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Adipose tissues serve as an energy reservoir and endocrine organ, yet the mechanisms that coordinate these functions remain elusive. Here, we show that the transcriptional coregulators, YAP and TAZ, uncouple fat mass from leptin levels and regulate adipocyte plasticity to maintain metabolic homeostasis. Activating YAP/TAZ signalling in adipocytes by deletion of the upstream regulators Lats1 and Lats2 results in a profound reduction in fat mass by converting mature adipocytes into delipidated progenitor-like cells, but does not cause lipodystrophy-related metabolic dysfunction, due to a paradoxical increase in circulating leptin levels. Mechanistically, we demonstrate that YAP/TAZ-TEAD signalling upregulates leptin expression by directly binding to an upstream enhancer site of the leptin gene. We further show that YAP/TAZ activity is associated with, and functionally required for, leptin regulation during fasting and refeeding. These results suggest that adipocyte Hippo-YAP/TAZ signalling constitutes a nexus for coordinating adipose tissue lipid storage capacity and systemic energy balance through the regulation of adipocyte plasticity and leptin gene transcription.
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Proteínas Adaptadoras de Transdução de Sinal , Adipócitos , Tecido Adiposo , Metabolismo Energético , Via de Sinalização Hippo , Leptina , Proteínas Serina-Treonina Quinases , Transdução de Sinais , Proteínas de Sinalização YAP , Animais , Leptina/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Camundongos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas de Sinalização YAP/metabolismo , Tecido Adiposo/metabolismo , Adipócitos/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional/metabolismo , Fosfoproteínas/metabolismo , Fosfoproteínas/genética , Proteínas Supressoras de Tumor/metabolismo , Proteínas Supressoras de Tumor/genética , Transativadores/metabolismo , Transativadores/genéticaRESUMO
Leucine-rich repeat kinase 2 (LRRK2), a large GTP-regulated serine/threonine kinase, is well-known for its mutations causing late-onset Parkinson's disease. However, the role of LRRK2 in glioblastoma (GBM) carcinogenesis has not yet been fully elucidated. Here, we discovered that LRRK2 was overexpressed in 40% of GBM patients, according to tissue microarray analysis, and high LRRK2 expression correlated with poor prognosis in GBM patients. LRRK2 and stemness factors were highly expressed in various patient-derived GBM stem cells, which are responsible for GBM initiation. Canonical serum-induced differentiation decreased the expression of both LRRK2 and stemness factors. Given that LRRK2 is a key regulator of glioma stem cell (GSC) stemness, we developed DNK72, a novel LRRK2 kinase inhibitor that penetrates the blood-brain barrier. DNK72 binds to the phosphorylation sites of active LRRK2 and dramatically reduced cell proliferation and stemness factors expression in in vitro studies. Orthotopic patient-derived xenograft mouse models demonstrated that LRRK2 inhibition with DNK72 effectively reduced tumor growth and increased survival time. We propose that LRRK2 plays a significant role in regulating the stemness of GSCs and that suppression of LRRK2 kinase activity leads to reduced GBM malignancy and proliferation. In the near future, targeting LRRK2 in patients with high LRRK2-expressing GBM could offer a superior therapeutic strategy and potentially replace current clinical treatment methods.
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OBJECTIVE: The purpose of this study was to analyze the clinical effects of orbicularis oculi muscle suspension in conjunction with transcutaneous blepharoplasty in patients 60 years of age or older. METHODS: A retrospective study was conducted on patients aged 60 or older who underwent orbicularis oculi flap suspension in conjunction with transcutaneous blepharoplasty to treat lower lid festoons, lower lid fat prolapse, and lid laxity at Korea University Guro Hospital. The outcomes were surgical success rate, recurrence rate, postoperative complications, and patient satisfaction. RESULTS: In total, 39 patients (18 males and 21 females) were included, with an average age of 67.5 years (60-86 y) and a follow-up period of 145.7 days (95-195 d). In all patients, eyelid laxity was effectively corrected, and there was no recurrence of eyelid laxity after surgery. Three patients suffered postoperative complications, transient skin edema in 2 patients, and transient ectropion in 1 patient. Subjective satisfaction score following surgery was high, with an average score of 2.56 out of 3 points. CONCLUSIONS: Orbicularis oculi flap suspension with transcutaneous blepharoplasty can shorten operation time due to surgical ease. It also has the advantage of not involving conjunctival-related complications such as conjunctival edema since it does not pass through the conjunctiva. Finally, it can successfully treat lid laxity and could be used in lower blepharoplasty procedures.
