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1.
Haematologica ; 109(4): 1171-1183, 2024 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-37646663

RESUMEN

The transcription factor MYC is a well-described oncogene with an important role in lymphomagenesis, but its significance for clinical outcome in mantle cell lymphoma (MCL) remains to be determined. We performed an investigation of the expression of MYC protein in a cohort of 251 MCL patients complemented by analyses of structural aberrations and mRNA, in a sub-cohort of patients. Fourteen percent (n=35) of patients showed high MYC protein expression with >20% positive cells (MYChigh), among whom only one translocation was identified, and 86% (n=216) of patients showed low MYC protein expression. Low copy number gains of MYC were detected in ten patients, but with no correlation to MYC protein levels. However, MYC mRNA levels correlated significantly to MYC protein levels with a R2 value of 0.76. Patients with a MYChigh tumor had both an independent inferior overall survival and an inferior progression-free survival (hazard ratio [HR]=2.03, 95% confidence interval [95% CI]: 1.2-3.4 and HR=2.2, 95% CI: 1.04-4.6, respectively) when adjusted for additional high-risk features. Patients with MYChigh tumors also tended to have additional high-risk features and to be older at diagnosis. A subgroup of 13 patients had concomitant MYChigh expression and TP53/p53 alterations and a substantially increased risk of progression (HR=16.9, 95% CI: 7.4-38.3) and death (HR=7.8, 95% CI: 4.4-14.1) with an average overall survival of only 0.9 years. In summary, we found that at diagnosis a subset of MCL patients (14%) overexpressed MYC protein, and had a poor prognosis but that MYC rearrangements were rare. Tumors with concurrent MYC overexpression and TP53/p53 alterations pinpointed MCL patients with a dismal prognosis with a median overall survival of less than 3 years. We propose that MYC needs to be assessed beyond the current high-risk factors in MCL in order to identify cases in need of alternative treatment.


Asunto(s)
Linfoma de Células del Manto , Adulto , Humanos , Proliferación Celular , Linfoma de Células del Manto/diagnóstico , Linfoma de Células del Manto/genética , Pronóstico , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo , ARN Mensajero , Translocación Genética , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo
2.
J Biol Chem ; 294(11): 4119-4136, 2019 03 15.
Artículo en Inglés | MEDLINE | ID: mdl-30622137

RESUMEN

TGFß signaling via SMAD proteins and protein kinase pathways up- or down-regulates the expression of many genes and thus affects physiological processes, such as differentiation, migration, cell cycle arrest, and apoptosis, during developmental or adult tissue homeostasis. We here report that NUAK family kinase 1 (NUAK1) and NUAK2 are two TGFß target genes. NUAK1/2 belong to the AMP-activated protein kinase (AMPK) family, whose members control central and protein metabolism, polarity, and overall cellular homeostasis. We found that TGFß-mediated transcriptional induction of NUAK1 and NUAK2 requires SMAD family members 2, 3, and 4 (SMAD2/3/4) and mitogen-activated protein kinase (MAPK) activities, which provided immediate and early signals for the transient expression of these two kinases. Genomic mapping identified an enhancer element within the first intron of the NUAK2 gene that can recruit SMAD proteins, which, when cloned, could confer induction by TGFß. Furthermore, NUAK2 formed protein complexes with SMAD3 and the TGFß type I receptor. Functionally, NUAK1 suppressed and NUAK2 induced TGFß signaling. This was evident during TGFß-induced epithelial cytostasis, mesenchymal differentiation, and myofibroblast contractility, in which NUAK1 or NUAK2 silencing enhanced or inhibited these responses, respectively. In conclusion, we have identified a bifurcating loop during TGFß signaling, whereby transcriptional induction of NUAK1 serves as a negative checkpoint and NUAK2 induction positively contributes to signaling and terminal differentiation responses to TGFß activity.


