RESUMO
T-cell acute lymphocytic leukemia protein 1 (TAL1) is one of the most frequently deregulated oncogenes in T-cell acute lymphoblastic leukemia (T-ALL). Its deregulation can occur through diverse cis-alterations, including SIL-TAL1 microdeletions, translocations with T-cell Receptor loci, and more recently described upstream intergenic non-coding mutations. These mutations consist of recurrent focal microinsertions that create an oncogenic neo-enhancer accompanied by activating epigenetic marks. This observation laid the groundwork for an innovative paradigm concerning the activation of proto-oncogenes via genomic alterations of non-coding intergenic regions. However, for the majority of T-ALL expressing TAL1 (TAL1+), the deregulation mechanism remains 'unresolved'. We took advantage of H3K27ac and H3K4me3 chromatin immunoprecipitation sequencing data of eight cases of T-ALL, including five TAL1+ cases. We identified a putative novel oncogenic neo-enhancer downstream of TAL1 in an unresolved monoallelic TAL1+ case. A rare but recurrent somatic heterozygous microinsertion within this region creates a de novo binding site for MYB transcription factor. Here we demonstrate that this mutation leads to increased enhancer activity, gain of active epigenetic marks, and TAL1 activation via recruitment of MYB. These results highlight the diversity of non-coding mutations that can drive oncogene activation.
Assuntos
Elementos Facilitadores Genéticos , Leucemia-Linfoma Linfoblástico de Células T Precursoras , Proteína 1 de Leucemia Linfocítica Aguda de Células T , Humanos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Mutação , Proteínas de Fusão Oncogênica/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Proteína 1 de Leucemia Linfocítica Aguda de Células T/genética , Linfócitos T/metabolismo , Fatores de Transcrição/genéticaRESUMO
Liver sinusoidal endothelial cells (LSECs) control clearance of Transforming growth factor, beta-induced, 68kDa (TGFBi) and Periostin (POSTN) through scavenger receptors Stabilin-1 (Stab1) and Stabilin-2 (Stab2). Stabilin inhibition can ameliorate atherosclerosis in mouse models, while Stabilin-double-knockout leads to glomerulofibrosis. Fibrotic organ damage may pose a limiting factor in future anti-Stabilin therapies. While Stab1-deficient (Stab1-/-) mice were shown to exhibit higher liver fibrosis levels upon challenges, fibrosis susceptibility has not been studied in Stab2-deficient (Stab2-/-) mice. Wildtype (WT), Stab1-/- and Stab2-/- mice were fed experimental diets, and local ligand abundance, hepatic fibrosis, and ligand plasma levels were measured. Hepatic fibrosis was increased in both Stab1-/- and Stab2-/- at baseline. A pro-fibrotic short Methionine-Choline-deficient (MCD) diet induced slightly increased liver fibrosis in Stab1-/- and Stab2-/- mice. A Choline-deficient L-amino acid-defined (CDAA) diet induced liver fibrosis of similar distribution and extent in all genotypes (WT, Stab1-/- and Stab2-/-). A hepatic abundance of Stabilin ligand TGFBi correlated very highly with liver fibrosis levels. In contrast, plasma levels of TGFBi were increased only in Stab2-/- mice after the CDAA diet but not the MCD diet, indicating the differential effects of these diets. Here we show that a single Stabilin deficiency of either Stab1 or Stab2 induces mildly increased collagen depositions under homeostatic conditions. Upon experimental dietary challenge, the local abundance of Stabilin ligand TGFBi was differentially altered in Stabilin-deficient mice, indicating differentially affected LSEC scavenger functions. Since anti-Stabilin-directed therapies are in clinical evaluation for the treatment of diseases, these findings bear relevance to treatment with novel anti-Stabilin agents.
