RESUMO
Human mutations in PQBP1, a molecule involved in transcription and splicing, result in a reduced but architecturally normal brain. Examination of a conditional Pqbp1-knockout (cKO) mouse with microcephaly failed to reveal either abnormal centrosomes or mitotic spindles, increased neurogenesis from the neural stem progenitor cell (NSPC) pool or increased cell death in vivo. Instead, we observed an increase in the length of the cell cycle, particularly for the M phase in NSPCs. Corresponding to the developmental expression of Pqbp1, the stem cell pool in vivo was decreased at E10 and remained at a low level during neurogenesis (E15) in Pqbp1-cKO mice. The expression profiles of NSPCs derived from the cKO mouse revealed significant changes in gene groups that control the M phase, including anaphase-promoting complex genes, via aberrant transcription and RNA splicing. Exogenous Apc4, a hub protein in the network of affected genes, recovered the cell cycle, proliferation, and cell phenotypes of NSPCs caused by Pqbp1-cKO. These data reveal a mechanism of brain size control based on the simple reduction of the NSPC pool by cell cycle time elongation. Finally, we demonstrated that in utero gene therapy for Pqbp1-cKO mice by intraperitoneal injection of the PQBP1-AAV vector at E10 successfully rescued microcephaly with preserved cortical structures and improved behavioral abnormalities in Pqbp1-cKO mice, opening a new strategy for treating this intractable developmental disorder.
Assuntos
Terapia Genética , Microcefalia/genética , Microcefalia/terapia , Células-Tronco Neurais/fisiologia , Proteínas Nucleares/deficiência , Adenoviridae/genética , Animais , Subunidade Apc4 do Ciclossomo-Complexo Promotor de Anáfase/metabolismo , Apoptose/genética , Encéfalo/patologia , Proteínas de Transporte/genética , Moléculas de Adesão Celular/metabolismo , Ciclo Celular , Proliferação de Células , Proteínas de Ligação a DNA , Modelos Animais de Doenças , Embrião de Mamíferos , Feminino , Humanos , Masculino , Camundongos , Camundongos Knockout , Microcefalia/patologia , Nestina/genética , Nestina/metabolismo , Neurogênese , Proteínas Nucleares/genética , Sinapsinas/genética , Sinapsinas/metabolismoRESUMO
BACKGROUND: The MYC oncogene has long been established as a central driver in many types of human cancers including colorectal cancer. However, the realization of MYC-targeting therapies remains elusive; as a result, synthetic lethal therapeutic approaches are alternatively being explored. A synthetic lethal therapeutic approach aims to kill MYC-driven tumors by targeting a certain co-regulator on the MYC pathway. PATIENTS AND METHODS: We analyzed copy number and expression profiles from 130 colorectal cancer tumors together with publicly available datasets to identify co-regulators on the MYC pathway. Candidates were functionally tested by in vitro assays using colorectal cancer and normal fibroblast cell lines. Additionally, survival analyses were carried out on another 159 colorectal cancer patients and public datasets. RESULTS: Our in silico screening identified two MYC co-regulator candidates, AURKA and TPX2, which are interacting mitotic regulators located on chromosome 20q. We found the two candidates showed frequent co-amplification with the MYC locus while expression levels of MYC and the two genes were positively correlated with those of MYC downstream target genes across multiple cancer types. In vitro, the aberrant expression of MYC, AURKA and TPX2 resulted in more aggressive anchorage-independent growth in normal fibroblast cells. Furthermore, knockdown of AURKA or TPX2, or treatment with an AURKA-specific inhibitor effectively suppressed the proliferation of MYC-expressing colorectal cancer cells. Additionally, combined high expression of MYC, AURKA and TPX2 proved to be a poor prognostic indicator of colorectal cancer patient survival. CONCLUSIONS: Through bioinformatic analyses and experiments, we proposed TPX2 and AURKA as novel co-regulators on the MYC pathway. Inhibiting the AURKA/TPX2 axis would be a novel synthetic lethal therapeutic approach for MYC-driven cancers.
