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1.
Lancet Haematol ; 7(3): e238-e246, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31879230

RESUMO

BACKGROUND: Data to inform surveillance and treatment for leukaemia predisposition syndromes are scarce and recommendations are largely based on expert opinion. This study aimed to investigate the clinical features and outcomes of patients with myelodysplastic syndrome or acute myeloid leukaemia and Shwachman-Diamond syndrome, an inherited bone marrow failure disorder with high risk of developing myeloid malignancies. METHODS: We did a multicentre, retrospective, cohort study in collaboration with the North American Shwachman-Diamond Syndrome Registry. We reviewed patient medical records from 17 centres in the USA and Canada. Patients with a genetic (biallelic mutations in the SBDS gene) or clinical diagnosis (cytopenias and pancreatic dysfunction) of Shwachman-Diamond syndrome who developed myelodysplastic syndrome or acute myeloid leukaemia were eligible without additional restriction. Medical records were reviewed between March 1, 2001, and Oct 5, 2017. Masked central review of bone marrow pathology was done if available to confirm leukaemia or myelodysplastic syndrome diagnosis. We describe the clinical features and overall survival of these patients. FINDINGS: We initially identified 37 patients with Shwachman-Diamond syndrome and myelodysplastic syndrome or acute myeloid leukaemia. 27 patients had samples available for central pathology review and were reclassified accordingly (central diagnosis concurred with local in 15 [56%] cases), 10 had no samples available and were classified based on the local review data, and 1 patient was excluded at this stage as not eligible. 36 patients were included in the analysis, of whom 10 (28%) initially presented with acute myeloid leukaemia and 26 (72%) initially presented with myelodysplastic syndrome. With a median follow-up of 4·9 years (IQR 3·9-8·4), median overall survival for patients with myelodysplastic syndrome was 7·7 years (95% CI 0·8-not reached) and 0·99 years (95% CI 0·2-2·4) for patients with acute myeloid leukaemia. Overall survival at 3 years was 11% (95% CI 1-39) for patients with leukaemia and 51% (29-68) for patients with myelodysplastic syndrome. Management and surveillance were variable. 18 (69%) of 26 patients with myelodysplastic syndrome received upfront therapy (14 haematopoietic stem cell transplantation and 4 chemotherapy), 4 (15%) patients received no treatment, 2 (8%) had unavailable data, and 2 (8%) progressed to acute myeloid leukaemia before receiving treatment. 12 patients received treatment for acute myeloid leukaemia-including the two patients initially diagnosed with myelodysplastic who progressed- two (16%) received HSCT as initial therapy and ten (83%) received chemotherapy with intent to proceed with HSCT. 33 (92%) of 36 patients (eight of ten with leukaemia and 25 of 26 with myelodysplastic syndrome) were known to have Shwachman-Diamond syndrome before development of a myeloid malignancy and could have been monitored with bone marrow surveillance. Bone marrow surveillance before myeloid malignancy diagnosis was done in three (33%) of nine patients with leukaemia for whom surveillance status was confirmed and 11 (46%) of 24 patients with myelodysplastic syndrome. Patients monitored had a 3-year overall survival of 62% (95% CI 32-82; n=14) compared with 28% (95% CI 10-50; n=19; p=0·13) without surveillance. Six (40%) of 15 patients with available longitudinal data developed myelodysplastic syndrome in the setting of stable blood counts. INTERPRETATION: Our results suggest that prognosis is poor for patients with Shwachman-Diamond syndrome and myelodysplastic syndrome or acute myeloid leukaemia owing to both therapy-resistant disease and treatment-related toxicities. Improved surveillance algorithms and risk stratification tools, studies of clonal evolution, and prospective trials are needed to inform effective prevention and treatment strategies for leukaemia predisposition in patients with Shwachman-Diamond syndrome. FUNDING: National Institute of Health.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Transplante de Células-Tronco Hematopoéticas/mortalidade , Leucemia Mieloide Aguda/mortalidade , Síndromes Mielodisplásicas/mortalidade , /mortalidade , Adolescente , Adulto , Criança , Pré-Escolar , Terapia Combinada , Feminino , Seguimentos , Humanos , Lactente , Recém-Nascido , Leucemia Mieloide Aguda/patologia , Leucemia Mieloide Aguda/terapia , Masculino , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/patologia , Síndromes Mielodisplásicas/terapia , Prognóstico , Estudos Retrospectivos , /terapia , Taxa de Sobrevida , Adulto Jovem
2.
Sensors (Basel) ; 19(11)2019 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-31159317