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Blefaroplastia , Masculino , Feminino , Humanos , Idoso , Pessoa de Meia-Idade , Blefaroplastia/métodos , Estudos Retrospectivos , Pálpebras/cirurgia , Edema/cirurgia , Complicações Pós-Operatórias/prevenção & controle , Complicações Pós-Operatórias/cirurgiaRESUMO
Lateral tarsal strip (LTS) is a simple surgical technique that can correct eyelid malposition. Clinical indications vary from involutional entropion and ectropion, lower eyelid laxity, and lower eyelid retraction to punctal and paralytic ectropion. Lateral tarsal strip mainly treats eyelid malposition by correcting horizontal laxity. Herein, the authors report the clinical indications for and effects of LTS. The authors retrospectively reviewed medical records of patients who underwent LTS by a single oculoplastic surgeon with 8 years of experience performing the procedure at Korea University Guro Hospital. The authors included 247 patients with 325 treated eyelids. Involutional entropion was the most common indication, accounting for 69 patients (27.94%) and 88 eyelids (27.08%). The second most common indication was lower eyelid laxity [n=44 patients (17.81%) and 68 eyelids (20.92%)]. Simultaneous surgery in addition to LTS was performed in 167 patients (67.6%) and 219 eyelids (67.4%); the most common was endoscopic dacryocystorhinostomy (DCR), which was performed in 50 patients (29.94%) and 80 eyelids (36.53%). Lateral tarsal strip can be performed alone or simultaneously with oculoplastic procedures for various indications. Overcorrection with fixation above the canthal angle is useful to reduce recurrence. This study aims to explain the clinical importance of the LTS procedure.
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Obesity-mediated hypoxic stress underlies inflammation, including interferon (IFN)-γ production by natural killer (NK) cells in white adipose tissue. However, the effects of obesity on NK cell IFN-γ production remain obscure. Here, we show that hypoxia promotes xCT-mediated glutamate excretion and C-X-C motif chemokine ligand 12 (CXCL12) expression in white adipocytes, resulting in CXCR4+ NK cell recruitment. Interestingly, this spatial proximity between adipocytes and NK cells induces IFN-γ production in NK cells by stimulating metabotropic glutamate receptor 5 (mGluR5). IFN-γ then triggers inflammatory activation of macrophages and augments xCT and CXCL12 expression in adipocytes, forming a bidirectional pathway. Genetic or pharmacological inhibition of xCT, mGluR5, or IFN-γ receptor in adipocytes or NK cells alleviates obesity-related metabolic disorders in mice. Consistently, patients with obesity showed elevated levels of glutamate/mGluR5 and CXCL12/CXCR4 axes, suggesting that a bidirectional pathway between adipocytes and NK cells could be a viable therapeutic target in obesity-related metabolic disorders.