Assuntos
Aurora Quinase A/metabolismo , Proteínas de Ciclo Celular/metabolismo , Neoplasias Colorretais/enzimologia , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Transdução de Sinais , Antineoplásicos/uso terapêutico , Aurora Quinase A/antagonistas & inibidores , Aurora Quinase A/genética , Proteínas de Ciclo Celular/genética , Proliferação de Células , Sobrevivência Celular , Cromossomos Humanos Par 20 , Cromossomos Humanos Par 8 , Neoplasias Colorretais/genética , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Biologia Computacional , Amplificação de Genes , Dosagem de Genes , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Células HCT116 , Humanos , Proteínas Associadas aos Microtúbulos/genética , Proteínas Nucleares/genética , Prognóstico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-myc/genética , Interferência de RNA , Transdução de Sinais/efeitos dos fármacos , Análise de Sobrevida , Fatores de Tempo , TransfecçãoRESUMO
We simulated the early phase of the 2009 influenza A(H1N1) pandemic and assessed the effectiveness of public health interventions in Japan. We show that the detection rate of border quarantine was low and the timing of the intervention was the most important factor involved in the control of the pandemic, with the maximum reduction in daily cases obtained after interventions started on day 6 or 11. Early interventions were not always effective.
Assuntos
Surtos de Doenças/prevenção & controle , Vírus da Influenza A Subtipo H1N1 , Influenza Humana/prevenção & controle , Modelos Teóricos , Quarentena , Humanos , Influenza Humana/epidemiologia , Japão/epidemiologia , Saúde Pública , Fatores de TempoRESUMO
Autologous blood transfusion (ABT) is rarely employed in patients with end-stage renal disease (ESRD); these patients are usually anemic. Since 1998, we have attempted ABT for ESRD patients undergoing living-related kidney transplantation. Among 20 patients enrolled in this study the preoperative hemoglobin and hematocrit levels were 10.0 +/- 1.2 mg/dL (range, 8.1-11.7) and 30.0 +/- 3.7% (range, 24.7-34.3), respectively. Blood volume collected on each occasion was 235.7 +/- 57.7 mL (range, 200-400), and the number of blood collections was 2.45 +/- 0.9 (range, 1-4). Total collected volume was 567.5 +/- 157.5 mL (range, 400-800). Symptomatic hypotension was seen in two patients, but vital signs recovered spontaneously. No other problems related to blood collection were observed. Allogeneic transfusion was need in only one patient (5%). ABT was safe and efficacious in ESRD patients scheduled for living-related kidney transplantation.
Assuntos
Transfusão de Sangue Autóloga , Transplante de Rim/fisiologia , Adolescente , Adulto , Anemia/etiologia , Família , Feminino , Hematócrito , Humanos , Falência Renal Crônica/complicações , Falência Renal Crônica/cirurgia , Doadores Vivos , Masculino , Pessoa de Meia-Idade , Diálise RenalRESUMO
We have designed a protocol for ABO-incompatible kidney transplantations based on preoperative plasmapheresis with a tacrolimus/mycophenolate mofetil/methylprednisolone/basiliximab protocol using low-dose rituximab (200 mg/body) instead of splenectomy to prevent antibody-mediated acute rejection. Eight patients successfully received transplants with this protocol. The titers of anti-A and -B antibodies as well as the number of CD20(+) cells were readily maintained at a low level posttransplantation. There were no side effects. All patients have renal transplant function with a follow-up of 1-34 months.
Assuntos
Sistema ABO de Grupos Sanguíneos , Anticorpos Monoclonais/uso terapêutico , Fatores Imunológicos/uso terapêutico , Transplante de Rim/imunologia , Troca Plasmática , Adulto , Anticorpos Monoclonais Murinos , Antígenos CD20/sangue , Antígenos CD20/imunologia , Incompatibilidade de Grupos Sanguíneos , Creatinina/sangue , Quimioterapia Combinada , Feminino , Seguimentos , Rejeição de Enxerto/epidemiologia , Teste de Histocompatibilidade/métodos , Humanos , Imunossupressores/uso terapêutico , Isoanticorpos/sangue , Doadores Vivos , Masculino , Pessoa de Meia-Idade , Núcleo Familiar , Plasmaferese , RituximabRESUMO
Basiliximab is potent, relatively safe immunosuppressive induction agent used in transplantation. Prophylactic use at the time of transplantation has been advocated to improve allograft outcomes. We report two cases of kidney transplant recipients with anaphylactic reactions after initial exposure to Basiliximab.