Asunto(s)
Proteínas Quinasas/genética , Proteínas Serina-Treonina Quinasas/genética , Proteínas Represoras/genética , Transducción de Señal/genética , Factor de Crecimiento Transformador beta/metabolismo , Células Cultivadas , Perfilación de la Expresión Génica , Humanos , Proteínas Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteínas Represoras/metabolismo
3.
Cell Commun Signal ; 16(1): 64, 2018 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-30285764

RESUMEN

BACKGROUND: Mammalian target of rapamycin (mTOR) is a master regulator of various cellular responses by forming two functional complexes, mTORC1 and mTORC2. mTOR signaling is frequently dysregulated in pancreatic neuroendocrine tumors (PNETs). mTOR inhibitors have been used in attempts to treat these lesions, and prolonged progression free survival has been recorded. If this holds true also for the multiple endocrine neoplasia type 1 (MEN1) associated PNETs is yet unclear. We investigated the relationship between expression of the MEN1 protein menin and mTOR signaling in the presence or absence of the mTOR inhibitor rapamycin. METHODS: In addition to use of menin wild type and menin-null mouse embryonic fibroblasts (MEFs), menin was silenced by siRNA in pancreatic neuroendocrine tumor cell line BON-1. Panels of protein phosphorylation, as activation markers downstream of PI3k-mTOR-Akt pathways, as well as menin expression were evaluated by immunoblotting. The impact of menin expression in the presence and absence of rapamycin was determinate upon Wound healing, migration and proliferation in MEFs and BON1 cells. RESULTS: PDGF-BB markedly increased phosphorylation of mTORC2 substrate Akt, at serine 473 (S473) and threonine 450 (T450) in menin-/- MEFs but did not alter phosphorylation of mTORC1 substrates ribosomal protein S6 or eIF4B. Acute rapamycin treatment by mTORC1-S6 inhibition caused a greater enhancement of Akt phosphorylation on S473 in menin-/- cells as compared to menin+/+ MEFs (116% vs 38%). Chronic rapamycin treatment, which inhibits both mTORC1and 2, reduced Akt phosphorylation of S473 to a lesser extent in menin-/- MEFs than menin+/+ MEFs (25% vs 75%). Silencing of menin expression in human PNET cell line (BON1) also enhanced Akt phosphorylation at S473, but not activation of mTORC1. Interestingly, silencing menin in BON1 cells elevated S473 phosphorylation of Akt in both acute and chronic treatments with rapamycin. Finally, we show that the inhibitory effect of rapamycin on serum mediated wound healing and cell migration is impaired in menin-/- MEFs, as well as in menin-silenced BON1 cells. CONCLUSIONS: Menin is involved in regulatory mechanism between the two mTOR complexes, and its reduced expression is accompanied with increased mTORC2-Akt signaling, which consequently impairs anti-migratory effect of rapamycin.


Asunto(s)
Regulación de la Expresión Génica/efectos de los fármacos , Diana Mecanicista del Complejo 2 de la Rapamicina/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Transducción de Señal/efectos de los fármacos , Sirolimus/farmacología , Animales , Línea Celular , Técnicas de Silenciamiento del Gen , Humanos , Ratones , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas/deficiencia , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , ARN Interferente Pequeño/genética
4.
Int J Gynecol Cancer ; 26(6): 1027-32, 2016 07.
Artículo en Inglés | MEDLINE | ID: mdl-27177282

RESUMEN

OBJECTIVE: This study aimed to perform an integrative genetic analysis of patients with matched serous ovarian cancer having long-term or short-term survival using formalin fixed paraffin-embedded (FFPE) tissue samples. METHODS: All patients with serous ovarian carcinoma who underwent surgery between 1998 and 2007 at the Department of Gynaecology, Uppsala University Hospital, Sweden were considered. From this cohort, we selected biomaterial from 2 groups of patients with long-term and short-term survival matched for age, stage, histologic grade, and outcome of surgery. Genomic DNA from FFPE sample was analyzed with SNP array and targeted next-generation sequencing of 26 genes. RESULTS: Forty-three samples (primary tumors and metastases) from 23 patients were selected for genomic profiling, the survival in the subgroups were 134 and 36 months, respectively. We observed a tendency toward increased genomic instability in those with long-term survival with higher proportion of somatic copy number alterations (P = 0.083) and higher average ploidy (P = 0.037). TP53 mutations were found in 50% of the patients. Frequency of TP53 mutations did not differ between the survival groups (P = 0.629). CONCLUSIONS: We validated both previous genomic findings in ovarian cancer and the proposed association between increased genomic instability and better survival. These results exemplify that analysis of genomic biomarkers is feasible on archived FFPE tissue.