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Adipócitos Brancos , Ácido Glutâmico , Interferon gama , Obesidade , Animais , Humanos , Camundongos , Adipócitos Brancos/metabolismo , Ácido Glutâmico/metabolismo , Interferon gama/metabolismo , Células Matadoras Naturais/metabolismo , Obesidade/metabolismo , Sistema y+ de Transporte de Aminoácidos/metabolismoRESUMO
BACKGROUND & AIMS: Mitochondrial dysfunction disrupts the synthesis and secretion of digestive enzymes in pancreatic acinar cells and plays a primary role in the etiology of exocrine pancreas disorders. However, the transcriptional mechanisms that regulate mitochondrial function to support acinar cell physiology are poorly understood. Here, we aim to elucidate the function of estrogen-related receptor γ (ERRγ) in pancreatic acinar cell mitochondrial homeostasis and energy production. METHODS: Two models of ERRγ inhibition, GSK5182-treated wild-type mice and ERRγ conditional knock-out (cKO) mice, were established to investigate ERRγ function in the exocrine pancreas. To identify the functional role of ERRγ in pancreatic acinar cells, we performed histologic and transcriptome analysis with the pancreas isolated from ERRγ cKO mice. To determine the relevance of these findings for human disease, we analyzed transcriptome data from multiple independent human cohorts and conducted genetic association studies for ESRRG variants in 2 distinct human pancreatitis cohorts. RESULTS: Blocking ERRγ function in mice by genetic deletion or inverse agonist treatment results in striking pancreatitis-like phenotypes accompanied by inflammation, fibrosis, and cell death. Mechanistically, loss of ERRγ in primary acini abrogates messenger RNA expression and protein levels of mitochondrial oxidative phosphorylation complex genes, resulting in defective acinar cell energetics. Mitochondrial dysfunction due to ERRγ deletion further triggers autophagy dysfunction, endoplasmic reticulum stress, and production of reactive oxygen species, ultimately leading to cell death. Interestingly, ERRγ-deficient acinar cells that escape cell death acquire ductal cell characteristics, indicating a role for ERRγ in acinar-to-ductal metaplasia. Consistent with our findings in ERRγ cKO mice, ERRγ expression was significantly reduced in patients with chronic pancreatitis compared with normal subjects. Furthermore, candidate locus region genetic association studies revealed multiple single nucleotide variants for ERRγ that are associated with chronic pancreatitis. CONCLUSIONS: Collectively, our findings highlight an essential role for ERRγ in maintaining the transcriptional program that supports acinar cell mitochondrial function and organellar homeostasis and provide a novel molecular link between ERRγ and exocrine pancreas disorders.
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Pâncreas Exócrino , Pancreatite Crônica , Células Acinares/patologia , Animais , Estrogênios/metabolismo , Humanos , Camundongos , Camundongos Knockout , Pâncreas/patologia , Pâncreas Exócrino/metabolismo , Pancreatite Crônica/patologiaRESUMO
Insulin resistance is a cornerstone of obesity-related complications such as type 2 diabetes, metabolic syndrome, and nonalcoholic fatty liver disease. A high rate of lipolysis is known to be associated with insulin resistance, and inhibiting adipose tissue lipolysis improves obesity-related insulin resistance. Here, we demonstrate that inhibition of serotonin (5-hydroxytryptamine [5-HT]) signaling through serotonin receptor 2B (HTR2B) in adipose tissues ameliorates insulin resistance by reducing lipolysis in visceral adipocytes. Chronic high-fat diet (HFD) feeding increased Htr2b expression in epididymal white adipose tissue, resulting in increased HTR2B signaling in visceral white adipose tissue. Moreover, HTR2B expression in white adipose tissue was increased in obese humans and positively correlated with metabolic parameters. We further found that adipocyte-specific Htr2b-knockout mice are resistant to HFD-induced insulin resistance, visceral adipose tissue inflammation, and hepatic steatosis. Enhanced 5-HT signaling through HTR2B directly activated lipolysis through phosphorylation of hormone-sensitive lipase in visceral adipocytes. Moreover, treatment with a selective HTR2B antagonist attenuated HFD-induced insulin resistance, visceral adipose tissue inflammation, and hepatic steatosis. Thus, adipose HTR2B signaling could be a potential therapeutic target for treatment of obesity-related insulin resistance.
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Resistência à Insulina , Gordura Intra-Abdominal/metabolismo , Obesidade/metabolismo , Receptor 5-HT2B de Serotonina/metabolismo , Serotonina/metabolismo , Adipócitos/citologia , Adipócitos Brancos , Tecido Adiposo , Tecido Adiposo Branco/metabolismo , Adulto , Animais , Dieta Hiperlipídica , Epididimo , Feminino , Glicerol/metabolismo , Humanos , Inflamação , Insulina/metabolismo , Lipólise , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Fosforilação , Transdução de Sinais , Adulto JovemRESUMO
Secretory proteins are an essential component of interorgan communication networks that regulate animal physiology. Current approaches for identifying secretory proteins from specific cell and tissue types are largely limited to in vitro or ex vivo models which often fail to recapitulate in vivo biology. As such, there is mounting interest in developing in vivo analytical tools that can provide accurate information on the origin, identity, and spatiotemporal dynamics of secretory proteins. Here, we describe iSLET (in situ Secretory protein Labeling via ER-anchored TurboID) which selectively labels proteins that transit through the classical secretory pathway via catalytic actions of Sec61b-TurboID, a proximity labeling enzyme anchored in the ER lumen. To validate iSLET in a whole-body system, we express iSLET in the mouse liver and demonstrate efficient labeling of liver secretory proteins which could be tracked and identified within circulating blood plasma. Furthermore, proteomic analysis of the labeled liver secretome enriched from liver iSLET mouse plasma is highly consistent with previous reports of liver secretory protein profiles. Taken together, iSLET is a versatile and powerful tool for studying spatiotemporal dynamics of secretory proteins, a valuable class of biomarkers and therapeutic targets.
