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1.
Cancer Cell ; 33(5): 937-948.e8, 2018 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-29681510

RESUMO

Somatic genetic alterations of IKZF1, which encodes the lymphoid transcription factor IKAROS, are common in high-risk B-progenitor acute lymphoblastic leukemia (ALL) and are associated with poor prognosis. Such alterations result in the acquisition of stem cell-like features, overexpression of adhesion molecules causing aberrant cell-cell and cell-stroma interaction, and decreased sensitivity to tyrosine kinase inhibitors. Here we report coding germline IKZF1 variation in familial childhood ALL and 0.9% of presumed sporadic B-ALL, identifying 28 unique variants in 45 children. The majority of variants adversely affected IKZF1 function and drug responsiveness of leukemic cells. These results identify IKZF1 as a leukemia predisposition gene, and emphasize the importance of germline genetic variation in the development of both familial and sporadic ALL.


Assuntos
Mutação em Linhagem Germinativa , Fator de Transcrição Ikaros/genética , Fator de Transcrição Ikaros/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Animais , Criança , Feminino , Mutação da Fase de Leitura , Predisposição Genética para Doença , Humanos , Masculino , Camundongos , Transplante de Neoplasias , Linhagem , Leucemia-Linfoma Linfoblástico de Células Precursoras B/metabolismo , Análise de Sequência de DNA
2.
Arch Pharm (Weinheim) ; 349(12): 925-933, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27862215

RESUMO

Janus kinases (JAKs) and their gain-of-function mutants have been implicated in a range of oncological, inflammatory, and autoimmune conditions, which has sparked great research interest in the discovery and development of small-molecule JAK inhibitors. Two molecules are currently marketed as JAK inhibitors, but due to the displayed side effects (owing to their suboptimal selectivities among the various JAK subtypes) new JAK inhibitors are still sought after. We present the results of an extensive virtual screening campaign based on a multi-step screening protocol involving ligand docking. The screening yielded five new, experimentally validated inhibitors of JAK1 with 8-hydroxyquinoline as a novel hinge-binding scaffold. The compounds did not only display favorable potencies in a JAK1V658F -driven cell-based assay but were also shown to be non-cytotoxic on rat liver cells.


Assuntos
Janus Quinase 1/antagonistas & inibidores , Oxiquinolina/análogos & derivados , Oxiquinolina/farmacologia , Animais , Morte Celular/efeitos dos fármacos , Células Cultivadas , Camundongos , Simulação de Acoplamento Molecular , Mutação , Oxiquinolina/síntese química , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/farmacologia , Ratos , Relação Estrutura-Atividade
3.
Artigo em Inglês | MEDLINE | ID: mdl-28781887

RESUMO

Variants of Uncertain Significance (VUS) are genetic variants whose association with a disease phenotype has not been established. They are a common finding in sequencing-based genetic tests and pose a significant clinical challenge. The objective of this study was to assess the use of functional data to classify variants according to pathogenicity. We conduct functional analysis of a large set of BRCA1 VUS combining a validated functional assay with VarCall, a Bayesian hierarchical model to estimate the likelihood of pathogenicity given the functional data. The results from the functional assays were incorporated into a joint analysis of 214 BRCA1 VUS to predict their likelihood of pathogenicity (breast cancer). We show that applying the VarCall model (1.0 sensitivity; lower bound of 95% confidence interval (CI) = 0.75 and 1.0 specificity; lower bound of 95% CI = 0.83) to the current set of BRCA1 variants, use of the functional data would significantly reduce the number of VUS associated with the C-terminal region of the BRCA1 protein by ~ 87%. We extend this work developing yeast-based functional assays for two other genes coding for BRCT domain containing proteins, MCPH1 and MDC1. Analysis of missense variants in MCPH1 and MDC1 shows that structural inference based on the BRCA1 data set can aid in prioritising variants for further analysis. Taken together our results indicate that systematic functional assays can provide a robust tool to aid in clinical annotation of VUS. We propose that well-validated functional assays could be used for clinical annotation even in the absence of additional sources of evidence.

