Biology subjects

Goodspeed, A.

Publications and source records attributed to Goodspeed, A..

14 recordsLinked to original sources

bioRxiv2026

Cytokine signaling regulates multiple stages of gammaherpesvirus infection in myeloid cells

The gammaherpesviruses, including Epstein-Barr Virus and Kaposis Sarcoma-associated Herpesvirus, establish lifelong infections by maintaining a balance between lytic and latent infection states. How this balance is regulated in myeloid cells, an important but understudied cell type that can achieve both infection states, remains unclear. Using the murine gammaherpesvirus 68 (MHV68) model, we show that IFNg; and IL-4 reciprocally regulate lytic infection in macrophages in a time- and JAK/STAT signaling dependent-manner, with basal JAK/STAT signaling further restricting lytic infection. Using a combination of MHV68 reporter viruses, cellular and molecular techniques, we demonstrate that IFNg blocks lytic replication through at least two distinct mechanisms, restricting delivery to or stability of the viral genome in the nucleus and potently restricting lytic cycle progression downstream of the immediate-early viral transactivator, RTA. These findings define specific points during lytic infection where IFNg; to enforce restricted, latent-like infection in myeloid cells.

immunology↗

bioRxiv2026

Estradiol Reprograms Microglia to Create an Immune-Suppressed Niche Permissive to Breast Cancer Brain Metastasis

BackgroundYoung age is an independent risk factor for the development of breast cancer brain metastases (BM). Prior work showed that 17{beta}-estradiol (E2), the predominant premenopausal hormone, promotes BM of tumors intrinsically unresponsive to E2, in part through modulating estrogen receptor-alpha expressing (ER) glial cells. However, how E2 reshapes the brain tumor microenvironment (TME), particularly microglia-mediated immunity, and its impact to BM progression remains unclear. MethodsscRNA sequencing and multiparametric flow cytometry were used to define the impact of E2 and E2-suppression on brain immune-cell populations across different stages of BM progression using spontaneous and experimental models of BM. Depletion of microglia and T cell co-cultures were used to study microglias role in E2-induced BM. The effects of E2-suppression alone or in combination with whole brain radiotherapy were tested in preclinical models mimicking late-stage BM. ResultsE2 repressed immune surveillance and immune activation programs in microglia from early to late stages of brain metastatic progression, suppressing recruitment of effector immune cells to BM. Estrogen suppression, in turn reactivated anti-tumoral signaling in microglia and increased recruitment of effector immune cells to the brain. Microglia from E2-stimulated BM-bearing mice showed decreased ability to induce interferon cytotoxic function and expansion of activated T cells. Conversely, E2-suppression reactivated an effective anti-tumoral response and synergized with RT to significantly decrease BM progression. ConclusionThese findings reveal a previously unrecognized mechanism by which E2 accelerates BC-BM progression through microglial immunosuppression and support evaluation of endocrine therapies as adjunct treatments for ER- brain metastases. Importance of the StudyStandard of care for BM includes stereotactic radiosurgery (SRS) alone or in combination with surgery, systemic chemotherapy or targeted therapies. Our studies show that ovariectomy (which eliminates ovarian E2) and aromatase inhibitors (AIs, which eliminate peripheral E2 synthesis) reduce progression of BM when used in combination with WBRT and in immuno-competent models. We demonstrate E2 promotes an immunosuppressive brain microenvironment from early stages of metastatic progression, in part through modulation of myeloid cells and repression of recruitment of effector T cells to the brain. Thus, these studies suggest that FDA-approved E2-depletion therapies (aromatase inhibitors and selective-estrogen modulators) could be used in combination with brain irradiation to decrease BM progression.

cancer biology↗

bioRxiv2025

HUMAN FETAL CIRCULATING FACTORS FROM PREGNANCIES COMPLICATED BY MATERNAL OBESITY INDUCE HYPERTROPHY IN NEONATAL RAT CARDIOMYOCYTES