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Retículo Endoplasmático/metabolismo , Canais de Translocação SEC/metabolismo , Via Secretória/fisiologia , Animais , Células HEK293 , Células HeLa , Células Hep G2 , Humanos , Fígado/metabolismo , Fígado/patologia , Camundongos , Camundongos Endogâmicos C57BL , Células NIH 3T3 , Proteoma/metabolismo , ProteômicaRESUMO
PTEN is one of the most frequently altered tumor suppressor genes in malignant tumors. The dominant-negative effect of PTEN alteration suggests that the aberrant function of PTEN mutation might be more disastrous than deletion, the most frequent genomic event in glioblastoma (GBM). This study aimed to understand the functional properties of various PTEN missense mutations and to investigate their clinical relevance. The genomic landscape of PTEN alteration was analyzed using the Samsung Medical Center GBM cohort and validated via The Cancer Genome Atlas dataset. Several hotspot mutations were identified, and their subcellular distributions and phenotypes were evaluated. We established a library of cancer cell lines that overexpress these mutant proteins using the U87MG and patient-derived cell models lacking functional PTEN. PTEN mutations were categorized into two major subsets: missense mutations in the phosphatase domain and truncal mutations in the C2 domain. We determined the subcellular compartmentalization of four mutant proteins (H93Y, C124S, R130Q, and R173C) from the former group and found that they had distinct localizations; those associated with invasive phenotypes ('edge mutations') localized to the cell periphery, while the R173C mutant localized to the nucleus. Invasive phenotypes derived from edge substitutions were unaffected by an anti-PI3K/Akt agent but were disrupted by microtubule inhibitors. PTEN mutations exhibit distinct functional properties regarding their subcellular localization. Further, some missense mutations ('edge mutations') in the phosphatase domain caused enhanced invasiveness associated with dysfunctional cytoskeletal assembly, thus suggesting it to be a potent therapeutic target.
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Glioblastoma/genética , Oncogenes/genética , PTEN Fosfo-Hidrolase/metabolismo , Humanos , MutaçãoRESUMO
In patients with advanced-stage cancer, cancer-associated anorexia affects treatment success and patient survival. However, the underlying mechanism is poorly understood. Here, we show that Dilp8, a Drosophila homologue of mammalian insulin-like 3 peptide (INSL3), is secreted from tumour tissues and induces anorexia through the Lgr3 receptor in the brain. Activated Dilp8-Lgr3 signalling upregulated anorexigenic nucleobinding 1 (NUCB1) and downregulated orexigenic short neuropeptide F (sNPF) and NPF expression in the brain. In the cancer condition, the protein expression of Lgr3 and NUCB1 was significantly upregulated in neurons expressing sNPF and NPF. INSL3 levels were increased in tumour-implanted mice and INSL3-treated mouse hypothalamic cells showed Nucb2 upregulation and Npy downregulation. Food consumption was significantly reduced in intracerebrospinal INSL3-injected mice. In patients with pancreatic cancer, higher serum INSL3 levels increased anorexia. These results indicate that tumour-derived Dilp8/INSL3 induces cancer anorexia by regulating feeding hormones through the Lgr3/Lgr8 receptor in Drosophila and mammals.
