Publications by authors named "Thomas Look"

Genome-wide gene expression analysis is a commonly used method to quantitatively examine the transcriptional signature of any tissue or cell state. Standard bulk cell RNA sequencing (RNA-seq) quantifies RNAs in the cells of the tissue type of interest through massive parallel sequencing of cDNA synthesized from the cellular RNA. The subsequent analysis of global RNA expression and normalization of RNA expression levels between two or more samples generally assumes that cells from all samples produce equivalent amounts of RNA per cell.

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  • Anti-PD-1 antibodies have transformed cancer treatment by achieving lasting remission in some patients, leading researchers to seek ways to enhance their effectiveness with new biomarkers and drugs.
  • This study introduces F8(scDb)-IL7, a novel fusion protein targeting EDA-FN, which is present in most tumors but rare in healthy tissue, potentially improving cancer therapy at the disease site.
  • F8(scDb)-IL7 was shown to boost T Cell Factor 1 expression in CD8+T cells and effectively eradicates sarcoma lesions when used alongside anti-PD-1 treatment, suggesting a promising combination for cancer patients.
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Synthetic mRNA produced by in vitro transcription (ivt mRNA) is the active pharmaceutical ingredient of approved anti-COVID-19 vaccines and of many drugs under development. Such synthetic mRNA typically contains several hundred bases of non-coding "untranslated" regions (UTRs) that are involved in the stabilization and translation of the mRNA. However, UTRs are often complex structures, which may complicate the entire production process.

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Anaplastic large cell lymphoma (ALCL) is an aggressive, CD30 T cell lymphoma of children and adults. ALK fusion transcripts or mutations in the JAK-STAT pathway are observed in most ALCL tumors, but the mechanisms underlying tumorigenesis are not fully understood. Here, we show that dysregulated STAT3 in ALCL cooccupies enhancers with master transcription factors BATF3, IRF4, and IKZF1 to form a core regulatory circuit that establishes and maintains the malignant cell state in ALCL.

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Cytokine-based therapeutics have been shown to mediate objective responses in certain tumor entities but suffer from insufficient selectivity, causing limiting toxicity which prevents dose escalation to therapeutically active regimens. The antibody-based delivery of cytokines significantly increases the therapeutic index of the corresponding payload but still suffers from side effects associated with peak concentrations of the product in blood upon intravenous administration. Here we devise a general strategy (named "Intra-Cork") to mask systemic cytokine activity without impacting anti-cancer efficacy.

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Deciphering the cell-state transitions underlying immune adaptation across time is fundamental for advancing biology. Empirical in vivo genomic technologies that capture cellular dynamics are currently lacking. We present Zman-seq, a single-cell technology recording transcriptomic dynamics across time by introducing time stamps into circulating immune cells, tracking them in tissues for days.

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The Large Plasma Device (LAPD) at UCLA (University of California, Los Angeles) produces an 18 m long, magnetized, quiescent, and uniform plasma at a high repetition rate to enable studies of fundamental plasma physics. Here, we report on a major upgrade to the LAPD plasma source that allows for more robust operation and significant expansion of achievable plasma parameters. The original plasma source made use of a heated barium oxide (BaO) coated nickel sheet as an electron emitter.

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Neuroblastoma originates from developing neural crest and can interconvert between the mesenchymal (MES) and adrenergic (ADRN) states, each of which are controlled by different sets of transcription factors forming the core regulatory circuit (CRC). However, the roles of CRC factors in induction and maintenance of specific state are poorly understood. Here, we demonstrate that overexpression of ASCL1, an ADRN CRC factor, in MES neuroblastoma cells opens closed chromatin at the promoters of key ADRN genes, accompanied by epigenetic activation and establishment of enhancer-promoter interactions, initiating the ADRN gene expression program.

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Introduction: Myelodysplastic syndrome (MDS) is a heterogeneous group of clonal hematopoietic disorders characterized by ineffective hematopoiesis, cytopenias, and dysplasia. The gene encoding ten-eleven translocation 2 (2), a dioxygenase enzyme that catalyzes the conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine, is a recurrently mutated tumor suppressor gene in MDS and other myeloid malignancies. Previously, we reported a stable zebrafish line with a loss-of-function mutation in the gene.

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  • The text outlines a protocol designed for the efficient and reproducible expansion of immune cells like T cells, NK cells, and macrophages, which is important for cancer research.
  • It includes specific steps for preparing surfaces with antibodies, isolating immune cells from mouse spleens, and techniques for growing T and NK cells.
  • Additionally, the protocol provides guidance on isolating bone marrow and differentiating macrophages for further studies.
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Glioblastoma is the most aggressive primary brain tumor with an unmet need for more effective therapies. Here, we investigated combination therapies based on L19TNF, an antibody-cytokine fusion protein based on tumor necrosis factor that selectively localizes to cancer neovasculature. Using immunocompetent orthotopic glioma mouse models, we identified strong anti-glioma activity of L19TNF in combination with the alkylating agent CCNU, which cured the majority of tumor-bearing mice, whereas monotherapies only had limited efficacy.