4.
Nat Protoc ; 11(1): 46-60, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26658467

RESUMO

This protocol provides a rapid, streamlined and scalable strategy to systematically scan genomic regions for the presence of transcriptional regulatory regions that are active in a specific cell type. It creates genomic tiles spanning a region of interest that are subsequently cloned by recombination into a luciferase reporter vector containing the simian virus 40 promoter. Tiling clones are transfected into specific cell types to test for the presence of transcriptional regulatory regions. The protocol includes testing of different single-nucleotide polymorphism (SNP) alleles to determine their effect on regulatory activity. This procedure provides a systematic framework for identifying candidate functional SNPs within a locus during functional analysis of genome-wide association studies. This protocol adapts and combines previous well-established molecular biology methods to provide a streamlined strategy, based on automated primer design and recombinational cloning, allowing one to rapidly go from a genomic locus to a set of candidate functional SNPs in 8 weeks.


Assuntos
Elementos Facilitadores Genéticos/genética , Loci Gênicos/genética , Genômica/métodos , Sequência de Bases , Linhagem Celular , Genômica/instrumentação , Humanos , Polimorfismo de Nucleotídeo Único/genética
5.
Sci Rep ; 5: 17367, 2015 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-26610392

RESUMO

Glioma is the most common malignant primary brain tumor and is associated with poor prognosis. Genetic factors contributing to glioma risk have recently been investigated through genome-wide association studies (GWAS), implicating seven independent glioma risk loci in six chromosomal regions. Here, we performed an in-depth functional analysis of the risk locus proximal to the PHLDB1 gene on 11q23.3. We retrieved all SNPs in linkage disequilibrium (r(2) ≥ 0.2) with the glioma-associated SNP (rs498872) and performed a comprehensive bioinformatics and experimental functional analysis for the region. After testing candidate SNPs for allele-specific activity in a luciferase-based enhancer scanning assay, we established a subset of 10 functional SNPs in the promoters of PHLDB1 and DDX6, and in a putative enhancer element. Chromatin conformation capture (3C) identified a physical interaction between the enhancer element containing a functional SNP (rs73001406) and the promoter of the DDX6 gene. Knockdown experiments in cell culture and 3D assays to evaluate the role of PHLDB1 and DDX6 suggest that both genes may contribute to the phenotype. These studies reveal the functional landscape of the 11q23.3 glioma susceptibility locus and identify a network of functional SNPs in regulatory elements and two target genes as a possible mechanism driving glioma risk association.


Assuntos
Neoplasias Encefálicas/genética , RNA Helicases DEAD-box/genética , Predisposição Genética para Doença , Glioma/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas do Tecido Nervoso/genética , Proteínas Proto-Oncogênicas/genética , Alelos , Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Cromatina/química , Cromatina/metabolismo , Cromossomos Humanos Par 11 , Biologia Computacional , RNA Helicases DEAD-box/antagonistas & inibidores , RNA Helicases DEAD-box/metabolismo , Elementos Facilitadores Genéticos , Expressão Gênica , Técnicas de Silenciamento de Genes , Loci Gênicos , Estudo de Associação Genômica Ampla , Glioma/diagnóstico , Glioma/metabolismo , Glioma/patologia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/metabolismo , Fenótipo , Polimorfismo de Nucleotídeo Único , Prognóstico , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteínas Proto-Oncogênicas/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Risco
6.
Eur J Hum Genet ; 23(1): 132-4, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24755950

RESUMO

Genome-wide association studies have recently identified a cancer susceptibility locus at 10p12 mapping to MLLT10 associated with the onset of diverse tumors. We genotyped two tightly linked single-nucleotide polymorphisms (SNPs) at MLLT10 associated with meningioma (rs12770228) or ovarian cancer (rs1243180), and tested for associations among 295 meningioma cases, 606 glioma cases and 646 noncancer controls, all of European descent. The variant 'A' allele in MLLT10 rs12770228 was associated with an increased risk of meningioma (per allele odds ratio: 1.25; 95% confidence interval: 1.02, 1.53; P=0.031). Similar associations were observed for rs1243180. MLLT10 variants were unrelated to glioma. Functional investigation identified 22 candidate functional SNPs mapping to this region. The present study further validates 10p12 as a meningioma risk locus.


