26 research outputs found
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Mediator Kinase Inhibition Further Activates Super-Enhancer Associated Genes in AML
Super-enhancers (SEs), which are composed of large clusters of enhancers densely loaded with the Mediator complex, transcription factors (TFs), and chromatin regulators, drive high expression of genes implicated in cell identity and disease, such as lineage-controlling TFs and oncogenes 1, 2. BRD4 and CDK7 are positive regulators of SE-mediated transcription3,4,5. In contrast, negative regulators of SE-associated genes have not been well described. Here we report that Mediator-associated kinases cyclin-dependent kinase 8 (CDK8) and CDK19 restrain increased activation of key SE-associated genes in acute myeloid leukaemia (AML) cells. We determined that the natural product cortistatin A (CA) selectively inhibited Mediator kinases, had antileukaemic activity in vitro and in vivo, and disproportionately induced upregulation of SE-associated genes in CA-sensitive AML cell lines but not in CA-insensitive cell lines. In AML cells, CA upregulated SE-associated genes with tumour suppressor and lineage-controlling functions, including the TFs CEBPA, IRF8, IRF1 and ETV6 6, 7, 8. The BRD4 inhibitor I-BET151 downregulated these SE-associated genes, yet also has antileukaemic activity. Individually increasing or decreasing expression of these TFs suppressed AML cell growth, providing evidence that leukaemia cells are sensitive to dosage of SE-associated genes. Our results demonstrate that Mediator kinases can negatively regulate SE-associated gene expression in specific cell types and can be pharmacologically targeted as a therapeutic approach to AML
Elective Cancer Surgery in COVID-19-Free Surgical Pathways During the SARS-CoV-2 Pandemic: An International, Multicenter, Comparative Cohort Study.
PURPOSE: As cancer surgery restarts after the first COVID-19 wave, health care providers urgently require data to determine where elective surgery is best performed. This study aimed to determine whether COVID-19-free surgical pathways were associated with lower postoperative pulmonary complication rates compared with hospitals with no defined pathway. PATIENTS AND METHODS: This international, multicenter cohort study included patients who underwent elective surgery for 10 solid cancer types without preoperative suspicion of SARS-CoV-2. Participating hospitals included patients from local emergence of SARS-CoV-2 until April 19, 2020. At the time of surgery, hospitals were defined as having a COVID-19-free surgical pathway (complete segregation of the operating theater, critical care, and inpatient ward areas) or no defined pathway (incomplete or no segregation, areas shared with patients with COVID-19). The primary outcome was 30-day postoperative pulmonary complications (pneumonia, acute respiratory distress syndrome, unexpected ventilation). RESULTS: Of 9,171 patients from 447 hospitals in 55 countries, 2,481 were operated on in COVID-19-free surgical pathways. Patients who underwent surgery within COVID-19-free surgical pathways were younger with fewer comorbidities than those in hospitals with no defined pathway but with similar proportions of major surgery. After adjustment, pulmonary complication rates were lower with COVID-19-free surgical pathways (2.2% v 4.9%; adjusted odds ratio [aOR], 0.62; 95% CI, 0.44 to 0.86). This was consistent in sensitivity analyses for low-risk patients (American Society of Anesthesiologists grade 1/2), propensity score-matched models, and patients with negative SARS-CoV-2 preoperative tests. The postoperative SARS-CoV-2 infection rate was also lower in COVID-19-free surgical pathways (2.1% v 3.6%; aOR, 0.53; 95% CI, 0.36 to 0.76). CONCLUSION: Within available resources, dedicated COVID-19-free surgical pathways should be established to provide safe elective cancer surgery during current and before future SARS-CoV-2 outbreaks
Elective cancer surgery in COVID-19-free surgical pathways during the SARS-CoV-2 pandemic: An international, multicenter, comparative cohort study
PURPOSE As cancer surgery restarts after the first COVID-19 wave, health care providers urgently require data to determine where elective surgery is best performed. This study aimed to determine whether COVID-19–free surgical pathways were associated with lower postoperative pulmonary complication rates compared with hospitals with no defined pathway. PATIENTS AND METHODS This international, multicenter cohort study included patients who underwent elective surgery for 10 solid cancer types without preoperative suspicion of SARS-CoV-2. Participating hospitals included patients from local emergence of SARS-CoV-2 until April 19, 2020. At the time of surgery, hospitals were defined as having a COVID-19–free surgical pathway (complete segregation of the operating theater, critical