Asunto(s)
Cistadenocarcinoma Seroso/genética , Cistadenocarcinoma Seroso/mortalidad , Neoplasias Ováricas/genética , Neoplasias Ováricas/mortalidad , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Cohortes , Cistadenocarcinoma Seroso/patología , Femenino , Formaldehído , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Persona de Mediana Edad , Neoplasias Ováricas/patología , Adhesión en Parafina , Polimorfismo de Nucleótido Simple , Estudios Retrospectivos , Sobrevivientes , Suecia/epidemiología , Fijación del Tejido
5.
Cell Commun Signal ; 11(1): 3, 2013 Jan 11.
Artículo en Inglés | MEDLINE | ID: mdl-23311350

RESUMEN

Mammalian target of rapamycin (mTOR) can be found in two multi-protein complexes, i.e. mTORC1 (containing Raptor) and mTORC2 (containing Rictor). Here, we investigated the mechanisms by which mTORC1 and mTORC2 are activated and their downstream targets in response to platelet-derived growth factor (PDGF)-BB treatment. Inhibition of phosphatidylinositol 3-kinase (PI3K) inhibited PDGF-BB activation of both mTORC1 and mTORC2. We found that in Rictor-null mouse embryonic fibroblasts, or after prolonged rapamycin treatment of NIH3T3 cells, PDGF-BB was not able to promote phosphorylation of Ser473 in the serine/threonine kinase Akt, whereas Thr308 phosphorylation was less affected, suggesting that Ser473 in Akt is phosphorylated in an mTORC2-dependent manner. This reduction in Akt phosphorylation did not influence the phosphorylation of the S6 protein, a well established protein downstream of mTORC1. Consistently, triciribine, an inhibitor of the Akt pathway, suppressed PDGF-BB-induced Akt phosphorylation without having any effect on S6 phosphorylation. Thus, mTORC2 does not appear to be upstream of mTORC1. We could also demonstrate that in Rictor-null cells the phosphorylation of phospholipase Cγ1 (PLCγ1) and protein kinase C (PKC) was impaired, and the PKCα protein levels strongly reduced. Furthermore, interfering with the PLCγ/Ca2+/PKC pathway inhibited PDGF-BB-induced Akt phosphorylation. In addition, PDGF-BB-induced activation of mTORC1, as measured by phosphorylation of the downstream S6 protein, was dependent on phospholipase D (PLD). It has been shown that Erk1/2 MAP-kinase directly phosphorylates and activates mTORC1; in partial agreement with this finding, we found that a Mek1/2 inhibitor delayed S6 phosphorylation in response to PDGF-BB, but it did not block it. Thus, whereas both mTORC1 and mTORC2 are activated in a PI3K-dependent manner, different additional signaling pathways are needed. mTORC1 is activated in a PLD-dependent manner and promotes phosphorylation of the S6 protein, whereas mTORC2, in concert with PLCγ signaling, promotes Akt phosphorylation.

6.
Sci Rep ; 11(1): 14772, 2021 07 20.
Artículo en Inglés | MEDLINE | ID: mdl-34285285

RESUMEN

Adrenocortical carcinoma is a rare aggressive disease commonly recurring regardless of radical surgery. Although data on genomic alterations in malignant tumors are accumulating, knowledge of molecular events of importance for initiation of adrenocortical transformation is scarce. In an attempt to recognize early molecular alterations, we used adrenals from young multiple endocrine neoplasia type 1 conventional knock-out mice (Men1+/-) closely mimicking the human MEN1 trait (i.e. transformation of pituitary, parathyroid, endocrine pancreatic, and adrenocortical cells). MicroRNA array and hierarchical clustering showed a distinct pattern. Twenty miRNAs were significantly upregulated and eleven were downregulated in Men1+/- compared to wild type littermates. The latter included the known suppressor miRNA miR-486-3p, which was chosen for transfection in human adrenocortical carcinoma cell lines H295R and SW13. Cell growth decreased in miR-486-3p overexpressing clones and levels of the predicted target gene fatty acid synthase (FASN) and its downstream product, palmitic acid, were lowered. In conclusion, heterozygous inactivation of Men1 in adrenals results in distinct miRNA profile regulating expression of genes with impact on tumorigenesis, e.g. transcription, nucleic acid and lipid metabolism. Low levels of miR-486-3p in the early stages of transformation may contribute to proliferation by increasing FASN and thus fatty acid production. FASN as a potentially druggable target for treatment of the devastating disease adrenocortical carcinoma warrants further studies.