Assuntos
Neoplasias Encefálicas/genética , Loci Gênicos , Variação Genética , Células Germinativas/metabolismo , Fatores de Transcrição/genética , Adulto , Idoso , Alelos , Neoplasias Encefálicas/epidemiologia , Feminino , Estudos de Associação Genética , Predisposição Genética para Doença , Genótipo , Humanos , Desequilíbrio de Ligação , Masculino , Pessoa de Meia-Idade , Razão de Chances , Polimorfismo de Nucleotídeo Único , Risco
7.
PLoS One ; 9(8): e105568, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25162558

RESUMO

Glioblastoma multiforme (GBM) is the most common and the most aggressive form of primary brain tumor. Jak2 is a non-receptor tyrosine kinase that is involved in proliferative signaling through its association with various cell surface receptors. Hyperactive Jak2 signaling has been implicated in numerous hematological disorders as well as in various solid tumors including GBM. Our lab has developed a Jak2 small molecule inhibitor known as G6. It exhibits potent efficacy in vitro and in several in vivo models of Jak2-mediated hematological disease. Here, we hypothesized that G6 would inhibit the pathogenic growth of GBM cells expressing hyperactive Jak2. To test this, we screened several GBM cell lines and found that T98G cells express readily detectable levels of active Jak2. We found that G6 treatment of these cells reduced the phosphorylation of Jak2 and STAT3, in a dose-dependent manner. In addition, G6 treatment reduced the migratory potential, invasive potential, clonogenic growth potential, and overall viability of these cells. The effect of G6 was due to its direct suppression of Jak2 function and not via off-target kinases, as these effects were recapitulated in T98G cells that received Jak2 specific shRNA. G6 also significantly increased the levels of caspase-dependent apoptosis in T98G cells, when compared to cells that were treated with vehicle control. Lastly, when T98G cells were injected into nude mice, G6 treatment significantly reduced tumor volume and this was concomitant with significantly decreased levels of phospho-Jak2 and phospho-STAT3 within the tumors themselves. Furthermore, tumors harvested from mice that received G6 had significantly less vimentin protein levels when compared to tumors from mice that received vehicle control solution. Overall, these combined in vitro and in vivo results indicate that G6 may be a viable therapeutic option against GBM exhibiting hyperactivation of Jak2.


Assuntos
Antineoplásicos/farmacologia , Neoplasias Encefálicas/tratamento farmacológico , Regulação Neoplásica da Expressão Gênica , Glioblastoma/tratamento farmacológico , Janus Quinase 2/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Animais , Antineoplásicos/síntese química , Apoptose , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Caspases/genética , Caspases/metabolismo , Linhagem Celular Tumoral , Feminino , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Camundongos , Camundongos Nus , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/síntese química , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais , Bibliotecas de Moléculas Pequenas/síntese química , Carga Tumoral/efeitos dos fármacos , Vimentina/antagonistas & inibidores , Vimentina/genética , Vimentina/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Am J Pathol ; 181(3): 858-65, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22796437