care, and inpatient ward areas) or no defined pathway (incomplete or no segregation, areas shared with patients with COVID-19). The primary outcome was 30-day postoperative pulmonary complications (pneumonia, acute respiratory distress syndrome, unexpected ventilation). RESULTS Of 9,171 patients from 447 hospitals in 55 countries, 2,481 were operated on in COVID-19–free surgical pathways. Patients who underwent surgery within COVID-19–free surgical pathways were younger with fewer comorbidities than those in hospitals with no defined pathway but with similar proportions of major surgery. After adjustment, pulmonary complication rates were lower with COVID-19–free surgical pathways (2.2% v 4.9%; adjusted odds ratio [aOR], 0.62; 95% CI, 0.44 to 0.86). This was consistent in sensitivity analyses for low-risk patients (American Society of Anesthesiologists grade 1/2), propensity score–matched models, and patients with negative SARS-CoV-2 preoperative tests. The postoperative SARS-CoV-2 infection rate was also lower in COVID-19–free surgical pathways (2.1% v 3.6%; aOR, 0.53; 95% CI, 0.36 to 0.76). CONCLUSION Within available resources, dedicated COVID-19–free surgical pathways should be established to provide safe elective cancer surgery during current and before future SARS-CoV-2 outbreaks
Transcription elongation factors represent in vivo cancer dependencies in glioblastoma
Glioblastoma is a universally lethal cancer with a median survival of approximately 15 months1. Despite substantial efforts to define druggable targets, there are no therapeutic options that meaningfully extend glioblastoma patient lifespan. While previous work has largely focused on in vitro cellular models, here we demonstrate a more physiologically relevant approach to target discovery in glioblastoma. We adapted pooled RNA interference (RNAi) screening technology2–4 for use in orthotopic patient-derived xenograft (PDX) models, creating a high-throughput negative selection screening platform in a functional in vivo tumour microenvironment. Using this approach, we performed parallel in vivo and in vitro screens and discovered that the chromatin and transcriptional regulators necessary for cell survival in vivo are non-overlapping with those required in vitro. We identified transcription pause-release and elongation factors as one set of in vivo-specific cancer dependencies and determined that these factors are necessary for enhancer-mediated transcriptional adaptations that enable cells to survive the tumour microenvironment. Our lead hit, JMJD6, mediates the upregulation of in vivo stress and stimulus response pathways through enhancer-mediated transcriptional pause-release, promoting cell survival specifically in vivo. Targeting JMJD6 or other identified elongation factors extends survival in orthotopic xenograft mouse models, supporting targeting the transcription elongation machinery as a therapeutic strategy for glioblastoma. More broadly, this study demonstrates the power of in vivo phenotypic screening to identify new classes of ‘cancer dependencies’ not identified by previous in vitro approaches, which could supply untapped opportunities for therapeutic intervention
A Unified Strategy for the Synthesis of 7‑Membered-Ring-Containing <i>Lycopodium</i> Alkaloids
A unique subset of
the <i>LycoÂpodium</i> alkaloid
natural products share a 7-membered-ring substructure and may potentially
arise from a common biosynthetic precursor. To both explore and exploit
these structural relationships, we sought to develop a unified biosynthetically
inspired strategy to efficiently access these complex polycyclic alkaloids
through the use of a cascade sequence. In pursuit of these goals,
the first total synthesis of (+)-fastigiatine (<b>2</b>) was
accomplished via a series of cascade reactions; we describe herein
a full account of our efforts. Insight from these endeavors led to
critical modifications of our synthetic strategy, which enabled the
first total syntheses of (−)-himeradine A (<b>1</b>),
(−)-lycoÂpecurine (<b>3</b>), and (−)-dehydroÂlycoÂpecurine
(<b>4</b>), as well as the syntheses of (+)-lycoÂnadin
A (<b>5</b>) and (−)-lycoÂnadin B (<b>6</b>). Our approach features a diastereoÂselective one-pot sequence
for constructing the common 7-membered-ring core system, followed
by either a biomimetic transannular Mannich reaction to access himeradine
A (<b>1</b>), lycoÂpecurine (<b>3</b>), and dehydroÂlycoÂpecurine
(<b>4</b>) or an imine reduction for lycoÂnadins A (<b>5</b>) and B (<b>6</b>). This strategy may potentially enable
access to all 7-membered-ring-containing <i>LycoÂpodium</i> alkaloids and provides additional insight into their biosynthetic
origin
Total Syntheses of HMP-Y1, Hibarimicinone, and HMP-P1
Total syntheses of HMP-Y1, atrop-HMP-Y1, hibarimicinone,
atrop-hibarimicinone,
and HMP-P1 are described using a two-directional synthesis strategy.