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Anorexia/metabolismo , Encéfalo/metabolismo , Proteínas de Drosophila/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Neoplasias/metabolismo , Neuropeptídeos/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Sequência de Aminoácidos , Animais , Anorexia/etiologia , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Linhagem Celular Tumoral , Modelos Animais de Doenças , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Drosophila melanogaster/metabolismo , Neoplasias Oculares/patologia , Comportamento Alimentar , Humanos , Hipotálamo/metabolismo , Insulina/sangue , Insulina/química , Insulina/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/química , Peptídeos e Proteínas de Sinalização Intercelular/genética , Camundongos Endogâmicos C57BL , Neoplasias/complicações , Neurônios/metabolismo , Neoplasias Pancreáticas/sangue , Neoplasias Pancreáticas/complicações , Proteínas/química , Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de SinaisRESUMO
Diffusely infiltrating gliomas (DIGs) are difficult to completely resect and are associated with a high rate of tumor relapse and progression from low- to high-grade glioma. In particular, optimized short-term culture-enriching patient-derived glioma stem cells (GSCs) are essential for customizing the therapeutic strategy based on clinically feasible in vitro drug screening for a wide range of DIGs, owing to the high inter-tumoral heterogeneity. Herein, we constructed a novel high-throughput culture condition screening platform called 'GFSCAN', which evaluated the cellular growth rates of GSCs for each DIG sample in 132 serum-free combinations, using 13 previously reported growth factors closely associated with glioma aggressiveness. In total, 72 patient-derived GSCs with available genomic profiles were tested in GFSCAN to explore the association between cellular growth rates in specific growth factor combinations and genomic/molecular backgrounds, including isocitrate dehydrogenase 1 (IDH1) mutation, chromosome arm 1p and 19q co-deletion, ATRX chromatin remodeler alteration, and transcriptional subtype. GSCs were clustered according to the dependency on epidermal growth factor and basic fibroblast growth factor (E&F), and isocitrate dehydrogenase 1 (IDH1) wild-type GSCs showed higher E&F dependencies than IDH1 mutant GSCs. More importantly, we elucidated optimal combinations for IDH1 mutant glioblastoma and lower grade glioma GSCs with low dependencies on E&F, which could be an aid in clinical decision-making for these DIGs. Thus, we demonstrated the utility of GFSCAN in personalizing in vitro cultivation to nominate personalized therapeutic options, in a clinically relevant time frame, for individual DIG patients, where standard clinical options have been exhausted.
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PURPOSE: Targeted next-generation sequencing (NGS) panels for solid tumors have been useful in clinical framework for accurate tumor diagnosis and identifying essential molecular aberrations. However, most cancer panels have been designed to address a wide spectrum of pan-cancer models, lacking integral prognostic markers that are highly specific to gliomas. MATERIALS AND METHODS: To address such challenges, we have developed a glioma-specific NGS panel, termed "GliomaSCAN," that is capable of capturing single nucleotide variations and insertion/deletion, copy number variation, and selected promoter mutations and structural variations that cover a subset of intron regions in 232 essential glioma-associated genes. We confirmed clinical concordance rate using pairwise comparison of the identified variants from whole exome sequencing (WES), immunohistochemical analysis, and fluorescence in situ hybridization. RESULTS: Our panel demonstrated high sensitivity in detecting potential genomic variants that were present in the standard materials. To ensure the accuracy of our targeted sequencing panel, we compared our targeted panel to WES. The comparison results demonstrated a high correlation. Furthermore, we evaluated clinical utility of our panel in 46 glioma patients to assess the detection capacity of potential actionable mutations. Thirty-two patients harbored at least one recurrent somatic mutation in clinically actionable gene. CONCLUSION: We have established a glioma-specific cancer panel. GliomaSCAN highly excelled in capturing somatic variations in terms of both sensitivity and specificity and provided potential clinical implication in facilitating genome-based clinical trials. Our results could provide conceptual advance towards improving the response of genomically guided molecularly targeted therapy in glioma patients.
