A transgenic mouse model of Down syndrome acute lymphoblastic leukemia identifies targetable vulnerabilities.

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CD57 defines a novel cancer stem cell that drive invasion of diffuse pediatric-type high grade gliomas.

BACKGROUND: Diffuse invasion remains a primary cause of treatment failure in pediatric high-grade glioma (pHGG). Identifying cellular driver(s) of pHGG invasion is needed for anti-invasion therapies. METHODS: Ten highly invasive patient-derived orthotopic xenograft (PDOX) models of pHGG were subjected to isolation of matching pairs of invasive (HGGINV) and tumor core (HGGTC) cells. RESULTS: pHGGINV cells were intrinsically more invasive than their matching pHGGTC cells. CSC profiling revealed co-positivity of CD133 and CD57 and identified CD57+CD133- cells as the most abundant CSCs in the invasive front. In addition to discovering a new order of self-renewal capacities, i.e., CD57+CD133- > CD57+CD133+ > CD57-CD133+ > CD57-CD133- cells, we showed that CSC hierarchy was impacted by their spatial locations, and the highest self-renewal capacities were found in CD57+CD133- cells in the HGGINV front (HGGINV/CD57+CD133- cells) mediated by NANOG and SHH over-expression. Direct implantation of CD57+ (CD57+/CD133- and CD57+/CD133+) cells into mouse brains reconstituted diffusely invasion, while depleting CD57+ cells (i.e., CD57-CD133+) abrogated pHGG invasion. CONCLUSION: We revealed significantly increased invasive capacities in HGGINV cells, confirmed CD57 as a novel glioma stem cell marker, identified CD57+CD133- and CD57+CD133+ cells as a new cellular driver of pHGG invasion and suggested a new dual-mode hierarchy of HGG stem cells.

Targeting neddylation and sumoylation in chemoresistant triple negative breast cancer.

Triple negative breast cancer (TNBC) accounts for 15-20% of breast cancer cases in the United States. Systemic neoadjuvant chemotherapy (NACT), with or without immunotherapy, is the current standard of care for patients with early-stage TNBC. However, up to 70% of TNBC patients have significant residual disease once NACT is completed, which is associated with a high risk of developing recurrence within two to three years of surgical resection. To identify targetable vulnerabilities in chemoresistant TNBC, we generated longitudinal patient-derived xenograft (PDX) models from TNBC tumors before and after patients received NACT. We then compiled transcriptomes and drug response profiles for all models. Transcriptomic analysis identified the enrichment of aberrant protein homeostasis pathways in models from post-NACT tumors relative to pre-NACT tumors. This observation correlated with increased sensitivity in vitro to inhibitors targeting the proteasome, heat shock proteins, and neddylation pathways. Pevonedistat, a drug annotated as a NEDD8-activating enzyme (NAE) inhibitor, was prioritized for validation in vivo and demonstrated efficacy as a single agent in multiple PDX models of TNBC. Pharmacotranscriptomic analysis identified a pathway-level correlation between pevonedistat activity and post-translational modification (PTM) machinery, particularly involving neddylation and sumoylation targets. Elevated levels of both NEDD8 and SUMO1 were observed in models exhibiting a favorable response to pevonedistat compared to those with a less favorable response in vivo. Moreover, a correlation emerged between the expression of neddylation-regulated pathways and tumor response to pevonedistat, indicating that targeting these PTM pathways may prove effective in combating chemoresistant TNBC.

Mother trees, altruistic fungi, and the perils of plant personification.

There are growing doubts about the true role of the common mycorrhizal networks (CMN or wood wide web) connecting the roots of trees in forests. We question the claims of a substantial carbon transfer from 'mother trees' to their offspring and nearby seedlings through the CMN. Recent reviews show that evidence for the 'mother tree concept' is inconclusive or absent. The origin of this concept seems to stem from a desire to humanize plant life but can lead to misunderstandings and false interpretations and may eventually harm rather than help the commendable cause of preserving forests. Two recent books serve as examples: The Hidden Life of Trees and Finding the Mother Tree.

Increase of Severe Pulmonary Infections in Adults Caused by M1UK Streptococcus pyogenes, Central Scotland, UK.

We characterized the epidemiology, host-pathogen characteristics, and outcomes of severe adult pulmonary Streptococcus pyogenes infections that coincided with a high community caseload in central Scotland, UK. The pulmonary infections had high illness and death rates and were associated with socioeconomic deprivation, influenza A co-infection, and the M1UK lineage of S. pyogenes.