RESUMO

In this paper, we consider the use of wearable sensors for providing affect-based adaptation in Ambient Intelligence (AmI) systems. We begin with discussion of selected issues regarding the applications of affective computing techniques. We describe our experiments for affect change detection with a range of wearable devices, such as wristbands and the BITalino platform, and discuss an original software solution, which we developed for this purpose. Furthermore, as a test-bed application for our work, we selected computer games. We discuss the state-of-the-art in affect-based adaptation in games, described in terms of the so-called affective loop. We present our original proposal of a conceptual design framework for games, called the affective game design patterns. As a proof-of-concept realization of this approach, we discuss some original game prototypes, which we have developed, involving emotion-based control and adaptation. Finally, we comment on a software framework, that we have previously developed, for context-aware systems which uses human emotional contexts. This framework provides means for implementing adaptive systems using mobile devices with wearable sensors.


Assuntos
Dispositivos Eletrônicos Vestíveis , Inteligência Artificial , Técnicas Biossensoriais
3.
Haematologica ; 104(10): 1974-1983, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30948484

RESUMO

Quality of response to immunosuppressive therapy and long-term outcomes for pediatric severe aplastic anemia remain incompletely characterized. Contemporary evidence to inform treatment of relapsed or refractory severe aplastic anemia for pediatric patients is also limited. The clinical features and outcomes for 314 children treated from 2002 to 2014 with immunosuppressive therapy for acquired severe aplastic anemia were analyzed retrospectively from 25 institutions in the North American Pediatric Aplastic Anemia Consortium. The majority of subjects (n=264) received horse anti-thymocyte globulin (hATG) plus cyclosporine (CyA) with a median 61 months follow up. Following hATG/CyA, 71.2% (95%CI: 65.3,76.6) achieved an objective response. In contrast to adult studies, the quality of response achieved in pediatric patients was high, with 59.8% (95%CI: 53.7,65.8) complete response and 68.2% (95%CI: 62.2,73.8) achieving at least a very good partial response with a platelet count ≥50×109L. At five years post-hATG/CyA, overall survival was 93% (95%CI: 89,96), but event-free survival without subsequent treatment was only 64% (95%CI: 57,69) without a plateau. Twelve of 171 evaluable patients (7%) acquired clonal abnormalities after diagnosis after a median 25.2 months (range: 4.3-71 months) post treatment. Myelodysplastic syndrome or leukemia developed in 6 of 314 (1.9%). For relapsed/refractory disease, treatment with a hematopoietic stem cell transplant had a superior event-free survival compared to second immunosuppressive therapy treatment in a multivariate analysis (HR=0.19, 95%CI: 0.08,0.47; P=0.0003). This study highlights the need for improved therapies to achieve sustained high-quality remission for children with severe aplastic anemia.

4.
Hum Mol Genet ; 28(4): 572-583, 2019 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-30335132

RESUMO

Schwannomas are common, highly morbid and medically untreatable tumors that can arise in patients with germ line as well as somatic mutations in neurofibromatosis type 2 (NF2). These mutations most commonly result in the loss of function of the NF2-encoded protein, Merlin. Little is known about how Merlin functions endogenously as a tumor suppressor and how its loss leads to oncogenic transformation in Schwann cells (SCs). Here, we identify nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)-inducing kinase (NIK) as a potential drug target driving NF-κB signaling and Merlin-deficient schwannoma genesis. Using a genomic approach to profile aberrant tumor signaling pathways, we describe multiple upregulated NF-κB signaling elements in human and murine schwannomas, leading us to identify a caspase-cleaved, proteasome-resistant NIK kinase domain fragment that amplifies pathogenic NF-κB signaling. Lentiviral-mediated transduction of this NIK fragment into normal SCs promotes proliferation, survival, and adhesion while inducing schwannoma formation in a novel in vivo orthotopic transplant model. Furthermore, we describe an NF-κB-potentiated hepatocyte growth factor (HGF) to MET proto-oncogene receptor tyrosine kinase (c-Met) autocrine feed-forward loop promoting SC proliferation. These innovative studies identify a novel signaling axis underlying schwannoma formation, revealing new and potentially druggable schwannoma vulnerabilities with future therapeutic potential.