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  • Childhood neuroblastomas can switch between two cell states: an undifferentiated mesenchymal state and a more mature sympathetic adrenergic state, influenced by specific genetic factors.
  • * The expression of the LMO1 gene, linked to neuroblastoma risk, is affected by a single nucleotide polymorphism (G/T) that alters its regulatory sequence.
  • * Research demonstrated that zebrafish with the GATA genotype developed neuroblastoma, while those with the protective TATA allele had lower tumor initiation rates, suggesting a conserved regulatory mechanism across species.
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  • Transcription factors are crucial for cell lineage commitment and differentiation, with IRF8 playing a key role in dendritic cell development and function.
  • The study identified a novel long non-coding RNA that interacts with the IRF8 promoter, showing different epigenetic patterns in plasmacytoid dendritic cells compared to classical dendritic cells.
  • A specific sequence element in the promoter is essential for differentiation and provides feedback inhibition, shedding light on the mechanisms regulating IRF8 autoactivation during dendritic cell development.
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  • Childhood neuroblastomas can shift between two cell states: an undifferentiated "mesenchymal" state and a more mature "adrenergic" state, influenced by core regulatory circuitries that aid in neural development.
  • LMO1 is crucial for establishing the adrenergic identity in neuroblastomas, with its expression levels linked to a specific genetic variation (G/T polymorphism) that alters an important motif in its gene.
  • Research using zebrafish shows that changing the G ATA genotype to a T ATA variant lowers the chance of neuroblastoma development by preventing the formation of the adrenergic cell state, indicating a preserved evolutionary mechanism between zebrafish and humans.
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TET2 inactivating mutations serve as initiating genetic lesions in the transformation of haematopoietic stem and progenitor cells (HSPCs). In this study, we analysed known drugs in zebrafish embryos for their ability to selectively kill tet2-mutant HSPCs in vivo. We found that the exportin 1 (XPO1) inhibitors, selinexor and eltanexor, selectively kill tet2-mutant HSPCs.

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  • * A Phase I trial tested the safety and effectiveness of the MET inhibitor merestinib combined with the FGFR inhibitor LY2874455, with initial findings showing manageable side effects and some patients achieving stable disease or complete remission.
  • * The study suggests that targeting MET and FGFR pathways may offer a promising treatment approach for AML, although further research is needed due to supply issues with one of the drugs in the combination therapy.
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Background: In this study, we describe the generation of a fully human monoclonal antibody (named '7NP2') targeting human fibroblast activation protein (FAP), an antigen expressed in the microenvironment of different types of solid neoplasms.

Methods: 7NP2 was isolated from a synthetic antibody phage display library and was improved by one round of mutagenesis-based affinity maturation. The tumor recognition properties of the antibody were validated by immunofluorescence procedures performed on cancer biopsies from human patients.

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Loss of the gene SMARCB1 drives the development of malignant rhabdoid tumors, epithelioid sarcomas, and other malignancies. The SMARCB1 protein is a core component of the SWI/SNF (SWItch/Sucrose Non-Fermentable) family of chromatin remodeling complexes, which are important regulators of gene expression and cell differentiation. Here, we use CRISPR-Cas9 to create germline smarcb1 loss of function in zebrafish.

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Purpose: Most chimeric antigen receptor (CAR) T-cell strategies against glioblastoma have demonstrated only modest therapeutic activity and are based on persistent gene modification strategies that have limited transgene capacity, long manufacturing processes, and the risk for uncontrollable off-tumor toxicities. mRNA-based T-cell modifications are an emerging safe, rapid, and cost-effective alternative to overcome these challenges, but are underexplored against glioblastoma.

Experimental Design: We generated mouse and human mRNA-based multifunctional T cells coexpressing a multitargeting CAR based on the natural killer group 2D (NKG2D) receptor and the proinflammatory cytokines IL12 and IFNα2 and assessed their antiglioma activity in vitro and in vivo.

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  • Differential leukocyte counts can now be determined using cell-type-specific DNA methylation, offering potential benefits for small or frozen blood samples, but clinical application needs refinement.
  • The study optimized targeted DNA methylation assays using a range of blood samples from healthy individuals and patients with hematological diseases, comparing these results to conventional counts.
  • The findings showed a strong correlation between the new DNA methylation methods and traditional leukocyte counting methods, although some optimizations are still needed for accurate absolute quantification, especially in patients with blood cancers.
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HoxB8 multipotent progenitors (MPP) are obtained by expression of the estrogen receptor hormone binding domain (ERHBD) HoxB8 fusion gene in mouse BM cells. HoxB8 MPP generate (i) the full complement of DC subsets (cDC1, cDC2, and pDC) in vitro and in vivo and (ii) allow CRISPR/Cas9-mediated gene editing, for example, generating homozygous deletions in cis-acting DNA elements at high precision, and (iii) efficient gene repression by dCas9-KRAB for studying gene regulation in DC differentiation.

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  • Gene expression regulation in neuroblastoma is primarily influenced by the histone acetyltransferase EP300, while CBP plays a minimal role, especially in high-risk cases.
  • A novel compound, JQAD1, was developed to target and degrade EP300, leading to decreased enhancer acetylation and increased apoptosis of neuroblastoma cells.
  • JQAD1 shows limited toxicity to normal cells and its effectiveness is dependent on the expression of cereblon (CRBN) in neuroblastoma cells.
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Neuroblastoma cell identity depends on a core regulatory circuit (CRC) of transcription factors that collaborate with to drive the oncogenic gene expression program. For neuroblastomas dependent on the adrenergic CRC, treatment with retinoids can inhibit cell growth and induce differentiation. Here, we show that when -amplified neuroblastoma cells are treated with retinoic acid, histone H3K27 acetylation and methylation become redistributed to decommission super-enhancers driving the expression of and , together with the activation of new super-enhancers that drive high levels of and expression.

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