Assuntos
Anafilaxia/induzido quimicamente , Anticorpos Monoclonais/efeitos adversos , Imunossupressores/efeitos adversos , Transplante de Rim/imunologia , Proteínas Recombinantes de Fusão/efeitos adversos , Adolescente , Adulto , Basiliximab , Feminino , Humanos , Diálise Peritoneal Ambulatorial Contínua , Resultado do TratamentoRESUMO
The transcription factor nuclear factor-κB (NF-κB) has important roles for tumorigenesis, but how it regulates cancer stem cells (CSCs) remains largely unclear. We identified insulin-like growth factor 2 (IGF2) is a key target of NF-κB activated by HER2/HER3 signaling to form tumor spheres in breast cancer cells. The IGF2 receptor, IGF1 R, was expressed at high levels in CSC-enriched populations in primary breast cancer cells. Moreover, IGF2-PI3K (IGF2-phosphatidyl inositol 3 kinase) signaling induced expression of a stemness transcription factor, inhibitor of DNA-binding 1 (ID1), and IGF2 itself. ID1 knockdown greatly reduced IGF2 expression, and tumor sphere formation. Finally, treatment with anti-IGF1/2 antibodies blocked tumorigenesis derived from the IGF1Rhigh CSC-enriched population in a patient-derived xenograft model. Thus, NF-κB may trigger IGF2-ID1-IGF2-positive feedback circuits that allow cancer stem-like cells to appear. Then, they may become addicted to the circuits. As the circuits are the Achilles' heels of CSCs, it will be critical to break them for eradication of CSCs.
Assuntos
Neoplasias da Mama/patologia , Regulação Neoplásica da Expressão Gênica , Proteína 1 Inibidora de Diferenciação/metabolismo , Fator de Crescimento Insulin-Like II/metabolismo , Recidiva Local de Neoplasia/patologia , Células-Tronco Neoplásicas/patologia , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Carcinogênese , Feminino , Humanos , Proteína 1 Inibidora de Diferenciação/genética , Fator de Crescimento Insulin-Like I/genética , Fator de Crescimento Insulin-Like I/metabolismo , Fator de Crescimento Insulin-Like II/genética , Camundongos , Camundongos Nus , NF-kappa B/genética , NF-kappa B/metabolismo , Gradação de Tumores , Invasividade Neoplásica , Metástase Neoplásica , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/metabolismo , Células-Tronco Neoplásicas/metabolismo , Fosfatidilinositol 3-Quinase/genética , Fosfatidilinositol 3-Quinase/metabolismo , Prognóstico , Transdução de Sinais , Esferoides Celulares , Taxa de Sobrevida , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Recent genetic analysis has identified frequent mutations in ten-eleven translocation 2 (TET2), DNA methyltransferase 3A (DNMT3A), isocitrate dehydrogenase 2 (IDH2) and ras homolog family member A (RHOA) in nodal T-cell lymphomas, including angioimmunoblastic T-cell lymphoma and peripheral T-cell lymphoma, not otherwise specified. We examined the distribution of mutations in these subtypes of mature T-/natural killer cell neoplasms to determine their clonal architecture. Targeted sequencing was performed for 71 genes in tumor-derived DNA of 87 cases. The mutations were then analyzed in a programmed death-1 (PD1)-positive population enriched with tumor cells and CD20-positive B cells purified by laser microdissection from 19 cases. TET2 and DNMT3A mutations were identified in both the PD1+ cells and the CD20+ cells in 15/16 and 4/7 cases, respectively. All the RHOA and IDH2 mutations were confined to the PD1+ cells, indicating that some, including RHOA and IDH2 mutations, being specific events in tumor cells. Notably, we found that all NOTCH1 mutations were detected only in the CD20+ cells. In conclusion, we identified both B- as well as T-cell-specific mutations, and mutations common to both T and B cells. These findings indicate the expansion of a clone after multistep and multilineal acquisition of gene mutations.