Assuntos
Neurilemoma/genética , Neurofibromatose 2/genética , Neurofibromina 2/genética , Proteínas Serina-Treonina Quinases/genética , Animais , Comunicação Autócrina/genética , Carcinogênese/genética , Caspase 1/genética , Proliferação de Células/genética , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica , Fator de Crescimento de Hepatócito/genética , Humanos , Camundongos , Terapia de Alvo Molecular , NF-kappa B/genética , Neurilemoma/complicações , Neurilemoma/tratamento farmacológico , Neurilemoma/patologia , Neurofibromatose 2/complicações , Neurofibromatose 2/tratamento farmacológico , Neurofibromatose 2/patologia , Complexo de Endopeptidases do Proteassoma/genética , Proteínas Proto-Oncogênicas c-met/genética , Células de Schwann , Transdução de Sinais/genética
6.
Artigo em Inglês | MEDLINE | ID: mdl-30275003

RESUMO

Correct diagnosis of inherited bone marrow failure syndromes is a challenge because of the significant overlap in clinical presentation of these disorders. Establishing right genetic diagnosis is crucial for patients' optimal clinical management and family counseling. A nondysmorphic infant reported here developed severe transfusion-dependent anemia and met clinical criteria for diagnosis of Diamond-Blackfan anemia (DBA). However, whole-exome sequencing demonstrated that the child was a compound heterozygote for a paternally inherited pathogenic truncating variant (SPTA1 c.4975 C>T) and a novel maternally inherited missense variant of uncertain significance (SPTA1 c.5029 G>A) within the spectrin gene, consistent with hereditary hemolytic anemia due to disruption of red blood cell (RBC) cytoskeleton. Ektacytometry demonstrated abnormal membrane flexibility of the child's RBCs. Scanning electron microscopy revealed morphological aberrations of the patient's RBCs. Both parents were found to have mild hereditary elliptocytosis. Importantly, patients with severe RBC membrane defects may be successfully managed with splenectomy to minimize peripheral destruction of misshapen RBCs, whereas patients with DBA require lifelong transfusions, steroid therapy, or hematopoietic stem cell transplantation. As suggested by the WES findings, splenectomy rendered our patient transfusion-independent, improving the family's quality of life and preventing transfusion-related iron overload. This case illustrates the utility of whole-exome sequencing in clinical care of children with genetic disorders of unclear presentation.


Assuntos
Anemia/diagnóstico , Anemia/genética , Proteínas de Transporte/genética , Proteínas dos Microfilamentos/genética , Anemia de Diamond-Blackfan/diagnóstico , Anemia de Diamond-Blackfan/genética , Anemia Hemolítica Congênita/genética , Proteínas de Transporte/metabolismo , Erros de Diagnóstico , Eliptocitose Hereditária/genética , Membrana Eritrocítica/fisiologia , Eritrócitos/patologia , Exoma/genética , Feminino , Humanos , Lactente , Proteínas dos Microfilamentos/metabolismo , Mutação/genética , Linhagem , Qualidade de Vida , Espectrina/genética , Espectrina/fisiologia , Sequenciamento Completo do Exoma/métodos
7.
Nat Rev Cancer ; 18(3): 168-185, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29376519