Assuntos
Células Endoteliais , Cirrose Hepática , Camundongos , Animais , Células Endoteliais/metabolismo , Ligantes , Cirrose Hepática/genética , Cirrose Hepática/metabolismo , Fígado/metabolismo , Metionina/metabolismo , Fatores de Crescimento Transformadores/metabolismo , Colina/metabolismo , Camundongos Endogâmicos C57BL , Moléculas de Adesão Celular Neuronais/genética , Moléculas de Adesão Celular Neuronais/metabolismoRESUMO
Accurate classification of melanocytic tumors is important for prognostic evaluation, treatment and follow-up protocols of patients. The majority of melanocytic proliferations can be classified solely based on clinical and pathological criteria, however in select cases a definitive diagnostic assessment remains challenging and additional diagnostic biomarkers would be advantageous. We analyzed melanomas, nevi, Spitz nevi and atypical spitzoid tumors using parallel sequencing (exons of 611 genes and 507 gene translocation analysis) and methylation arrays (850k Illumina EPIC). By combining detailed genetic and epigenetic analysis with reference-based and reference-free DNA methylome deconvolution we compared Spitz nevi to nevi and melanoma and assessed the potential for these methods in classifying challenging spitzoid tumors. Results were correlated with clinical and histologic features. Spitz nevi were found to cluster independently of nevi and melanoma and demonstrated a different mutation profile. Multiple copy number alterations and TERT promoter mutations were identified only in melanomas. Genome-wide methylation in Spitz nevi was comparable to benign nevi while the Leukocytes UnMethylation for Purity (LUMP) algorithm in Spitz nevi was comparable to melanoma. Histologically difficult to classify Spitz tumor cases were assessed which, based on methylation arrays, clustered between Spitz nevi and melanoma and in terms of genetic profile or copy number variations demonstrated worrisome features suggesting a malignant neoplasm. Comprehensive sequencing and methylation analysis verify Spitz nevi as an independent melanocytic entity distinct from both nevi and melanoma. Combined genetic and methylation assays can offer additional insights in diagnosing difficult to classify Spitzoid tumors.
Assuntos
Melanoma , Nevo de Células Epitelioides e Fusiformes , Paraganglioma , Neoplasias Cutâneas , Variações do Número de Cópias de DNA , Diagnóstico Diferencial , Humanos , Melanoma/diagnóstico , Melanoma/genética , Melanoma/patologia , Metilação , Nevo de Células Epitelioides e Fusiformes/diagnóstico , Nevo de Células Epitelioides e Fusiformes/genética , Neoplasias Cutâneas/diagnóstico , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , SíndromeRESUMO
Stabilin-1 (Stab1) and Stabilin-2 (Stab2) are scavenger receptors expressed by liver sinusoidal endothelial cells (LSECs). The Stabilin-mediated scavenging function is responsible for regulating the molecular composition of circulating blood in mammals. Stab1 and Stab2 have been shown to influence fibrosis in liver and kidneys and to modulate inflammation in atherosclerosis. In this context, circulating and localized TGFBi and POSTN are differentially controlled by the Stabilins as their receptors. To assess Stab1 and Stab2 functions in inflammatory and fibrotic skin disease, topical Imiquimod (IMQ) was used to induce psoriasis-like skin lesions in mice and Bleomycin (BLM) was applied subcutaneously to induce scleroderma-like effects in the skin. The topical treatment with IMQ, as expected, led to psoriasis-like changes in the skin of mice, including increased epidermal thickness and significant weight loss. Clinical severity was reduced in Stab2-deficient compared to Stab1-deficient mice. We did not observe differential effects in the skin of Stabilin-deficient mice after bleomycin injection. Interestingly, treatment with IMQ led to a significant increase of Stabilin ligand TGFBi plasma levels in Stab2-/- mice, treatment with BLM resulted in a significant decrease in TGFBi levels in Stab1-/- mice. Overall, Stab1 and Stab2 deficiency resulted in minor alterations of the disease phenotypes accompanied by alterations of circulating ligands in the blood in response to the disease models. Stabilin-mediated clearance of TGFBi was altered in these disease processes. Taken together our results suggest that Stabilin deficiency-associated plasma alterations may interfere with preclinical disease severity and treatment responses in patients.
RESUMO
Acute myeloid leukemia (AML) with the t(7;12)(q36;p13) translocation occurs only in very young children and has a poor clinical outcome. The expected oncofusion between breakpoint partners (MNX1 and ETV6) has only been reported in a subset of cases. However, a universal feature is the strong transcript and protein expression of MNX1, a homeobox transcription factor that is normally not expressed in hematopoietic cells. Here, we map the translocation breakpoints on chromosomes 7 and 12 in affected patients to a region proximal to MNX1 and either introns 1 or 2 of ETV6. The frequency of MNX1 overexpression in pediatric AML (n=1556, own and published data) is 2.4% and occurs predominantly in t(7;12)(q36;p13) AML. Chromatin interaction assays in a t(7;12)(q36;p13) iPSC cell line model unravel an enhancer-hijacking event that explains MNX1 overexpression in hematopoietic cells. Our data suggest that enhancer-hijacking may be a more widespread consequence of translocations where no oncofusion product was identified, including e.g. t(1;3) or t(4;12) AML.