Assuntos
Biomarcadores Tumorais , Linfoma Extranodal de Células T-NK/genética , Mutação , Alelos , Substituição de Aminoácidos , Linfócitos B/metabolismo , Linfócitos B/patologia , DNA Metiltransferase 3A , Rearranjo Gênico do Linfócito T , Predisposição Genética para Doença , Humanos , Cadeias Pesadas de Imunoglobulinas/genética , Imuno-Histoquímica , Imunofenotipagem , Linfoma Extranodal de Células T-NK/metabolismo , Linfoma Extranodal de Células T-NK/patologia , Especificidade de Órgãos/genética , Fenótipo , Análise de Sequência de DNA , Recombinação V(D)J , Proteína rhoA de Ligação ao GTP/genética , Proteína rhoA de Ligação ao GTP/metabolismoRESUMO
In the treatment of childhood acute lymphoblastic leukemia (ALL), excess shortening of maintenance therapy resulted in high relapse rate, as shown by our previous trial, TCCSG L92-13, in which maintenance therapy was terminated at 1 year from initiation of treatment. In this study, we aimed to confirm the long-term outcome of L92-13, and to identify who can or cannot be cured by shorter duration of maintenance therapy. To obtain sentinel cytogenetics information that had been missed before, we performed genetic analysis with genomic microarray and target intron-capture sequencing from diagnostic bone marrow smear. Disease-free survival (DFS) at 10 years from the end of therapy was 66.0±2.8%. Females (n=138) had better DFS (74.6±3.7%) than males (n=142, 57.5±4.2%, P=0.002). Patients with TCF3-PBX1 (n=11) and ETV6-RUNX1 (n=16) had excellent DFS (90.9±8.7% and 93.8±6.1%, respectively), whereas high hyperdiploidy (n=23) was the most unfavorable subgroup, with 56.6±10.3% of DFS. Short duration of therapy can cure more than half of pediatric ALL, especially females, TCF3-PBX1 and ETV6-RUNX1. Our retrospective observations suggest a gender/karyotype inhomogeneity on the impact of brief therapy.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/mortalidade , Adolescente , Criança , Pré-Escolar , Feminino , Predisposição Genética para Doença , Humanos , Lactente , Recém-Nascido , Quimioterapia de Manutenção , Masculino , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Prognóstico , Recidiva , Fatores de Risco , Análise de Sobrevida , Fatores de Tempo , Translocação Genética , Resultado do TratamentoRESUMO
Although tyrosine kinase inhibitors (TKIs) have significantly improved the prognosis of chronic myeloid leukemia (CML), the ability of TKIs to eradicate CML remains uncertain and patients must continue TKI therapy for indefinite periods. In this study, we performed whole-exome sequencing to identify somatic mutations in 24 patients with newly diagnosed chronic phase CML who were registered in the JALSG CML212 study. We identified 191 somatic mutations other than the BCR-ABL1 fusion gene (median 8, range 1-17). Age, hemoglobin concentration and white blood cell counts were correlated with the number of mutations. Patients with mutations ⩾6 showed higher rate of achieving major molecular response than those<6 (P=0.0381). Mutations in epigenetic regulator, ASXL1, TET2, TET3, KDM1A and MSH6 were found in 25% of patients. TET2 or TET3, AKT1 and RUNX1 were mutated in one patient each. ASXL1 was mutated within exon 12 in three cases. Mutated genes were significantly enriched with cell signaling and cell division pathways. Furthermore, DNA copy number analysis showed that 2 of 24 patients had uniparental disomy of chromosome 1p or 3q, which disappeared major molecular response was achieved. These mutations may play significant roles in CML pathogenesis in addition to the strong driver mutation BCR-ABL1.
Assuntos
Proteínas de Ligação a DNA/genética , Dioxigenases/genética , Histona Desmetilases/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Repressoras/genética , Fatores Etários , Variações do Número de Cópias de DNA/genética , Resistencia a Medicamentos Antineoplásicos/genética , Epigênese Genética/genética , Feminino , Proteínas de Fusão bcr-abl/genética , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/sangue , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Contagem de Leucócitos , Masculino , Mutação , Inibidores de Proteínas Quinases/administração & dosagem , Transdução de Sinais , Sequenciamento do ExomaRESUMO
Partial tandem duplication of MLL (MLL-PTD) characterizes acute myeloid leukemia (AML) patients often with a poor prognosis. To understand the order of occurrence of MLL-PTD in relation to other major AML mutations and to identify novel mutations that may be present in this unique AML molecular subtype, exome and targeted sequencing was performed on 85 MLL-PTD AML samples using HiSeq-2000. Genes involved in the cohesin complex (STAG2), a splicing factor (U2AF1) and a poorly studied gene, MGA were recurrently mutated, whereas NPM1, one of the most frequently mutated AML gene, was not mutated in MLL-PTD patients. Interestingly, clonality analysis suggests that IDH2/1, DNMT3A, U2AF1 and TET2 mutations are clonal and occur early, and MLL-PTD likely arises after these initial mutations. Conversely, proliferative mutations (FLT3, RAS), typically appear later, are largely subclonal and tend to be unstable. This study provides important insights for understanding the relative importance of different mutations for defining a targeted therapeutic strategy for MLL-PTD AML patients.