An assessment of the downstream implications of blood culture collection and transit.

The implications of the variables within the pre-analytical phase of blood culture processing are poorly understood. This study aims to explore the effect of transit times (TT) and culture volume, on time to microbiological diagnosis and patient outcomes. Blood cultures received between 1st March and 31st July 2020/21 were identified. TT, time in incubator (TII), and for positive samples, request to positivity times (RPT) were calculated. Demographic details were recorded for all samples, and culture volume, length of stay (LoS), and 30-day mortality for patients with positive samples. Statistical analysis examined how culture volume and TT effected culture positivity and outcome; in the context of the 4-h national TT target. Totally, 14,375 blood culture bottles were received from 7367 patients; 988 (13.4%) were positive for organisms. There was no significant difference between TT of negative and positive samples. The RPT was significantly lower for samples with TT < 4 h (p < 0.001). Culture bottle volume did not affect RPT (p = 0.482) or TII (p = 0.367). A prolonged TT was associated with a longer length-of-stay in those with a bacteraemia with a significant organism (p = 0.001). We found shorter blood culture transportation time was associated with a significantly faster time of positive culture reporting, while optimal blood culture volume did not make a significant impact. Delays in reporting for significant organisms correspond to a prolonged LoS. Laboratory centralisation makes achieving the 4-h target a logistical challenge; however, this data suggests such targets have significant microbiological and clinical impacts.

Cloning a profibrotic stem cell variant in idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a progressive, irreversible, and rapidly fatal interstitial lung disease marked by the replacement of lung alveoli with dense fibrotic matrices. Although the mechanisms initiating IPF remain unclear, rare and common alleles of genes expressed in lung epithelia, combined with aging, contribute to the risk for this condition. Consistently, single-cell RNA sequencing (scRNA-seq) studies have identified lung basal cell heterogeneity in IPF that might be pathogenic. We used single-cell cloning technologies to generate "libraries" of basal stem cells from the distal lungs of 16 patients with IPF and 10 controls. We identified a major stem cell variant that was distinguished from normal stem cells by its ability to transform normal lung fibroblasts into pathogenic myofibroblasts in vitro and to activate and recruit myofibroblasts in clonal xenografts. This profibrotic stem cell variant, which was shown to preexist in low quantities in normal and even fetal lungs, expressed a broad network of genes implicated in organ fibrosis and showed overlap in gene expression with abnormal epithelial signatures identified in previously published scRNA-seq studies of IPF. Drug screens highlighted specific vulnerabilities of this profibrotic variant to inhibitors of epidermal growth factor and mammalian target of rapamycin signaling as prospective therapeutic targets. This profibrotic stem cell variant in IPF was distinct from recently identified profibrotic stem cell variants in chronic obstructive pulmonary disease and may extend the notion that inappropriate accrual of minor and preexisting stem cell variants contributes to chronic lung conditions.

Epigenetic Alterations of Repeated Relapses in Patient-matched Childhood Ependymomas.

Recurrence is frequent in pediatric ependymoma (EPN). Our longitudinal integrated analysis of 30 patient-matched repeated relapses (3.67 ± 1.76 times) over 13 years (5.8 ± 3.8) reveals stable molecular subtypes (RELA and PFA) and convergent DNA methylation reprogramming during serial relapses accompanied by increased orthotopic patient derived xenograft (PDX) (13/27) formation in the late recurrences. A set of differentially methylated CpGs (DMCs) and DNA methylation regions (DMRs) are found to persist in primary and relapse tumors (potential driver DMCs) and are acquired exclusively in the relapses (potential booster DMCs). Integrating with RNAseq reveals differentially expressed genes regulated by potential driver DMRs (CACNA1H, SLC12A7, RARA in RELA and HSPB8, GMPR, ITGB4 in PFA) and potential booster DMRs (PLEKHG1 in RELA and NOTCH, EPHA2, SUFU, FOXJ1 in PFA tumors). DMCs predicators of relapse are also identified in the primary tumors. This study provides a high-resolution epigenetic roadmap of serial EPN relapses and 13 orthotopic PDX models to facilitate biological and preclinical studies.

deepOrganoid: A brightfield cell viability model for screening matrix-embedded organoids.