Asunto(s)
Neoplasias de la Corteza Suprarrenal/genética , Carcinoma Corticosuprarrenal/genética , Regulación hacia Abajo , Acido Graso Sintasa Tipo I/genética , MicroARNs/genética , Proteínas Proto-Oncogénicas/genética , Neoplasias de la Corteza Suprarrenal/metabolismo , Carcinoma Corticosuprarrenal/metabolismo , Animales , Línea Celular Tumoral , Proliferación Celular , Aprendizaje Profundo , Acido Graso Sintasa Tipo I/metabolismo , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Ratones , Ratones Noqueados , Análisis de Secuencia por Matrices de Oligonucleótidos
7.
Sci Rep ; 10(1): 14572, 2020 09 03.
Artículo en Inglés | MEDLINE | ID: mdl-32884006

RESUMEN

Among patients with the rare diagnosis of pancreatic neuroendocrine tumor (P-NET), a substantial proportion suffer from the inherited cancer syndrome multiple endocrine neoplasia type 1 (MEN1), which is caused by germline mutations of the MEN1 suppressor gene. Somatic mutations and loss of the MEN1 protein (menin) are frequently also found in sporadic P-NETs. Thus, a human neuroendocrine pancreatic cell line with biallelic inactivation of MEN1 might be of value for studying tumorigenesis. We used the polyclonal human P-NET cell line BON1, which expresses menin, serotonin, chromogranin A and neurotensin, to generate a monoclonal stable MEN1 knockout BON1 cell line (MEN1-KO-BON1) by CRISPR/Cas9 editing. Changes in morphology, hormone secretion, and proliferation were analyzed, and proteomics were assessed using nanoLC-MS/MS and Ingenuity Pathway Analysis (IPA). The menin-lacking MEN1-KO-BON1 cells had increased chromogranin A production and were smaller, more homogenous, rounder and grew faster than their control counterparts. Proteomic analysis revealed 457 significantly altered proteins, and IPA identified biological functions related to cancer, e.g., posttranslational modification and cell death/survival. Among 39 proteins with at least a two-fold difference in expression, twelve are relevant in glucose homeostasis and insulin resistance. The stable monoclonal MEN1-KO-BON1 cell line was found to have preserved neuroendocrine differentiation, increased proliferation, and an altered protein profile.


Asunto(s)
Biomarcadores de Tumor/metabolismo , Sistemas CRISPR-Cas , Edición Génica , Tumores Neuroendocrinos/patología , Neoplasias Pancreáticas/patología , Proteoma/análisis , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Regulación Neoplásica de la Expresión Génica , Humanos , Tumores Neuroendocrinos/metabolismo , Neoplasias Pancreáticas/metabolismo , Proteínas Proto-Oncogénicas/genética , Células Tumorales Cultivadas
8.
Sci Rep ; 8(1): 748, 2018 01 15.
Artículo en Inglés | MEDLINE | ID: mdl-29335487

RESUMEN

Multiple endocrine neoplasia type 1 (MEN1) is an endocrine tumor syndrome caused by heterozygous mutations in the MEN1 tumor suppressor gene. The MEN1 pancreas of the adolescent gene carrier frequently contain diffusely spread pre-neoplasias and microadenomas, progressing to macroscopic and potentially malignant pancreatic neuroendocrine tumors (P-NET), which represents the major death cause in MEN1. The unveiling of the molecular mechanism of P-NET which is not currently understood fully to allow the optimization of diagnostics and treatment. Glucagon-like peptide 1 (GLP-1) pathway is essential in islet regeneration, i.e. inhibition of ß-cell apoptosis and enhancement of ß-cell proliferation, yet involvement of GLP-1 in MEN1 related P-NET has not yet been demonstrated. The objective of this work was to investigate if normal sized islets of Men1 heterozygous mice have increased Glucagon-like peptide-1 receptor (GLP-1R) expression compared to wild type islets, and if this increase is detectable in vivo with positron emission tomography (PET) using [68Ga]Ga-DO3A-VS-Cys40-Exendin-4 (68Ga-Exendin-4). 68Ga-Exendin-4 showed potential for early lesion detection in MEN1 pancreas due to increased GLP1R expression.


Asunto(s)
Carcinoma Neuroendocrino/diagnóstico por imagen , Receptor del Péptido 1 Similar al Glucagón/análisis , Heterocigoto , Neoplasias Pancreáticas/diagnóstico por imagen , Tomografía de Emisión de Positrones/métodos , Proteínas Proto-Oncogénicas/genética , Animales , Carcinoma Neuroendocrino/patología , Ratones , Neoplasias Pancreáticas/patología
10.
Cell Signal ; 24(3): 635-40, 2012 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22100392