Assuntos
Histona-Lisina N-Metiltransferase/genética , Leucemia Mieloide Aguda/genética , Mutação , Proteína de Leucina Linfoide-Mieloide/genética , Proliferação de Células/genética , Células Clonais , Exoma , Humanos , Taxa de Mutação , Nucleofosmina , Sequências de Repetição em Tandem , Fatores de TempoRESUMO
Metastasis is a critical factor contributing to poor prognosis in cancer, but the underlying mechanisms of metastasis are still poorly understood. We established a highly metastatic cell subline (HOC313-LM) derived from an oral squamous cell carcinoma cell line (HOC313) for uncovering the mechanisms of metastasis, and identified deoxyhypusine synthase (DHPS) as a metastasis-associated gene within the specific amplification at 19p13.2-p13.13 in HOC313-LM. DHPS-mediated hypusine-modification of eukaryotic translation factor 5A facilitated the translation of RhoA, resulting in the activation of the RhoA signaling pathway and leading to not only increased cell motility, invasion and metastasis of cancer cells in vitro, but also increased tumor growth in vivo. Moreover, the use of N1-Guanyl-1,7-diaminoheptane, a DHPS inhibitor, resulted in a significant decrease in tumor formation in vivo. In patients with esophageal squamous cell carcinoma (ESCC), overexpression of DHPS in ESCC tumors was significantly associated with worse recurrence-free survival, and correlated with distant metastasis. The elucidation of these molecular mechanisms within the hypusine cascade suggests opportunities for novel therapeutic targets in SCC.
Assuntos
Carcinoma de Células Escamosas/tratamento farmacológico , Neoplasias Esofágicas/tratamento farmacológico , Lisina/análogos & derivados , Oxirredutases atuantes sobre Doadores de Grupo CH-NH/genética , Proteína rhoA de Ligação ao GTP/genética , Adulto , Idoso , Animais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Diaminas/administração & dosagem , Progressão da Doença , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Lisina/biossíntese , Lisina/genética , Masculino , Camundongos , Pessoa de Meia-Idade , Metástase Neoplásica , Oxirredutases atuantes sobre Doadores de Grupo CH-NH/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacosRESUMO
UNLABELLED: Essentials Two groups recently reported GFI1B as a novel causative gene for congenital macrothrombocytopenia. We performed functional analysis of a novel GFI1B mutation and previous mutations. An immunofluorescence analysis of the platelet CD34 expression can be useful as a screening test. Mutant-transduced megakaryocytes produced enlarged proplatelet tips which were reduced in number. SUMMARY: Background GFI1B is an essential transcription factor for megakaryocyte and erythrocyte development. Two groups have recently identified GFI1B as a novel causative gene for congenital macrothrombocytopenia associated with α-granule deficiency. Methods We performed whole exome sequencing and identified a novel GFI1B p.G272fsX274 mutation in a family with macrothrombocytopenia, and a decreased number of platelet α-granules and abnormally shaped red blood cells. p.G272fsX274 and the previous two mutations all predicted disruption of an essential DNA-binding domain in GFI1B. We therefore performed functional studies to characterize the biochemical and biological effects of these three patient-derived mutations. Results An immunofluorescence analysis revealed decreased thrombospondin-1 and increased CD34 expression in platelets from our patient. Consistent with the previous studies, the three patient-derived mutants were unable to repress the expression of the reporter gene and had a dominant-negative effect over wild-type GFI1B. In addition, the three mutations abolished recognition of a consensus-binding site in gel shift assays. Furthermore, transduction of mouse fetal liver-derived megakaryocytes with the three GFI1B mutants resulted in the production of abnormally large proplatelet tips, which were reduced in number. Conclusions Our study provides further proof of concept that GFI1B is an essential protein for the normal development of the megakaryocyte lineage.
