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In chronic lymphocytic leukemia (CLL), epigenetic alterations are considered to centrally shape the transcriptional signatures that drive disease evolution and underlie its biological and clinical subsets. Characterizations of epigenetic regulators, particularly histone-modifying enzymes, are very rudimentary in CLL. In efforts to establish effectors of the CLL-associated oncogene T-cell leukemia 1A (TCL1A), we identified here the lysine-specific histone demethylase KDM1A to interact with the TCL1A protein in B cells in conjunction with an increased catalytic activity of KDM1A. We demonstrate that KDM1A is upregulated in malignant B cells. Elevated KDM1A and associated gene expression signatures correlated with aggressive disease features and adverse clinical outcomes in a large prospective CLL trial cohort. Genetic Kdm1a knockdown in Eµ-TCL1A mice reduced leukemic burden and prolonged animal survival, accompanied by upregulated p53 and proapoptotic pathways. Genetic KDM1A depletion also affected milieu components (T, stromal, and monocytic cells), resulting in significant reductions in their capacity to support CLL-cell survival and proliferation. Integrated analyses of differential global transcriptomes (RNA sequencing) and H3K4me3 marks (chromatin immunoprecipitation sequencing) in Eµ-TCL1A vs iKdm1aKD;Eµ-TCL1A mice (confirmed in human CLL) implicate KDM1A as an oncogenic transcriptional repressor in CLL which alters histone methylation patterns with pronounced effects on defined cell death and motility pathways. Finally, pharmacologic KDM1A inhibition altered H3K4/9 target methylation and revealed marked anti-B-cell leukemic synergisms. Overall, we established the pathogenic role and effector networks of KDM1A in CLL via tumor-cell intrinsic mechanisms and its impacts in cells of the microenvironment. Our data also provide rationales to further investigate therapeutic KDM1A targeting in CLL.
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Leucemia Linfocítica Crónica de Células B , Humanos , Ratones , Animales , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Histonas/metabolismo , Lisina , Estudios Prospectivos , Histona Demetilasas/genética , Histona Demetilasas/metabolismo , Microambiente TumoralRESUMEN
BACKGROUND: Testicular germ cell tumours (TGCTs) have a high metastasis rate. However, the mechanisms related to their invasion, progression and metastasis are unclear. Therefore, we investigated gene expression changes that might be linked to metastasis in seminomatous testicular germ cell tumour (STGCT) patients. METHODS: Defined areas [invasive tumour front (TF) and tumour centre (TC)] of non-metastatic (with surveillance and recurrence-free follow-up >2 years) and metastatic STGCTs were collected separately using laser capture microdissection. The expression of 760 genes related to tumour progression and metastasis was analysed using nCounter technology and validated with quantitative real-time PCR and enzyme-linked immunosorbent assay. RESULTS: Distinct gene expression patterns were observed in metastatic and non-metastatic seminomas with respect to both the TF and TC. Comprehensive pathway analysis showed enrichment of genes related to tumour functions such as inflammation, angiogenesis and metabolism at the TF compared to the TC. Remarkably, prominent inflammatory and cancer-related pathways, such as interleukin-6 (IL-6) signalling, integrin signalling and nuclear factor-κB signalling, were significantly upregulated in the TF of metastatic vs non-metastatic tumours. CONCLUSIONS: IL-6 signalling was the most significantly upregulated pathway in metastatic vs non-metastatic tumours and therefore could constitute a therapeutic target for future personalised therapy. In addition, this is the first study showing intra- and inter-tumour heterogeneity in STGCT.
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Neoplasias de Células Germinales y Embrionarias , Seminoma , Neoplasias Testiculares , Perfilación de la Expresión Génica , Humanos , Inmunidad , Masculino , Neoplasias de Células Germinales y Embrionarias/genética , Seminoma/genética , Seminoma/metabolismo , Neoplasias Testiculares/patología , Regulación hacia ArribaRESUMEN
Cancer is one of the leading causes of death worldwide and well known for its complexity. Cancer cells within the same tumor or from different tumors are highly heterogeneous. Furthermore, stromal and immune cells within tumor microenvironment interact with cancer cells to play important roles in how tumors progress and respond to different treatments. Recent advances in single cell technologies, especially massively parallel single cell sequencing, have made it possible to analyze cancer cells and cells in its tumor microenvironment in parallel with unprecedented high resolution. In this chapter, we will review recent developments in single cell sequencing technologies and their applications in cancer research. We will also explain how insights generated from single cell sequencing can be used to develop novel diagnostic and therapeutic approaches to conquer cancer.
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Neoplasias/diagnóstico , Neoplasias/terapia , Análisis de Secuencia , Análisis de la Célula Individual , Humanos , Neoplasias/genética , Microambiente TumoralRESUMEN
We describe here use of a cell-permeable Cre to efficiently convert the EUCOMM/KOMP-CSD tm1a allele to the tm1b form in preimplantation mouse embryos in a high-throughput manner, consistent with the requirements of the International Mouse Phenotyping Consortium-affiliated NIH KOMP2 project. This method results in rapid allele conversion and minimizes the use of experimental animals when compared to conventional Cre transgenic mouse breeding, resulting in a significant reduction in costs and time with increased welfare benefits.