RESUMO

Fanconi anaemia (FA) is a genetic disorder that is characterized by bone marrow failure (BMF), developmental abnormalities and predisposition to cancer. Together with other proteins involved in DNA repair processes and cell division, the FA proteins maintain genome homeostasis, and germline mutation of any one of the genes that encode FA proteins causes FA. Monoallelic inactivation of some FA genes, such as FA complementation group D1 (FANCD1; also known as the breast and ovarian cancer susceptibility gene BRCA2), leads to adult-onset cancer predisposition but does not cause FA, and somatic mutations in FA genes occur in cancers in the general population. Carcinogenesis resulting from a dysregulated FA pathway is multifaceted, as FA proteins monitor multiple complementary genome-surveillance checkpoints throughout interphase, where monoubiquitylation of the FANCD2-FANCI heterodimer by the FA core complex promotes recruitment of DNA repair effectors to chromatin lesions to resolve DNA damage and mitosis. In this Review, we discuss how the FA pathway safeguards genome integrity throughout the cell cycle and show how studies of FA have revealed opportunities to develop rational therapeutics for this genetic disease and for malignancies that acquire somatic mutations within the FA pathway.


Assuntos
Anemia de Fanconi/genética , Mutação , Neoplasias/genética , Proteína BRCA1/genética , Proteína BRCA2/genética , Carcinoma de Células Escamosas/genética , Reparo do DNA , Proteína do Grupo de Complementação D2 da Anemia de Fanconi/genética , Predisposição Genética para Doença , Instabilidade Genômica , Humanos , Leucemia Mieloide Aguda/genética , Neoplasias/terapia
8.
Pediatr Blood Cancer ; 65(1)2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28868793

RESUMO

Diagnosis of bone marrow failure (BMF) disorders is challenging but essential for optimal patient management. Here, we report a young adult from nonconsanguineous parents with progressive pancytopenia since childhood, bone pain, increased bone density, and haphazard ossification replacing hematopoiesis within the bone marrow. Sequencing revealed two novel biallelic variants of unknown significance within the thromboxane A synthase gene, TBXAS1 (c.266T > C; c.989T > C), bioinformatically predicted to disrupt the protein. TBXAS1 mutations result in Ghosal hematodiaphyseal dysplasia (OMIM 231095), the autosomal recessive syndrome associated with abnormal bone structure and BMF. Identification of the genetic defect prompted steroid therapy leading to resolution of symptoms.


Assuntos
Anemia Refratária , Densidade Óssea/genética , Osteocondrodisplasias , Pancitopenia , Mutação Puntual , Tromboxano-A Sintase/deficiência , Anemia Refratária/enzimologia , Anemia Refratária/genética , Anemia Refratária/patologia , Doença Crônica , Feminino , Humanos , Lactente , Osteocondrodisplasias/enzimologia , Osteocondrodisplasias/genética , Osteocondrodisplasias/patologia , Pancitopenia/enzimologia , Pancitopenia/genética , Pancitopenia/patologia
10.
Haematologica ; 102(6): 1017-1027, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28341737

RESUMO

Fanconi anemia is a complex heterogeneous genetic disorder with a high incidence of bone marrow failure, clonal evolution to acute myeloid leukemia and mesenchymal-derived congenital anomalies. Increasing evidence in Fanconi anemia and other genetic disorders points towards an interdependence of skeletal and hematopoietic development, yet the impact of the marrow microenvironment in the pathogenesis of the bone marrow failure in Fanconi anemia remains unclear. Here we demonstrated that mice with double knockout of both Fancc and Fancg genes had decreased bone formation at least partially due to impaired osteoblast differentiation from mesenchymal stem/progenitor cells. Mesenchymal stem/progenitor cells from the double knockout mice showed impaired hematopoietic supportive activity. Mesenchymal stem/progenitor cells of patients with Fanconi anemia exhibited similar cellular deficits, including increased senescence, reduced proliferation, impaired osteoblast differentiation and defective hematopoietic stem/progenitor cell supportive activity. Collectively, these studies provide unique insights into the physiological significance of mesenchymal stem/progenitor cells in supporting the marrow microenvironment, which is potentially of broad relevance in hematopoietic stem cell transplantation.