AimObesity in pregnant women increases offspring cardiovascular risk and causes fetal cardiac dysfunction. The underpinning mechanisms remain unclear. We hypothesised that circulating factors in serum from fetuses of women with obesity induce pathological cardiomyocyte hypertrophy. MethodsPregnant women with obesity or healthy weight were recruited at term and provided umbilical cord serum and placentas, which were used for isolation of primary trophoblast cells. Primary cardiomyocytes were isolated from neonatal rats. ResultsCompared to serum-free medium, supplementing cardiomyocytes with umbilical cord serum increased their mRNA expression of atrial natriuretic factor (Anf) and brain natriuretic peptide (Bnp), hallmarks of pathological hypertrophy, but did not alter their size. Cord serum from women with obesity further upregulated cardiomyocyte Anf and Bnp, and increased the ratio of beta- to alpha-myosin heavy chain expression (Myh7:Myh6), compared to cord serum from healthy weight women. This effect was prevented by treating the cord serum with heat- freeze cycling and DNase or RNase digestion. Conditioned medium from trophoblast cells from women with obesity also increased cardiomyocyte Anf, Bnp and Myh7:Myh6 expression. MicroRNAs miR-142 and miR-17, which are associated with cardiac function, were increased in abundance in extracellular vesicles isolated from cord serum from women with obesity. However, miR-142-3p, miR-142-5p and miR-17-5p did not increase Anf, Bnp or Myh7:Myh6 expression when they were transfected into cardiomyocytes. ConclusionThe results show that human fetal circulating and placenta-derived factors induce hallmarks of pathological hypertrophy in cardiomyocytes and may mediate cardiac dysfunction in children of women with obesity.

physiology↗

bioRxiv2025

Tumor-associated stroma shapes the spatial tumor immune microenvironment of primary Ewing sarcomas

To date, few studies have detailed the tumor microenvironment (TME) of Ewing sarcoma (EwS). The TME has a vital role in cancer survival and progression with implications in drug resistance and immune escape. By performing spatially resolved transcriptomic analysis of primary treatment-naive EwS samples, we discovered greater stromal enrichment in localized EwS tumors compared to metastatic EwS tumors. Through spatial ligand-receptor analysis, we show that the stromal enriched regions harbor unique extracellular matrix related cytokines, immune recruitment and proinflammatory microenvironmental signals, implying EwS stroma may play an anti-tumorigenic role by acting as an immune recruitment center. All EwS tumors expressed pro-tumorigenic MIF-CD74 immune signaling, suggesting a potential immune-evasive mechanism and immunotherapy target. Our findings provide insight into tumor cell/stromal cell interactions in EwS and serve as a valuable resource for further investigations in the tumor immune microenvironment of EwS.

cancer biology↗

bioRxiv2024

Inflammation promotes aging-associated oncogenesis in the lung.

BackgroundLung cancer is the leading cause of cancer death in the world. While cigarette smoking is the major preventable factor for cancers in general and lung cancer in particular, old age is also a major risk factor. Aging-related chronic, low-level inflammation, termed inflammaging, has been widely documented; however, it remains unclear how inflammaging contributes to increased lung cancer incidence. AimTo establish connections between aging-associated changes in the lungs and cancer risk. MethodsWe analyzed public databases of gene expression for normal and cancerous human lungs and used mouse models to understand which changes were dependent on inflammation, as well as to assess the impact on oncogenesis. ResultsAnalyses of GTEx and TCGA databases comparing gene expression profiles from normal lungs, lung adenocarcinoma, lung squamous cell carcinoma of subjects across age groups revealed upregulated pathways such as inflammatory response, TNFA signaling via NF{kappa}B, and interferon-gamma response. Similar pathways were identified comparing the gene expression profiles of young and old mouse lungs. Transgenic expression of alpha 1 antitrypsin (AAT) partially reverses increases in markers of aging-associated inflammation and immune deregulation. Using an orthotopic model of lung cancer using cells derived from EML4-ALK fusion-induced adenomas, we demonstrated an increased tumor outgrowth in lungs of old mice while NLRP3 knockout in old mice decreased tumor volumes, suggesting that inflammation contributes to increased lung cancer development in aging organisms. ConclusionsThese studies reveal how expression of an anti-inflammatory mediator (AAT) can reduce some but not all aging-associated changes in mRNA and protein expression in the lungs. We further show that aging is associated with increased tumor outgrowth in the lungs, which may relate to an increased inflammatory microenvironment.

cancer biology↗

bioRxiv2024

Single cell RNA-sequencing of Ewing sarcoma tumors demonstrates transcriptional heterogeneity and clonal evolution.