High-throughput viability screens are commonly used in the identification and development of chemotherapeutic drugs. These systems rely on the fidelity of the cellular model systems to recapitulate the drug response that occurs in vivo. In recent years, there has been an expansion in the utilization of patient-derived materials as well as advanced cell culture techniques, such as multi-cellular tumor organoids, to further enhance the translational relevance of cellular model systems. Simple quantitative analysis remains a challenge, primarily due to the difficulties of robust image segmentation in heterogenous 3D cultures. However, explicit segmentation is not required with the advancement of deep learning, and it can be used for both continuous (regression) or categorical classification problems. Deep learning approaches are additionally benefited by being fully data-driven and highly automatable, thus they can be established and run with minimal to no user-defined parameters. In this article, we describe the development and implementation of a regressive deep learning model trained on brightfield images of patient-derived organoids and use the terminal viability readout (CellTiter-Glo) as training labels. Ultimately, this has led to the generation of a non-invasive and label-free tool to evaluate changes in organoid viability.

Mammary-specific expression of Trim24 establishes a mouse model of human metaplastic breast cancer.

Conditional overexpression of histone reader Tripartite motif containing protein 24 (TRIM24) in mouse mammary epithelia (Trim24COE) drives spontaneous development of mammary carcinosarcoma tumors, lacking ER, PR and HER2. Human carcinosarcomas or metaplastic breast cancers (MpBC) are a rare, chemorefractory subclass of triple-negative breast cancers (TNBC). Comparison of Trim24COE metaplastic carcinosarcoma morphology, TRIM24 protein levels and a derived Trim24COE gene signature reveals strong correlation with human MpBC tumors and MpBC patient-derived xenograft (PDX) models. Global and single-cell tumor profiling reveal Met as a direct oncogenic target of TRIM24, leading to aberrant PI3K/mTOR activation. Here, we find that pharmacological inhibition of these pathways in primary Trim24COE tumor cells and TRIM24-PROTAC treatment of MpBC TNBC PDX tumorspheres decreased cellular viability, suggesting potential in therapeutically targeting TRIM24 and its regulated pathways in TRIM24-expressing TNBC.

Land use planning to support climate change adaptation in threatened plant communities.

Among the many causes of habitat loss, urbanization coupled with climate change has produced some of the greatest local extinction rates and has led to the loss of many native species. Managing native vegetation in a rapidly expanding urban setting requires land management strategies that are cognizant of these impacts and how species and communities may adapt to a future climate. Here, we demonstrate how identifying climate refugia for threatened vegetation communities in an urban matrix can be used to support management decisions by local government authorities under the dual pressures of urban expansion and climate change. This research was focused on a local government area in New South Wales, Australia, that is undergoing significant residential, commercial and agricultural expansion resulting in the transition of native forest to other more intensive land-uses. Our results indicate that the key drivers of change from native vegetation to urban and agriculture classes were population density and the proximity to urban areas. We found two of the most cleared vegetation community types are physically restricted to land owned or managed by council, suggesting their long-term ecological viability is uncertain under a warming climate. We propose that land use planning decisions must recognize the compounding spatial and temporal pressures of urban development, land clearing and climate change, and how current policy responses, such as biodiversity offsetting, can respond positively to habitat shifts in order to secure the longevity of important ecological communities.

Pharmacologic profiling of patient-derived xenograft models of primary treatment-naïve triple-negative breast cancer.

Triple-negative breast cancer (TNBC) accounts for 15-20% of breast cancer cases in the United States, lacks targeted therapeutic options, and is associated with a 40-80% risk of recurrence. Thus, identifying actionable targets in treatment-naïve and chemoresistant TNBC is a critical unmet medical need. To address this need, we performed high-throughput drug viability screens on human tumor cells isolated from 16 patient-derived xenograft models of treatment-naïve primary TNBC. The models span a range of TNBC subtypes and exhibit a diverse set of putative driver mutations, thus providing a unique patient-derived, molecularly annotated pharmacologic resource that is reflective of TNBC. We identified therapeutically actionable targets including kinesin spindle protein (KSP). The KSP inhibitor targets the mitotic spindle through mechanisms independent of microtubule stability and showed efficacy in models that were resistant to microtubule inhibitors used as part of the current standard of care for TNBC. We also observed subtype selectivity of Prima-1Met, which showed higher levels of efficacy in the mesenchymal subtype. Coupling pharmacologic data with genomic and transcriptomic information, we showed that Prima-1Met activity was independent of its canonical target, mutant p53, and was better associated with glutathione metabolism, providing an alternate molecularly defined biomarker for this drug.

Reliable tumor detection by whole-genome methylation sequencing of cell-free DNA in cerebrospinal fluid of pediatric medulloblastoma.