Assuntos
Plaquetas/metabolismo , Megacariócitos/citologia , Proteínas Proto-Oncogênicas/genética , Proteínas Repressoras/genética , Trombocitopenia/congênito , Trombocitopenia/genética , Adolescente , Animais , Antígenos CD34/sangue , Antígenos CD34/metabolismo , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Plaquetas/citologia , Linhagem da Célula , Deformação Eritrocítica , Eritrócitos/citologia , Exoma , Feminino , Genes Dominantes , Humanos , Masculino , Camundongos , Microscopia de Fluorescência , Mutação , Linhagem , Contagem de Plaquetas , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Repressoras/metabolismo , Análise de Sequência de DNA , Trombospondina 1/sangue , Trombospondina 1/genética , Trombospondina 1/metabolismoRESUMO
In this study, we identify signaling network of necrotic cell death induced by transcriptional repression (TRIAD) by α-amanitin (AMA), the selective RNA polymerase II inhibitor, as a model of neurodegenerative cell death. We performed genetic screen of a knockdown (KD) fly library by measuring the ratio of transformation from pupa to larva (PL ratio) under TRIAD, and selected the cell death-promoting genes. Systems biology analysis of the positive genes mapped on protein-protein interaction databases predicted the signaling network of TRIAD and the core pathway including heterogeneous nuclear ribonucleoproteins (hnRNPs) and huntingtin (Htt). RNA sequencing revealed that AMA impaired transcription and RNA splicing of Htt, which is known as an endoplasmic reticulum (ER)-stabilizing molecule. The impairment in RNA splicing and PL ratio was rescued by overexpresion of hnRNP that had been also affected by transcriptional repression. Fly genetics with suppressor or expresser of Htt and hnRNP worsened or ameliorated the decreased PL ratio by AMA, respectively. Collectively, these results suggested involvement of RNA splicing and a regulatory role of the hnRNP-Htt axis in the process of the transcriptional repression-induced necrosis.
Assuntos
Apoptose , Proteínas de Drosophila/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas/metabolismo , Proteína Huntingtina/metabolismo , Amanitinas/farmacologia , Animais , Apoptose/efeitos dos fármacos , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Células Cultivadas , Drosophila/crescimento & desenvolvimento , Drosophila/metabolismo , Proteínas de Drosophila/genética , Embrião de Mamíferos/citologia , Ribonucleoproteínas Nucleares Heterogêneas Grupo A-B/genética , Ribonucleoproteínas Nucleares Heterogêneas Grupo A-B/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas/genética , Proteína Huntingtina/antagonistas & inibidores , Proteína Huntingtina/genética , Larva/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Pupa/metabolismo , Splicing de RNA/efeitos dos fármacos , Ratos , Ratos Wistar , Ribonucleoproteínas/genética , Ribonucleoproteínas/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transcrição Gênica/efeitos dos fármacos , Proteínas Contendo Repetições de beta-Transducina/genética , Proteínas Contendo Repetições de beta-Transducina/metabolismo , Quinase 1 Polo-LikeRESUMO
Acute promyelocytic leukemia (APL) is a subtype of myeloid leukemia characterized by differentiation block at the promyelocyte stage. Besides the presence of chromosomal rearrangement t(15;17), leading to the formation of PML-RARA (promyelocytic leukemia-retinoic acid receptor alpha) fusion, other genetic alterations have also been implicated in APL. Here, we performed comprehensive mutational analysis of primary and relapse APL to identify somatic alterations, which cooperate with PML-RARA in the pathogenesis of APL. We explored the mutational landscape using whole-exome (n=12) and subsequent targeted sequencing of 398 genes in 153 primary and 69 relapse APL. Both primary and relapse APL harbored an average of eight non-silent somatic mutations per exome. We observed recurrent alterations of FLT3, WT1, NRAS and KRAS in the newly diagnosed APL, whereas mutations in other genes commonly mutated in myeloid leukemia were rarely detected. The molecular signature of APL relapse was characterized by emergence of frequent mutations in PML and RARA genes. Our sequencing data also demonstrates incidence of loss-of-function mutations in previously unidentified genes, ARID1B and ARID1A, both of which encode for key components of the SWI/SNF complex. We show that knockdown of ARID1B in APL cell line, NB4, results in large-scale activation of gene expression and reduced in vitro differentiation potential.