RESUMO
Feather pecking and cannibalism are prominent problems in modern laying hen husbandry. Among the various approaches to address this issue, environmental enrichment plays a crucial role. In this on-farm study, four winter gardens of an organic farm henhouse were equipped with an automatic enrichment device. Different quantities of downpipes dosing grain on rough-coated pecking plates (PPs) were tested. One group served as a control (CG) without an automatic enrichment device, while the others were offered different numbers of PPs, with one equipped with a doubled amount of PPs (DEG) compared to the other two groups (SEG). Video analyses of the hens' duration of stay and behaviors in the recorded winter garden area and around the PPs were performed, and regular assessments of the plumage condition were conducted. By the end of the production cycle, no hens with intact plumage were found, with hens in the CG and DEG showing worse scores and earlier deterioration in the plumage condition than in the SEG. The offer of PPs showed a significant influence on the duration of stay in the filmed area. Hens in the DEG stayed significantly longer (mean: 129 s, SD: 126 s) than those in the CG (mean: 79 s, SD: 91 s; p < 0.05) and SEG (mean: 75 s, SD: 83 s; p < 0.005). On the performed behavioral bouts per hen and minute (CG mean bouts/minute (SD): 5.47 (2.92); SEG mean (SD): 5.33 (2.76); SEG mean (SD): 5.81 (3.24)), no significant influences were detected. Environmental pecking was the behavior most frequently observed in all winter gardens, where, particularly around the PPs, pecking at the device was observed. Therefore, the enrichment device can be assessed as well accepted by the hens in winter gardens. The effect of the device on the plumage condition remains unclear, with external factors probably showing a greater influence than the enrichment.
RESUMO
Stabilin-1 (Stab1) and Stabilin-2 (Stab2) are two major scavenger receptors of liver sinusoidal endothelial cells that mediate removal of diverse molecules from the plasma. Double-knockout mice (Stab-DKO) develop impaired kidney function and a decreased lifespan, while single Stabilin deficiency or therapeutic inhibition ameliorates atherosclerosis and Stab1-inhibition is subject of clinical trials in immuno-oncology. Although POSTN and TFGBI have recently been described as novel Stabilin ligands, the dynamics and functional implications of these ligands have not been comprehensively studied. Immunofluorescence, Western Blotting and Simple Western™ as well as in situ hybridization (RNAScope™) and qRT-PCR were used to analyze transcription levels and tissue distribution of POSTN and TGFBI in Stab-KO mice. Stab-POSTN-Triple deficient mice were generated to assess kidney and liver fibrosis and function in young and aged mice. TGFBI and POSTN protein accumulated in liver tissue in Stab-DKO mice and age-dependent in glomeruli of Stabilin-deficient mice despite unchanged transcriptional levels. Stab-POSTN-Triple KO mice showed glomerulofibrosis and a reduced lifespan comparable to Stab-DKO mice. However, alterations of the glomerular diameter and vascular density were partially normalized in Stab-POSTN-Triple KO. TGFBI and POSTN are Stabilin-ligands that are deposited in an age-dependent manner in the kidneys and liver due to insufficient scavenging in the liver. Functionally, POSTN might partially contribute to the observed renal phenotype in Stab-DKO mice. This study provides details on downstream effects how Stabilin dysfunction affects organ function on a molecular and functional level.