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Desarrollo Embrionario/genética , Integrasas/genética , Alelos , Animales , Cruzamiento , Embrión de Mamíferos , Células Madre Embrionarias , Ratones , Ratones TransgénicosRESUMEN
The Sanger Mouse Genetics Project generates knockout mice strains using the EUCOMM/KOMP-CSD embryonic stem (ES) cell collection and characterizes the consequences of the mutations using a high-throughput primary phenotyping screen. Upon achieving germline transmission, new strains are subject to a panel of quality control (QC) PCR- and qPCR-based assays to confirm the correct targeting, cassette structure, and the presence of the 3' LoxP site (required for the potential conditionality of the allele). We report that over 86 % of the 731 strains studied showed the correct targeting and cassette structure, of which 97 % retained the 3' LoxP site. We discuss the characteristics of the lines that failed QC and postulate that the majority of these may be due to mixed ES cell populations which were not detectable with the original screening techniques employed when creating the ES cell resource.
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Células Madre Embrionarias/citología , Células Germinativas/citología , Ratones Mutantes/genética , Animales , Cruzamiento , Ratones , Control de CalidadRESUMEN
INTRODUCTION: The underlying molecular mechanisms of parotid gland carcinomas (PGC) are still unknown. Knowledge about the tumor-driving signaling pathways is necessary either for diagnostics or developing new therapeutic options in this heterogeneous and rare entity. MATERIAL AND METHODS: 94 matching RNA formalin-fixed and paraffin-embedded tissue samples from PGC and the corresponding non-tumor area, RNA quality and quantity were sufficient for gene expression profiling of 770 genes using the NanoString's nCounter technology. Oncogenic and tumor suppressor genes were examined in the three common PGC tumor entities: adenoid cystic carcinoma (ACC), adenocarcinoma NOS (AC-NOS), and mucoepidermoid carcinoma (MEC). RESULTS: Expression profiling and subsequent hierarchical cluster analysis clearly differentiated between non-tumor gland tissue samples and PGC. In addition expression pattern of all three entities differed. The extensive pathway analysis proved a prominent dysregulation of the Wnt signaling pathway in the three PGC entities. Moreover, transcript upstream analysis demonstrated a pronounced activation of the PI3K pathway in ACC and MEC. DISCUSSION: Our findings revealed divergent molecular expression profiles in MEC, ACC and AC-NOS that are presently studied for their potential application in PGC diagnostics. Importantly, identification of Wnt and PI3K signaling in PGC revealed novel options of PGC therapy.
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BACKGROUND: Neuropathic pain (NP) is a debilitating condition that may result from spinal cord injury (SCI). Nearly 75% of all SCI results in NP affecting 17,000 new individuals in the United States every year, and an estimated 7-10% of people worldwide. It is caused by damaged or dysfunctional nerve fibers sending aberrant signals to pain centers in the central nervous system causing severe pain that affects daily life and routine. The mechanisms underlying NP are not fully understood, making treatment difficult. Identification of specific molecular pathways that are involved in pain syndromes and finding effective treatments has become a major priority in current SCI research. Yoga has therapeutic applications may prove beneficial in treating subjects suffering chronically with SCI induced NP, chronic back and associated pains if necessary experimental data is generated. SUMMARY: This review aims to discuss the implications of various mechanistic approaches of yoga which can be tested by new study designs around various nociceptive molecules including matrix metalloproteinases (MMPs), cation-dependent chloride transporter (NKCC1) etc in SCI induced NP patients. KEY MESSAGES: Thus, yogic practices could be used in managing SCI induced NP pain by regulating the action of various mechanisms and its associated molecules. Modern prescriptive treatment strategies combined with alternative approaches like yoga should be used in rehabilitation centers and clinics in order to ameliorate chronic NP. We recommend practical considerations of careful yoga practice as part of an integrative medicine approach for NP associated with SCI.
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Lysine-specific demethylase 1 (LSD1) is a histone modifier that is highly overexpressed in lung adenocarcinoma, which results in aggressive tumor biology. Tumor cell proliferation and migration analysis after LSD1 inhibition in the lung adenocarcinoma cell line PC9, using the LSD1 inhibitor HCI-2509 and siRNA, demonstrated that LSD1 activity was essential for proliferation and migration capacities of tumor cells. Moreover, reduced proliferation rates after LSD1 inhibition were shown to be associated with a cell-cycle arrest of the tumor cells in the G2-M-phase. Expression profiling followed by functional classification and pathway analysis indicated prominent repression of the polo-like kinase 1 (PLK1) pathway upon LSD1 inhibition. In contrast, transient overexpression of exogenous PLK1 plasmid rescued the LSD1 inhibition-mediated downregulation of PLK1 pathway genes. Mechanistically, LSD1 directly regulates expression of PLK1 by binding to its promoter region that subsequently affects expression of its downstream target genes. Notably, using lung adenocarcinoma TCGA datasets a significant correlation between LSD1 and PLK1 along with its downstream targets was observed. Furthermore, the LSD1/PLK1 linkage was confirmed by IHC analysis in a clinical lung adenocarcinoma cohort (n = 43). Conclusively, this is the first study showing a direct transcriptional link between LSD1 and PLK1. IMPLICATIONS: These findings point to a role of LSD1 in regulating PLK1 and thus efficient G2-M-transition-mediating proliferation of tumor cells and suggest targeting the LSD1/PLK1 axis as a novel therapeutic approach for lung adenocarcinoma treatment.