Assuntos
Análise Mutacional de DNA/métodos , Leucemia Promielocítica Aguda/genética , Diferenciação Celular , Proteínas de Ligação a DNA/genética , Exoma/genética , Perfilação da Expressão Gênica , Humanos , Proteínas Nucleares/genética , Recidiva , Fatores de Transcrição/genéticaRESUMO
MOTIVATION: Sigma factors regulate the expression of genes in Bacillus subtilis at the transcriptional level. We assess the accuracy of a fold-change analysis, Bayesian networks, dynamic models and supervised learning based on coregulation in predicting gene regulation by sigma factors from gene expression data. To improve the prediction accuracy, we combine sequence information with expression data by adding their log-likelihood scores and by using a logistic regression model. We use the resulting score function to discover currently unknown gene regulations by sigma factors. RESULTS: The coregulation-based supervised learning method gave the most accurate prediction of sigma factors from expression data. We found that the logistic regression model effectively combines expression data with sequence information. In a genome-wide search, highly significant logistic regression scores were found for several genes whose transcriptional regulation is currently unknown. We provide the corresponding RNA polymerase binding sites to enable a straightforward experimental verification of these predictions.
Assuntos
Bacillus subtilis/metabolismo , Proteínas de Bactérias/genética , Mapeamento Cromossômico/métodos , Interpretação Estatística de Dados , Regulação da Expressão Gênica/fisiologia , Modelos Biológicos , Fator sigma/fisiologia , Algoritmos , Simulação por Computador , Perfilação da Expressão Gênica , Modelos EstatísticosRESUMO
BACKGROUND: In kidney transplant recipients, the most widely used method for the reconstruction of the urinary pathway is ureteroneocystostomy, which may be difficult in cases with disused atrophic bladder. In this study, we evaluated kidney transplant recipients who underwent uretero-ureteral end-to-side anastomosis (UUA) in urinary reconstruction due to disused atrophic bladder. METHODS: To clarify the effectiveness of this method, we retrospectively reviewed the clinical records of kidney transplant recipients in our hospital. RESULTS: A total of 9 recipients with urinary reconstruction using UUA were evaluated. All of these patients had a history of long-term hemodialysis before transplantation, accompanied by complete anuria and small capacity of the bladder. In 4 patients, cranial native ureter was ligated, whereas it was not ligated in the remaining 5 patients. In 2 of 4 patients with cranial ligation, hydronephrosis developed in the native kidney with no further treatment being required. No patients experienced urinary tract complications including hydronephrosis in the graft, urine extravasation, or urinary tract infection in the follow-up period (757.6 ± 491.3 days). Allograft function was maintained well in all patients (serum creatinine level, 1.08 ± 0.23 mg/dL). CONCLUSIONS: Although UUA is not a routine method of urinary reconstruction in kidney transplantation, it can be safely performed and should be a surgical option, especially for recipients with disused atrophic bladder. The ligation of cranial native ureter may lead to hydronephrosis of the native kidney, and it is tentatively concluded that UUA without native ureteral ligation is clinically feasible.
Assuntos
Falência Renal Crônica/terapia , Transplante de Rim , Procedimentos de Cirurgia Plástica/métodos , Ureter/cirurgia , Bexiga Urinária/patologia , Bexiga Urinária/cirurgia , Adulto , Anastomose Cirúrgica , Atrofia/etiologia , Atrofia/patologia , Atrofia/cirurgia , Feminino , Humanos , Falência Renal Crônica/etiologia , Falência Renal Crônica/patologia , Ligadura , Masculino , Pessoa de Meia-Idade , Diálise Renal/efeitos adversos , Estudos RetrospectivosRESUMO
Allogeneic hematopoietic stem cell transplantation (HSCT) is one of curative treatment options for patients with hematologic malignancies. Although GVHD mediated by the donor's T lymphocytes remains the most challenging toxicity of allo-HSCT, graft-versus-leukemia (GVL) effect targeting leukemic cells, has an important role in affecting the overall outcome of patients with AML. Here we comprehensively characterized the TCR repertoire in patients who underwent matched donor or haplo-cord HSCT using next-generation sequencing approach. Our study defines the functional kinetics of each TCRA and TCRB clone, and changes in T-cell diversity (with identification of CDR3 sequences) and the extent of clonal expansion of certain T-cells. Using this approach, our study demonstrates that higher percentage of cord-blood cells at 30 days after transplant was correlated with higher diversity of TCR repertoire, implicating the role of cord-chimerism in enhancing immune recovery. Importantly, we found that GVHD and relapse, exclusive of each other, were correlated with lower TCR repertoire diversity and expansion of certain T-cell clones. Our results highlight novel insights into the balance between GVHD and GVL effect, suggesting that higher diversity early after transplant possibly implies lower risks of both GVHD and relapse following the HSCT transplantation.