RESUMO

Philadelphia chromosome-negative myeloproliferative neoplasms, including polycythemia vera, essential thrombocytosis, and myelofibrosis, are disorders characterized by abnormal hematopoiesis. Among these myeloproliferative neoplasms, myelofibrosis has the most unfavorable prognosis. Furthermore, currently available therapies for myelofibrosis have little to no efficacy in the bone marrow and hence, are palliative. We recently developed a Janus kinase 2 (Jak2) small molecule inhibitor called G6 and found that it exhibits marked efficacy in a xenograft model of Jak2-V617F-mediated hyperplasia and a transgenic mouse model of Jak2-V617F-mediated polycythemia vera/essential thrombocytosis. However, its efficacy in Jak2-mediated myelofibrosis has not previously been examined. Here, we hypothesized that G6 would be efficacious in Jak2-V617F-mediated myelofibrosis. To test this, mice expressing the human Jak2-V617F cDNA under the control of the vav promoter were administered G6 or vehicle control solution, and efficacy was determined by measuring parameters within the peripheral blood, liver, spleen, and bone marrow. We found that G6 significantly reduced extramedullary hematopoiesis in the liver and splenomegaly. In the bone marrow, G6 significantly reduced pathogenic Jak/STAT signaling by 53%, megakaryocytic hyperplasia by 70%, and the Jak2 mutant burden by 68%. Furthermore, G6 significantly improved the myeloid to erythroid ratio and significantly reversed the myelofibrosis. Collectively, these results indicate that G6 is efficacious in Jak2-V617F-mediated myelofibrosis, and given its bone marrow efficacy, it may alter the natural history of this disease.


Assuntos
Janus Quinase 2/metabolismo , Mielofibrose Primária/tratamento farmacológico , Mielofibrose Primária/enzimologia , Inibidores de Proteínas Quinases/uso terapêutico , Bibliotecas de Moléculas Pequenas/uso terapêutico , Estilbenos/uso terapêutico , Substituição de Aminoácidos/genética , Animais , Medula Óssea/efeitos dos fármacos , Medula Óssea/patologia , Modelos Animais de Doenças , Hematopoese Extramedular/efeitos dos fármacos , Humanos , Hiperplasia , Janus Quinase 2/antagonistas & inibidores , Megacariócitos/efeitos dos fármacos , Megacariócitos/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Células Mieloides/efeitos dos fármacos , Células Mieloides/patologia , Fosforilação/efeitos dos fármacos , Mielofibrose Primária/sangue , Mielofibrose Primária/fisiopatologia , Inibidores de Proteínas Quinases/farmacologia , Reticulina/efeitos dos fármacos , Reticulina/metabolismo , Fator de Transcrição STAT5/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Baço/efeitos dos fármacos , Baço/patologia , Baço/fisiopatologia , Esplenomegalia/complicações , Esplenomegalia/tratamento farmacológico , Esplenomegalia/patologia , Esplenomegalia/fisiopatologia , Estilbenos/farmacologia
9.
Bioorg Med Chem Lett ; 22(3): 1402-7, 2012 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-22227213

RESUMO

In this study, we analyzed the structure-activity relationship properties of the small molecule Jak2 inhibitor G6. We synthesized a set of derivatives containing the native para-hydroxyl structure or an alternative meta-hydroxyl structure and examined their Jak2 inhibitory properties. We found that the para-hydroxyl derivative known as NB15 had excellent Jak2 inhibitory properties in silico, in vitro, and ex vivo when compared with meta-hydroxyl derivatives. These results indicate that NB15 is a potent derivative of the Jak2 inhibitor G6, and that maintaining the para-hydroxyl orientation of G6 is critical for its Jak2 inhibitory potential.


Assuntos
Benzilaminas/química , Benzilaminas/farmacologia , Janus Quinase 2/antagonistas & inibidores , Modelos Moleculares , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Hidroxilação , Camundongos , Camundongos Transgênicos , Estrutura Molecular , Estilbenos/química , Estilbenos/farmacologia , Relação Estrutura-Atividade
10.
Cell Signal ; 24(2): 435-42, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21963429