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Biomarcadores Tumorais , Testes Genéticos , Glioma/diagnóstico , Glioma/genética , Sequenciamento de Nucleotídeos em Larga Escala , Mutação , Alelos , Variações do Número de Cópias de DNA , Diagnóstico Diferencial , Feminino , Frequência do Gene , Estudos de Associação Genética , Predisposição Genética para Doença , Testes Genéticos/métodos , Genômica/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Imuno-Histoquímica , Hibridização in Situ Fluorescente , Masculino , Sequenciamento do ExomaRESUMO
Glioblastoma (GBM) is the most lethal primary brain tumor with few treatment options. The survival of glioma-initiating cells (GICs) is one of the major factors contributing to treatment failure. GICs frequently produce and respond to their own growth factors that support cell proliferation and survival. In this study, we aimed to identify critical autocrine factors mediating GIC survival and to evaluate the anti-GBM effect of antagonizing these factors. Proteomic analysis was performed using conditioned media from two different patient-derived GBM tumor spheres under a growth factor-depleted status. Then, the antitumor effects of inhibiting an identified autocrine factor were evaluated by bioinformatic analysis and molecular validation. Proteins secreted by sphere-forming GICs promote cell proliferation/survival and detoxify reactive oxygen species (ROS). Among these proteins, we focused on midkine (MDK) as a clinically significant and pathologically relevant autocrine factor. Antagonizing MDK reduced the survival of GBM tumor spheres through the promotion of cell cycle arrest and the consequent apoptotic cell death caused by oxidative stress-induced DNA damage. We also identified PCBP4, a novel molecular predictor of resistance to anti-MDK treatment. Collectively, our results indicate that MDK inhibition is an important therapeutic option by suppressing GIC survival through the induction of ROS-mediated cell cycle arrest and apoptosis.
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Sistema Nervoso Central/metabolismo , Glioblastoma/metabolismo , Midkina/metabolismo , Proteínas de Ligação a RNA/metabolismo , Apoptose/genética , Apoptose/fisiologia , Ciclo Celular/genética , Ciclo Celular/fisiologia , Biologia Computacional , Dano ao DNA/genética , Dano ao DNA/fisiologia , Humanos , Técnicas In Vitro , Espécies Reativas de Oxigênio/metabolismo , Análise de Sequência de RNARESUMO
Glioblastoma is a highly aggressive and lethal brain tumor, with limited treatment options. Abnormal activation of the neddylation pathway is observed in glioblastoma, and the NEDD8-activating enzyme (NAE) inhibitor, MLN4924, was previously shown to be effective in glioblastoma cell line models. However, its effect has not been tested in patient-derived glioblastoma stem cells. We first analyzed public data to determine whether NEDD8 pathway proteins are important in glioblastoma development and patient survival. NAE1 and UBA3 levels increased in glioblastoma patients; high NEDD8 levels were associated with poor clinical outcomes. Immunohistochemistry results also supported this result. The effects of MLN4924 were evaluated in 4 glioblastoma cell lines and 15 patient-derived glioblastoma stem cells using high content analysis. Glioblastoma cell lines and patient-derived stem cells were highly susceptible to MLN4924, while normal human astrocytes were resistant. In addition, there were various responses in 15 patient-derived glioblastoma stem cells upon MLN4924 treatment. Genomic analyses indicated that MLN4924 sensitive cells exhibited enrichment of Extracellular Signal Regulated Kinase (ERK) and Protein kinase B (AKT, also known as PKB) signaling. We verified that MLN4924 inhibits ERK and AKT phosphorylation in MLN4924 sensitive cells. Our findings suggest that patient-derived glioblastoma stem cells in the context of ERK and AKT activation are sensitive and highly regulated by neddylation inhibition.
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As glioblastomas are mostly localized infiltrative lesions, gene therapy based on the retroviral replicating vector (RRV) system is considered an attractive strategy. Combinations of multiple suicide genes can circumvent the limitations associated with each gene, achieving direct and synergistic cytotoxic effects, along with bystander cell killing. In this study, we constructed a semi-and pseudotyped-RRV (sp-RRV) system harboring two suicide genes-herpes simplex virus type 1 thymidine kinase (TK) and yeast cytosine deaminase (CD)-to verify the dissemination and antitumor efficacy of our sp-RRV system (spRRVe-sEF1α-TK/sRRVgp-sEF1α-CD) in seven patient-derived glioblastoma stem-like cells (GSCs). Flow cytometry and high-content analysis revealed a wide range of transduction efficiency and good correlation between the delivery of therapeutic genes and susceptibility to the prodrugs ganciclovir and 5-fluorocytosine in patient-derived GSCs in vitro. Intra-tumoral delivery of spRRVe-sEF1α-TK/sRRVgp-sEF1α-CD, combined with prodrug treatment, synergistically inhibited cell proliferation and angiogenesis while increasing apoptosis and the depletion of tumor-associated macrophages in orthotopic glioblastoma xenografts. Genomic profiling of patient-derived GSCs revealed that the key genes preventing sp-RRV infection and transmission were associated with cell adhesion, migration, development, differentiation, and proliferation. This is the first report demonstrating that a novel sp-RRV-mediated TK/CD double suicide gene transfer system has high oncolytic power against extremely heterogeneous and treatment-refractory glioblastomas.