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Proteínas de Ciclo Celular/genética , Proliferación Celular/genética , Histona Demetilasas/genética , Mitosis/genética , Proteínas Serina-Treonina Quinasas/genética , Proteínas Proto-Oncogénicas/genética , Adenocarcinoma del Pulmón/genética , Adenocarcinoma del Pulmón/patología , Puntos de Control del Ciclo Celular/genética , Línea Celular Tumoral , Movimiento Celular/genética , Regulación hacia Abajo/genética , Puntos de Control de la Fase G2 del Ciclo Celular/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Regiones Promotoras Genéticas/genética , ARN Interferente Pequeño/genética , Transcripción Genética/genética , Quinasa Tipo Polo 1RESUMEN
Lung adenocarcinoma (LUAD) is the most prevalent subtype of non-small cell lung cancer. Despite the development of novel targeted and immune therapies, the 5-year survival rate is still only 21%, indicating the need for more efficient treatment regimens. Lysine-specific demethylase 1 (LSD1) is an epigenetic eraser that modifies histone 3 methylation status, and is highly overexpressed in LUAD. Using representative human cell culture systems and two autochthonous transgenic mouse models, we investigated inhibition of LSD1 as a novel therapeutic option for treating LUAD. The reversible LSD1 inhibitor HCI-2509 significantly reduced cell growth with an IC50 of 0.3-5 µmin vitro, which was linked to an enhancement of histone 3 lysine methylation. Most importantly, growth arrest, as well as inhibition of the invasion capacities, was independent of the underlying driver mutations. Subsequent expression profiling revealed that the cell cycle and replication machinery were prominently affected after LSD1 inhibition. In addition, our data provide evidence that LSD1 blockade significantly interferes with EGFR downstream signaling. Finally, our in vitro results were confirmed by preclinical therapeutic approaches, including the use of two autochthonous transgenic LUAD mouse models driven by either EGFR or KRAS mutations. Importantly, LSD1 inhibition resulted in significantly lower tumor formation and a strong reduction in tumor progression, which were independent of the underlying mutational background of the mouse models. Hence, our findings provide substantial evidence indicating that tumor growth of LUAD can be markedly decreased by HCI-2509 treatment, suggesting its use as a single agent maintenance therapy or combined therapeutical application in novel concerted drug approaches.
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Adenocarcinoma del Pulmón/tratamiento farmacológico , Inhibidores Enzimáticos/farmacología , Histona Demetilasas/antagonistas & inhibidores , Neoplasias Pulmonares/tratamiento farmacológico , Proteínas de Neoplasias/antagonistas & inhibidores , Neoplasias Experimentales/tratamiento farmacológico , Adenocarcinoma del Pulmón/enzimología , Adenocarcinoma del Pulmón/genética , Adenocarcinoma del Pulmón/patología , Animales , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Histona Demetilasas/genética , Histona Demetilasas/metabolismo , Humanos , Neoplasias Pulmonares/enzimología , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Ratones , Ratones Transgénicos , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias Experimentales/enzimología , Neoplasias Experimentales/genética , Neoplasias Experimentales/patologíaRESUMEN
A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.
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The epigenetic writer lysine-specific demethylase 1 (LSD1) is aberrantly upregulated in many cancer types and its overexpression correlates with poor survival and tumor progression. In this study, we analysed LSD1 function in non-small cell lung cancer adenocarcinomas. Expression profiling of 182 cases of lung adenocarcinoma proved a significant correlation of LSD1 overexpression with lung adenocarcinoma progression and metastasis. KRAS-mutated lung cancer cell clones were stably silenced for LSD1 expression. RNA-seq and comprehensive pathway analysis revealed, that genes related to a recently described non-canonical integrin ß3 pathway, were significantly downregulated by LSD1 silencing. Hence, invasion and self-renewal capabilities were strongly decreased. Notably, this novel defined LSD1/integrin ß3 axis, was also detected in human lung adenocarcinoma specimens. Furthermore, the linkage of LSD1 to an altered expression pattern of lung-lineage specific transcription factors and genes, which are involved in alveolar epithelial differentiation, was demonstrated. Thus, our findings point to a LSD1-integrin ß3 axis, conferring attributes of invasiveness and tumor progression to lung adenocarcinoma.