Medulloblastoma (MB), the most common form of pediatric brain malignancy, has a low frequency of oncogenic mutations but pronouncedly abnormal DNA methylation changes. Epigenetic analysis of circulating cell-free tumor DNA (ctDNA) by liquid biopsy offers an approach for real-time monitoring of tumor status without tumor dissection. In this study, we identified 6598 differentially methylated CpGs in both MB tumors and the ctDNA isolated from matched cerebrospinal fluid (CSF) compared with normal cerebellum, which could be used to detect MB tumor occurrence and determine its subtype. Furthermore, DNA methylation changes in serial CSF samples could be used to monitor the treatment response and tumor recurrence. Integrating our data with large public datasets, we identified reliable MB DNA methylation signatures in ctDNA that have potential diagnostic and prognostic values. Our study sets the stage for exploiting epigenetic markers in CSF to improve the clinical management of patients with MB.

A combination strategy targeting enhancer plasticity exerts synergistic lethality against BETi-resistant leukemia cells.

Primary and acquired drug resistance imposes a major threat to achieving optimized clinical outcomes during cancer treatment. Aberrant changes in epigenetic modifications are closely involved in drug resistance of tumor cells. Using BET inhibitor (BETi) resistant leukemia cells as a model system, we demonstrated herein that genome-wide enhancer remodeling played a pivotal role in driving therapeutic resistance via compensational re-expression of pro-survival genes. Capitalizing on the CRISPR interference technology, we identified the second intron of IncRNA, PVT1, as a unique bona fide gained enhancer that restored MYC transcription independent of BRD4 recruitment in leukemia. A combined BETi and CDK7 inhibitor treatment abolished MYC transcription by impeding RNAPII loading without affecting PVT1-mediated chromatin looping at the MYC locus in BETi-resistant leukemia cells. Together, our findings have established the feasibility of targeting enhancer plasticity to overcome drug resistance associated with epigenetic therapies.

Seasonal changes in amino acids and phenolic compounds in fruits from hybrid cross populations of American grapes differing in disease resistance.

The production of wine grapes in upstate New York (USA) is limited by diseases that are promoted by the cool and sometimes rainy climate. A breeding program has been introducing disease resistance from related species into the cultivated stock. Previous work has indicated that such resistance may be based on biochemical reactions rather than on a hypersensitive reaction. We therefore undertook metabolic profiling of amino acids and phenolic compounds in berries from collections of susceptible and resistant hybrids over the course of berry development to determine whether any of these compounds could be causal in disease resistance. The most abundant amino acids were GLN, ARG, PRO and THR. The amount of amino acids in ripe berries was from 3 to 4.7-fold higher compared to earlier stages. The concentrations of total phenolics were variable through the season with no consistent trend between susceptible and resistant fruits. Notable changes in phenolic compounds, especially anthocyanins, were recorded, especially during the ripening phase, when phenolics and anthocyanins increased following veraison. The most abundant phenolic compounds were catechin and epi-catechin; the most abundant anthocyanin was delphinidin-3-glucoside, which had a slightly greater concentration in resistant fruit at harvest, followed by malvidin-3-glucoside and petunidin-3-glucoside. The content of both amino acids and phenolic compounds in white-fruited parent cv. Horizon was equal to several-fold lower than the progeny plants, whether susceptible or resistant, depending on the harvest time. While no major differences between susceptible and resistant lines were found, multivariate analyses showed that it is possible to discriminate the susceptibility or resistance of grapes by analyzing their combined concentrations of amino acids, polyphenols and anthocyanins. Therefore, these compounds are influenced by the resistance capacity of grapes and could be used as a chemical fingerprint of this ability. However, it is likely that these are associations with disease resistance rather than their cause as no major consistent differences were noted.

Tracing natural and industrial contamination and lead isotopic compositions in an Australian native bee species.

