolated from AD patients. Whether these altered functions are inherent to the AD disease state is unknown. The goal of this study was to determine if neutrophil functions are altered in AD mice and if these changes occur only after symptoms appear. To address this hypothesis, we used a primary neuronal culture model, generated from 3xTg perinatal mice, since AD is considered a CNS disease. The 3xTg primary neuronal culture gradually increase the release of A{beta} (40 and 42) as the culture ages. To assess neutrophil functions, neutrophils isolated from young male/female mice (3-6 months of age) or aged male/female mice (16-18 months of age) of WT or 3xTg mice were exposed to 3xTg primary neuronal cultures. To assess phagocytosis, we characterized the effect of neutrophils on pathogenic amyloid beta (A{beta}) 42 levels. To assess the levels of immune modulators (cytokines, chemokines, growth factors, NETosis, and neutrophil granular content), culture media were assessed using Luminex multiplex assay. Our results show that neutrophils from young AD mice have impaired phagocytosis, as observed in a decreased ability to remove A{beta} and cellular debris in vitro. Neutrophils from young AD mice also increased release of pro-inflammatory cytokines, granule content, and NETs in 3xTg primary neuronal cultures. Interestingly, neutrophils from aged 3xTg mice decreased A{beta} levels in culture and expression of proinflammatory cytokines when compared to neutrophils from aged WT mice. These neutrophils increased their release of granule content and NETs in 3xTg primary neuronal cultures. These data show that in AD neutrophil function is altered both prodromal (young mice) and diseased (old mice) stage.

immunology↗

bioRxiv2025

Overcoming myeloid-driven resistance to CAR T therapy by targeting SPP1

Chimeric antigen receptor (CAR) T cell therapy faces notable limitations in treatment of solid tumors. The suppressive tumor microenvironment (TME), characterized by complex interactions among immune and stromal cells, is gaining recognition in conferring resistance to CAR T cell therapy. Despite the abundance and diversity of macrophages in the TME, their intricate involvement in modulating responses to CAR T cell therapies remains poorly understood. Here, we conducted single-cell RNA sequencing (scRNA-seq) on tumors from 41 glioma patients undergoing IL13R2-targeted CAR T cell therapy, identifying elevated suppressive SPP1 signatures predominantly in macrophages from patients who were resistant to treatment. Further integrative scRNA-seq analysis of high-grade gliomas as well as an interferon-signaling deficient syngeneic mouse model--both resistant to CAR T therapy--demonstrated the role of congruent suppressive pathways in mediating resistance to CAR T cells and a dominant role for SPP1+ macrophages. SPP1 blockade with an anti-SPP1 antibody abrogates the suppressive TME effects and substantially prolongs survival in IFN signaling-deficient and glioma syngeneic mouse models resistant to CAR T cell therapy. These findings illuminate the role of SPP1+ macrophages in fueling a suppressive TME and driving solid tumor resistance to CAR cell therapies. Targeting SPP1 may serve as a universal strategy to reprogram immune dynamics in solid tumors mitigating resistance to CAR T cell therapies.

cancer biology↗

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↗