Assuntos
Transplante de Células-Tronco de Sangue do Cordão Umbilical , Transplante de Células-Tronco Hematopoéticas , Leucemia Mieloide Aguda , Síndromes Mielodisplásicas , Receptores de Antígenos de Linfócitos T alfa-beta , Linfócitos T/imunologia , Adulto , Idoso , Aloenxertos , Regiões Determinantes de Complementaridade/genética , Regiões Determinantes de Complementaridade/imunologia , Feminino , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/terapia , Masculino , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/genética , Síndromes Mielodisplásicas/imunologia , Síndromes Mielodisplásicas/terapia , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Receptores de Antígenos de Linfócitos T alfa-beta/imunologiaRESUMO
Mutations of spliceosome components are common in myeloid neoplasms. One of the affected genes, PRPF8, encodes the most evolutionarily conserved spliceosomal protein. We identified either recurrent somatic PRPF8 mutations or hemizygous deletions in 15/447 and 24/450 cases, respectively. Fifty percent of PRPF8 mutant and del(17p) cases were found in AML and conveyed poor prognosis. PRPF8 defects correlated with increased myeloblasts and ring sideroblasts in cases without SF3B1 mutations. Knockdown of PRPF8 in K562 and CD34+ primary bone marrow cells increased proliferative capacity. Whole-RNA deep sequencing of primary cells from patients with PRPF8 abnormalities demonstrated consistent missplicing defects. In yeast models, homologous mutations introduced into Prp8 abrogated a block experimentally produced in the second step of the RNA splicing process, suggesting that the mutants have defects in proof-reading functions. In sum, the exploration of clinical and functional consequences suggests that PRPF8 is a novel leukemogenic gene in myeloid neoplasms with a distinct phenotype likely manifested through aberrant splicing.
Assuntos
Neoplasias Hematológicas/genética , Splicing de RNA , Proteínas de Ligação a RNA/genética , Sequência de Aminoácidos , Proliferação de Células , Deleção de Genes , Técnicas de Silenciamento de Genes , Neoplasias Hematológicas/patologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Dados de Sequência Molecular , Proteínas de Ligação a RNA/química , Reação em Cadeia da Polimerase em Tempo Real , Homologia de Sequência de AminoácidosRESUMO
Mutations in isocitrate dehydrogenase 1/2 (IDH1/2(MT)) are drivers of a variety of myeloid neoplasms. As they yield the same oncometabolite, D-2-hydroxyglutarate, they are often treated as equivalent, and pooled. We studied the validity of this approach and found IDH1/2 mutations in 179 of 2119 myeloid neoplasms (8%). Cross-sectionally, the frequencies of these mutations increased from lower- to higher risk disease, thus suggesting a role in clinical progression. Variant allelic frequencies indicated that IDH1(MT) and IDH2(MT) are ancestral in up to 14/74 (19%) vs 34/99 (34%; P=0.027) of cases, respectively, illustrating the pathogenic role of these lesions in myeloid neoplasms. IDH1/2(MT) was associated with poor overall survival, particularly in lower risk myelodysplastic syndromes. Ancestral IDH1(MT) cases were associated with a worse prognosis than subclonal IDH1(MT) cases, whereas the position of IDH2(MT) within clonal hierarchy did not impact survival. This may relate to distinct mutational spectra with more DNMT3A and NPM1 mutations associated with IDH1(MT) cases, and more ASXL1, SRSF2, RUNX1, STAG2 mutations associated with IDH2(MT) cases. Our data demonstrate important clinical and biological differences between IDH1(MT) and IDH2(MT) myeloid neoplasms. These mutations should be considered separately as their differences could have implications for diagnosis, prognosis and treatment with IDH1/2(MT) inhibitors of IDH1/2(MT) patients.