Assuntos
Medula Óssea/patologia , Microambiente Celular , Anemia de Fanconi/patologia , Animais , Osso e Ossos/anormalidades , Osso e Ossos/fisiopatologia , Linhagem da Célula , Anemia de Fanconi/fisiopatologia , Proteína do Grupo de Complementação C da Anemia de Fanconi/genética , Hematopoese , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Mesenquimais/patologia , Camundongos , Camundongos Knockout
11.
J Allergy Clin Immunol ; 139(1): 142-151.e5, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27554818

RESUMO

BACKGROUND: Atopic dermatitis (AD) is characterized by intense pruritis and is a common childhood inflammatory disease. Many factors are known to affect AD development, including the pleiotropic cytokine IL-4. Yet little is known regarding the direct effects of IL-4 on keratinocyte function. OBJECTIVE AND METHODS: In this report RNA sequencing and functional assays were used to define the effect of the allergic environment on primary keratinocyte function and wound repair in mice. RESULTS: Acute or chronic stimulation by IL-4 modified expression of more than 1000 genes expressed in human keratinocytes that are involved in a broad spectrum of nonoverlapping functions. Among the IL-4-induced changes, repression of fibronectin critically impaired the human keratinocyte wound response. Moreover, in mouse models of spontaneous and induced AD-like lesions, there was delayed re-epithelialization. Importantly, topical treatment with fibronectin restored the epidermal repair response. CONCLUSION: Keratinocyte gene expression is critically shaped by IL-4, altering cell fate decisions, which are likely important for the clinical manifestations and pathology of allergic skin disease.


Assuntos
Fibronectinas/imunologia , Interleucina-4/imunologia , Queratinócitos/imunologia , Cicatrização/imunologia , Animais , Células Cultivadas , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fator de Transcrição STAT6/genética , Pele/imunologia , Transcriptoma/efeitos dos fármacos , Cicatrização/genética
12.
Mol Cell Biol ; 37(6)2017 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-28031327

RESUMO

The partially understood phosphoinositide signaling cascade regulates multiple aspects of cellular metabolism. Previous studies revealed that INPP5E, the inositol polyphosphate-5-phosphatase that is mutated in the developmental disorders Joubert and MORM syndromes, is essential for the function of the primary cilium and maintenance of phosphoinositide balance in nondividing cells. Here, we report that INPP5E further contributes to cellular homeostasis by regulating cell division. We found that silencing or genetic knockout of INPP5E in human and murine cells impairs the spindle assembly checkpoint, centrosome and spindle function, and maintenance of chromosomal integrity. Consistent with a cell cycle regulatory role, we found that INPP5E expression is cell cycle dependent, peaking at mitotic entry. INPP5E localizes to centrosomes, chromosomes, and kinetochores in early mitosis and shuttles to the midzone spindle at mitotic exit. Our findings identify the previously unknown, essential role of INPP5E in mitosis and prevention of aneuploidy, providing a new perspective on the function of this phosphoinositide phosphatase in health and development.


Assuntos
Instabilidade Genômica , Mitose/genética , Monoéster Fosfórico Hidrolases/metabolismo , Animais , Ciclo Celular , Núcleo Celular/metabolismo , Fibroblastos/metabolismo , Técnicas de Silenciamento de Genes , Células HeLa , Homeostase , Humanos , Interfase , Pontos de Checagem da Fase M do Ciclo Celular , Camundongos , Modelos Biológicos , Fosfatidilinositol 4,5-Difosfato/metabolismo , Transporte Proteico , Frações Subcelulares/metabolismo , Especificidade por Substrato
14.
Exp Hematol ; 44(5): 352-7, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26860989

RESUMO

Fanconi anemia (FA) is an inherited disorder of genomic instability associated with high risk of myelodysplasia and acute myeloid leukemia (AML). Young mice deficient in FA core complex genes do not naturally develop cancer, hampering preclinical studies on malignant hematopoiesis in FA. Here we describe that aging Fancc(-/-) mice are prone to genomically unstable AML and other hematologic neoplasms. We report that aneuploidy precedes malignant transformation during Fancc(-/-) hematopoiesis. Our observations reveal that Fancc(-/-) mice develop hematopoietic chromosomal instability followed by leukemia in an age-dependent manner, recapitulating the clinical phenotype of human FA and providing a proof of concept for future development of preclinical models of FA-associated leukemogenesis.