Ewing sarcoma is the second most common bone cancer in children, accounting for 2% of pediatric cancer diagnoses. Patients who present with metastatic disease at the time of diagnosis have a dismal prognosis, compared to the >70% 5-year survival of those with localized disease. Here, we utilized single cell RNA-sequencing to characterize the transcriptional landscape of primary Ewing sarcoma tumors and surrounding tumor microenvironment (TME). Copy-number analysis identified subclonal evolution within patients prior to treatment. Primary tumor samples demonstrate a heterogenous transcriptional landscape with several conserved gene expression programs, including those composed of genes related to proliferation and EWS targets. Single cell RNA-sequencing and immunofluorescence of circulating tumor cells at the time of diagnosis identified TSPAN8 as a novel therapeutic target.

cancer biology↗

bioRxiv2023

Immunosuppressive tumor microenvironment of osteosarcoma

Osteosarcoma is the most common malignant bone tumor in children, characterized by a high degree of genomic instability, resulting in copy-number alterations and genomic rearrangements without disease-defining recurrent mutations. Clinical trials based on molecular characterization have failed to find new effective therapies or improve outcomes over the last 40 years. To better understand the immune microenvironment of osteosarcoma, we performed single-cell RNA sequencing on six tumor biopsy samples, combined with a previously-published cohort of six samples. Additional osteosarcoma samples were profiled using spatial transcriptomics for validation of discovered subtypes and to add spatial context. Analysis revealed immunosuppressive cells, including myeloid-derived suppressor cells (MDSCs), regulatory and exhausted T-cells, and LAMP3+ dendritic cells. Using cell-cell communication modeling, we identified robust interactions between MDSCs and other cells, leading to NF-{kappa}B upregulation and an immunosuppressive microenvironment, as well as interactions involving regulatory T-cells and osteosarcoma cells that promoted tumor progression and a proangiogenic niche. Statement of SignificanceOsteosarcoma patient survival has remained stagnant for several decades due to lack of successful therapy for high-risk patients, including those with metastatic disease. Identifying novel therapeutics including immunotherapies is of great clinical importance. Our study highlights several important immunosuppressive mechanisms within osteosarcoma that should be considered when developing future immunotherapies.

cancer biology↗

bioRxiv2023

Murine Gammaherpesvirus 68 Efficiently Infects Myeloid Cells Resulting In An Atypical, Restricted Form Of Infection

The gammaherpesviruses establish a lifelong infection, with the cellular outcome of infection intimately regulated by cell type. Though myeloid cells are an early infection target, infection results in widely divergent outcomes without a clear explanation. Here we use murine gammaherpesvirus 68 to demonstrate that macrophages are readily infectable, resulting in three divergent infection outcomes dictated by viral and host factors. Infection in vivo and in a cell culture model results in a high frequency of viable cells characterized by restricted viral transcription and unexpected transcription of the ORF75 locus, with rare cells initiating but failing to complete lytic replication. Restricted infection can be fully converted to productive lytic replication by pre-treatment with the host cytokine IL-4. Finally, infection with a high viral dose also results in productive infection, but triggers increased cell death. These studies demonstrate stepwise regulation of virus infection and identify virus and host factors that dictate divergent outcomes in a single cell type.

microbiology↗

bioRxiv2022

Hematopoietic stem cells are a long-term reservoir for trained immunity in a mouse model of chronic autoimmune disease