RESUMO

The serum- and glucocorticoid-inducible kinase 1 (SGK1) is known to regulate a wide variety of cellular processes, including renal sodium retention and cell survival. Angiotensin II (Ang II) is one of the many signaling molecules capable of regulating SGK1 expression, and is also known to impact cell survival. Here, we examined the role of SGK1 in Ang II-mediated cell survival. We hypothesized that Ang II protects cells from apoptosis by upregulating and activating SGK1. To test this, we examined the effects of Ang II stimulation on SGK1 expression and downstream signaling. We also examined the effects of Ang II treatment and siRNA-mediated SGK1 knockdown on apoptosis after serum starvation. We found that after 2h of Ang II treatment, SGK1 mRNA expression was increased approximately 2-fold. This induction was sensitive to reductions in intracellular calcium levels after pretreatment with BAPTA-AM, but insensitive to the L-type calcium channel blocker verapamil. SGK1 induction was also sensitive to the tyrosine kinase inhibitor genistein. Ang II treatment also caused a rapid increase in the level of phosphorylation of SGK1 at Ser422 and Thr256, and Ser422 phosphorylation was rapamycin-sensitive. We found that Ang II treatment was protective against serum starvation-induced apoptosis, and this protective effect was significantly blunted when SGK1 was silenced via siRNA. Lastly, Ang II induced FOXO3A phosphorylation in an SGK1-dependent manner, thereby reducing the pro-apoptotic actions of FOXO3A. Overall, these results indicate that Ang II upregulates and activates SGK1, leading to increased cell survival via multiple, non-redundant mechanisms.


Assuntos
Angiotensina II/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Fibrossarcoma/enzimologia , Proteínas Imediatamente Precoces/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Cálcio , Bloqueadores dos Canais de Cálcio/metabolismo , Canais de Cálcio Tipo L/metabolismo , Linhagem Celular Tumoral , Ácido Egtázico/análogos & derivados , Ácido Egtázico/metabolismo , Fibrossarcoma/genética , Fibrossarcoma/patologia , Proteína Forkhead Box O3 , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Genisteína/farmacologia , Humanos , Proteínas Imediatamente Precoces/antagonistas & inibidores , Proteínas Imediatamente Precoces/genética , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , RNA Mensageiro/análise , RNA Mensageiro/biossíntese , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/efeitos dos fármacos , Verapamil/farmacologia
11.
Exp Hematol ; 40(1): 22-34, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22019628

RESUMO

Hyperkinetic Jak2 tyrosine kinase signaling has been implicated in several hematological disorders, including myeloproliferative neoplasms. Effective Jak2 inhibitors can have significant therapeutic potential. Here, using structure-based virtual screening, we identified a benzothiophene-derived Jak2 inhibitor named A46. We hypothesized that this compound would inhibit Jak2-V617F-mediated pathologic cell growth. To test this, A46 was analyzed for its ability to inhibit recombinant Jak2 protein catalysis; suppress Jak2-mediated pathogenic cell growth in vitro; inhibit the aberrant ex vivo growth of Jak2-V617F-expressing primary human bone marrow cells; and inhibit Jak2-mediated pathogenesis in vivo. To this end, we found that A46 selectively inhibited Jak2-V617F protein when compared to wild-type Jak2 protein. The drug also selectively inhibited the proliferation of Jak2-V617F-expressing cells in both a time- and dose-dependent manner, and this correlated with decreased Jak2 and signal transducers and activators of transcription 5 phosphorylation within treated cells. The Jak2-V617F cell growth inhibition correlated with an induction of cell cycle arrest and promotion of apoptosis. A46 also inhibited the pathologic growth of primary Jak2-V617F-expressing bone marrow cells ex vivo. Lastly, using a mouse model of Jak2-V617F-mediated myeloproliferative neoplasia. A46 significantly reduced the splenomegaly and megakaryocytic hyperplasia in the spleens of treated mice and the levels of interleukin-6 in the plasma. Collectively, our data demonstrate that the benzothiophene-based compound, A46, suppresses Jak2-mediated pathogenesis, thereby making it a potential candidate drug against Jak2-mediated disorders.