Assuntos
Envelhecimento/genética , Instabilidade Cromossômica , Proteína do Grupo de Complementação C da Anemia de Fanconi/genética , Leucemia Mieloide/genética , Doença Aguda , Fatores Etários , Aneuploidia , Animais , Anemia de Fanconi/genética , Anemia de Fanconi/metabolismo , Proteína do Grupo de Complementação C da Anemia de Fanconi/deficiência , Hematopoese/genética , Humanos , Estimativa de Kaplan-Meier , Leucemia Mieloide/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout
15.
Pediatr Blood Cancer ; 63(5): 917-21, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26713410

RESUMO

We describe a child with dyserythropoietic anemia, thrombocytosis, functional platelet defect, and megakaryocyte dysplasia. We show that (i) this constellation of hematopoietic abnormalities was due to a germline mutation within the 5' untranslated region (5'UTR) of globin transcription factor 1 (GATA1); (ii) the mutation impaired a 5'UTR GATA1 splicing site, with promoted production of the shortened GATA1 isoform lacking the N-terminus; and (iii) expression of the GATA1 N-terminus is restricted to erythroblasts and megakaryocytes in normal marrow, consistent with the patient's abnormal erythropoiesis and megakaryopoiesis. Our findings provide insights into the clinically relevant in vivo function of the N-terminal domain of GATA1 in human hematopoiesis.


Assuntos
Regiões 5' não Traduzidas , Anemia Diseritropoética Congênita/genética , Fator de Transcrição GATA1/genética , Megacariócitos , Sítios de Splice de RNA , Processamento Alternativo , Anemia Diseritropoética Congênita/metabolismo , Pré-Escolar , Fator de Transcrição GATA1/biossíntese , Humanos , Masculino , Isoformas de Proteínas/biossíntese , Isoformas de Proteínas/genética
16.
Exp Hematol ; 43(12): 1031-1046.e12, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26366677

RESUMO

The Fanconi anemia (FA/BRCA) signaling network controls multiple genome-housekeeping checkpoints, from interphase DNA repair to mitosis. The in vivo role of abnormal cell division in FA remains unknown. Here, we quantified the origins of genomic instability in FA patients and mice in vivo and ex vivo. We found that both mitotic errors and interphase DNA damage significantly contribute to genomic instability during FA-deficient hematopoiesis and in nonhematopoietic human and murine FA primary cells. Super-resolution microscopy coupled with functional assays revealed that FANCA shuttles to the pericentriolar material to regulate spindle assembly at mitotic entry. Loss of FA signaling rendered cells hypersensitive to spindle chemotherapeutics and allowed escape from the chemotherapy-induced spindle assembly checkpoint. In support of these findings, direct comparison of DNA crosslinking and anti-mitotic chemotherapeutics in primary FANCA-/- cells revealed genomic instability originating through divergent cell cycle checkpoint aberrations. Our data indicate that FA/BRCA signaling functions as an in vivo gatekeeper of genomic integrity throughout interphase and mitosis, which may have implications for future targeted therapies in FA and FA-deficient cancers.


Assuntos
Proteína do Grupo de Complementação A da Anemia de Fanconi/metabolismo , Anemia de Fanconi/metabolismo , Hematopoese , Interfase , Mitose , Animais , Anemia de Fanconi/genética , Anemia de Fanconi/patologia , Anemia de Fanconi/terapia , Proteína do Grupo de Complementação A da Anemia de Fanconi/genética , Feminino , Humanos , Masculino , Camundongos , Camundongos Knockout , Fuso Acromático/genética , Fuso Acromático/metabolismo , Fuso Acromático/patologia
17.
Sci Rep ; 5: 11450, 2015 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-26095125