This study investigates trace element concentrations (arsenic (As), manganese (Mn), lead (Pb) and zinc (Zn)) and Pb isotopic compositions in an Australian native bee species, Tetragonula carbonaria, and its products of honey and wax. Co-located soil and dust samples were simultaneously analysed with the objective of determining if the bees or their products had potential application as a proxy for monitoring environmental contamination. The most significant relationships were found between Pb concentrations in honey (r = 0.814, p = 0.014) and wax (r = 0.883, p = 0.004) and those in co-located dust samples. In addition, Zn concentrations in honey and soil were significantly associated (r = 0.709, p = 0.049). Lead isotopic compositions of native bee products collected from background sites adjacent to Sydney national parks (206Pb/207Pb = 1.144, 208Pb/207Pb = 2.437) corresponded to local geogenic rock and soil values (206Pb/207Pb = 1.123-1.176, 208Pb/207Pb = 2.413-2.500). By contrast, inner Sydney metropolitan samples, including native bees and wax (206Pb/207Pb = 1.072-1.121, 208Pb/207Pb = 2.348-2.409), co-located soil and dust (206Pb/207Pb = 1.090-1.122, 208Pb/207Pb = 2.368-2.403), corresponded most closely to aerosols collected during the period of leaded petrol use (206Pb/207Pb = 1.067-1.148, 208Pb/207Pb = 2.341-2.410). A large range of Pb isotopic compositions in beehive samples suggests that other legacy sources, such as Pb-based paints and industrials, may have also contributed to Pb contamination in beehive samples. Native bee data were compared to corresponding samples from the more common European honey bee (Apis mellifera). Although Pb isotopic compositions were similar in both species, significant differences in trace element concentrations were evident across the trace element suite, the bees and their products. The statistical association between T. carbonaria and co-located environmental contaminant concentrations were stronger than those in European honey bees, which may be attributable to its smaller foraging distance (0.3-0.7 km versus 5-9 km, respectively). This implies that T. carbonaria may be more suitable for assessing small spatial scale variations of trace element concentrations than European honey bees.

Identifying Sources of Environmental Contamination in European Honey Bees (Apis mellifera) Using Trace Elements and Lead Isotopic Compositions.

Trace element concentrations (As, Mn, Pb, and Zn) and Pb isotopic compositions were analyzed in honey bees, wax, and honey along with co-located soil and dust samples from Sydney metropolitan and Broken Hill, Australia. Compared with the other trace elements, Pearson correlations show that Pb concentrations in soil and dust had the strongest relationship to corresponding values in honey bees and their products. Dust Pb was not only highly correlated to corresponding soil values (r = 0.806, p = 0.005), it was the strongest predictor of Pb concentrations in honey bees, wax, and honey (p = 0.001, 0.007, 0.017, respectively). Lead isotopic compositions (206Pb/207Pb and 208Pb/207Pb) showed that honey bees and their products from Broken Hill were nearly identical (95-98%) to the composition of the local ore body. Samples of honey bees and their products collected from background sites adjacent to national parks in Sydney had Pb isotopic compositions (206Pb/207Pb = 1.138-1.159, 208Pb/207Pb = 2.417-2.435) corresponding to local geogenic values (206Pb/207Pb = 1.123-1.176, 208Pb/207Pb = 2.413-2.500). By contrast, honey bees and their products from Sydney metropolitan (206Pb/207Pb = 1.081-1.126, 208Pb/207Pb = 2.352-2.408) were similar to aerosols measured during the period of leaded petrol use (206Pb/207Pb = 1.067-1.148, 208Pb/207Pb = 2.341-2.410). These measurements show Pb concentrations and its isotopic compositions of honey bees, and their products can be used to trace both legacy and contemporary environmental contamination, particularly where sources are well documented. Moreover, this study demonstrates that legacy Pb emissions continue to be remobilized in dust, contaminating both food and ecological systems.

River sediment quality assessment using sediment quality indices for the Sydney basin, Australia affected by coal and coal seam gas mining.

Coal mining activities in the Sydney basin have been historically associated with significant environmental impacts. The region is facing more recent coal seam gas extraction activities and the synergetic environmental impacts of the new mining activities are still largely unknown. The aim of this study was to provide environmental assessment of river sediments comparing upstream to downstream areas relative to industrial-discharge sites associated with coal and coal-seam-gas extraction within the Sydney basin. Various contaminants were measured to determine the sediment quality according to the Australian and New Zealand Environment and Conservation Council (ANZECC) guidelines. Arsenic, nickel and zinc were the main sediment contaminants in downstream samples exceeding the ANZECC guidelines. Degree of contamination (Cd), geoaccumulation index (Igeo), enrichment factor (EF), pollution load index (PLI) and sediment environmental toxicity quotients' increment in downstream sediment were estimated for the studied areas. Toxicology indices of metals present in the sediments near industrial discharge sites were used as an additional tool to compare the level of environmental effects with their increment. The study revealed that the sediments from coal mining sites were highly affected by increased concentrations of manganese, zinc, cobalt, nickel and barium. The sediments associated with coal mining activities were found to be substantially more affected than the sediments near coal seam gas production sites, mainly attributed to the different wastewater discharge licencing requirements. The approach applied in this study can be used as an additional model to assess the contribution of industrial and mining activities on aquatic environments.