Trained immunity (TI) has been speculated to serve as a contributor to autoimmune disease (AD) pathogenesis via generation of hyper-inflammatory myeloid cells. Using a mouse model of systemic lupus erythematosus (SLE), we show that hematopoietic stem cells (HSC) constitute a transplantable, long-term reservoir for macrophages that exhibit features of TI, including increased Mycobacterium avium killing, inflammatory cytokine production, and augmented capacity to co-stimulate naive T cells. Strikingly, hematopoietic progenitor cells derived from these HSC exhibit unique molecular features characterized by reduced chromatin accessibility and transcription of metabolic genes, accompanied by reduced glycolysis and central carbon metabolism. Altogether, our data identify HSC as a functional unit of TI in chronic AD, establish increased T-cell costimulatory activity as a potentially pathogenic feature of TI in this setting, and show that macrophages inherit reduced metabolic activity from AD-exposed HSC, suggesting metabolic activation and TI can be decoupled. Our findings thus implicate HSC as a long-term reservoir for TI that could contribute to AD.

immunology↗

bioRxiv2022

Therapeutic resistance to FLT3 inhibition is driven by a novel ATM/mTOR pathway regulating oxidative phosphorylation

While leukemic cells are susceptible to various therapeutic insults, residence in the bone marrow microenvironment typically confers protection from a wide range of drugs. Thus, understanding the unique molecular changes elicited by the marrow is of critical importance towards improving therapeutic outcomes. In the present study, we demonstrate that aberrant activation of oxidative phosphorylation serves to induce therapeutic resistance in FLT3 mutant human AML cells challenged with FLT3 inhibitor drugs. Importantly, our findings show that AML cells are protected from apoptosis following FLT3 inhibition due to marrow-mediated activation of ATM, which in turn up-regulates oxidative phosphorylation via mTOR signaling. mTOR is required for the bone marrow stroma-dependent maintenance of protein translation, with selective polysome enrichment of oxidative phosphorylation transcripts, despite FLT3 inhibition. To investigate the therapeutic significance of this finding, we tested the mTOR inhibitor everolimus in combination with the FLT3 inhibitor quizartinib in primary human AML xenograft models. While marrow resident AML cells were highly resistant to quizartinib alone, the addition of everolimus induced profound reduction in tumor burden and prevented relapse. Taken together, these data provide a novel mechanistic understanding of marrow-based therapeutic resistance, and a promising strategy for improved treatment of FLT3 mutant AML patients.

cancer biology↗

bioRxiv2021

The M1 aminopeptidase NPEPPS is a novel regulator of cisplatin sensitivity

There is an unmet need to improve efficacy of platinum-based cancer chemotherapy. Using multi-omic assessment of cisplatin-responsive and -resistant human bladder cancer cell lines and whole-genome CRISPR screens, we identified Puromycin-Sensitive Aminopeptidase, NPEPPS, as a novel driver of cisplatin resistance. NPEPPS depletion sensitizes resistant bladder cancer cells to cisplatin in vitro and in vivo. Conversely, overexpression of NPEPPS in sensitive cells increased cisplatin resistance. We show that NPEPPS affects treatment response by regulating intracellular cisplatin concentrations. Patient-derived organoids (PDOs) generated from bladder cancer samples before and after cisplatin-based treatment, and from patients who did not receive cisplatin, were evaluated for sensitivity to cisplatin and they were found to be concordant with clinical response. In PDOs, shRNA depletion or pharmacologic inhibition of NPEPPS led to increased cisplatin sensitivity, while NPEPPS overexpression had the opposite effect. Our data present NPEPPS as a novel and druggable driver of cisplatin resistance by regulating intracellular cisplatin concentrations, along with providing the preclinical data to support clinical trials combining NPEPPS inhibition with cisplatin.

cancer biology↗

bioRxiv2020

Chronic Interleukin-1 exposure triggers selective expansion of Cebpa-deficient multipotent hematopoietic progenitors