A novel benzyl fluoride Michael–Claisen reaction sequence was
developed to construct the complete carbon skeleton of HMP-Y1 and
atrop-HMP-Y1 via a symmetrical, two-directional, double annulation.
Through efforts to convert HMP-Y1 derivatives to hibarimicinone and
HMP-P1, a biomimetic mono-oxidation to desymmetrize protected HMP-Y1
was realized. A two-directional unsymmetrical double annulation and
biomimetic etherification was developed to construct the polycyclic
and highly oxidized skeleton of hibarimicinone, atrop-hibarimicinone,
and HMP-P1. The use of a racemic biaryl precursor allowed for the
synthesis of both hibarimicinone atropisomers and provides the first
confirmation of the structure of atrop-hibarimicinone. Additionally,
this work documents the first reported full characterization of atrop-hibarimicinone,
HMP-Y1, atrop-HMP-Y1, and HMP-P1. Last, a pH-dependent rotational
barrier about the C2–C2′ bond of hibarimicinone was
discovered, which provides valuable information necessary to achieve
syntheses of the glycosylated congeners of hibarimicinone
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Insulator dysfunction and oncogene activation in IDH mutant gliomas
Gain-of-function IDH mutations are initiating events that define major clinical and prognostic classes of gliomas1,2. Mutant IDH protein produces a novel onco-metabolite, 2-hydroxyglutarate (2-HG), that interferes with iron-dependent hydroxylases, including the TET family of 5′-methylcytosine hydroxylases3–7. TET enzymes catalyze a key step in the removal of DNA methylation8,9. IDH mutant gliomas thus manifest a CpG island methylator phenotype (G-CIMP)10,11, though the functional significance of this altered epigenetic state remains unclear. Here we show that IDH mutant gliomas exhibit hyper-methylation at CTCF binding sites, compromising binding of this methylation-sensitive insulator protein. Reduced CTCF binding is associated with loss of insulation between topological domains and aberrant gene activation. We specifically demonstrate that loss of CTCF at a domain boundary permits a constitutive enhancer to aberrantly interact with the receptor tyrosine kinase gene PDGFRA, a prominent glioma oncogene. Treatment of IDH mutant gliomaspheres with demethylating agent partially restores insulator function and down-regulates PDGFRA. Conversely, CRISPR-mediated disruption of the CTCF motif in IDH wildtype gliomaspheres up-regulates PDGFRA and increases proliferation. Our study suggests that IDH mutations promote gliomagenesis by disrupting chromosomal topology and allowing aberrant regulatory interactions that induce oncogene expression
Polycomb-lamina antagonism partitions heterochromatin at the nuclear periphery
Here the authors developed ‘Lamina-Inducible Methylation and Hi-C’ (LIMe-Hi-C) to simultaneously measure chromosome conformation, DNA methylation, and nuclear lamina positioning. Application of the method revealed dynamic changes upon PRC2 inhibition and an essential function of H3K27me3 in regulating sub-compartments and lamina association
Characterizing Protein Glycosylation through On-Chip Glycan Modification and Probing
Glycans
are critical to protein biology and are useful as disease
biomarkers. Many studies of glycans rely on clinical specimens, but
the low amount of sample available for some specimens limits the experimental
options. Here we present a method to obtain information about protein
glycosylation using a minimal amount of protein. We treat proteins
that were captured or directly spotted in small microarrays (2.2 mm
× 2.2 mm) with exoglycosidases to successively expose underlying
features, and then we probe the native or exposed features using a
panel of lectins or glycan-binding reagents. We developed an algorithm
to interpret the data and provide predictions about the glycan motifs
that are present in the sample. We demonstrated the efficacy of the
method to characterize differences between glycoproteins in their
sialic acid linkages and N-linked glycan branching, and we validated
the assignments by comparing results from mass spectrometry and chromatography.
The amount of protein used on-chip was about 11 ng. The method also
proved effective for analyzing the glycosylation of a cancer biomarker
in human plasma, MUC5AC, using only 20 μL of the plasma. A glycan
on MUC5AC that is associated with cancer had mostly 2,3-linked sialic
acid, whereas other glycans on MUC5AC had a 2,6 linkage of sialic
acid. The on-chip glycan modification and probing (on-chip GMAP) method
provides a platform for analyzing protein glycosylation in clinical
specimens and could complement the existing toolkit for studying glycosylation
in disease