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BACKGROUND AND PURPOSE: To find genetic markers associated with response to radiotherapy (RT) in glioblastoma (GB) patients. MATERIALS AND METHODS: From Jan 2009 to Dec 2016, 161 patients with newly diagnosed IDH-wild type GB were treated with surgery and adjuvant concurrent chemoradiotherapy with the Stupp's regimen, and then genomic research proceeded with their surgical specimens. Among the 161 patients, 49 with clinically measurable disease on postoperative MRI were analyzed. The response evaluation to RT was based on Response Assessment in Neuro-Oncology (RANO) criteria. For genomic analyses to compare between patients with progression and non-progression, Fisher test for DNA mutations and copy number alterations and the Gene Set Enrichment Analysis (GSEA) were performed. RESULTS: RT responses were non-progressive and progressive disease (PD) in 22 (44.9%) and 27 patients (55.1%), respectively. After three months, seven of PD exhibited pseudoprogression. For true response adjusting pseudoprogression from PD, 1-year progression-free survival for true Non-Responders (tNR-group) and true Responders (tR-group) were 0% and 45.4% (pâ¯<â¯0.001), and overall survival were 52.5% and 81.1% (pâ¯=â¯0.046), respectively. In genomic analyses, the tNR-group had more CDKN2A deletions (94.4% vs. 55.6%, pâ¯=â¯0.013), EGFR mutations (33.3% vs. 3.7%, pâ¯=â¯0.012) and less TP53 mutations (22.2% vs. 40.7%, pâ¯=â¯0.333) than the tR-group. In GSEA, immune-related gene sets were enriched in the tNR-group, and in contrast, some gene sets related with cell cycle were enriched in tR-groups. CONCLUSION: The RANO criteria were feasible for the short-term response evaluation from RT despite of pseudoprogression. Genomic differences such as CDKN2A deletion, EGFR mutation, and immune- or inflammation-related related gene sets enrichment were found to be potentially predictive markers of RT.
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Neoplasias Encefálicas/genética , Neoplasias Encefálicas/radioterapia , Glioblastoma/genética , Glioblastoma/radioterapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/cirurgia , Quimiorradioterapia Adjuvante , Progressão da Doença , Feminino , Marcadores Genéticos , Glioblastoma/tratamento farmacológico , Glioblastoma/cirurgia , Humanos , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Medicina de Precisão , Adulto JovemRESUMO
OBJECTIVE: Isocitrate dehydrogenase 1 (IDH1) mutation status is an independent favorable prognostic factor for glioblastoma (GBM) and is usually determined by sequencing or immunohistochemistry. An accurate prediction of IDH1 mutation status via noninvasive methods helps establish the appropriate treatment strategy. We aimed to predict IDH1 mutation status using quantitative radiomic data in patients with GBM. METHODS: Between May 2010 and June 2015, we retrospectively identified 88 patients with newly diagnosed GBM. After semiautomatic segmentation of the lesions, we extracted 31 features from preoperative multiparametric magnetic resonance images. We also determined IDH1 mutation status using targeted sequencing and immunohistochemistry. A training cohort (n = 88) was used to train machine learning-based classifiers, with internal validation. The machine-learning technique was then validated in an external dataset of 35 patients with GBM. RESULTS: We detected the IDH1 mutation in 12 out of 88 GBMs. Multiparametric radiomic profiles revealed that the IDH1 mutation was associated with a smaller enhancing area volume and a larger necrotic area volume. Using the machine learning-based classification algorithms, we identified 70.3%-87.3% of prediction rate of IDH1 mutation status and found 66.3%-83.4% accuracy in the external validation set. CONCLUSIONS: We demonstrate that machine learning algorithms can predict IDH1 mutation status in GBM using preoperative multiparametric magnetic resonance images.