The early events that drive hematologic disorders like clonal hematopoiesis, myelodysplastic syndrome, myeloproliferative neoplasm, and acute myeloid leukemia are not well understood. Most studies focus on the cell-intrinsic genetic changes that occur in these disorders and how they impact cell fate decisions. We consider how chronic exposure to the pro-inflammatory cytokine, interleukin-1{beta} (IL-1{beta}), impacts Cebpa-deficient hematopoietic stem and progenitor cells (HSPC) in competitive settings. We surprisingly found that Cebpa-deficient HSPC did not have a hematopoietic cell intrinsic competitive advantage; rather chronic IL-1{beta} exposure engendered potent selection for Cebpa loss. Chronic IL-1{beta} augments myeloid lineage output by activating differentiation and repressing stem cell gene expression programs in a Cebpa-dependent manner. As a result, Cebpa-deficient HSPC are resistant to the pro-differentiative effects of chronic IL-1{beta}, and competitively expand. These findings have important implications for the earliest events that drive hematologic disorders, suggesting that chronic inflammation could be an important driver of leukemogenesis and a potential target for intervention. SummaryHiga et al. show that chronic interleukin-1{beta} exposure primes hematopoietic stem and progenitor cells for myelopoiesis by upregulating myeloid differentiation programs and repressing stem gene programs in a Cebpa-dependent manner. Consequently, interleukin-1 potently selects for Cebpa loss in hematopoietic stem and progenitor cells.

cancer biology↗

bioRxiv2018

Low MSH2 protein levels identify muscle-invasive bladder cancer resistant to cisplatin

BackgroundThe response to first-line, platinum-based treatment of muscle-invasive bladder cancer has not improved in three decades.\n\nObjectiveThe objective of this study is to identify genes that predict cisplatin resistance in bladder cancer.\n\nDesignWe performed a whole-genome, CRISPR-based screen in a bladder cancer cell line treated with cisplatin to identify genes that mediate response to cisplatin. Targeted validation was performed in vitro across two bladder cancer cell lines. The top gene candidate was validated in a publicly available bladder cancer dataset containing 340 bladder cancer patients with treatment, protein, and survival information.\n\nResults and limitationsThe cisplatin resistance screen suggested the mismatch repair pathway through the loss of MSH2 and MLH1 contribute to cisplatin resistance. Bladder cancer cells depleted of MSH2 are resistant to cisplatin in vitro, in part due to a reduction in apoptosis. These cells maintain sensitivity to the cisplatin-analog, oxaliplatin. Bladder tumors with low protein levels of MSH2 have poorer overall survival when treated with cisplatin- or carboplatin-based therapy.\n\nConclusionsWe generated in vitro and clinical support that bladder cancer cell lines and tumors with low levels of MSH2 are more resistant to cisplatin-based therapy. Further studies are warranted to determine the ability of MSH2 protein levels to serve as a prospective biomarker of chemotherapy response in bladder cancer.\n\nPatient summaryWe report the first evidence that the protein level of MSH2 may contribute to chemotherapy resistance observed in bladder cancer. MSH2 levels has the potential to serve as a biomarker of treatment response.

cancer biology↗

bioRxiv2018

GSEA-InContext: Identifying novel and common patterns in expression experiments

MotivationGene Set Enrichment Analysis (GSEA) is routinely used to analyze and interpret coordinate changes in transcriptomics experiments. For an experiment where less than seven samples per condition are compared, GSEA employs a competitive null hypothesis to test significance. A gene set enrichment score is tested against a null distribution of enrichment scores generated from permuted gene sets, where genes are randomly selected from the input experiment. Looking across a variety of biological conditions, however, genes are not randomly distributed with many showing consistent patterns of up- or down-regulation. As a result, common patterns of positively and negatively enriched gene sets are observed across experiments. Placing a single experiment into the context of a relevant set of background experiments allows us to identify both the common and experiment-specific patterns of gene set enrichment.\n\nResultsWe compiled a compendium of 442 small molecule transcriptomic experiments and used GSEA to characterize common patterns of positively and negatively enriched gene sets. To identify experiment-specific gene set enrichment, we developed the GSEA-InContext method that accounts for gene expression patterns within a user-defined background set of experiments to identify statistically significantly enriched gene sets. We evaluated GSEA-InContext on experiments using small molecules with known targets and show that it successfully prioritizes gene sets that are specific to each experiment, thus providing valuable insights that complement standard GSEA analysis.\n\nAvailability and ImplementationGSEA-InContext is implemented in Python. Code, the background expression compendium, and results are available at: https://github.com/CostelloLab/GSEA-InContext

bioinformatics↗