RESUMO

Dense fibrotic stroma associated with pancreatic ductal adenocarcinoma (PDAC) is a major obstacle for drug delivery to the tumor bed and plays a crucial role in pancreatic cancer progression. Current, anti-stromal therapies have failed to improve tumor response to chemotherapy and patient survival. Furthermore, recent studies show that stroma impedes tumor progression, and its complete ablation accelerates PDAC progression. In an effort to understand the molecular mechanisms associated with tumor-stromal interactions, using in vitro and in vivo models and PDAC patient biopsies, we show that the loss of miR-29 is a common phenomenon of activated pancreatic stellate cells (PSCs)/fibroblasts, the major stromal cells responsible for fibrotic stromal reaction. Loss of miR-29 is correlated with a significant increase in extracellular matrix (ECM) deposition, a major component in PDAC stroma. Our in vitro miR-29 gain/loss-of-function studies document the role of miR-29 in PSC-mediated ECM stromal protein accumulation. Overexpression of miR-29 in activated stellate cells reduced stromal deposition, cancer cell viability, and cancer growth in co-culture. Furthermore, the loss of miR-29 in TGF-ß1 activated PSCs is SMAD3 dependent. These results provide insights into the mechanistic role of miR-29 in PDAC stroma and its potential use as a therapeutic agent to target PDAC.


Assuntos
Carcinoma Ductal Pancreático/genética , Matriz Extracelular/metabolismo , Fibrose/patologia , MicroRNAs/genética , Neoplasias Pancreáticas/genética , Animais , Antineoplásicos/uso terapêutico , Carcinoma Ductal Pancreático/tratamento farmacológico , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Ativação Enzimática/genética , Matriz Extracelular/genética , Fibroblastos/citologia , Fibrose/genética , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Pâncreas/metabolismo , Pâncreas/patologia , Neoplasias Pancreáticas/tratamento farmacológico , Células Estreladas do Pâncreas/citologia , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Microambiente Tumoral
18.
J Biol Chem ; 290(7): 4075-85, 2015 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-25548279

RESUMO

Cyclin E-CDK2 is a key regulator in G1/S transition. Previously, we identified a number of CDK2-interacting proteins, including PHF8 (plant homeodomain finger protein 8). In this report, we confirmed that PHF8 is a novel cyclin E-CDK2 substrate. By taking the approach of mass spectrometry, we identified that PHF8 Ser-844 is phosphorylated by cyclin E-CDK2. Immunoblotting analysis indicated that WT PHF8 demethylates histone H3K9me2 more efficiently than the cyclin E-CDK2 phosphorylation-deficient PHF8-S844A mutant. Furthermore, flow cytometry analysis showed that WT PHF8 promotes S phase progression more robustly than PHF8-S844A. Real-time PCR results demonstrated that PHF8 increases transcription of cyclin E, E2F3, and E2F7 to significantly higher levels compared with PHF8-S844A. Further analysis by ChIP assay indicated that PHF8 binds to the cyclin E promoter stronger than PHF8-S844A and reduces the H3K9me2 level at the cyclin E promoter more efficiently than PHF8-S844A. In addition, we found that cyclin E-CDK2-mediated phosphorylation of PHF8 Ser-844 promotes PHF8-dependent rRNA transcription in luciferase reporter assays and real-time PCR. Taken together, these results indicate that cyclin E-CDK2 phosphorylates PHF8 to stimulate its demethylase activity to promote rRNA transcription and cell cycle progression.


Assuntos
Ciclina E/metabolismo , Quinase 2 Dependente de Ciclina/metabolismo , Regulação da Expressão Gênica , Histona Desmetilases/metabolismo , Histonas/metabolismo , Proteínas Oncogênicas/metabolismo , Fase S/fisiologia , Fatores de Transcrição/metabolismo , Western Blotting , Proliferação de Células , Células Cultivadas , Imunoprecipitação da Cromatina , Ciclina E/genética , Quinase 2 Dependente de Ciclina/genética , Metilação de DNA , DNA Ribossômico/genética , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Células HeLa , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases/genética , Humanos , Imunoprecipitação , Proteínas Oncogênicas/genética , Fosforilação , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genética
19.
Hum Mol Genet ; 24(1): 1-8, 2015 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-25113746