Assuntos
Moléculas de Adesão Celular Neuronais , Células Endoteliais , Animais , Camundongos , Moléculas de Adesão Celular Neuronais/genética , Moléculas de Adesão Celular Neuronais/metabolismo , Células Endoteliais/metabolismo , Rim/metabolismo , Ligantes , Fígado/metabolismo , Camundongos Knockout , Receptores Depuradores/metabolismoRESUMO
To characterize the genomic landscape and leukemogenic pathways of older, newly diagnosed, non-intensively treated patients with AML and to study the clinical implications, comprehensive genetics analyses were performed including targeted DNA sequencing of 263 genes in 604 patients treated in a prospective Phase III clinical trial. Leukemic trajectories were delineated using oncogenetic tree modeling and hierarchical clustering, and prognostic groups were derived from multivariable Cox regression models. Clonal hematopoiesis-related genes (ASXL1, TET2, SRSF2, DNMT3A) were most frequently mutated. The oncogenetic modeling algorithm produced a tree with five branches with ASXL1, DDX41, DNMT3A, TET2, and TP53 emanating from the root suggesting leukemia-initiating events which gave rise to further subbranches with distinct subclones. Unsupervised clustering mirrored the genetic groups identified by the tree model. Multivariable analysis identified FLT3 internal tandem duplications (ITD), SRSF2, and TP53 mutations as poor prognostic factors, while DDX41 mutations exerted an exceptionally favorable effect. Subsequent backwards elimination based on the Akaike information criterion delineated three genetic risk groups: DDX41 mutations (favorable-risk), DDX41wildtype/FLT3-ITDneg/TP53wildtype (intermediate-risk), and FLT3-ITD or TP53 mutations (high-risk). Our data identified distinct trajectories of leukemia development in older AML patients and provide a basis for a clinically meaningful genetic outcome stratification for patients receiving less intensive therapies.
Assuntos
Leucemia Mieloide Aguda , Nucleofosmina , Humanos , Idoso , Estudos Prospectivos , Leucemia Mieloide Aguda/terapia , Leucemia Mieloide Aguda/tratamento farmacológico , Mutação , Prognóstico , Genômica , Fatores de Transcrição/genética , Tirosina Quinase 3 Semelhante a fms/genética , Tirosina Quinase 3 Semelhante a fms/uso terapêuticoRESUMO
Melanocytic neoplasms have been genetically characterized in detail during the last decade. Recurrent CTNNB1 exon 3 mutations have been recognized in the distinct group of melanocytic tumors showing deep penetrating nevus-like morphology. In addition, they have been identified in 1-2% of advanced melanoma. Performing a detailed genetic analysis of difficult-to-classify nevi and melanomas with CTNNB1 mutations, we found that benign tumors (nevi) show characteristic morphological, genetic and epigenetic traits, which distinguish them from other nevi and melanoma. Malignant CTNNB1-mutant tumors (melanomas) demonstrated a different genetic profile, instead grouping clearly with other non-CTNNB1 melanomas in methylation assays. To further evaluate the role of CTNNB1 mutations in melanoma, we assessed a large cohort of clinically sequenced melanomas, identifying 38 tumors with CTNNB1 exon 3 mutations, including recurrent S45 (n = 13, 34%), G34 (n = 5, 13%), and S27 (n = 5, 13%) mutations. Locations and histological subtype of CTNNB1-mutated melanoma varied; none were reported as showing deep penetrating nevus-like morphology. The most frequent concurrent activating mutations were BRAF V600 (n = 21, 55%) and NRAS Q61 (n = 13, 34%). In our cohort, four of seven (58%) and one of nine (11%) patients treated with targeted therapy (BRAF and MEK Inhibitors) or immune-checkpoint therapy, respectively, showed disease control (partial response or stable disease). In summary, CTNNB1 mutations are associated with a unique melanocytic tumor type in benign tumors (nevi), which can be applied in a diagnostic setting. In advanced disease, no clear characteristics distinguishing CTNNB1-mutant from other melanomas were observed; however, studies of larger, optimally prospective, cohorts are warranted.
RESUMO
Intercellular communication plays an essential role in lung cancer (LC). One of the major players in cell-cell-communication is small extracellular vesicles (sEV). SEV trigger various biological responses by transporting cellular cargo to target cells. One essential sEV component are microRNAs (miRs), whose transport has recently attracted increasing research interest. We report that prostaglandin E2 (PGE2 ), a key inflammatory lipid mediator, specifically induces the sorting of miR-574-5p in sEV of A549 and 2106T cells. We found that sEV-derived miR-574-5p activates Toll-like receptors (TLR) 7/8, thereby decreasing PGE2 -levels. In contrast, intracellular miR-574-5p induces PGE2 -biosynthesis. Consequently, the combination of intracellular and sEV-derived miR-574-5p controls PGE2 -levels via a feedback loop. This was only observed in adeno- but not in squamous cell carcinoma, indicating a cell-specific response to sEV-derived miRs, which might be due to unique tetraspanin compositions. Hence, we describe a novel function of miR-574-5p unique to adenocarcinoma. Intracellular miR-574-5p induces PGE2 and thus the secretion of sEV-derived miR-574-5p, which in turn decreases PGE2 -biosynthesis in recipient cells.