RESUMEN

MAP kinase phosphatase-3 (MKP3), also known as DUSP6 or Pyst1, is a dual specificity phosphatase considered to selectively dephosphorylate extracellular-signal-regulated kinase 1/2 (Erk1/2). Here, we report that in NIH3T3 cells, MKP3 is induced in response to platelet-derived growth factor (PDGF)-BB treatment in an Erk1/2- and phosphatidylinositol 3-kinase (PI3K)-dependent manner, but independently of Erk5 expression. Silencing of MKP3 expression did not affect PDGF-BB-induced Erk1/2 or p38 phosphorylation; however, their basal level of phosphorylation was elevated. Furthermore, we found that PDGF-BB-mediated activation of Erk5 and Akt was enhanced when the MKP3 expression was reduced. Interfering with Mek1/2 or PI3K using the inhibitors CI-1040 and LY-294002, respectively, inhibited PDGF-BB-induced MKP3 expression. Functionally, we found that MKP3 silencing did not affect cell proliferation, but enhanced the chemotactic response toward PDGF-BB. Although both Akt and Erk5 have been linked to increased cell survival, downregulation of MKP3 did not alter the ability of PDGF-BB to protect NIH3T3 cells from starvation-induced apoptosis. However, we observed an increased apoptosis in untreated cells with reduced MKP3 expression. In summary, our data indicate that there is negative cross-talk between Erk1/2 and Erk5 that involves regulation of MKP3 expression, and that PI3K in addition to promoting Akt phosphorylation also negatively modulates Akt, through MKP3 expression.


Asunto(s)
Quimiotaxis/efectos de los fármacos , Fosfatasa 6 de Especificidad Dual/metabolismo , Proteína Quinasa 7 Activada por Mitógenos/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-sis/farmacología , Animales , Apoptosis , Becaplermina , Movimiento Celular , Proliferación Celular , Fosfatasa 6 de Especificidad Dual/antagonistas & inhibidores , Fosfatasa 6 de Especificidad Dual/genética , Ratones , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Células 3T3 NIH , Fosfatidilinositol 3-Quinasas/metabolismo , Fosforilación/efectos de los fármacos , Interferencia de ARN , ARN Interferente Pequeño
11.
Br J Haematol ; 133(3): 315-22, 2006 May.
Artículo en Inglés | MEDLINE | ID: mdl-16643434

RESUMEN

Diabetes mellitus (DM) and hyperglycaemia are associated with platelet activation. The present study was designed to investigate how high glucose levels influence platelet function. Fasting human blood was incubated with different concentrations of D-glucose (5, 15 and 30 mmol/l) and other sugars without or with in vitro stimuli. Platelet activation was monitored by whole blood flow cytometry. High glucose levels enhanced adenosine diphosphate (ADP)- and thrombin receptor-activating peptide (TRAP)-induced platelet P-selectin expression, and TRAP-induced platelet fibrinogen binding. Similar effects were seen with 30 mmol/l L-glucose, sucrose and galactose. Hyperglycaemia also increased TRAP-induced platelet-leucocyte aggregation. Protein kinase C (PKC) blockade did not counteract the enhancement of platelet P-selectin expression, but abolished the enhancement of TRAP-induced platelet fibrinogen binding by hyperglycaemia. Superoxide anion scavenging by superoxide dismutase (SOD) attenuated the hyperglycaemic enhancement of platelet P-selectin expression, but did not counteract the enhancement of TRAP-induced platelet fibrinogen binding. Hyperglycaemia did not alter platelet intracellular calcium responses to agonist stimulation. Blockade of cyclo-oxygenase (COX), phosphotidylinositol-3 (PI3) kinase, or nitric oxide synthase, or the addition of insulin did not influence the effect of hyperglycaemia. In conclusion, high glucose levels enhanced platelet reactivity to agonist stimulation through elevated osmolality. This occurred via superoxide anion production, which enhanced platelet P-selectin expression (secretion), and PKC signalling, which enhanced TRAP-induced fibrinogen binding (aggregablity).


Asunto(s)
Glucosa/farmacología , Activación Plaquetaria/efectos de los fármacos , Adenosina Difosfato/farmacología , Adulto , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Calcio/sangre , Células Cultivadas , Relación Dosis-Respuesta a Droga , Femenino , Fibrinógeno/metabolismo , Citometría de Flujo , Galactosa/farmacología , Humanos , Hiperglucemia/sangre , Masculino , Persona de Mediana Edad , Concentración Osmolar , Selectina-P/sangre , Fragmentos de Péptidos/farmacología , Activación Plaquetaria/fisiología , Agregación Plaquetaria/efectos de los fármacos , Proteína Quinasa C/fisiología , Sacarosa/farmacología , Superóxidos/metabolismo
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