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Neoplasias Encefálicas/genética , Glioblastoma/genética , Isocitrato Desidrogenase/genética , Aprendizado de Máquina , Adulto , Idoso , Idoso de 80 Anos ou mais , Algoritmos , Neoplasias Encefálicas/patologia , Feminino , Glioblastoma/patologia , Glioma/genética , Glioma/patologia , Humanos , Imageamento por Ressonância Magnética/métodos , Masculino , Pessoa de Meia-Idade , Estudos RetrospectivosRESUMO
BACKGROUND: Cancer is a complex disease with profound genomic alterations and extensive heterogeneity. Recent studies on large-scale genomics have shed light on the impact of core oncogenic pathways, which are frequently dysregulated in a wide spectrum of cancer types. Aberrant activation of the hepatocyte growth factor (HGF) signaling axis has been associated with promoting various oncogenic programs during tumor initiation, progression, and treatment resistance. As a result, HGF-targeted therapy has emerged as an attractive therapeutic approach. However, recent clinical trials involving HGF-targeted therapies have demonstrated rather disappointing results. Thus, an alternative, in-depth assessment of new patient stratification is necessary to shift the current clinical course. METHODS: To address such challenges, we have evaluated the therapeutic efficacy of YYB-101, an HGF-neutralizing antibody, in a series of primary glioblastoma stem cells (GSCs) both in vitro and in vivo. Furthermore, we performed genome and transcriptome analysis to determine genetic and molecular traits that exhibit therapeutic susceptibility to HGF-mediated therapy. RESULTS: We have identified several differentially expressed genes, including MET, KDR, and SOX3, which are associated with tumor invasiveness, malignancy, and unfavorable prognosis in glioblastoma patients. We also demonstrated the HGF-MET signaling axis as a key molecular determinant in GSC invasion, and we discovered that a significant association in HGF expression existed between mesenchymal phenotype and immune cell recruitment. CONCLUSIONS: Upregulation of MET and mesenchymal cellular state are essential in generating HGF-mediated therapeutic responses. Our results provide an important framework for evaluating HGF-targeted therapy in future clinical settings.
Assuntos
Anticorpos Monoclonais Humanizados/farmacologia , Biomarcadores Tumorais/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Genômica/métodos , Glioblastoma/tratamento farmacológico , Fator de Crescimento de Hepatócito/antagonistas & inibidores , Transcriptoma , Animais , Apoptose , Movimento Celular , Proliferação de Células , Feminino , Glioblastoma/genética , Glioblastoma/patologia , Fator de Crescimento de Hepatócito/imunologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Fenótipo , Transdução de Sinais , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Skeletal muscle regeneration in mice has traditionally been studied using local freeze burn or snake venom injection models. More recently, a barium chloride (BaCl2)-induced muscle injury model has been established and is gaining popularity due to the relatively simple procedure and accessibility to required reagents. Here we sought to characterize the local and systemic effects of BaCl2-induced muscle injury. For this study, a 1.2% BaCl2 solution was locally administered to the tibialis anterior (TA) muscle and local and systemic phenotypes were analyzed at different timepoints. When 50 µL of the solution was injected unilaterally in the TA muscle, no mortality was observed. However, when 100 µL of the solution was injected, 50% of the mice died within 24 h. Serum analysis of the mice injected with 50 µL of BaCl2 solution at days 1 and 7 revealed changes resembling rhabdomyolysis. At day 1 post-injection of 50 µL of the BaCl2 solution, acute suppurative inflammation was observed in gross examination of the TA muscle, while extensive hemorrhagic necrosis was revealed on histological examination. At day 7, regenerated myofibers with centralized nuclei appeared with the resolution of acute inflammatory infiltration and the muscle tissue displayed molecular signatures consistent with myofiber differentiation. The overall muscle injury and regeneration phenotypes in the BaCl2-induced muscle injury model were similar to those of the well-established freeze burn or snake venom injection models. Taken together, the BaCl2-induced muscle injury model is comparable to conventional muscle injury and regeneration models, with considerations for possible systemic effects.