RESUMO

Neurofibromatosis type 2 (NF2) is an autosomal dominant genetic disorder resulting from germline mutations in the NF2 gene. Bilateral vestibular schwannomas, tumors on cranial nerve VIII, are pathognomonic for NF2 disease. Furthermore, schwannomas also commonly develop in other cranial nerves, dorsal root ganglia and peripheral nerves. These tumors are a major cause of morbidity and mortality, and medical therapies to treat them are limited. Animal models that accurately recapitulate the full anatomical spectrum of human NF2-related schwannomas, including the characteristic functional deficits in hearing and balance associated with cranial nerve VIII tumors, would allow systematic evaluation of experimental therapeutics prior to clinical use. Here, we present a genetically engineered NF2 mouse model generated through excision of the Nf2 gene driven by Cre expression under control of a tissue-restricted 3.9kbPeriostin promoter element. By 10 months of age, 100% of Postn-Cre; Nf2(flox/flox) mice develop spinal, peripheral and cranial nerve tumors histologically identical to human schwannomas. In addition, the development of cranial nerve VIII tumors correlates with functional impairments in hearing and balance, as measured by auditory brainstem response and vestibular testing. Overall, the Postn-Cre; Nf2(flox/flox) tumor model provides a novel tool for future mechanistic and therapeutic studies of NF2-associated schwannomas.


Assuntos
Moléculas de Adesão Celular/genética , Gânglios Espinais/patologia , Neurofibromatose 2/genética , Neurofibromina 2/genética , Neuroma Acústico/fisiopatologia , Nervo Vestibulococlear/patologia , Animais , Modelos Animais de Doenças , Éxons , Audição , Humanos , Camundongos , Camundongos Transgênicos , Mutação , Neurofibromatose 2/complicações , Neurofibromatose 2/fisiopatologia , Neuroma Acústico/genética , Neuroma Acústico/patologia
20.
PLoS One ; 9(10): e111611, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25360622

RESUMO

CDKN3 (cyclin-dependent kinase inhibitor 3), a dual specificity protein phosphatase, dephosphorylates cyclin-dependent kinases (CDKs) and thus functions as a key negative regulator of cell cycle progression. Deregulation or mutations of CDNK3 have been implicated in various cancers. However, the role of CDKN3 in Bcr-Abl-mediated chronic myelogenous leukemia (CML) remains unknown. Here we found that CDKN3 acts as a tumor suppressor in Bcr-Abl-mediated leukemogenesis. Overexpression of CDKN3 sensitized the K562 leukemic cells to imanitib-induced apoptosis and dramatically inhibited K562 xenografted tumor growth in nude mouse model. Ectopic expression of CDKN3 significantly reduced the efficiency of Bcr-Abl-mediated transformation of FDCP1 cells to growth factor independence. In contrast, depletion of CDKN3 expression conferred resistance to imatinib-induced apoptosis in the leukemic cells and accelerated the growth of xenograph leukemia in mice. In addition, we found that CDKN3 mutant (CDKN3-C140S) devoid of the phosphatase activity failed to affect the K562 leukemic cell survival and xenografted tumor growth, suggesting that the phosphatase of CDKN3 was required for its tumor suppressor function. Furthermore, we observed that overexpression of CDKN3 reduced the leukemic cell survival by dephosphorylating CDK2, thereby inhibiting CDK2-dependent XIAP expression. Moreover, overexpression of CDKN3 delayed G1/S transition in K562 leukemic cells. Our results highlight the importance of CDKN3 in Bcr-Abl-mediated leukemogenesis, and provide new insights into diagnostics and therapeutics of the leukemia.


Assuntos
Carcinogênese/metabolismo , Carcinogênese/patologia , Proteínas Inibidoras de Quinase Dependente de Ciclina/metabolismo , Fosfatases de Especificidade Dupla/metabolismo , Proteínas de Fusão bcr-abl/metabolismo , Animais , Carcinogênese/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Transformação Celular Neoplásica/efeitos dos fármacos , Transformação Celular Neoplásica/patologia , Quinase 2 Dependente de Ciclina/metabolismo , Feminino , Fase G1/efeitos dos fármacos , Humanos , Mesilato de Imatinib/farmacologia , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Células K562 , Camundongos Nus , Fosforilação/efeitos dos fármacos , Fase S/efeitos dos fármacos , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
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