Assuntos
Células da Medula Óssea/citologia , Células da Medula Óssea/efeitos dos fármacos , Janus Quinase 2/antagonistas & inibidores , Tiofenos/farmacologia , Animais , Apoptose/efeitos dos fármacos , Biocatálise , Células da Medula Óssea/patologia , Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Janus Quinase 2/metabolismo , Camundongos , Camundongos Transgênicos , Relação Estrutura-Atividade , Tiofenos/química , Células Tumorais Cultivadas
12.
J Biol Chem ; 285(41): 31399-407, 2010 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-20667821

RESUMO

Somatic mutations in the Jak2 protein, such as V617F, cause aberrant Jak/STAT signaling and can lead to the development of myeloproliferative neoplasms. This discovery has led to the search for small molecule inhibitors that target Jak2. Using structure-based virtual screening, our group recently identified a novel small molecule inhibitor of Jak2 named G6. Here, we identified a structure-function correlation of this compound. Specifically, five derivative compounds of G6 having structural similarity to the original lead compound were obtained and analyzed for their ability to (i) inhibit Jak2-V617F-mediated cell growth, (ii) inhibit the levels of phospho-Jak2, phospho-STAT3, and phospho-STAT5; (iii) induce apoptosis in human erythroleukemia cells; and (iv) suppress pathologic cell growth of Jak2-V617F-expressing human bone marrow cells ex vivo. Additionally, we computationally examined the interactions of these compounds with the ATP-binding pocket of the Jak2 kinase domain. We found that the stilbenoid core-containing derivatives of G6 significantly inhibited Jak2-V617F-mediated cell proliferation in a time- and dose-dependent manner. They also inhibited phosphorylation of Jak2, STAT3, and STAT5 proteins within cells, resulting in higher levels of apoptosis via the intrinsic apoptotic pathway. Finally, the stilbenoid derivatives inhibited the pathologic growth of Jak2-V617F-expressing human bone marrow cells ex vivo. Collectively, our data demonstrate that G6 has a stilbenoid core that is indispensable for maintaining its Jak2 inhibitory potential.


Assuntos
Janus Quinase 2/antagonistas & inibidores , Policitemia Vera/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Estilbenos/farmacologia , Substituição de Aminoácidos , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Mutação de Sentido Incorreto , Policitemia Vera/enzimologia , Policitemia Vera/genética , Inibidores de Proteínas Quinases/química , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/metabolismo , Estilbenos/química , Relação Estrutura-Atividade
13.
Colloids Surf B Biointerfaces ; 62(2): 238-42, 2008 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-18035524

RESUMO

The aggregation behavior of the bile salts taurodeoxycholate (NaTDC) and sodium cholate (NaC), are followed at concentrations below critical micelle concentrations (CMCs) using the environment sensitive, fluorescent-labeled phospholipid, 2-(6-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)hexanoyl-1-hexadecanoyl-sn-glycero-3-phosphocholine (NBD-C(6)-HPC). A buffer solution containing NBD-C(6)-HPC is titrated with increasing NaC or NaTDC and the fluorescence changes followed. Both bile salts induced fluorescence changes below their critical micelle concentration indicating the presence of a bile salt-phospholipid aggregate. A critical control experiment using 6-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino) hexanoic acid (NBD-X) shows that the bile salts are interacting with the longer, C16 hydrocarbon tail, not the NBD probe. The fluorescence curves were fitted to the Hill equation as a model for cooperative aggregation. The cooperativity model provides a minimum estimate for the number of bile salts to give maximal fluorescence. This number was calculated for NaC and NaTDC to have a minimum value of approximately 2. A small aggregation number supports the existence of primary micellar aggregates at submicellar concentrations for bile salt-phospholipid aqueous solutions.


Assuntos
Ácidos e Sais Biliares/química , Fosfolipídeos/química , Algoritmos , Fluorescência , Corantes Fluorescentes , Luz , Micelas , Espalhamento de Radiação , Colato de Sódio/química , Ácido Taurodesoxicólico/química
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